Gene-editing should be abolished (due 40 hours)

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Should gene-editing be abolished?

Analury Sanchez

Professor Ocxanne Jean, Ph.D.

Advance Writing and Research-DL-B

Apr 02, 2022

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Should gene-editing be abolished?

I. Introduction: In the recent years, humans have witnessed technological developments

whereby tomatoes ripen slowly, cattle without horns, and even mosquitos that cannot transmit

malaria. This has been necessitated by gene editing. According to Ayanoğlu, Elçin & Elçin (2020),

gene editing is a technology that provides scientists with an opportunity of making changes to the

DNA of an organism. Thanks to gene editing, it is possible to edit a particular disease out of an

individual. The increase in ethical controversy of gene editing can be attributed to its potential of

asserting some significant control over the kind of future for humans. This topic emphasizes why

gene editing should be abolished due to the unprecedented health implications of genetically

modified humans.

II. Background: According to Abuhammad, Khabour and Alzoubi (2021), genetic

modifications can lead to the creation of super-humans and “designer babies” while also

perpetrating fundamental alteration of the human species. As a matter of fact, genomic research

may potentially be weaponized towards targeting as well as harming particular population groups.

The moral, ethical, and legal boundaries of utilizing genetic technologies are largely unclear, which

creates opportunities for their abuse and misuse. On the other hand, Howard et al. (2018) argued

that gene-editing technologies are associated with diverse ethical concerns, particularly when the

process is utilized towards addressing a given genetic diagnosis of an unborn child due to the

potential evolution of off-target edits.

Gene editing can result in unprecedented and unwanted heritable genetic alterations that

may contribute to long-term risks in clinical space (Conboy, 2018). Accessing gene therapies for

combating diseases, for instance, could be limited to those who can afford them, which increases

health inequality outcomes across and within countries. Ethically, there are safety concerns to the

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side effects of the technology, including lack of informed consent for germline as the affected

clients by the edits are not yet born (Holm, 2019). Many countries, particularly the developing and

underdeveloped nations, may fail to afford the technology, which increases the inequality gap in

society. Should gene-editing be abolished? Gene editing should be abolished due to the potential

impact on society and serious ethical concerns associated with the technology.

III. Arguments: Medicine has recently reached a turning point with major changes highly

likely to be experienced, particularly with the growth of disruptive technologies like cell therapies,

RNA, and gene enabling scientists to approach diseases in ways that have never been witnessed

before. From a scientific perspective, medical researchers are keen on establishing the risks and

opportunities of gene editing. In this regard, critics of gene editing argue that the technology could

be associated with unpredictable implications on the environment and human health, especially

fears of creating “designer humans.”

a. Reason 1: Altering the genes of a child before birth implies that such alterations are

passed on to future generations, meaning that the DNA of the child’s body is permanently and

irreversibly changed.

i. Evidence 1a: There is a need to preserve the human right to an open future and

bodily integrity. According to Davies (2019), there is a high potential for errors being

experienced in the process of gene editing.

ii. Evidence 1b: Gene editing could have errors associated with devastating effects

like accidentally deleting a gene, thus leading to developmental defects in the unborn child

(Davies, 2019).

iii. Evidence 1c: There is a possibility that germline editing and adverse effects

may be passed on from one generation to another (Davies, 2019).

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b. Reason 2: Gene editing is bound to reinforce inequalities in society as the commercial

and social dynamics whereby modifying the human germline may exacerbate global disparities and

take structural inequality to greater heights.

i. Evidence 2a: When humans are presented with an opportunity of accessing the

technology, there could be serious challenges in that attempt to control what it is used for,

thus creating a slippery slope. In this regard, parents-to-be could utilize the technologies in

what may be termed as racist or sexist (Khan, 2019).

ii. Evidence 2b: If parents are given an opportunity of choosing the sex of their

baby, it could lead to sexism.

iii. Evidence 2c: The ability to choose the physical characteristics of a child so that

s/he is more attractive could lead to racism (Khan, 2019).

c. Reason 3: Gene editing entails a change of cellular structure.

i. Evidence 3a: A slight change of cells can result in new creatures that can

threaten societal existence (Abuhammad et.al, 2021).

ii. Evidence 3b: A small error in gene editing is likely to lead to an undesired

outcome. Some of the experiments are meant to create diseases resistant human beings

(Conboy, 2018).

iii Evidence 3c: There is some likelihood of creating some deadlier diseases in the

process (Conboy, 2018). Therefore, gene editing is a dangerous experiment.

III. Refuting Opponents’ Arguments

a. Opposing view 1: A. Those supporting gene editing have argued that technology is

instrumental in dealing with the most severe and deadly diseases.

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i. Evidence 1a. Diverse genetic mutations affecting millions of people globally

could end if humans are actively involved in genetically engineering the next generation

(Conboy, 2018).

ii. Evidence 1b. Genetic modification in mice has been shown to have

unanticipated long-term adverse effects (Conboy, 2018).

iii. Evidence 1c. CRISPR Gene Editing has been shown to increase the risk of

developing cancer cells and affect healthy cells faster (Conboy, 2018).

b. Opposing view 2: Gene editing can extend the human lifespan as diseases and illnesses

that shorten the lifespan of many people are eliminated.

i. Evidence 2a: To this end, genetic editing can reverse the most fundamental

reasons for the natural decline of the human body on a cellular level (Holms, 2019).

ii. Evidence 2b: Drastically improving both the quality of life and span (Holms,

2019).

IV. Conclusion: Gene editing is a technology that should not be embraced anywhere due

to the increasing uncertainty of the side effects and implications on future generations. There is a

need for more research on the topic towards establishing the potential benefits, opportunities, and

risks associated with the technology for it to be advanced. This topic is important because changing

the genetic inheritance of the human species may potentially provoke a backlash, which implies

that people need to condemn pernicious genetic technologies while encouraging those that can

benefit the human species.

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References

Abuhammad, S., Khabour, O. F., & Alzoubi, K. H. (2021). Researchers views about perceived

harms and benefits of gene editing: A study from the MENA region. Heliyon, 7(4), e06860.

https://doi.org/10.1016/j.heliyon.2021.e06860

Ayanoğlu, F. B., Elçin, A. E., & Elçin, Y. M. (2020). Bioethical issues in genome editing by

CRISPR-Cas9 technology. Turkish Journal of Biology, 44(2), 110-120.

https://doi.org/10.3906/biy-1912-52

Conboy, I. (2018). Faculty opinions recommendation of CRISPR-Cas9 genome editing induces a

p53-mediated DNA damage response. Faculty Opinions – Post-Publication Peer Review of

the Biomedical Literature. https://doi.org/10.3410/f.733427168.793553934

Davies, B. (2019). The technical risks of human gene editing. Human Reproduction, 34(11), 2104-

2111. https://doi.org/10.1093/humrep/dez162

Holm, S. (2019). Let us assume that gene editing is safe—the role of safety arguments in the gene-

editing debate. Cambridge Quarterly of Healthcare Ethics, 28(1), 100-111.

https://doi.org/10.1017/S0963180118000439

Howard, H. C., van El, C. G., Forzano, F., Radojkovic, D., Rial-Sebbag, E., de Wert, G., … &

Cornel, M. C. (2018). One small edit for humans, one giant edit for humankind? Points and

questions to consider for a responsible way forward for gene editing in humans. European

Journal of Human Genetics, 26(1), 1-11. https://doi.org/10.1038/s41431-017-0024-z

Khan, S. H. (2019). Genome-editing technologies: concept, pros, and cons of various genome-

editing techniques and bioethical concerns for clinical application. Molecular Therapy-

Nucleic Acids, 16, 326-334. https://doi.org/10.1016/j.omtn.2019.02.027

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http://journals.tubitak.gov.tr/biology/

Turkish Journal of Biology Turk J Biol
(2020) 44: 110-120
© TÜBİTAK
doi:10.3906/biy-1912-52

Bioethical issues in genome editing by CRISPR-Cas9 technology

Fatma Betül AYANOĞLU1, Ayşe Eser ELÇİN1, Yaşar Murat ELÇİN1,2,*
1Tissue Engineering, Biomaterials and Nanobiotechnology Laboratory, Ankara University Faculty of Science,

Ankara University Biotechnology Institute, Ankara University Stem Cell Institute, Ankara, Turkey
2Biovalda Health Technologies, Inc., Ankara, Turkey

* Correspondence: [email protected]

1. Introduction
For many years, molecular biologists have sought ways
to use cellular repair mechanisms to manipulate DNA
through genome editing. In this way, they would have the
power to change the genome by correcting a mutation or
introducing a new function (Rodriguez, 2016). For this
purpose, genome editing technologies were developed
(Memi et al., 2018). In recent years, clustered regularly
interspaced short palindromic repeats technology
(CRISPR-Cas9) has become the most preferred method of
gene editing. This technology has advantages such as high
accuracy, easy handling, and relatively low cost compared to
previous technologies, such as zinc-finger nuclease (ZFN)
and transcription activator-like effector nuclease (TALEN).
Thanks to these benefits, CRISPR-Cas9 technology can be
easily applied in any molecular biology laboratory.

Genome editing technologies are used in the formation
of human disease models in experimental animals and for
the understanding of basic gene functions. They also have
great therapeutic potential for future treatment of untreated
diseases such as certain cancers, genetic disorders, and

HIV/AIDS. Today, genome editing in somatic cells is one
of the promising areas of therapeutic development (Otieno,
2015). However, various bioethical issues have arisen due
to the potential impact of these technologies on the safety
of food stocks and clinical applications (Hundleby and
Harwood, 2018; Hirch et al., 2019). This review discusses
the challenges, possible consequences, and bioethical
issues of CRISPR-Cas9 in detail.

2. Biology and function of CRISPR-Cas9 technology
Genome editing technologies often work by creating
fractures in chromosomal DNA. ZFN, TALEN, and
CRISPR-Cas9 are all based solely on nucleases (Kim
and Kim, 2014; Roh et al., 2018). The strength of these
technologies stems from the ability to create fractures in the
desired region of a specific target sequence as determined
by the researcher. This allows researchers to modify the
genome in practice in any region (Memi et al., 2018).

The creation of changes in the genome depends mainly
on the DNA repair capacity of the cells (Lau et al., 2018).
All cells have two basic mechanisms for the repair of double

Abstract: Genome editing technologies have led to fundamental changes in genetic science. Among them, CRISPR-Cas9 technology
particularly stands out due to its advantages such as easy handling, high accuracy, and low cost. It has made a quick introduction in
fields related to humans, animals, and the environment, while raising difficult questions, applications, concerns, and bioethical issues
to be discussed. Most concerns stem from the use of CRISPR-Cas9 to genetically alter human germline cells and embryos (called
germline genome editing). Germline genome editing leads to serial bioethical issues, such as the occurrence of undesirable changes
in the genome, from whom and how informed consent is obtained, and the breeding of the human species (eugenics). However, the
bioethical issues that CRISPR-Cas9 technology could cause in the environment, agriculture and livestock should also not be forgotten.
In order for CRISPR-Cas9 to be used safely in all areas and to solve potential issues, worldwide legislation should be prepared, taking
into account the opinions of both life and social scientists, policy makers, and all other stakeholders of the sectors, and CRISPR-Cas9
applications should be implemented according to such legislations. However, these controls should not restrict scientific freedom. Here,
various applications of CRISPR-Cas9 technology, especially in medicine and agriculture, are described and ethical issues related to
genome editing using CRISPR-Cas9 technology are discussed. The social and bioethical concerns in relation to human beings, other
organisms, and the environment are addressed.

Key words: Genome editing, CRISPR-Cas9 technology, bioethical issues, bioethics

Received: 15.12.2019 Accepted/Published Online: 26.02.2020 Final Version: 02.04.2020

Review Article

This work is licensed under a Creative Commons Attribution 4.0 International License.

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chain breaks on DNA. One of them is nonhomologous
end joining (NHEJ) and the other is the homologous
dependent repair (HDR) mechanism. In NHEJ, the ends of
the fractures are quickly connected directly to each other,
regardless of the sequence homology, while HDR requires
homology to repair the damaged DNA site. In order to
achieve homology, the undamaged sister chromatid is
used as a template and DNA damage is repaired (Urnov,
2018).

CRISPR-Cas9 is a naturally occurring defense system in
prokaryotic organisms that provides resistance to foreign
genetic elements such as plasmids and bacteriophages
(Barrangau et al., 2007; Marraffini and Sontheimer, 2008).
When the virus or plasmid enters a bacterial cell, CRISPR-
Cas9 allows the addition of short viral DNA molecules
to the CRISPR site. CRISPR sequences (CRISPRs) are
short DNA repeats of viral or plasmid origin found in the
genomes of bacteria and are defined as clustered regularly
interspaced short palindromic repeats. Cas genes (CRISPR-
related) are genes that encode nuclease or helicase proteins
associated with CRISPR repeat sequences that have the
function of cutting or dissolving DNA (Jansen et al., 2002).
Cas9, a member of the Cas gene family, was isolated from
Streptococcus pyogenes and is an endonuclease capable of
cutting DNA from two active cut regions at both ends of
the DNA double helix (Doudna and Charpentier, 2014;
Rodriguez, 2016). The CRISPR-Cas system recognizes the
DNA of the invading virus or bacterium and directs the
Cas protein to destroy foreign DNA (Otieno, 2015).

In the following years, it was discovered that the
CRISPR-Cas system can be programmed to find and cut
specific target DNA regions, thereby providing genome
editing (Jinek et al., 2012; Hsu et al., 2013). As a result of
understanding that the human genome can be edited by
CRISPR-Cas9, it became clear that genome editing could
also be used for therapeutic purposes, and a new era in
genetic engineering began (Lau et al., 2018; Roh et al.,
2018).

3. Application areas
3.1. Animal models
CRISPR-Cas9 can be used to create animal models
to mimic human diseases and to understand disease
development by mutating or silencing genes. A mouse
model has been developed to determine the harmful
effects of mutations in cancer by making mutations that
cause the loss of function in tumor suppressor genes or
give functions to protooncogenes (Chin, 2015).

Conventional genetically modified (GM) mouse
models are produced by gene targeting in embryonic
stem cells or transgenesis, which are time-consuming
and highly expensive. With CRISPR-Cas9, GM mice can
be efficiently produced in a much shorter time (Mei et

al., 2016). It can be applied to nonhuman primates such
as monkeys. Nonhuman primates are more similar to
humans in anatomical, physiological, and genetic terms
than rodents (Zhang et al., 2014). Therefore, they are more
suitable models than rodents in understanding human
biology and disease development (Xin et al., 2016). The
first successful application of CRISPR-Cas9 in nonhuman
primates, from which a knockout monkey was produced,
was realized in 2014 (Niu et al., 2014). However, genome
sequences of many nonhuman primates are not yet fully
identified. This makes it difficult to design selected single-
guide RNAs (sgRNAs) (Gou and Li, 2015; Lou et al., 2016).
Therefore, the application of CRISPR-Cas9 in nonhuman
primates is still at an early stage.
3.2. Genome editing in specific tissues
Researchers have been able to modify the genomes of
specific tissues such as liver and brain tissues using
hydrodynamic injection and adeno-associated virus
(AAV) (Rodriguez et al., 2014; Senis et al., 2014). In a
study, CRISPR-Cas9 has been successfully and effectively
applied to the mammalian nervous system. A mixture
of green fluorescent protein (GFP)-labeled AAV-spCas9
and AAV-spGuide plasmids was transferred in vivo to
the hippocampal toothed brain folds of adult male mice
(Swiech et al., 2015). It is thought that the number of
such applications will grow in the fields of cancer and
neuroscience in the following years (Mei et al., 2016).
3.3. Multiple gene mutations
CRISPR-Cas9 can be used to generate mutants for target
genes. In the first such study by Li et al., six sgRNAs
targeting Cas9 mRNA and six different genomic regions
encoding the Tet1, Tet2, and Tet3 genes were transferred
to the cytoplasm of rat embryos (Li et al., 2013). Findings
showed that all three Tet genes carried the desired
mutations in 59% of newborn rat pups. Successful results
were also reported in studies with zebrafish embryos and
Arabidopsis as well (Ota et al., 2014; Wang et al., 2015).
3.4. Epigenome studies
Epigenome studies can be performed in two different ways:
genome or epigenome editing (Chen et al., 2014; Huisman
et al., 2015). In genome editing, nuclease is used to modify
the DNA sequence, whereas in epigenome editing, an
effector domain is used and the DNA sequence is not
changed. This function is achieved by catalytic inactivation
of the Cas9-associated effector domain by replacing
the Cas9 protein. Altered effector proteins are used to
activate or suppress transcription (Lau and Davie, 2016).
In epigenome editing, the epigenome can be modified
by changing the proteins that maintain and protect the
epigenome. Suppression of DNA methylation as a result
of degradation of catalytic domains that accelerate the loss
of spherical DNA methylation in human cells and lead to

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cell death is a good example of epigenome studies using
CRISPR-Cas9 (Liao et al., 2015).

Another application is the editing of long nonencoded
RNAs (lncRNAs) and enhancer RNAs (eRNAs) that can
control gene expression and epigenome processes. In a
study, eRNA-expression factors and IncRNA-expression
enhancers were suppressed by stimulating deletion
mutations in a lymphoma cell line using CRISPR-Cas9
(Pefanis et al., 2015). It is predicted that CRISPR-Cas9
may allow different levels of epigenome modification and
facilitate further changes to humans (Liao et al., 2015; Mei
et al., 2016).
3.5. Treatment of diseases
CRISPR-Cas9 can be applied to cells in vivo or ex vivo. In
the in vivo approach, CRISPR-Cas9 is directly transferred
to cells in the body using either viral or nonviral methods.
In the ex vivo approach, first the cells are removed from
the body; then CRISPR is applied to the cells and they
are transferred back to the body (Roh et al., 2018). This
approach has great potential to develop tissue-based
therapies (Rath et al., 2015). Using CRISPR-Cas9, the
mutation in the dystrophin protein responsible for the
most common form of Duchenne muscular dystrophy
was successfully removed (Amoasii et al., 2018; Duchêne
et al., 2018; Koo et al., 2018; Long et al., 2018). There are
studies to prevent and treat AIDS by inhibiting the entry
of HIV into the cell or by removing the HIV genome
integrated into the host genome using CRISPR-Cas9
(Saayman et al., 2015). Induced pluripotent stem cells
(iPSCs) were successfully produced from cystic fibrosis
patients with confirmed F508 deletion in the cystic fibrosis
transmembrane regulator (CFTR) gene by CRISPR-Cas9
(Firth et al., 2015). There are also studies for cataracts
(Wu et al., 2015; Yang et al., 2016) and Parkinson’s disease
(Yang et al., 2016). However, recent studies have shown
that CRISPR-Cas9 activates the type 1 interferon (INF)
pathway, causing a type 1 INF-mediated immune response
(Kim et al., 2018; Charleswort et al., 2019). These findings
currently limit the use of CRISPR-Cas9 in treatment.
3.6. Industrial uses
CRISPR was first used for commercial purposes to make
bacterial cultures used in cheese and yogurt production
resistant to viral infections (van Erp et al., 2015). One of
the applications in agriculture is to produce GM crops
(Hundleby and Harwood, 2019). There are attempts to
increase the yield in the livestock industry (van Erp et
al., 2015). It can be used to control invasive pest species
to reverse pesticide and herbicide resistance in insects
and weeds or to prevent disease spread (Esvelt et al.,
2014). Researchers have succeeded in preventing the
spread of genes protecting mosquitoes from harmful
malaria parasites (Gantz et al., 2015) and making female
mosquitoes infertile in the laboratory (Hammond et al.,

2016). Vaccine development is another significant area
of interest. The smallpox virus vector (VACV) is used
in the eradication and vaccination of smallpox. Using
CRISPR-Cas9, the efficiency of marker-free VACV vectors
has been increased (Yuan et al., 2015). Another example
is the hepatitis B vaccine. In order to prevent viral gene
expression and replication, specific regions of the hepatitis
B genome were targeted and cut by CRISPR-Cas9
(Ramanan et al., 2015).
3.7. RNA editing
Single-stranded RNA (ssRNA) sequences can also be
edited by CRISPR-Cas9. In RNA editing, CRISPR-Cas9
consists of a DNA oligonucleotide presenting the PAM
(protospacer adjacent motif ) region (PAMmer), ssRNA,
guide RNA (gRNA), and Cas9 protein. PAMmer acts as
a PAM region specifically recognized by Cas9 and directs
Cas9 to bind and cut the target ssRNA. 5’-Elongated
PAMmers containing bases paired with different ssRNAs
and immediately in front of PAM are required for specific
binding of target ssRNAs. Since RNA molecules have
different functions than DNA, CRISPR-Cas9 can offer a
much more flexible application than other genome editing
methods (Mei et al., 2016).
3.8. Military applications
One of the lesser-discussed application areas of CRISPR-
Cas9 technology is its use for military purposes. As is
known, a substantial portion of genome editing studies
are supported by the defense ministries of the countries.
These studies are commonly focused on increasing
the tolerance of soldiers against biological or chemical
warfare. This technology has the potential to influence
human performance optimization (Greene and Master,
2018). Studies are usually concentrated on discovering
different genes that can be harnessed from other species
(Gracheva et al., 2010) and identifying new genes that can
be associated with posttraumatic stress disorder, which is
frequently experienced by soldiers (Cornelis et al., 2010).
In a study by Zou et al. (2015), researchers developed dog
embryos with higher muscle mass using CRISPR-Cas9.
Another interesting study showed that the CMG2 gene,
known to cause low sensitivity to anthrax toxin when
expressed in small amounts, could be silenced by this
technology (Arévalo et al., 2014). However, it should be
noted that far more research needs to be conducted for
using CRISPR technology in humans as a defense tool
against biological and chemical weapons (Greene and
Master, 2018).
3.9. DNA replacement in human embryos (germline
genome therapy)
The most controversial usage of CRISPR-Cas9 is the
modification of human embryo DNA, or, in other words,
its use for germline genome therapy. In 2015, a group of

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Chinese researchers led by Junjiu Huang applied CRISPR-
Cas9 to remove a mutation that causes β-thalassemia,
which is a fatal blood disease, from the human β-globulin
(HBB) gene in the germline of human embryos. In this
research, six abnormal embryos not suitable for in vitro
fertilization were used. The mutation could be corrected
in only one of the embryos. Although the mutation could
be corrected in two other embryos, nontarget effects
occurred in other genes. In the other three embryos, the
mutation could not be corrected. It has been reported
that this technique is not ready for clinical use because
of nontarget effects on different genes (Roh et al., 2018;
Carroll, 2019). Modifications that occur in germline cells
can be transferred to future generations. Scientists think
that they can extract genes that cause diseases in the
population using CRISPR-Cas9 (Cai et al., 2018; Memi et
al., 2018).

4. Bioethical issues
The fact that CRISPR-Cas9 is among the important
discoveries of the 21st century is widely accepted in the
scientific community and related industries. However,
the rapid rise of CRISPR-Cas9 has led to new bioethical,
social, and legal issues in medicine, agriculture, livestock,
and the environment. Possible risks and bioethical issues
related to CRISPR-Cas9 are summarized in the Table.
4.1. Ecological imbalance
In studies using RNA-targeted gene editing methods based
on CRISPR-Cas9, nontarget effects should be examined in

depth. Since gene drift will persist in a population, possible
off-target mutations will continue in each generation. In
addition, the number and effect of mutations may increase
as generations progress (Rodriguez, 2016; Hundleby
and Harwood, 2019). Another concern is the possibility
that genes can be transferred to other species in the
environment. Transferring the regulated sequences to
other species may result in the transmission of negative
characteristics to the associated organisms (Esvelt et al.,
2014). The distribution of the properties of the entrained
genes among the populations can make control very
difficult.
4.2. Regulations for consumers
The use of CRISPR-Cas9 to obtain the desired genetic
modifications makes it very difficult to identify and regulate
genetically modified organisms (GMOs) in the market after
they leave the laboratory. Therefore, regulatory agencies,
such as the US Food and Drug Administration (FDA),
European Medicines Agency (EMA), and others, should
consider whether any GMOs are suitable for consumers.
However, it is not known exactly how to evaluate the
possibilities of a growing market with CRISPR-Cas9
(Ledford, 2015; Hundleby and Harwood, 2019).

One of the dilemmas of CRISPR-Cas9 that concerns
all the humanity is patenting. As is known, transgenic
organisms of industrial use and also some human gene
sequences for clinical purposes have been patented
(Rodriguez, 2016; Sherkow, 2018). As technologies such
as CRISPR continue to evolve, patent-related issues in

Table. Possible risks and bioethical issues related to CRISPR-Cas9 technology.

Organism Risks Bioethical issues References

Bacteria
Nontarget mutations
Gene drifts

Ecological imbalance
Rodriguez, 2016
Hundleby and Harwood, 2019
Esvelt et al., 2014

Plants
Nontarget mutations
Gene drifts

Ecological imbalance
Patenting

Shinwari et al., 2017
Hundleby and Harwood, 2019

Animals /
chimeric animals

Nontarget mutations

Ecological imbalance
Patenting
Animal welfare and dignity
Threatening of human dignity and identity

Rodriguez, 2016
Polcz and Lewis, 2016
Rodriguez, 2017 Eriksson et al., 2018
Koplin, 2019 Degrazia, 2019
de Graeff et al., 2019

Humans

Nontarget mutations
Side effects
Cost
Genetic mosaicism

Eugenics
Informed consent
Enhancement
Accessibility
Patenting
Safety
Incomplete or over legislations

Otieno, 2015
Rodriguez, 2016
Duardo-Sánchez, 2017
Shinwari et al., 2017
Greene and Master, 2018
Sherkow, 2018
Cathomen et al., 2019
Hirsch et al., 2019

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many areas of biotechnology will continue to increase
in the upcoming years. Even today, there are many such
cases of patenting. The best-known case is the patent right
case between Zhang and Doudna and Charpentier for the
therapeutic use of CRISPR-Cas9 in human cells. In the
case concluded on 2 December 2016, it was decided to
grant the patent to Caribou Biosciences, which Doudna
was the founder of (Donohoue et al., 2018).
4.3. Genome editing for enhancement
The editing of human germline cells with CRISPR-Cas9,
which will be discussed later in more detail, is prohibited
for various safety reasons. However, the rate of application
of CRISPR-Cas9 to somatic cells is gradually increasing
in order to transfer the desired characteristics to our
lives. Many phenotypic characteristics have a genetic
component independent of the environment. By utilizing
this feature, CRISPR-Cas9 can be used to improve the
performance of athletes, to prevent violent behavior,
or to reduce dependence (Rodriguez, 2016). Although
gene therapy is often used to treat patients for their own
benefit, the criminal justice system may require repeater or
dangerous offenders to correct the genes associated with
violence by genome editing technologies in the future.
One of the biggest dilemmas here is to obtain informed
consent for an underage person if the intervention is
made during the development of the zygote. This will give
parents or guardians the right to make decisions on behalf
of minors for nonhealth reasons. Furthermore, when
socially assessed, some genetically improved populations
or individuals may have some advantages in comparison
to others in terms of various features such as mental and
physical capacity (Brokowski, 2018). Therefore, the use of
CRISPR-Cas9 in genome enhancement should be seriously
discussed both socially and morally.
4.4. Military research
The use of CRISPR technology for military purposes is
generally considered within the scope of nontherapeutic
enhancement and is covered similarly. From this point of
view, related bioethical issues are commonly discussed
in terms of concepts of benefit/risk, informed consent,
and accessibility (Greene and Master, 2018). A notable
bioethical problem is the off-target mutations that have
been mentioned in relation to other topics. Off-target
mutations can cause many undesirable changes in the
genome or even lead to fatal diseases. Current information
obtained from studies on off-target mutations caused by
CRISPR on the genome is very limited. Therefore, the
benefit/risk relationship needs to be evaluated carefully.
In addition, the possibility that this technology can be
used for the production of new biological weapons is
frightening.

Another ambiguous issue that needs to be discussed in
military enhancement applications is informed consent.

It could be difficult to obtain informed consent forms
independently without any interaction among individuals
due to military training methods, strict norms, and
chains of command. Additionally, some soldiers may have
difficulty in understanding the concepts of gene therapy
and genome editing, as well as the potential risks and
benefits of the applications (Greene and Master, 2018).

One important ethical issue is that the use of such
technologies will support ongoing inequalities among
military parties (Amoroso and Wenger, 2003). CRISPR
is currently an expensive technology. Some developed
countries might think of using this technology to further
strengthen their defenses and even attack underdeveloped
or developing countries. This situation could cause
a constant tension, making it difficult to provide an
environment of peace and stability worldwide.
4.5. Generation of chimeric animals for organ
transplantation
Organ transplantation is the replacement of an organ that
cannot function in an individual’s body with a healthy
organ from a living donor or cadaver. The primary purpose
is to save the life of the patient, who is in danger of organ
failure, and to increase the lifespan and quality of life
(Black et al., 2018). The development of chimeric animals
may prevent patients from spending precious time waiting
for an appropriate donor.

Bioethical issues in the generation of chimeric animals
arise from the fact that chimeras contain human nerve and
germ cells (Polcz and Lewis, 2016). The two main issues
can be summarized as defining the order of nature and
the moral disorders caused by how the organism is treated
depending on whether the organism is accepted as human
or animal. Some people think that chimeric embryos will
affect human dignity and identity because they have the
power to develop organisms with human-derived cells and
tissues. The others state that chimeric organisms containing
human cells cannot turn into humans and therefore will
not affect human dignity. They also argue that the human-
like features imparted to chimeras will neither affect the
biological environment nor the moral status of animals
and will never reach human consciousness (Koplin, 2019;
Degrazia, 2019).
4.6. Animal welfare and dignity
Animal welfare is another bioethical concern encountered
during the application of genome editing technologies on
animals. First of all, the possibility of off-target mutations
in the genome can lead to diseases or different side effects
in animals (Ishii, 2017a; Schultz-Bergin, 2018; de Graeff
et al., 2019). Such a situation will adversely affect animal
welfare (Rodriguez, 2017).

The second bioethical issue to be discussed could be the
concerns about “animal dignity” (Eriksson et al., 2018) and
alterations in their natural environments and physiological

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needs (Manesh et al., 2014). Some studies have stated that
the use of animals as objects only serving for humans
is not ethically or morally acceptable (Martinelli et al.,
2014; Fung and Kerridge, 2016; Greenfield, 2017), and
such practices can lead to greater control over humans on
animals (Ishii, 2017a; de Graeff et al., 2019). Some others
think that animals are not bound by any moral law and
therefore there is no need for a discussion regarding animal
dignity (Heeger, 2015; Shriver and McConnachie, 2018).
Schultz-Bergin (2017) stated that animal rights, welfare,
and dignity will not be adversely affected since these
animals will occur through genome editing technologies.
The existence of contrary opinions on this matter indicates
that the mentioned bioethical issues will be on the agenda
for a long time.
4.7. CRISPR-Cas9 for human germline
The potential for using CRISPR-Cas9 for genome editing
in the human germline has raised serious ethical debates.
Until 2015, all therapeutic applications in humans
were performed in somatic cells using genome editing
technologies. However, in 2015, the editing of the human
germline performed by Chinese scientist Huang and
his team with CRISPR-Cas9 raised new social, moral,
and bioethical issues (Liang et al., 2015; Ormond et al.,
2017). Bioethical issues caused by genome editing in the
germline can be classified into two main topics depending
on the success and failure of genome editing technologies
(Ormond et al., 2017; Greely, 2019).
4.7.1. Issues that may occur in the failure of germline
genome editing
Some of the ethical dilemmas of genome editing in the
germline arise from the fact that changes in the genome can
be transferred to the next generations. Therapeutic genome
editing in somatic cells generally does not cause significant
concerns when assessing the risk/benefit balance and the
use of informed consent. The application of CRISPR-Cas9
in the germline is considered more problematic because
of the risk of causing various mutations and side effects
and transferring undesirable changes to future generations
(Cyranoski and Reardon, 2015; Brokowski, 2018; Cai et al.,
2018; Halpern et al., 2019). In fact, Huang and his team
found that nontarget mutations in the genome occurred
and the study was terminated earlier than planned (Liang
et al., 2015). Nontarget mutations are unintentional
mutations in the genome and may have harmful effects
on the organism as these mutations can lead to cell death
or transformation (Zhang et al., 2014). Frighteningly,
researchers have found that mutations caused by CRISPR-
Cas9 in embryos are much more common than in mouse
or human adult cells (Cyranoski and Reardon, 2015). In a
study performed with human embryos, it was stated that
nontarget mutations occur only in the exon regions and
therefore the number of mutations may be much higher

than expected (Liang et al., 2015). Due to the high risk of
nontarget mutations, some scientists argue that genome
editing studies in germline cells should be terminated and
its future should be discussed (Cyranoski and Reardon,
2015). Some scientists state that newly developed CRISPR-
Cas9 could reduce or even prevent the number of nontarget
mutations. In this method, the efficiency of CRISPR-Cas9
was increased by using Cas9-regulated human iPSCs in
region-specific gene targeting (Yumlu et al., 2019).

Another bioethical dilemma is the cost of germline
genome editing. Genome editing is an expensive
technology (Wilson and Carroll, 2019). While families
in rich countries may have the power to cover this cost,
families in developing countries may not. This situation
may cause children born in developed countries to have an
unfair advantage in terms of various characteristics such
as intelligence and physical state compared to children in
other countries (Otieno, 2015).

CRISPR-Cas9 is based on the use of nuclease enzymes.
The nuclease enzymes used may not be as effective as
desired and not be able to cut all copies of the target gene,
or the cell may begin to divide before genome editing is
completed. As a result, a condition called genetic mosaicism
can occur (Lanphier et al., 2015). Genetic mosaicism is the
presence of genetically different somatic cell populations
in an organism and is often masked. Mosaicism can also
lead to major phenotypic changes, the formation of fatal
genetic mutations (Capalbo and Rienzi, 2017), and some
genetic diseases such as Down, Klinefelter, and Turner
Syndromes (Otieno, 2015). Therefore, the nuclease
cleavage sites should be exactly confirmed and the
possibility of mosaicism should be completely eliminated.

One of the important bioethical issues is side effects
in embryos. It is pointed out that the possible side effects
cannot be predicted before birth and the consequences
are not clearly known (Otieno, 2015; Brokowski, 2018).
Controls can only be performed in a small group of cells.
This limitation causes the effects of genome editing on
embryos to be unknown and unprevented until birth.
In fact, it should be considered that it may take years for
many potential problems to emerge (Lanphier et al., 2015;
Halpern et al., 2019).
4.7.2. Issues that may occur in the successful application
of germline genome editing
The first of the bioethical issues of successful germline
genome editing is the use for nontherapeutic changes
(Lanphier et al., 2015; Greely, 2019). Such uses will lead
to new questions about breeding (eugenics) of the human
species and its position in the universe (Yang, 2015). In
one study, the fur color of rats was successfully changed by
genome editing (Yoshimi et al., 2014). It is possible that the
skin color of people could be changed in the future. Since
the characteristics of individuals can be determined by

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genome editing rather than blood relations, the possibility
that children with similar physical and mental health can
be born in the same way should be considered (Ishi, 2015).

The second bioethical issue is what the fate of
children born using genome editing will be. From whom
or where informed consent will be obtained in the
case of undesirable effects on behalf of genome-edited
children and whether informed consent will give detailed
information are important questions (Beriain and del
Cano, 2018; Neuhaus and Zacharias, 2018; Sykora, 2018;
Knoppers and Kleiderman, 2019). While clear informed
consent can be given for genome-edited somatic cells to be
used in clinical trials, it is an enigma to whom and how to
give precise information about the potential risks involved
in germline editing (Lanphier et al., 2015; Neuhaus and
Zacharias, 2018; Knoppers and Kleiderman, 2019).

In December 2015, the International Summit on
Human Gene Editing was convened to discuss the
social, moral, and bioethical issues caused by genome
editing in the human germline. The results of the summit
concluded that basic and clinical investigations should
be continued in accordance with the appropriate legal
and ethical regulations; however, genome editing on
gametocytes and embryos that would cause hereditary
changes in humans was found to be irresponsible. It was
therefore emphasized that the use of CRISPR-Cas9 on the
human germline should be postponed until a solution is
found for existing bioethical, social, legal, and technical
concerns and issues (Baltimore et al., 2015). In addition,
it was agreed to establish an international forum where
such concerns could be addressed continuously and the
studies in different countries could be organized together
(Baltimore et al., 2015; Lanphier et al., 2015; Olson,
2015). The NIH announced that genome editing studies
in human embryos will not be financially supported
(Collins, 2015). In spite of the joint decisions that were
made, in February 2016, British scientists were allowed
to use CRISPR-Cas9 and similar technologies in human
embryos for research purposes only (https://www.bbc.
com/news/health-35459054). In March 2017, the US
National Academy of Sciences and the American Society
of Human Genetics published a position statement stating
that they should be aware of the scientific and bioethical
issues that can be caused by germline genome editing, but,
on the other hand, research should continue (Ormond et
al., 2017). As of January 2020, 24 countries have forbidden
genome editing in human embryos by law and 9 countries
have banned it by guidelines. However, there are countries
that do not impose strict prohibitions on germline genome
editing (Ishii, 2017b; Lau et al., 2018; Macintosh, 2019).

5. Discussion and future directions
Thanks to its high accuracy, ease of use, and relatively low
cost, CRISPR-Cas9 offers a wide range of applications for

many people in the medical, agricultural, livestock, and
environmental sectors. Furthermore, its precision and
accuracy are much higher compared to older technologies
such as ZFN and TALEN (Mittal, 2019). The powerful
effects of CRISPR-Cas9 have raised many social, moral,
and bioethical issues.

Discussions have generally focused on the social,
bioethical, and legal consequences of using genome editing
technology in human germline cells. Scientists generally
agree that CRISPR-Cas9 should be allowed for use in the
creation of human disease models, and in understanding
the development and molecular mechanisms of diseases;
however, it should be prohibited for the purposes of
eugenics or enhancement. When ethical issues, safety
concerns, and application difficulties are considered
together, it is predicted that therapeutic genome editing
in human embryos will not be possible in the near future.
Thus, the risk of hereditary nontarget genetic mutation is
higher than the possible treatment benefits and it affects the
principle of intentional harm. Nevertheless, it is clear that
scientists will apply CRISPR-Cas9 in germline cells in the
future if solutions are found to the issues mentioned here
(Duardo-Sánchez, 2017; Hirsch et al., 2019). CRISPR-Cas9
must be fully reliable for therapeutic use in germline cells.
Social, legal, and bioethical issues should be discussed in
detail once genome editing technologies have reached the
permissible level of safety for clinical applications in the
prevention of genetic diseases (Rossant, 2018; Cathomen
et al., 2019). Subsequently, regulatory laws that may
eliminate breaches of germline genome editing will need
to be reassessed (Rodriguez, 2016; Duardo-Sánchez, 2017;
Cathomen et al., 2019; Macintosh, 2019). The therapeutic
use of CRISPR-Cas9 and its rapid rise in the medical
field are expected to continue. While studies on the use
of CRISPR-Cas9 for clinical purposes are continuing, the
necessary legal, social, and ethical legislation should be put
into practice as soon as possible and the public conscience
should not be ignored.

On the other hand, the potential effects of CRISPR-
Cas9 in other areas should not be forgotten. CRISPR-
Cas9 is not just about social and bioethical issues related
to people. Interactions with other organisms and the
environment, such as the consideration of the principle
of intentional harm in risk assessment, safety measures to
prevent ecological degradation, or potential use in genetic
enhancement of animals and agriculture products should
also be discussed (Rodriguez, 2016; Hirsch et al., 2019).
There are serious concerns about changes in the natural
ecosystem that may occur if the GMOs produced with
CRISPR-Cas9 are released to the ecosystem in a controlled
or uncontrolled manner. Considering the applications
of CRISPR-Cas9 that protect mosquitoes from malaria
parasites (Gantz et al., 2015) or make female mosquitoes

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117

infertile (Hammond et al., 2016), the effect of GM
mosquitoes on other organisms with which they are
associated in their ecosystems cannot be predicted. It is
clear that small-scale research in the laboratory does not
fully reflect possible changes in the natural ecosystem
(Carroll, 2017). In agriculture, another concern about
GMOs produced with CRISPR-Cas9 is whether they will
be accepted by the public. GMOs that were produced
using different technologies in the past faced harsh
public reactions (Carroll, 2017). Furthermore, the fact
that GMOs produced with CRISPR-Cas9 are difficult
to identify outside the laboratory raises safety concerns
(Shinwari et al., 2017). Before the launch of such products,
the necessary explanations and declarations should be
made by the authorities in a transparent and clear manner
in order to prevent misjudgments and questions that may
occur in the public, and precautions and arrangements
should be established to ensure the safety of the public.

Another issue to consider about CRISPR-Cas9 is
patenting. Patenting can considerably limit the application
of such technologies. Unilateral patenting can significantly
increase the profitability of biotechnology companies,
which may lead to a rise of bioethical issues. There is
disagreement in the scientific community regarding the
patenting or nonpatenting of GMOs to be used specifically
for therapeutic purposes (Shinwari et al., 2017; Sherkow,
2018). However, there are some who think that patenting
will help to eliminate and regulate the deficiencies in the

field (Rodriguez, 2016; Shinwari et al., 2017). It should
not be forgotten that the most important of these debates
about patenting is commercialization and the release of
only reliable products.

In recent years, deals between the scientific community
and the pharmaceutical and biotechnology sectors for
the therapeutic use of CRISPR-Cas9 have raised public
safety concerns (Shinwari et al., 2017; Carroll, 2019) The
guidelines and legislations that will regulate the content
and application of these deals should be prepared as
quickly as possible and shared with the public. Due to
the challenges and bioethical issues of CRISPR-Cas9,
the scientific community and other interested bioethical,
social, legal, and governmental parties should be provided
with a detailed guide for future processing and use of
this technology (Otieno, 2015; Shinwari et al., 2017;
Cathomen et al., 2019). In this way, a long-term policy can
be developed that will support the scientific development
of CRISPR-Cas9 technology together with the discussion
of the possible problems in advance and preparation of
solution plans.

Conflict of interest
The third author is the founder and shareholder of Biovalda
Health Technologies, Inc. (Ankara, Turkey). The authors
declare no competing financial interests in relation to this
article. The authors alone are responsible for the content
and writing of the paper.

References

Amoasii L, Hildyard JC, Li H, Sanchez-Ortiz E, Mireault A et al.
(2018). Gene editing restores dystrophin expression in a canine
model of Duchenne muscular dystrophy. Science 362 (6410):
86-91.

Amoroso PJ, Wenger LL (2003). The human volunteer in military
biomedical research. In: Beam TE, Sparacino LR, Pellegrino
ED, Hartle AE, Howe EG (editors). Military Medical Ethics.
Volume 2. Washington, DC, USA: Walter Reed Army Medical
Center, pp. 563-660.

Arévalo MT, Navarro A, Arico CD, Li J, Alkhatib O et al. (2014).
Targeted silencing of anthrax toxin receptors protects against
anthrax toxins. Journal of Biological Chemistry 289 (22):
15730-15738.

Baltimore D, Berg P, Botchan M, Carroll D, Charo RA et al. (2015).
A prudent path forward for genomic engineering and germline
gene modification. Science 348 (6230): 36-38.

Barrangou R, Fremaux C, Deveau H, Richards M, Boyaval P et al.
(2007). CRISPR provides acquired resistance against viruses in
prokaryotes. Science 315 (5819): 1709-1712.

Beriain IDM, del Cano AMM (2018). Gene editing in human
embryos. A comment on the ethical issues involved. In:
Soniewicka M (editor). The Ethics of Reproductive Genetics.
Cham, Switzerland: Springer, pp. 173-187.

Black CK, Termanini KM, Aguirre O, Hawksworth JS, Sosin M
(2018). Solid organ transplantation in the 21st century. Annals
of Translational Medicine 6 (20): 409.

Brokowski C (2018). Do CRISPR germline ethics statements cut it?
CRISPR Journal 1 (2): 115-125.

Cai L, Zheng LA, He L (2018). The forty years of medical genetics
in China. Journal of Genetics and Genomics 45 (11): 569-582.

Carroll D (2017). Focus: Genome editing: genome editing: past,
present, and future. Yale Journal of Biology and Medicine 90
(4): 653.

Cathomen T, Schüle S, Schüßler-Lenz M, Abou-El-Enein M (2019).
The human genome editing race: loosening regulatory
standards for commercial advantage? Trends in Biotechnology
37 (2): 120-123.

AYANOĞLU et al. / Turk J Biol

118

Charlesworth CT, Deshpande PS, Dever DP, Camarena J, Lemgart VT
et al. (2019). Identification of preexisting adaptive immunity to
Cas9 proteins in humans. Nature Medicine 25 (2): 249.

Chen H, Kazemier HG, de Groote ML, Ruiters MH, Xu GL et al.
(2013). Induced DNA demethylation by targeting ten-eleven
translocation 2 to the human ICAM-1 promoter. Nucleic Acids
Research 42 (3): 1563-1574.

Chin A (2015). CRISPR-Cas9 Therapeutics: A Technology Overview.
Oxford, UK: Biostars.

Collins FS (2015). Statement on NIH Funding of Research Using
Gene-Editing Technologies in Human Embryos. Bethesda, MD,
USA: National Institutes of Health.

Cornelis MC, Nugent NR, Amstadter AB, Koenen KC (2010). Genetics
of post-traumatic stress disorder: review and recommendations
for genome-wide association studies. Current Psychiatry
Reports 12 (4): 313-326.

Cyranoski D, Reardon S (2015). Chinese scientists genetically modify
human embryos. Nature News 346: 1258096.

de Graeff N, Jongsma KR, Johnston J, Hartley S, Bredenoord AL
(2019). The ethics of genome editing in non-human animals: a
systematic review of reasons reported in the academic literature.
Philosophical Transactions of the Royal Society B 374 (1772):
20180106.

Degrazia D (2019). Human-animal chimeras, “human” cognitive
capacities, and moral status. Hastings Center Report 49 (5): 33-
34.

Doudna JA, Charpentier E (2014). The new frontier of genome
engineering with CRISPR-Cas9. Science 346 (6213): 1258096.

Duardo-Sanchez A (2017). CRISPR-Cas in medicinal chemistry:
applications and regulatory concerns. Current Topics in
Medicinal Chemistry 17 (30): 3308-3315.

Duchêne BL, Cherif K, Iyombe-Engembe JP, Guyon A, Rousseau J
et al. (2018). CRISPR-induced deletion with saCas9 restores
dystrophin expression in dystrophic models in vitro and in vivo.
Molecular Therapy 26 (11): 2604-2616.

Eriksson S, Jonas E, Rydhmer L, Röcklinsberg H (2018). Invited review:
Breeding and ethical perspectives on genetically modified and
genome edited cattle. Journal of Dairy Science 101 (1): 1-17.

Esvelt KM, Smidler AL, Catteruccia F, Church GM (2014). Concerning
RNA-guided gene drives for the alteration of wild populations.
Elife 3: e03401.

Firth AL, Menon T, Parker GS, Qualls SJ, Lewis BM et al. 2015.
Functional gene correction for cystic fibrosis in lung epithelial
cells generated from patient iPSCs. Cell Reports 12 (9): 1385-
1390. doi: 10.1016/j.celrep.2015.07.062

Fung RKF, Kerridge IH (2016). Gene editing advance re-ignites debate
on the merits and risks of animal to human transplantation.
Internal Medicine Journal 46 (9): 1017-1022.

Gantz VM, Jasinskiene N, Tatarenkova O, Fazekas A, Macias VM
et al. (2015). Highly efficient Cas9-mediated gene drive for
population modification of the malaria vector mosquito
Anopheles stephensi. Proceedings of the National Academy of
Sciences of the USA 112 (49): E6736-E6743.

Gracheva EO, Ingolia NT, Kelly YM, Cordero-Morales JF, Hollopeter
G et al. (2010). Molecular basis of infrared detection by snakes.
Nature 464 (7291): 1006.

Greely HT (2019). Human germline genome editing: an assessment.
CRISPR Journal 2 (5): 253-265.

Greene M, Master Z (2018). Ethical issues of using CRISPR
technologies for research on military enhancement. Journal of
Bioethical Inquiry 15 (3): 327-335.

Greenfield A (2017). Editing mammalian genomes: ethical
considerations. Mammalian Genome 28 (7-8): 388-393.

Guo X, Li XJ (2015). Targeted genome editing in primate embryos.
Cell Research 25 (7): 767.

Halpern J, O’Hara SE, Doxzen KW, Witkowsky LB, Owen AL (2019).
Societal and ethical impacts of germline genome editing: How
can we secure human rights? CRISPR Journal 2 (5): 293-298.

Hammond A, Galizi R, Kyrou K, Simoni A, Siniscalchi C et al.
(2016). A CRISPR-Cas9 gene drive system targeting female
reproduction in the malaria mosquito vector Anopheles
gambiae. Nature Biotechnology 34 (1): 78.

Heeger R (2015). Dignity only for humans? A controversy. In: Duwell
M (editor). The Cambridge Handbook of Human Dignity:
Interdisciplinary Perspectives. Cambridge, UK: Cambridge
University Press, pp. 541-545.

Hermerén G (2015). Ethical considerations in chimera research.
Development 142 (1): 3-5.

Hirsch F, Iphofen R, Koporc Z (2019). Ethics assessment in research
proposals adopting CRISPR technology. Biochemia Medica 29
(2): 206-213.

Huisman C, van der Wijst MG, Falahi F, Overkamp J, Karsten G et al.
(2015). Prolonged re-expression of the hypermethylated gene
EPB41L3 using artificial transcription factors and epigenetic
drugs. Epigenetics 10 (5): 384-396.

Ishii T (2017a). Genome-edited livestock: ethics and social
acceptance. Animal Frontiers 7 (2): 24-32.

Ishii T (2017b). Germ line genome editing in clinics: the approaches,
objectives and global society. Briefings in Functional Genomics
16 (1): 46-56.

Kim H, Kim JS (2014). A guide to genome engineering with
programmable nucleases. Nature Reviews Genetics 15 (5):
321-334.

Kim S, Koo T, Jee HG, Cho HY, Lee G et al. (2018). CRISPR RNAs
trigger innate immune responses in human cells. Genome
Research 28 (3): 367-373.

Knoppers BM, Kleiderman E (2019). Heritable genome editing: Who
speaks for “future” children? CRISPR Journal 2 (5): 285-292.

Koo T, Lu-Nguyen NB, Malerba A, Kim E, Kim D et al. (2018).
Functional rescue of dystrophin deficiency in mice caused
by frameshift mutations using campylobacter jejuni Cas9.
Molecular Therapy 26 (6): 1529-1538.

Koplin JJ (2019). Human-animal chimeras: the moral insignificance
of uniquely human capacities. Hastings Center Report 49 (5):
23-32.

AYANOĞLU et al. / Turk J Biol

119

Lanphier E, Urnov F, Haecker SE, Werner M, Smolenski J (2015).
Don’t edit the human germ line. Nature News 519 (7544): 410.

Lau RW, Wang B, Ricardo SD (2018). Gene editing of stem cells
for kidney disease modelling and therapeutic intervention.
Nephrology 23 (11): 981-990.

Lau V, Davie JR (2016). The discovery and development of the
CRISPR system in applications in genome manipulation.
Biochemistry and Cell Biology 95 (2): 203-210.

Ledford H (2015). CRISPR, the disruptor. Nature 522 (7554): 20-24.

Li W, Teng F, Li T, Zhou Q (2013). Simultaneous generation and
germline transmission of multiple gene mutations in rat using
CRISPR-Cas systems. Nature Biotechnology 31 (8): 684.

Liang P, Xu Y, Zhang X, Ding C, Huang R et al. (2015). CRISPR/Cas9-
mediated gene editing in human tripronuclear zygotes. Protein
& Cell 6 (5): 363-372.

Liao J, Karnik R, Gu H, Ziller MJ, Clement K et al. (2015). Targeted
disruption of DNMT1, DNMT3A and DNMT3B in human
embryonic stem cells. Nature Genetics 47 (5): 469.

Long C, Li H, Tiburcy M, Rodriguez-Caycedo C, Kyrychenko V et al.
(2018). Correction of diverse muscular dystrophy mutations in
human engineered heart muscle by single-site genome editing.
Science Advances 4 (1): eaap9004.

Macintosh KL (2019). Heritable genome editing and the downsides
of a global moratorium. CRISPR Journal 2 (5): 272-279.

Manesh SB, Samani RO, Manesh SB (2014). Ethical issues of
transplanting organs from transgenic animals into human
beings. Cell Journal (Yakhteh) 16 (3): 353.

Marraffini LA, Sontheimer EJ (2008). CRISPR interference limits
horizontal gene transfer in staphylococci by targeting DNA.
Science 322 (5909): 1843-1845.

Martinelli L, Oksanen M, Siipi H (2014). De-extinction: a novel
and remarkable case of bio-objectification. Croatian Medical
Journal 55 (4): 423.

Mei Y, Wang Y, Chen H, Sun ZS, Ju XD (2016). Recent progress in
CRISPR/Cas9 technology. Journal of Genetics and Genomics
43 (2): 63-75.

Memi F, Ntokou A, Papangeli I (2018). CRISPR/Cas9 gene-editing:
Research technologies, clinical applications and ethical
considerations. Seminars in Perinatology 42 (8): 487-500.

Mittal RD (2019). Gene editing in clinical practice: where are we?
Indian Journal of Clinical Biochemistry 34 (1): 19-25.

Neuhaus CP, Zacharias RL (2018). Compassionate use of gene
therapies in pediatrics: an ethical analysis. Seminars in
Perinatology 42 (8): 508-514.

Niu YY, Shen B, Cui YQ, Chen YC, Wang JY et al. (2014). Generation
of gene-modified cynomolgus monkey via Cas9/RNA-
mediated gene targeting in one-cell embryos. Cell 156 (4): 836-
843.

Olson S (editor) (2016). International Summit on Human Gene
Editing: A Global Discussion. Washington, DC, USA: National
Academies Press.

Ormond KE, Mortlock DP, Scholes DT, Bombard Y, Brody LC et al.
(2017). Human germline genome editing. American Journal of
Human Genetics 101 (2): 167-176.

Ota S, Hisano Y, Ikawa Y, Kawahara A (2014). Multiple genome
modifications by the CRISPR/Cas9 system in zebrafish. Genes
to Cells 19 (7): 555-564.

Otieno MO (2015). CRISPR-Cas9 human genome editing: challenges,
ethical concerns and implications. Journal of Clinical Research
and Bioethics 6 (6): 253-255.

Oye KA, Esvelt K, Appleton E, Catteruccia F, Church G et al. (2014).
Regulating gene drives. Science 345 (6197): 626-628.

Polcz S, Lewis A (2016). CRISPR-Cas9 and the non-germline non-
controversy. Journal of Law and the Biosciences 3: 413-425. doi:
10.1093/jlb/lsw016

Ramanan V, Shlomai A, Cox DB, Schwartz RE, Michailidis E et
al. (2015). CRISPR/Cas9 cleavage of viral DNA efficiently
suppresses hepatitis B virus. Scientific Reports 5: 10833.

Rath D, Amlinger L, Rath A, Lundgren M (2015). The CRISPR-
Cas immune system: biology, mechanisms and applications.
Biochimie 117: 119-128.

Rodriguez E (2016). Ethical issues in genome editing using Crispr/
Cas9 system. Journal of Clinical Research and Bioethics 7 (2):
266.

Rodriguez E (2017). Ethical issues in genome editing for non-human
organisms using CRISPR/Cas9 system. Journal of Clinical
Research and Bioethics 8 (300): 10-4172.

Rodriguez E, Keiser M, McLoughlin H, Zhang F, Davidson BL (2014).
AAV-CRISPR: a new therapeutic approach to nucleotide repeat
diseases. Molecular Therapy 22: 94-94.

Roh DS, Li EBH, Liao EC (2018). CRISPR Craft: DNA editing the
reconstructive ladder. Plastic and Reconstructive Surgery 142
(5): 1355-1364.

Rossant J (2018). Gene editing in human development: ethical
concerns and practical applications. Development 145 (16):
dev150888.

Schultz-Bergin M (2017). The dignity of diminished animals: Species
norms and engineering to improve welfare. Ethical Theory and
Moral Practice 20 (4): 843-856.

Schultz-Bergin M (2018). Is CRISPR an ethical game changer? Journal
of Agricultural and Environmental Ethics 31 (2): 219-238.

Senis E, Fatouros C, Große S, Wiedtke E, Niopek D et al. (2014).
CRISPR/Cas9-mediated genome engineering: an adeno-
associated viral (AAV) vector toolbox. Biotechnology Journal 9
(11): 1402-1412.

Sherkow JS (2018). The CRISPR patent landscape: past, present, and
future. CRISPR Journal 1 (1): 5-9.

Shinwari ZK, Tanveer F, Khalil AT (2017). Ethical issues regarding
CRISPR-mediated genome editing. Current Issues in Molecular
Biology 26: 103-110. doi: 10.21775/9781910190630.09

Shriver A, McConnachie E (2018). Genetically modifying livestock
for improved welfare: a path forward. Journal of Agricultural
and Environmental Ethics 31 (2): 161-180.

AYANOĞLU et al. / Turk J Biol

120

Swiech L, Heidenreich M, Banerjee A, Habib N, Li Y et al. (2015). In
vivo interrogation of gene function in the mammalian brain
using CRISPR-Cas9. Nature Biotechnology 33 (1): 102.

Sýkora P (2018). Germline gene therapy in the era of precise genome
editing: how far should we go? In: Soniewicka M (editor). The
Ethics of Reproductive Genetics. Cham, Switzerland: Springer,
pp. 157-171.

Urnov FD (2018). Genome Editing BC (before CRISPR): lasting
lessons from the “old testament”. CRISPR Journal 1 (1): 34-46.

van Erp PB, Bloomer G, Wilkinson R, Wiedenheft B (2015). The
history and market impact of CRISPR RNA-guided nucleases.
Current Opinion in Virology 12: 85-90.

Wang ZP, Xing HL, Dong L, Zhang HY, Han CY et al. (2015). Egg
cell-specific promoter-controlled CRISPR/Cas9 efficiently
generates homozygous mutants for multiple target genes in
Arabidopsis in a single generation. Genome Biology 16 (1):
144.

Wilson RC, Carroll D (2019). The daunting economics of therapeutic
genome editing. CRISPR Journal 2 (5): 280-284.

Wu Y, Zhou H, Fan X, Zhang Y, Zhang M et al. (2015). Correction
of a genetic disease by CRISPR-Cas9-mediated gene editing in
mouse spermatogonial stem cells. Cell Research 25 (1): 67.

Xin L, Min L, Bing S (2016). Application of the genome editing tool
CRISPR/Cas9 in non-human primates. Zoological Research 37
(4): 241.

Yang A (2015). Thinking Towards the Future of CRISPR/Cas9.
London, UK: Brevia.

Yang W, Tu Z, Sun Q, Li XJ (2016). CRISPR/Cas9: implications for
modeling and therapy of neurodegenerative diseases. Frontiers
in Molecular Neuroscience 9: (30).

Yoshimi K, Kaneko T, Voigt B, Mashimo T (2014). Allele-specific
genome editing and correction of disease-associated
phenotypes in rats using the CRISPR–Cas platform. Nature
Communications 5: 4240.

Yuan M, Gao X, Chard LS, Ali Z, Ahmed J et al. (2015). A marker-
free system for highly efficient construction of vaccinia virus
vectors using CRISPR Cas9. Molecular Therapy-Methods &
Clinical Development 2: 15035.

Yumlu S, Bashir S, Stumm J, Kühn R (2019). Efficient gene editing
of human induced pluripotent stem cells using CRISPR/Cas9.
In: Luo Y (editor). CRISPR Gene Editing. New York, NY, USA:
Humana Press, pp. 137-151.

Zhang F, Wen Y, Guo X (2014). CRISPR/Cas9 for genome editing:
progress, implications and challenges. Human Molecular
Genetics 23 (1): 40-46.

Zhang XL, Wei P, Xin-Tian H, Jia-Li L, Yong-Gang Y et al. (2014).
Experimental primates and non-human primate (NHP)
models of human diseases in China: current status and
progress. Zoological Research 35 (6): 447.

Zou Q, Wang X, Liu Y, Ouyang Z, Long H et al. (2015). Generation
of gene-target dogs using CRISPR/Cas9 system. Journal of
Molecular Cell Biology 7 (6): 580-583.

Heliyon 7 (2021) e06860

Contents lists available at ScienceDirect

Heliyon

journal homepage: www.cell.com/heliyon

Research article

Researchers views about perceived harms and benefits of gene editing:
A study from the MENA region

Sawsan Abuhammad a,*, Omar F. Khabour b, Karem H. Alzoubi c

a Dept. of Maternal and Child Health, Jordan University of Science and Technology, Irbid 22110, Jordan
b Dept. of Medical Laboratory Sciences, Jordan University of Science and Technology, Irbid 22110, Jordan
c Dept. of Clinical Pharmacy, Jordan University of Science and Technology, Irbid 22110, Jordan

A R T I C L E I N F O

Keywords:
Gene editing
MENA
Ethics
Gene therapy
Enhancement

* Corresponding author.
E-mail address: [email protected] (S. A

https://doi.org/10.1016/j.heliyon.2021.e06860
Received 7 November 2020; Received in revised fo
2405-8440/© 2021 The Author(s). Published by Els
nc-nd/4.0/).

A B S T R A C T

Background: The development of gene editing technologies is very promising for the treatment of genetic diseases.
However, gene editing can be also used to enhance the characteristics of healthy individuals. This study aims to
determine ethical challenges that may face the constitution of gene editing in the Middle East and North Africa
(MENA) region.
Methods: An online discussion forum about the ethical challenges of applying gene editing technologies was held.
The participants were a group of researchers (n ¼ 28) from the MENA region.
Results: Most of the participants agreed on the importance of gene editing for the treatment of genetic diseases.
However, participants had concerns regarding the use of gene editing to enhance the characteristics of healthy
individuals such as athletic abilities and intelligence. Among ethical issues that were raised are justice, harm,
beneficence, discrimination, conflict with religion and culture, and lack of regulations.
Conclusion: Several ethical issues were raised for using gene editing technologies based on the perception of
biomedical researchers from the MENA region. Therefore, the scientific community and other interested
bioethical, social, legal, and governmental parties should be provided with a detailed guide from the scientists in
this area for future uses of this technology.

1. Introduction

The evolution of genetic technologies has made it possible to modify
somatic and germ cells (Sung et al., 2012; Kimbrel and Lanza 2020; van
Haasteren et al., 2020). Scientists have recently used gene editing tools to
efficiently edit the human embryonic genome (Huang et al., 2020; Kar-
imian et al., 2020). Gene editing focuses on a specific region in the
genome leading to the altering of harmful loci that cause diseases
(Mehravar et al., 2020). It is intended that the next generations should
inherit these alterations to eradicate mutated genes that cause diseases.
The applications of gene editing in the human zygote to correct genetic
diseases such as beta-globin gene disorders were reviewed (Tang et al.,
2017). According to Sharma and Scott (2015), the deliberations among
leading scientists concerning possible ethical issues of gene editing and
the way to impact coming generations, increasing the global concerns on
challenges such as, will gene editing produce designer babies?, who will
choose the destiny of a child resulting from the technology?, is there a
ground for the child to decide/consent?, and will people begin using such
techniques to improve their abilities?

buhammad).

rm 10 March 2021; Accepted 16
evier Ltd. This is an open access a

The extent of achievement of the scientists using gene editing in
humans highlights the need to develop ethical guidelines that regulate
this area of research (Lanphier et al., 2015; Vogel 2015; Peng et al.,
2016). Scientists currently suggest that a cure can be attained at the so-
matic cell level since using gene editing in germ cells might lead to un-
predictable results. According to Reardon (Kaye et al., 2009), there is an
urgent need to develop ethical guidelines for future human genetic en-
gineering research. In fact, due to the accelerated developments in gene
editing, there is more amplification of ethical issues with emerging new
questions (Kaye et al., 2009). Consequently, it is not strange to find
significant studies focusing on gene editing related ethical issues, such as
justice, harm, culture, religion, beneficence, discrimination and govern-
mental regulations (Doudna 2020; Niemiec and Howard 2020; Zhang
et al., 2020). Due to these reasons, this research aimed to determine
ethical challenges that may face the constitution of gene editing in the
Middle East and North Africa (MENA) region. Possible issues were
examined by a team of scholars from the region representing various
biomedical fields. The study highlights the importance of gene editing
and certain ethical challenges associated with this technology among

April 2021
rticle under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-

S. Abuhammad et al. Heliyon 7 (2021) e06860

researchers and health care professionals in the MENA region. All pre-
sented revelations contained few problems that may be adopted to build
an effective gene editing research in the MENA. Moreover, the re-
searchers agreed that such work could energize policymakers and
stockholders in creating an effective and legitimate genetic editing
structure.

2. Methods

2.1. Study plan

This study used a thematic qualitative design by imagining an ethical
situation. A situation in this study was to suppose that, in the future, gene
editing becomes routine clinical practice for the treatment of diseases.
This inspired the authors of this project to ask: What are ethical issues
that might be associated with gene editing technology? Should we
restrict gene therapy to the treatment of human genetic diseases? These
questions were asked to a group of researchers from Jordan and MENA
through an online discussion forum. The forum was part of the activities
of the “Ethics of Genetic and Clinical studies” course that was offered by
the Research Ethics Education Program in Jordan. All participants in this
program (n ¼ 28) were confronted in this forum, consisting of nation-
alities from the MENA region. These include Jordan, Iraq, Tunisia, Sudan,
Morocco, Yemen, Gaza, and Algeria. Few of the participants (n ¼ 4) were
research assistants with a master’s degree, whereas the rest were faculty
members with a Ph.D degree in health-related disciplines (pharmacy,
medicine, dentistry, nursing, and applied medical sciences). Among the
participants, four researchers were specialized in molecular genetics with
long experience in the conduction of human genetic studies. Researchers
were chosen from different biomedical fields to get a comprehensive
view of the studied topic.

The forum was inaugurated with a period of 14 days, and 92 posts
from 28 members were tallied. The main themes that resulted from these
posts were 1) Justice, 2) Harm, 3) Beneficence, 4) Discrimination, 5)
Respect of culture and religion, and 6) Legislations and laws. All posts
from the participants in this forum were followed and reviewed by
guiding researchers. An expert in qualitative studies checked each dis-
cussion. This observing procedure incorporated elevating the members to
be occupied with the conversation by utilizing a few posts like “your
point needs more clarification” and “could you explain your opinion
more” to confirm the partners’ perceptions. These questions increased the
truthiness of the collected information. To improve the investigation’s
objectivity, another investigator not taking part in the conversational
discussion checked the collected information.

The IRB at the Jordan University of Science and Technology (IRB-
JUST) endorsed this research. Alongside, IRB ensured that the ethical
principles are followed, which is based on the 1964 statement of Helsinki
and the principals’ further modifications.

3. Results

A selected team of researchers residing in the MENA region discussed
ethical issues related to gene editing using a web-based portal. The main
ethical issues that emerged from the discussion were justice, harm,
beneficence, discrimination, respect culture and religion, and regulations
and laws.

3.1. Justice

Another concern for using gene editing is the justice of using such
expensive technology. The male participant (MA, 38 years old and
Muslim) mentioned “gene editing may cause biases in benefit distribu-
tion, due to the limited convenience and high costs of such technology for
the public”. Another female participant (SA, 43 years old and Muslim)
added “genetic editing technology will be focusing on rich people and
ignoring poor people”. Furthermore, the male participant (AA, 36 years

2

old and Muslim) mentioned “applying gene editing technology will cause
rich people to have more advantages than poor people to treat their
diseases”. Another male participant (OK, 43 years old and Muslim) added
“if an athlete used gene therapy to enhance the strength of his muscles by
5–10%. This will make a big difference in sport competitions between
countries …”. Female (EM, 37 years old and Muslim) added “there is an
organization called The World Anti-Doping Agency (WADA) to detect the
use of this technique among athletes. However, this agency is facing a lot
of challenges in determining whether gene therapy was used or not.”

3.2. Beneficence

Most of the researchers emphasized the beneficence of gene editing
technology in the treatment of genetic diseases. The ALA participant (35
years old and Muslim) said “gene therapy could be very helpful in the
future. We could use this technology for either prevent or treat genetic
diseases, some types of cancers, and certain viral infections”. Another
participant (OA, female and Muslim) added: ”This technology as multiple
medications or surgical procedures that are used to make people behave
better, look better and so on. Use of gene therapy may look equivalent to
these medications and procedures”. Furthermore, the female participant
(SA, 43 years old and Muslim) mentioned “Applying “Gene Therapy” to
cure or prevent certain diseases after careful studies where benefits
overweigh risks is ethical.”

3.3. Harm

The associated risks of gene editing were a major ethical issue high-
lighted by most participants. For example, a male participant (AA, 36
years old and Muslim) said “I think that we should restrict gene therapy
to treatment of genetic diseases as that the entire world would agree on
that …. Gene editing for enhancement purposes might introduce a new
disease that could be more fatal than the one to be treated”. Another
female participant (HA, 35 years old and Muslim) said “There are people
who are willing to put themselves at risk of gene editing to get a 5%
modification in whatever area they are after, be it cognition, sports,
strength, etc…”. Another male participant (NM, 31 years old and
Muslim) mentioned that “there are attempts to alter or improve a ‘normal’
person by gene manipulation, which might not be ethical.” One male
participant (MF, 30 years old and Muslim) pointed that gene editing
might affect genetic diversity in the human population “Gene therapy
could be used to select some characteristics for the newborn babies that
are preferred in the community. This could result in having all human
looks the same and preventing the natural selection”.

3.4. Discrimination

Participants insisted that gene editing could lead to sex discrimina-
tion. For example, a female participant (ANA, 39 years old and Muslim)
said “Genetic could result in having a gender imbalance in some societies.
For example, boys are preferred over the girls in Arab countries”.

3.5. Respect culture and religion

Respect for culture and religion is a major issue facing using gene
editing. A female participant (TA, 34 years old and Muslim) mentioned
that “Genetics technology is not accepted or allowed in Islam”. Another
participant male (HA, 35 years old and Muslim) added “Religion along-
side culture in our society has to take a strong educated stance with
regards to such research, prior to its advancements infiltrating and
affecting our societies, to protect us and to recognize wrong from right
early on. A female participant (MA, 45 years old and Muslim) mentioned”
Most people are seeking for quick treatment for their diseases/condi-
tions, but new methods are not easy to be applied in our Arab countries, it
still needs more time to be understood.”

S. Abuhammad et al. Heliyon 7 (2021) e06860

3.6. Regulations and laws

Researchers highlighted the importance of having guidelines that
regulate research activities in Jordan and the MENA. A female partici-
pant (AYA, 37 years old and Muslim) mentioned “I believe that strict
regulations should be applied to gene therapy in order to restrict its use to
people who are in actual need of it”. A female participant (LA, 42 years
old and Muslim) added that “I would think that its use shall be regulated
as the use of already available techniques (ex, plastic surgery) without
total prevention or restriction for medical use only”.

4. Discussion

This was the first study in the MENA region regarding ethical issues of
gene editing. The ethical challenges that might face the establishment of
genetic editing in the MENA were discussed among a group of researchers
from the region. These researchers were chosen from various biomedical
fields to get a comprehensive view of the studied topic. Among the
highlighted ethical issues about gene editing were justice, harm, benefi-
cence, discrimination, respect culture and religion, and regulations and
laws. The results of this study provide the scientific community and other
interested bioethical, social, legal, and governmental parties with a
detailed guide for future processing and use of this technology.

According to the current study, many respondents (75%) stated that
the issues concerning gene editing are challenges associated with justice.
Biases in benefit distribution, due to the limited convenience and high
costs of gene-altering techniques for the public, may widen the disparity
between various groups, and increase the impacts of genetic variations
between individuals. Additionally, this may result in a dislike of genetic
research. The ‘justice’ principle is a central ethical and health equity
foundation that led the execution of gene editing studies and use in
clinical treatment. According to the National Academy of Sciences (NAS)
report (Neufeld and Scheck 2010), justice is a guideline that “requires
similar handling of similar instances, and equal distribution of risks and
benefits (distributive justice)”.

Responsibilities that emerge from adhering to this regulation include
equal sharing of research benefits and difficulties; and broad and equal
access to the clinical application benefits of editing the human gene.
Patients and families of individuals suffering from the sickle cell and
inherited blindness expressed equality and justice as a dominant theme
(Bonham and Smilan, 2018). In various studies, stakeholders were con-
cerned about who the actual beneficiary of gene editing would be
(Bonham et al., 2010; Bonham and Smilan 2018).

Another concern regarding harm is the application of genetic editing
on a person and society. Besides, respondents were concerned that this
technique could have unpredictable impacts on human health or could be
abused. Some respondents (34%) were afraid that unethical scientists or
physicians could exploit weak patients; they reiterated an oversight need
during this technology implementation. “I think this testing (gene edit-
ing) can produce some undesirable side effects which may cause worse
problems than what you are suffering from” (Hildebrandt and Marron
2018). Although it was apparent that the majority of discussion forum
respondents had limited genetics/gene-editing method knowledge, the
participants expressed similar ethical concerns to the cited published
literature on the general public attitude, including the progressing
consultation by academic, industry, and government partners (Haga and
Beskow, 2008).

One of the most concern in our study was beneficence in applying
gene editing to individuals and society. This was similar to the past work,
where research revealed that in general, there was approximately
60–70% level of acceptance for therapeutically centered gene editing
(Hoeyer et al., 2004; Veit, 2018). The key differences seem to lean on the
proposed gene editing applications (e.g., a therapeutic strategy compared

3

to nonmedical or improvements). For instance, the Pew research showed
that 72% backed gene-editing treatment of an acute illness will affect a
baby during birth; 60% will minimize the possibility of a severe disease
that may happen in an individual’s life; and 19% will increase the in-
telligence of a baby (Balica, 2019). A study in the UK revealed that 83%
of participants will back gene editing if were a carrier of a genetic dis-
order and there was a risk for the future generation inheriting the dis-
order. Yet, only 23% backed the use of gene editing ‘to improve the
intelligence of future children and 12% supported it to ‘alter the
appearance of future children (2014).

Lastly, in qualitative research meant to evoke gene editing opinions
on human embryos and somatic gene treatment, focus group respondents
in the upper Midwest USA produced similar outcomes in assessing the
wider public (Ormond et al., 2017). Generally, the discussion forum re-
spondents in the current study supported the application of gene editing
to cure severe or life-threatening congenital or adult-onset illness but
were more uncertain about applying gene editing to cure multifactorial
diseases that could be treated by modifying lifestyles.

The current study reported a concern that gene editing might lead to
sex discrimination as boys are preferred over girls in Arab countries
(Obermeyer and Cardenas, 1997). This might create an imbalance in the
population, which already started to arise in some Asian countries that
favor boys but is less likely in Western Europe and North America
(Macklin, 2010). Thus, gene editing technology should be regulated to
avoid such discriminations and to prevent undesirable consequences in
the communities.

Another ethical concern of gene editing that was discussed by other
studies and was not highlighted by the current study participants is
related to discrimination against disabled children (Giorgini et al., 2015;
Sparrow 2019). For example, gene editing procedures could affect evo-
lution, both socially and scientifically, and the negative effect of losing
societal heterogeneity, a kind of ‘counter-eugenic logic (Giorgini et al.,
2015; Sparrow, 2019).

Many participants in the current study stated religious and cultural
concerns about gene editing. Literature provides limited research con-
cerning the association between spiritual connections and providing gene
editing information (Sanderson et al., 2017). The two leading Jordanian
religions are Islam (primarily Sunni) then Christianity (usually Orthodox).
Individuals of these faiths seek religious guidance on their daily life issues
and focus on matters allowed versus not allowed in their religions (Ahram
et al., 2014). Strict Muslim researchers have revealed that Islam embraces
the exploration of gene editing foundation and highly credits the inde-
pendence and privacy code (Alahmad and Dierickx 2012). According to
the Islamic point of view, the main argument against embracing gene
editing is that it is changing God’s creation (taghy�ı rbi-Khalq All�ah). The
devil promises that “And I will instruct them, and they will change what
God has created!” (Qur’an, 4:119). Modifying God’s creation means
apparent disobedience of a proscription that originated from the Qur’an
(Lala 2020). Before delving into how and if gene editing changes God’s
creation, we must first be sure which verse is so firmly and steadfastly
adduced to in all issues of human alteration. Altering God’s creation in-
volves not only physical modifications, but also involves teleological, and
synderesis modifications that may attend the physical changes, and
cannot be eliminated. A fundamental interaction principle is that adjust-
ments, though physical, can have psychological impacts. An interest to
follow a particular direction can simultaneously result in a primordial
possibility, and a readiness (isti’d�ad) in Akbarian parlance (Lala, 2020),
which may be present in the genetic constitution. Thus, modifying human
genes not only alters who we are in a physical sense but also, results in an
ontological change that is against culture and religions for many faiths.

Many participants in the current study stated that it is necessary to
legislate and enact laws to monitor the use of genetic materials. Re-
searchers stated that there is an urgent need to track the moral issues and

S. Abuhammad et al. Heliyon 7 (2021) e06860

regulations concerned with gene editing. Many founders and gene edit-
ing staff, specialists, researchers, and research assistants in Western
countries such as the USA and UK have considered gene editing to be a
contentious investigation industry (2014). Currently, there is a lack of
systematic and extensive evaluation or quality appraisal for the decisions
that the individual research ethics board makes. Besides, there are no
jurisdictions for committees evaluating such research to use a particular
yardstick. Those who violate these guidelines and regulations do not face
a criminal penalty. However, some physicians, attorneys, and bio-
chemists are attempting to escalate these to low levels with related
criminal punishments.

Scientists in China have appealed for the enforcement of clear laws
and regulations to control research in human gene editing, as well as
determining the kinds of research that can or cannot be conducted. From
their perspective, they have been concerns that the right basic research
may promote ethical discussions and thus hindering research with
beneficial scientific and ethical quality.

Although restrictive laws regulating embryo research and gene alter-
ations still exist in many countries such as China, some nations permit
basic studies or have processes that allow such studies, e.g., the UK, China,
and Sweden (Callaway 2016). According to Reardon, the NIH in the US
has stressed its prohibition of utilizing federal financing for gene editing of
human embryos due to the issues surrounding the human embryos ethical
status. Such research is allowed without funding from the federal gov-
ernment. According to previous research, the US public appears to have a
conservative perspective towards gene editing (Winickoff, 2007).

This traditional perspective may lead to more restrictions in the US
compared to China and the UK. After the publication of the work of Dr.
Huang, a consensus conference was convened in Washington DC in
December 2015, with delegates from China, the USA, and the UK (Olson
et al., 2016). After the summit, a statement lifted a worldwide suspension
of all human genetic editing research, permitting fundamental research
progress, but agreed on the prematurity of any clinical use. Overall, the
continuing discussion regarding human gene-editing research for
non-reproductive use is majorly about gene editing safety issues, because
based on the existing biology knowledge; there is an extremely high ratio
of risk/benefit of editing the human gene (Olson et al., 2016).

There are still high rates of unintended modifications in gene editing
and other unintended impacts. Scientists are progressing on minimizing
gene editing risks, thus, there is an urgent need for gene editing regu-
lations. After the second international conference on gene editing, the
summit statement, conducted in Hong Kong in November 2018, offered
hopes of establishing thirteen ethical concerns in editing human genes: a
viewpoint and route towards the clinical application of embryo gene
editing (Olson et al., 2016).

5. Conclusion

The participants of the current study representing various countries
from the MENA region, and different biomedical fields agreed on the
importance of gene editing to treat genetic conditions. They also high-
lighted the need for regulations to prevent the misuse of gene editing
technology. Among the raised concerns regarding gene editing in the
MENA were justice, harm, beneficence, discrimination, and govern-
mental regulations. Therefore, the scientific community and other
interested bioethical, social, legal, and governmental parties should be
provided with a detailed guide from the scientists in this area for future
processing and use of this technology.

Declarations

Author contribution statement

Sawsan Abuhammad, Omar Khabour, Karem Alzoubi: Conceived and
designed the experiments; Performed the experiments; Analyzed and

4

interpreted the data; Contributed reagents, materials, analysis tools or
data; Wrote the paper.

Funding statement

This work was supported by the Foundation for the National Institutes
of Health, (1R25TW010026-01) and Jordan University of Science and
Technology.

Data availability statement

The data that has been used is confidential.

Declaration of interests statement

The authors declare no conflict of interest.

Additional information

No additional information is available for this paper.

References

Ahram, M., Othman, A., Shahrouri, M., Mustafa, E., 2014. Factors influencing public
participation in biobanking. Eur. J. Hum. Genet. 22 (4), 445–451.

Alahmad, G., Dierickx, K., 2012. What do Islamic institutional fatwas say about medical
and research confidentiality and breach of confidentiality? Develop. World Bioeth. 12
(2), 104–112.

Balica, R., 2019. Regulating human germline genome editing: medical counseling, ethical
permissibility, and potentially grave threats. Rev. Contemp. Philos. (18), 133–139.

Bonham, V.L., Smilan, L.E., 2018. Somatic genome editing in sickle cell disease: rewriting
a more just future. NCL Rev. 97, 1093.

Bonham, V.L., Dover, G.J., Brody, L.C., 2010. Screening student athletes for sickle cell
trait–a social and clinical experiment. N. Engl. J. Med. 363 (11), 997–999.

Callaway, E., 2016. UK scientists gain licence to edit genes in human embryos. Nature 530
(7588), 18.

Doudna, J.A., 2020. The promise and challenge of therapeutic genome editing. Nature
578 (7794), 229–236.

Giorgini, V., Mecca, J.T., Gibson, C., Medeiros, K., Mumford, M.D., Connelly, S.,
Devenport, L.D., 2015. Researcher perceptions of ethical guidelines and codes of
conduct. Account. Res. 22 (3), 123–138.

Haga, S.B., Beskow, L.M., 2008. Ethical, legal, and social implications of biobanks for
genetics research. Adv. Genet. 60, 505–544.

Hildebrandt, C.C., Marron, J.M., 2018. Justice in CRISPR/Cas9 research and clinical
applications. AMA J. Ethics 20 (9), E826–833.

Hoeyer, K., Olofsson, B.O., Mj€orndal, T., Lyn€oe, N., 2004. Informed consent and biobanks:
a population-based study of attitudes towards tissue donation for genetic research.
Scand. J. Publ. Health 32 (3), 224–229.

Huang, D., Miller, M., Ashok, B., Jain, S., Peppas, N.A., 2020. CRISPR/Cas systems to
overcome challenges in developing the next generation of T cells for cancer therapy.
Adv. Drug Deliv. Rev.

Karimian, A., Gorjizadeh, N., Alemi, F., Asemi, Z., Azizian, K., Soleimanpour, J.,
Malakouti, F., Targhazeh, N., Majidinia, M., Yousefi, B., 2020. CRISPR/Cas9 novel
therapeutic road for the treatment of neurodegenerative diseases. Life Sci. 259,
118165.

Kaye, J., Heeney, C., Hawkins, N., de Vries, J., Boddington, P., 2009. Data sharing in
genomics–re-shaping scientific practice. Nat. Rev. Genet. 10 (5), 331–335.

Kimbrel, E.A., Lanza, R., 2020. Next-generation stem cells – ushering in a new era of cell-
based therapies, 19 (7), 463–479.

Lala, I., 2020. Germ-inating solutions or gene-rating problems: an Islamic perspective on
human germline gene editing. J. Relig. Health 59 (4), 1855–1869.

Lanphier, E., Urnov, F., Haecker, S.E., Werner, M., Smolenski, J., 2015. Don’t edit the
human germ line. Nature 519 (7544), 410–411.

Macklin, R., 2010. The ethics of sex selection and family balancing. Semin. Reprod. Med.
28 (4), 315–321.

Mehravar, M., Roshandel, E., Salimi, M., Chegeni, R., Gholizadeh, M., Mohammadi, M.H.,
Hajifathali, A., 2020. Utilization of CRISPR/Cas9 gene editing in cellular therapies
for lymphoid malignancies. Immunol. Lett. 226, 71–82.

Neufeld, P., Scheck, B., 2010. Making forensic science more scientific. Nature 464 (7287),
351.

Niemiec, E., Howard, H.C., 2020. Ethical issues related to research on genome editing in
human embryos. Comput. Struct. Biotechnol. J. 18, 887–896.

Obermeyer, C.M., Cardenas, R., 1997. Son preference and differential treatment in
Morocco and Tunisia. Stud. Fam. Plann. 28 (3), 235–244.

Olson, S., C. on Science, E. National Academies of Sciences and Medicine, 2016.
International summit on Human Gene Editing: A Global Discussion. International
Summit on Human Gene Editing: A Global Discussion. National Academies Press
(US).

S. Abuhammad et al. Heliyon 7 (2021) e06860

Ormond, K.E., Mortlock, D.P., Scholes, D.T., Bombard, Y., Brody, L.C., Faucett, W.A.,
Garrison, N.A., Hercher, L., Isasi, R., Middleton, A., Musunuru, K., Shriner, D.,
Virani, A., Young, C.E., 2017. Human germline genome editing. Am. J. Hum. Genet.
101 (2), 167–176.

Peng, R., Lin, G., Li, J., 2016. Potential pitfalls of CRISPR/Cas9-mediated genome editing.
FEBS J. 283 (7), 1218–1231.

Sanderson, S.C., Brothers, K.B., Mercaldo, N.D., Clayton, E.W., Antommaria, A.H.M.,
Aufox, S.A., Brilliant, M.H., Campos, D., Carrell, D.S., Connolly, J., Conway, P.,
Fullerton, S.M., Garrison, N.A., Horowitz, C.R., Jarvik, G.P., Kaufman, D.,
Kitchner, T.E., Li, R., Ludman, E.J., McCarty, C.A., McCormick, J.B., McManus, V.D.,
Myers, M.F., Scrol, A., Williams, J.L., Shrubsole, M.J., Schildcrout, J.S., Smith, M.E.,
Holm, I.A., 2017. Public attitudes toward consent and data sharing in biobank
research: a large multi-site experimental survey in the US. Am. J. Hum. Genet. 100
(3), 414–427.

Sharma, A., Scott, C.T., 2015. The ethics of publishing human germline research. Nat.
Biotechnol. 33 (6), 590–592.

5

Sparrow, R., 2019. Yesterday’s child: how gene editing for enhancement will produce
obsolescence-and why it matters. Am. J. Bioeth. 19 (7), 6–15.

Sung, Y.H., Baek, I.J., Seong, J.K., Kim, J.S., Lee, H.W., 2012. Mouse genetics: catalogue
and scissors. BMB Rep. 45 (12), 686–692.

Tang, L., Zeng, Y., Du, H., Gong, M., Peng, J., Zhang, B., Lei, M., Zhao, F., Wang, W., Li, X.,
Liu, J., 2017. CRISPR/Cas9-mediated gene editing in human zygotes using Cas9
protein. Mol. Genet. Genom. 292 (3), 525–533.

van Haasteren, J., Li, J., Scheideler, O.J., Murthy, N., Schaffer, D.V., 2020. The delivery
challenge: fulfilling the promise of therapeutic genome editing, 38 (7), 845–855.

Veit, W., 2018. Procreative beneficence and genetic enhancement.
Vogel, G., 2015. Bioethics. Embryo engineering alarm. Science 347 (6228), 1301.
Winickoff, D.E., 2007. Partnership in U.K. Biobank: a third way for genomic property?

J. Law Med. Ethics 35 (3), 440–456.
Zhang, D., Hussain, A., Manghwar, H., Xie, K., 2020. Genome editing with the

CRISPR-Cas system: an art, ethics and global regulatory perspective, 18 (8),
1651–1669.

European Journal of Human Genetics (2018) 26:1–11
https://doi.org/10.1038/s41431-017-0024-z

POLICY

One small edit for humans, one giant edit for humankind? Points
and questions to consider for a responsible way forward for gene
editing in humans

Heidi C. Howard1 ● Carla G. van El2 ● Francesca Forzano3 ● Dragica Radojkovic4 ● Emmanuelle Rial-Sebbag5 ●

Guido de Wert7 ● Pascal Borry6 ● Martina C. Cornel 2 on behalf of the Public and Professional Policy
Committee of the European Society of Human Genetics

Received: 2 February 2017 / Revised: 24 September 2017 / Accepted: 29 September 2017 / Published online: 30 November 2017
© The Author(s) 2018. This article is published with open access

Abstract
Gene editing, which allows for specific location(s) in the genome to be targeted and altered by deleting, adding or
substituting nucleotides, is currently the subject of important academic and policy discussions. With the advent of efficient
tools, such as CRISPR-Cas9, the plausibility of using gene editing safely in humans for either somatic or germ line gene
editing is being considered seriously. Beyond safety issues, somatic gene editing in humans does raise ethical, legal and
social issues (ELSI), however, it is suggested to be less challenging to existing ethical and legal frameworks; indeed somatic
gene editing is already applied in (pre-) clinical trials. In contrast, the notion of altering the germ line or embryo such that
alterations could be heritable in humans raises a large number of ELSI; it is currently debated whether it should even be
allowed in the context of basic research. Even greater ELSI debates address the potential use of germ line or embryo gene
editing for clinical purposes, which, at the moment is not being conducted and is prohibited in several jurisdictions. In the
context of these ongoing debates surrounding gene editing, we present herein guidance to further discussion and
investigation by highlighting three crucial areas that merit the most attention, time and resources at this stage in the
responsible development and use of gene editing technologies: (1) conducting careful scientific research and disseminating
results to build a solid evidence base; (2) conducting ethical, legal and social issues research; and (3) conducting meaningful
stakeholder engagement, education and dialogue.

Introduction

Gene editing, which allows for specific location(s) in the
genome to be targeted and changed by deleting, adding or
substituting nucleotides, is currently the subject of much
academic, industry and policy discussions. While not new
per se, gene editing has become a particularly salient topic

primarily due to a relatively novel tool called CRISPR-
Cas9. This specific tool distinguishes itself from its coun-
terparts, (e.g., zinc-finger nucleases and TAL effector
nucleases (TALENs)) due to a mixture of increased effi-
ciency (number of sites altered), specificity (at the exact
location targeted), ease of use and accessibility for
researchers (e.g., commercially available kits), as well as a

* Heidi C. Howard
[email protected]

1 Centre for Research Ethics and Bioethics, Uppsala University,
Uppsala, Sweden

2 Department of Clinical Genetics, Section Community Genetics
and EMGO Institute for Health and Care Research, VU University
Medical Center, Amsterdam, The Netherlands

3 Department of Clinical Genetics, Great Ormond Street Hospital,
London, UK

4 Laboratory for Molecular Genetics, Institute of Molecular

Genetics and Genetic Engineering, University of Belgrade,
Belgrade, Serbia

5 UMR 1027, Inserm, Faculté de médecine Université Toulouse 3,
Paul Sabatier, Toulouse, France

6 Centre for Biomedical Ethics and Law, Department of Public
Health and Primary Care, Leuven Institute for Genomics and
Society, KU Leuven, Kapucijnenvoer 35 Box 7001, 3000
Leuven, Belgium

7 Department of Health, Ethics and Society, Research Schools
CAPHRI and GROW, Maastricht University, Maastricht, The
Netherlands

1
2
3
4
5
6
7
8
9
0

relatively affordable price [1]. These attributes make
CRISPR-Cas9 an extremely useful and powerful tool that
can (and has) been used in research in order to alter the
genes in cells from a large range of different organisms,
including plants, non-human animals and microorganisms,
as well as in human cells [2]. Ultimately, CRISPR-Cas9 is
becoming increasingly available to a larger number of sci-
entists, who have used it, or intend to use it for a myriad of
reasons in many different research domains. When such
powerful and potentially disruptive technologies or tools
(begin to) show a tendency to become widely used, it is
common for debate and discussion to erupt. Germane to this
debate is the fact that with the advent of CRISPR-Cas9 and
other similar tools (e.g., CRISPR Cpf1), the possibility of
using the technique of gene editing in a potentially safe and
effective manner in humans—whether for somatic or germ
line/heritable1gene editing—has become feasible in the near
to medium future.

With some clinical trials underway, somatic genetic
editing for therapeutic purposes is certainly much closer to
being offered in the clinic. For example, several clinical
trials on HIV are ongoing [3, 4]; in 2015 an infant with
leukaemia was treated with modified immunes cells (using
TALENs) from a healthy donor [5]. Moreover, in the
autumn of 2016, a Chinese group became ‘the first to inject
a person with cells that contain genes edited using the
CRISPR-Cas9 technique’ within the context of a clinical
trial for aggressive lung cancer [6]. With such tools, gene
editing is being touted as a feasible approach to treat or even
cure certain single-gene diseases such as beta-thalassaemia
and sickle-cell disease through somatic gene editing [3].

Beyond somatic cell gene editing, there is also discussion
that through the manipulation of germ line cells or embryos,
gene editing could be used to trans-generationally ‘correct’ or
avoid single-gene disorders entirely. Notably, (ethical) con-
cerns about heritable gene editing in humans were heigh-
tened when in April 2015, a group at Sun Yat-sen University
in Guangzhou, China, led by Dr. Junjiu Huang reported they
had successfully used gene editing in human embryos [7].
They used CRISPR-Cas9 to modify the beta-globin gene in
non-viable (triplonuclear) spare embryos from in vitro ferti-
lity treatments. The authors concluded that while the
experiments were successful overall, it is difficult to predict
all the intended and unintended outcomes of gene editing in
embryos (e.g., mosaicism, off-target events) and that ‘clinical
applications of the CRISPR-Cas9 system may be premature
at this stage’ [7]. Partly in anticipation/response to these
experiments and to the increasing use of CRISPR-Cas9 in

many different areas, a number of articles were published [2,
8–14] and meetings were organized [9, 10, 15–17] in order to
further discuss the scientific, ethical, legal, policy and social
issues of gene editing, particularly regarding heritable human
gene editing and the responsible way forward.

Internationally, some first position papers on human gene
editing were published in 2015 and 2016. Interestingly, these
different recommendations and statements do not entirely
concur with one another. The United Nations Educational,
Scientific and Cultural Organisation (UNESCO) called for a
temporary ban on any use of germ line gene editing [18]. The
Society for Developmental Biology ‘supports a voluntary
moratorium by members of the scientific community on all
manipulation of pre- implantation human embryos by gen-
ome editing’ [19]. The Washington Summit (2015) organi-
zers (National Academy of Sciences, the U.S. National
Academy of Medicine, the Chinese Academy of Sciences
and the U.K.’s Royal Society) recommended against any use
of it in the clinic at present [17] and specified that with
increasing scientific knowledge and advances, this stance
‘should be revisited on regular basis’ [17]. Indeed, this was
done, to some extent, in a follow-up report by the US
National Academy of Sciences and National Academy of
Medicine, in which the tone of the recommendations appear
much more open towards allowing germ line modifications in
the clinic [20, 21]. Meanwhile, the ‘Hinxton group’ also
stated that gene editing ‘is not sufficiently developed to
consider human genome editing for clinical reproductive
purposes at this time’ [22] and they proposed a set of general
recommendations to move the science of gene editing ahead
in an established and accepted regulatory framework. Despite
these differences, at least two arguments are consistent
throughout these guidance documents: (1) the recognition of
the need for further research regarding the risks and benefits;
and (2) the recognition of the need for on-going discussion
and/or education involving a wide range of stakeholders
(including lay publics) regarding the potential clinical use
and ethical and societal issues and impacts of heritable gene
editing. It should be noted, however, that in the 2017
National Academies of Science, and of Medicine Report, the
role of public engagement (PE) and dialogue was presented
within the context of having to discuss the use of gene
editing for enhancement vs. therapy (rather than somatic vs.
heritable gene editing, which was the case in the 2015 sum-
mit report) [20, 21].

Although many stakeholders, including scientists, clin-
icians and patients are enthusiastic about the present and
potential future applications of these more efficient tools in
both the research and clinical contexts, there are also
important concerns about moving forward with gene editing
technologies for clinical use in humans, and to some extent,
for use in the laboratory as well. As we have learned from
other ethically sensitive areas in the field of genetics and

1 In this category, we include the editing of germ line cells, or
embryonic cells, or even somatic cells that are edited and promoted to
then become germ line cells in such a way that the alterations would be
heritable.

2 H. C. Howard et al.

genomics, such as newborn screening, reproductive genet-
ics or return of results, normative positions held by different
stakeholders may be dissimilar and even completely
incompatible. This might be influenced by various factors,
such as commercial pressure, a technological imperative,
ideological or political views, or personal values. Further-
more, it is clear that associated values often differ between
different stakeholder groups, different cultures and coun-
tries (e.g., where some may be more/less liberal), making
widespread or global agreement on such criteria very dif-
ficult, if not impossible to reach [23, 24].

From this perspective, it was important to study the
opportunities and challenges created by the use of gene
editing (with CRISPR-Cas9 and other similar tools) within
the Public and Professional Policy Committee (PPPC)2of
the European Society of Human Genetics (ESHG; https://
www.eshg.org/pppc.0.html). Our committee advances that
ESHG members and related stakeholders should be aware
of, and if possible, take part in the current debates sur-
rounding gene editing. Although not all genetics researchers
will necessarily use gene editing in their research, and while
gene editing as a potential treatment strategy, may appear,
initially, somewhat separate from the diagnostics-focused
present day Genetics Clinic, we believe that these stake-
holders have an important role to play in the discussions
around the development of these tools. For one, their
expertise in the science of genetics and in dealing with
patients with genetic diseases makes them a rare set of
stakeholders who are particularly well placed to not only
understand the molecular aspects and critically assess the
scientific discourse, but also understand current clinic/hos-
pital/health system resources, as well as human/patient
needs. Furthermore, in more practical terms, one could
consider that clinical genetics laboratories could be
involved in the genome sequencing needed to verify for off-
target events in somatic gene editing; and that clinical
geneticists and/or genetic counsellors could be involved in
some way in the offer of such treatment, especially in any
counselling related to the genetic condition for which
treatment is sought.

The PPPC is an interdisciplinary group of clinicians and
researchers with backgrounds in different fields of expertise
including Genetics, Health Law, Bioethics, Philosophy,
Sociology, Health Policy, Psychology, as well as Health
Economics. As a first step, a sub-committee was assigned
the task to specifically study the subject of gene editing
(including attending international meetings on the subject)
and report back to the remaining members. Subsequently,
all PPPC members contributed to a collective discussion
during the January 2016 PPPC meeting in Zaandam, The

Netherlands (15–16 January 2016). At this meeting, a
decision was reached to develop an article outlining the
main areas that need to be addressed in order to proceed
responsibly with human gene editing, including a review of
the critical issues for a multidisciplinary audience and the
formulation of crucial questions that require answers as we
move forward. A first draft of the article was developed by
the sub-committee. This draft was further discussed during
the 2016 ESHG annual meeting in Barcelona (21–24 May
2016). A second draft was developed and sent out for
comments by all PPPC members and a final draft of the
article was concluded based on these comments. Although
the work herein acts as guidance for further discussion,
reflection and research, the ESHG will be publishing
separate recommendations on germ line gene editing
(accepted during the 2017 annual meeting in Copenhagen,
Denmark).

In the context of the ongoing discussion and debate
surrounding gene editing, we present herein three crucial
areas that merit the most attention at this stage in the
responsible development and use of these gene editing
technologies, particularly for uses that directly or indirectly
affect humans:

1. Conducting careful scientific research to build an
evidence base.

2. Conducting ethical, legal and social issues (ELSI)
research.

3. Conducting meaningful stakeholder engagement,
education, and dialogue (SEED).

Although the main focus of this discussion article is on
the use of gene editing in humans (or in human cells) in
research and in the clinic for both somatic and heritable
gene editing, we also briefly mention the use of gene editing
in non-humans as this will also affect humans indirectly.

Conduct ongoing responsible scientific
research to build a solid evidence base

The benefits, as well as risks and negative impacts
encountered when conducting gene editing in any research
context should be adequately monitored and information
about these should be made readily available. Particular
attention should be paid to the dissemination of the infor-
mation by reporting and/or publishing both the ‘successful’
and ‘unsuccessful’ experiments including the benefits and
risks involved in experiments using gene editing in both
human and non-human cells and organisms (Table 1).

An evidence base regarding actual (and potential) health
risks and benefits relevant to the use of gene editing in the
human context still needs to be built. Therefore, a discus-
sion needs to be held regarding what type of monitoring,

2 This group studies salient ethical, legal, social, policy and economic
aspects relating to genetics and genomics.

ELSI of gene editing 3

reporting and potential proactive search for any physically
based risks and benefits should be conducted by researchers
using gene editing. Hereby, various questions emerge: are
the current expectations and practices of sharing the results
of academic and commercial research adequate for the
current and future field of gene editing? Should there be a
specific system established for the (systematic) monitoring
of some types of basic and (pre-) clinical research? If so,
which stakeholders/agencies should or could be responsible
for this? How could or should an informative long-term
medical surveillance of human patients be organized? Fol-
lowing treatment, would patients be obliged to commit to
lifelong follow-up? And, if relevant, how could long-term
consequences be monitored for future generations? For
example, if heritable gene editing was allowed, from
logistical and ELSI perspectives, there would be many
challenges in attempting to ensure that the initial patients (in
whom gene editing was conducted), as well as their off-
spring would report for some form of follow-up medical
check-ups to assess the full impact of gene editing on future
generations while still respecting these individuals’
autonomy.

Although the availability of results and potential mon-
itoring are especially important in a biomedical context for
all experiments and assays conducted in human cells, and
especially in any ex vivo or in vivo trials with humans,
relevant and useful information (to the human context and/
or affecting humans) can also be gleaned from the results of
experiments with non-human animals and even plants.
Furthermore, as clearly explained by Caplan et al. [2], gene
editing in insects, plants and non-human animals are cur-
rently taking place and may have very concrete and
important impacts on human health long before any gene
editing experiments are used in any regular way in the
health-care setting. As such, while keeping a focus on
human use, there should also be monitoring of the results in
non-human and non-model organism experiments and
potential applications [2]. Effects might include change of
the ecosystem, of microbial environment, (including the
microbiome, of parasites and zoonosis, which can involve
new combinations with some disappearing, and/or new
unexpected ones appearing), change to vegetation, which
has a reflection on our vegetal food and on animals’ food
and natural niche [25]. All this will have an impact on the
environment, and consequently on organisms (including
humans) who are exposed to this altered environment,
hence the monitoring of risks and benefits is very important.
Especially with gene editing of organisms for human con-
sumption (in essence, genetically modified organisms), it
will be important to note that the absence of obvious harms
does not mean that there are no harms. Proper studies must
be conducted and information regarding these should be
made readily available.Ta

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4 H. C. Howard et al.

Ongoing reflection, research and dialogue
on the ELSI of gene editing as it pertains to
humans

Research on the ELSI and impacts of human gene editing
should be conducted in tandem with the basic scientific

research, as well as with any implementations of gene
editing in the clinic. Appropriate resources and priority
should be granted to support and promote ELSI research; it
should be performed unabated, in a meaningful way and
by individuals from a diverse range of disciplines
(Table 2).

Table 2 Example of questions needed to be addressed for the ethical, legal, and social issues research (ELSI) of gene editing

Area Example of questions

ELSI of somatic cell gene
editing

Does the current legal framework need amendments/additions to address somatic gene editing? If so, who will
further shape the legal framework for somatic gene editing?

Do present clinical trial principles and protocols suffice?

How will trials in somatic gene editing be conducted and evaluated?

Do we need particular protection or status for patients in such trials?

What procedures will be instilled for patients receiving such treatments (e.g., consent, genetic counselling,
follow-up monitoring)?

To what extent will commercial companies be able to, or be allowed to offer, potentially upon consumer
request, treatments based on techniques where so much uncertainty regarding harms remains?

Which health-care professionals will be involved in the provision of somatic gene therapy and the care of
patients who undergo such treatments?

How will we ensure fair access to such technology?

How will we ensure that the use is driven by need and not the technological imperative?

Who will decide on roles and responsibilities in this novel context?

Based on what criteria will the eligible diseases/populations to be treated be chosen?

How can we ensure that research funds are allocated to ELSI research proportional (in some way) to the amount
of research on gene editing.

ELSI of heritable gene editing

Should gene editing of human germ line cells, gametes and embryos be allowed in basic research—for the
further understanding of human biology (e.g., human development) and without the intention of being used for
creating modified human life?

Should gene editing of germ line cells, gametes or embryos or any other cell that results in a heritable alteration
be allowed in humans in a clinical setting?

What, if any principles or reasoning would justify the use of hereditary gene editing in humans in a clinical
context given the current ban on such techniques in many jurisdictions?

Why should we consider using heritable gene editing in the clinic if there are alternative ways for couples to
have healthy (biologically related) children? Who will decide? Based on what criteria?

Should we first understand the risks and benefits of somatic gene editing before even seriously considering
heritable gene editing?

What are the roles, and responsibilities of different actors in these decisions?

How do commercial incentives and the technological imperative play a role in these decisions?

If we do entertain its use, what, if any criteria, will be safe enough according to different stakeholders
(scientists, ethicists, clinicians, policy makers, patients, general public) for it to be legitimate to consider using
gene editing for reproductive use? Who will set this safety threshold and based on what risk/benefit
calculations?

If heritable gene editing was allowed, how would the fact that for the first time, a human (scientist or clinician)
would be directly editing the nuclear DNA of another human in a heritable way cause some form of segregation
of types of humans? Creators and the created?

If ever allowed, should heritable human gene editing be permitted only for specific medical purposes with a
particular high chance of developing a disease (e.g., only when parents have a-near-100% risk of having a child
affected with a serious disorder), and if so, would it matter if the risk is not 100%, but (much) lower?

How can we, or should we define/demarcate medical reasons from enhancement? And, as was posed above for
the use in somatic cells, for what medical conditions will gene editing be considered appropriate for use? What
will the criteria be and who will decide?

ELSI of gene editing 5

Ongoing research, reflection and dialogue should address
all ELSI3salient to gene editing. With respect to gene
editing in humans, both somatic and germ line/heritable
embryonic gene editing contexts should be addressed. As
stated above, we should also study the ELSI of gene editing
in non-human and non experimental/model organisms,
including issues surrounding the potential (legal and logis-
tical related to implementation) confusions surrounding the
use of the terms genetically modified organisms vs. the term
gene-edited organisms.

Somatic gene editing

Although somatic gene editing is not free from ethical, legal
and social implications—it is, in many respects, similar to
more traditional ‘gene therapy’ approaches in humans—it
has been suggested that in many cases, the use of somatic
gene editing does not challenge existing ethical, legal and
social frameworks as much as heritable gene editing.
However, as with any new experimental therapeutic, the
unknowns still outweigh what is known and issues of risk
assessment and safety, risk/benefit calculation, patient
monitoring (potentially for long periods), reimbursement,
equity in access to new therapies and the potential for the
unjustified draining of resources from more pressing (albeit
less novel) therapies, particular protection for vulnerable
populations (e.g., fetuses, children (lacking competencies)),
and informed consent remain important to study further
[26].

Furthermore, as with any new (disruptive) technology or
application, there often remains a gap to be filled between
the setting of abstract principles or guidelines and how to
apply these in practice. Indeed, important questions and
uncertainties surrounding somatic gene editing both in
research and in the clinic remain, including, but not limited
to: do the established (national and international) legal and
regulatory frameworks (e.g., Regulation (EC) no. 1394/
2007 on advanced therapy medicinal products) need further
shaping/revisions to appropriately address somatic gene
editing (including not just issues with the products per se

but also for issues related to potential health tourism)? And
if so, how would this best be accomplished? Do present
clinical trial principles and protocols suffice? How exactly
will trials in somatic gene editing be conducted and eval-
uated? Do we need particular protection or status for
patients in such trials? What procedures will be instilled for
patients receiving such treatments (e.g., consent, genetic
counselling, follow-up monitoring)? Furthermore, to what
extent will commercial companies be able to, or be allowed
to offer, potentially upon consumer request, treatments
based on techniques where so much uncertainty regarding
harms remains? Importantly, which health-care profes-
sionals will be involved in the provision of somatic gene
therapy and the care of patients who undergo such treat-
ments? Who will decide on roles and responsibilities in this
novel context? And, based on what criteria will the eligible
diseases/populations to be treated be chosen? Indeed, these
questions can also all be applied to the context of heritable
gene editing, which is discussed below.

Germ line/heritable gene editing

With respect to germ line or heritable gene editing in
humans, the ELSI are more challenging than for somatic
gene editing, yet they are not all new per se either. Some of
these previously discussed concerns include, but are not
limited to: issues addressing sanctity of human life, and
respect for human dignity, the moral status of the human
embryo, individual autonomy, respect and protection for
vulnerable persons, respect for cultural and biological
diversity and pluralism, disability rights, protection of
future generations, equitable access to new technologies and
health care, the potential reduction of human genetic var-
iation, stakeholder roles and responsibilities in decision
making, as well as how to conduct ‘globally responsible’
science [16, 2, 11, 18]. Discussions and debates over some
of these topics have been held numerous times in the last
three decades, especially within the context of in vitro fer-
tilization, transgenic animals, cloning, pre-implantation
genetic diagnosis (PGD), research with stem cells and
induced pluripotent stem cells, as well as related to the large
scope of discussion around ‘enhancement’ [13]. Although it
is important to identify and reflect on more general ELSI
linked with heritable gene editing and these different con-
texts, it is also vital to reflect on the ELSI that may be
(more) specific to this novel approach. For example, would
the fact that for the first time a human (scientist or clinician)
would be directly editing the nuclear DNA of another
human in a heritable way cause some form of segregation of
types of humans? Creators and the created? [27] Clearly, we
need time for additional reflection and discussion on such
topics. Distinguishing the ELSI between different yet rela-
ted contexts will allow for a deeper understanding of the

3 Herein, the terms ‘ethical’, ‘legal’ and ‘social’ are used in a broad
sense, where, for example, issues such as economic evaluations, public
health prioritization and other related areas would also be included.
Indeed the first goal of ‘SEED’ (see below) is also, to some extent, part
of ELSI research, however, given the paucity of meaningful PE in the
past, combined with strong consensus regarding the current need and
importance of such activities, we have chosen to highlight it sepa-
rately. We also wish to stress the difference between academic ELSI
research and the work of ethics review committees. Although both deal
with ethical and legal issues, the former has as a main goal to advance
research and does not act as a policing body, nor does it have an
agenda per se. Furthemore, ELSI research does not only identify issues
to be addressed but also works with scientists and policy makers to
address the issues responsibly.

6 H. C. Howard et al.

issues and the rationale behind their (un)acceptability by
different stakeholders.

A major contextual difference in the current discussions
regarding germ line/heritable gene editing is that we have
never been so close to having the technology to perform it
in humans in a potentially safe and effective manner.
Hence, as we move closer to this technical possibility and as
we work out the scientific issues of efficiency and safety,
the discussions orient themselves increasingly towards the
ELSI regarding whether or not we want to even use heri-
table gene editing in a laboratory or clinical setting, and if
so, how we want it to be used, by whom and based on
which criteria? This includes, but is not limited to the fol-
lowing questions: should gene editing of human germ line
cells, gametes and embryos be allowed in basic research—
for the further understanding of human biology (e.g., human
development) and without the intention of being used for
creating modified human life? Some jurisdictions, such as
the UK, have already answered this question, and are
allowing this technique in the research setting in human
cells in vitro (they will not be placed in a human body, the
research will only involve studying the human embryos
outside of the body) whereby researchers need to apply for
permission to conduct such research. Some believe that
allowing this will inevitably lead to the technology being
used in the clinic (the so-called ‘slippery slope’ argument).
This, then, brings us to the question at the centre of the
debate: should gene editing of germ line cells, gametes or
embryos or any other cell that results in a heritable altera-
tion be allowed in humans in a clinical setting? Germane to
this issue is another vital question: what, if any, principles
or reasoning would justify the use of hereditary gene editing
in humans in a clinical context given the current ban on
such techniques in many jurisdictions? The new EU clinical
trial Regulation (536/2014 Art 90 al.2.) does not allow germ
line modification in humans. Should there be leeway for
reconsidering this ban in the future in view of the possible
benefits of therapeutic germ line gene editing? Should we
first understand the risks and benefits of somatic gene
editing before even seriously considering heritable gene
editing? If we consider that it could be used in some
situations, should we only consider using germ line gene
editing in the clinic if there are absolutely no other alter-
natives? Should already established and potentially safer4

reproductive alternatives, like PGD, be the approaches of
choice before even considering germ line gene editing? If
we do entertain its use, what, if any criteria, will be safe
enough according to different stakeholders (scientists,
ethicists, clinicians, policy makers, patients, general public)
for it to be legitimate to consider using gene editing for

reproductive use? Who will set this safety threshold and
based on what risk/benefit calculations? Furthermore, if
ever allowed, should heritable human gene editing be per-
mitted only for specific medical purposes with a particular
high chance of developing a disease (e.g., only when par-
ents have a-near-100% risk of having a child affected with a
serious disorder), and if so, would it matter if the risk is not
100%, but (much) lower? In addition, how can we, or
should we define/demarcate medical reasons from
enhancement? And, as was posed above for the use in
somatic cells, for what medical conditions will gene editing
be considered appropriate for use? What will the criteria be
and who will decide?

Taking a step back and looking at the issues from a more
general perspective, such ELSI research and reflection will
need to address, among others, questions that fall under the
following themes:

the balance of risks and benefits for individual patients
and also for the larger community and ecosystem as a
whole;
the ethical, governance and legislative frameworks;
the motivations and interests ‘pushing’ gene editing to be
used;
the roles and responsibilities of different stakeholders in
ensuring the ethically acceptable use of gene editing,
including making sure that every stakeholder voice is
heard;
the commercial presence, influence, and impact on (the
use of) gene editing;
the rationale behind the allocation of resources for health
care and research and if and which kind of shift might be
expected with the new technologies on the rise.

Additional overarching issues relating to ELSI include
the need to take a historical perspective and consider pre-
vious attempts to deal with genetic technologies and what or
how we can learn from these; the need to consider how
group actors could or should accept a shared global
responsibility when it comes to the governance of gene
editing; the potential eugenic tendencies related to new
technologies used to eliminate disease phenotypes; the
responsibility of current society for future generations; the
way different stakeholders may perceive and desire to
eliminate (genetic) risk and/or uncertainty by using new
technologies such as gene editing; and the potential role(s)
different stakeholders, including ‘experts’, may inad-
vertently play in propagating a false sense of control over
human health.

Although the human context is where much of the atten-
tion currently resides, and is indeed, the focus of this article,
as mentioned above, we also stress that many concerns and
ELSI also stem from the use of gene editing in non-human
organisms (plants, insects and microorganisms), the study of

4 It is important to note that despite attempts at addressing these
issues, even for technologies such as PGD [28].

ELSI of gene editing 7

which, could inform the human context. More importantly,
given that the use of gene editing in these organisms is
currently taking place in laboratories and, if released, some of
these gene-edited organisms could have a large impact on the
environment and society [2], the ELSI of gene editing in non-
human organisms should also be seriously addressed. In this
respect, the current debates over definitions and whether
plants and non-human animals in which gene editing is
performed are considered (legally) genetically modified
organisms (GMOs) are particularly important to consider;
indeed, this legal stance may be a misleading way to describe
the scientific differences in practice. Moreover, the manip-
ulation of definitions may also be used to circumvent the
negative press and opinions surrounding GMOs in Europe.
Last, but not least, the use of gene editing for the creation of
biologic weapons is a possibility that must be discussed and
adequately managed [2].

In order to ensure that the appropriate ELSI research is
conducted to answer these myriad questions, ELSI
researchers must ensure adequate understanding of scien-
tific facts and possibilities of gene editing, ensure appro-
priate use of robust methods [29] to answer specific ELSI
questions, as well as learn from previous research on related
themes such as (traditional) gene therapy, reproductive
technologies, and GMOs. Furthermore, funding will have to
be prioritized for ELSI research. National and European
funding agencies should ensure that ELSI funding is given
in certain proportion to how much gene editing research is
being conducted in the laboratory and (pre) clinical domain.
In practice, this will mean ensuring that there are adequate
review panels for stand-alone ELSI grants, which do not
usually fall within any one traditional academic field (e.g.,
philosophy, law or social sciences). The requirement of

including ELSI work packages within science grants may
also be useful if such work packages are conducted by ELSI
experts (and this is verified by the funding agencies), that
they are given enough budget to conduct research and not
only offer services, and that the ELSI work package is not
co-opted by the science agenda. Spending money on ELSI
research has already allowed for the information to be used
in more applied ways. Among others, ELSI research has
contributed to helping individual researchers understand
what kind of research they are (not) allowed to do in certain
countries or regions; helped to design appropriate consent
forms for research and clinic; and has helped inform policy
decisions.

As ELSI are identified, studied and discussed, it will be
of utmost importance to communicate these with as many
publics as relevant and possible in a clear and comprehen-
sive way so that the largest number of different stakeholders
can understand and engage in a discussion about these
issues. With respect to engaging non-academic and non-
expert audiences in meaningful dialogue, the challenges are
greater. Yet, as this is a vital element of conducting science
and preparing clinical applications in a responsible manner
and stretches beyond the academic focus of ELSI we pro-
pose to distinguish a third domain dedicated to such sta-
keholder engagement, education and dialogue (SEED)
described below.

Stakeholder engagement, education and
dialogue (SEED)

To deliver socially responsible research (and health care),
an ongoing robust and meaningful multidisciplinary

Table 3 Examples of questions
to be answered regarding
stakeholder, engagement,
education and dialogue (SEED)
for gene editing

Example of questions

Planting SEEDs for
gene editing

What are the roles and responsibilities of different stakeholders in setting up and
maintaining responsible engagement, education and dialogue?

What will, and what should be the role of scientists and other academics in this
type of popular media communications, and engagement activties?

Since PE can have different purposes, before each activity, we must consider:
what are our goals? And, what method of engagement will best meet these goals?

How will the multitude of voices we want to involve in PE be ‘weighted’ against
each other?

What role will different stakeholders’ inputs and ‘preferences’ play in the debate
and in the decision-making process?

How do we make sure that all voices are heard?

How will these voices be weighed and considered, if at all, in policy making?

How can we ensure that public education will not be reduced to a token work
package in science grants and/or to campaigns that try to convince for or against
gene editing?

How can we make sure that such public education and engagement is accessible
to all, including in countries that may currently not have the resources to take on
such ‘SEED’ activities?

8 H. C. Howard et al.

dialogue among a diverse group of stakeholders, including
lay publics, should be initiated and maintained to discuss
scientific and ethically relevant issues related to gene edit-
ing. Publics must not only be asked to engage in the dis-
cussion, but they should also be given proper information
and education regarding the known facts, as well as the
uncertainties regarding the use of gene editing in research
and in the clinic. In this way, the two focal areas described
above will feed into these SEED goals. Stakeholders should
also be given the tools to be able to reflect on the ethically
relevant issues in order to help informed decision making.
Appropriate resources and prioritization should be granted
to support and promote SEED (Table 3).

As mentioned in the introduction, the statements
addressing gene editing published by different groups and
organizations have highlighted the need for an ongoing
discussion about human gene editing among all stake-
holders, including experts, and the general public(s) [8, 9,
17], In calling for an ‘ongoing international forum to discuss
the potential clinical uses of gene editing’, the organizing
committee of the International Summit on Human Gene
Editing stated that

‘The forum should be inclusive among nations and
engage a wide range of perspectives and expertise –
including from biomedical scientists, social scientists,
ethicists, health care providers, patients and their
families, people with disabilities, policymakers,
regulators, research funders, faith leaders, public
interest advocates, industry representatives, and mem-
bers of the general public’ [17].

Hence, this implies that not only should different
expertise be represented in this ongoing discussion, but lay
publics should also be included. For this to be a meaningful
and impactful endeavour, all stakeholders involved should
be appropriately informed and educated about the basic
science and possibilities of gene editing. Academic/profes-
sional silos, differences in language, definitions, approaches
and general lack of experience with multi- and inter-
disciplinary work are all barriers to involving different
expert stakeholders in meaningful exchange and dialogue.
Some first constructive steps have included the posting
online of meeting and conference presentations on gene
editing (e.g., the 3 days of the Washington Summit (http://
www.nationalacademies.org/gene-editing/Gene-Edit-
Summit/index.htm.), Eurordis webinars and meetings aimed
at informing patients, http://www.eurordis.org/tv). Beyond
this, one important barrier to having a truly meaningful and
inclusive multidisciplinary discussion about new technolo-
gies is the (potential) lack of knowledge and/or under-
standing of different publics [30]. Indeed, it is not
reasonable for experts to expect that all concerned stake-
holders are properly informed about the science and/or the

social and ethical issues, which are important requisites for
having meaningful and productive conversations about
responsible gene editing. Furthermore, a pitfall we must
avoid is using PE with the aim of persuading or gaining
acceptance of technologies instead of ‘true participation’
[31] and as a means to allow for supporting informed
opinions.

Another critical issue is the role and influence of dif-
ferent stakeholders, including the media, in educating and
informing the public. What are the roles and responsibilities
of different stakeholders in setting up and maintaining
responsible engagement and dialogue? What will, and what
should be the role of scientists in popular media commu-
nications and other SEED activities? Where will the funding
for these activities come from? Financial and temporal
resources will have to be reserved for such SEED regarding
gene editing. Resources will also be needed to conduct
further research on the best way to engage different publics
and to study whether engagement strategies are successful.

Moreover, before engaging different publics and asking
for their feedback, whichever stakeholders take on this task
must seriously reflect on the precise reasons for which lay
publics are being engaged. What is the goal? And, what
method of engagement will best meet these goals? There is
also a need for honest evaluation of engagement efforts to
report on their impacts and outcomes. Indeed, the purposes
of PE in science can vary widely, including, among others,
informing, consulting and/or collaborating; [32] clearly
each of these implies different levels of participation by
publics, and by extension, different levels of influence on a
topic. Importantly, there are a long list of questions that also
need to be answered for PE (Table 3), including but not
limited to how different voices will be weighed and if or
how they will be used in any policy or decision making.

The value of PE in the form of public dialogue in a
democratic society, (and we would specify its contribution
to responsible science) is very well summarized by Mohr
and Raman (2012) in a perspective piece on the UK Stem
Cell Dialogue: [31]

‘The value of public dialogue in a democratic society
is twofold. From a normative perspective, the process
of PE is in itself a good thing in that the public should
be consulted on decisions in which they have a stake.
From a substantive standpoint, PE generates manifold
perspectives, visions, and values that are relevant to
the science and technologies in question, and could
potentially lead to more socially robust outcomes
(which may differ from the outcomes envisaged by
sponsors or scientists)’ [31].

Particularly for the purposes of gene editing, we consider
SEED a way to try to ensure that decisions on a subject that
is filled with uncertainties, and could have important

ELSI of gene editing 9

implications for society for generations to come, is not left
in the hands of a few. We want to underline the need for: lay
publics to be informed to support transparency; lay publics
to be educated to support autonomy and informed opinion/
decision making; different voices and concerns to be heard
and considered through ongoing dialogue to help ensure
that no one stakeholder group pursue their interests
unchecked. Although it is beyond the scope of this article to
go into any detail, it is important to take the time to learn
from past and ongoing engagement efforts in science in
general [32], as well as in biomedicine, including areas like
stem cell research [31] and genetics [30, 33]. For example,
we can learn about: how PE can generate value and impact
for a society, as well as how to conceive of and evaluate a
PE programme [32]; the nuances around ‘representative
samples’ and if they really are representative [31]; how
letting citizens be the ‘architects’ rather than just participants
of engagement (activities) could help to ward against the
generation of ‘predetermined outcomes’ [31]; the utility of
deliberative PE to ‘offer useful information to policy makers
[30]. Given all the different reasons for PE, and given the
higher standards expected for PE in recent years [34] it is to
be expected that each PE activity will have to be adjusted
for the specific context. There are, also, useful tools for PE
from a European funded project called ‘PE2020, Public
Engagement Innovations for Horizon 2020’ [35], which has
as an aim to ‘to identify, analyse and refine innovative
public engagement (PE) tools and instruments for dynamic
governance in the field of Science in Society (SiS)’ [35].

As already mentioned above for ELSI research, funding
agencies will have to prioritize resources for these
SEED activities, and the strategies we outlined for ELSI,
could also apply for SEED.

Conclusion

In the midst of a plethora of debate over gene editing, dif-
ferent stakeholder views, preferences, agendas and mes-
sages, it is crucial to focus our limited resources, including
human resources, time and finances on the most important
areas that will enable and support the responsible use of
gene editing. We have identified the following three areas
that merit an equitable distribution of attention and resour-
ces in the immediate and medium-term future:

1. Conducting careful scientific research to build an
evidence base.

2. Conducting ELSI research.
3. Conducting meaningful stakeholder engagement,

education, and dialogue (SEED).

Indeed, one way to ensure that each of these three
important areas receive adequate financial support to

conduct the necessary work would be for international and
national funding agencies to announce specific funding calls
on gene editing. They could also encourage or require that
scientific projects focused on gene editing include ELSI and
SEED along with the scientific work packages. Further-
more, understandably, priorities need to be made with
respect to resource allocation in the biomedical sciences,
especially in such uncertain financial contexts, however, as
expressed at the World Science Forum in Budapest in
November 2011, we must ward against scarce funding
being funnelled to single disciplines since it is common
knowledge that much of the most valuable work is now
multidisciplinary [36]. Moreover, at such a time funding
entities must not ‘expel’ the social sciences ‘from the temple’
but rather, the hard sciences should ‘invite them in to help
public engagement’ [36].

Acknowledgements We thank all members of the Public and Profes-
sional Policy Committee of the ESHG for their valuable feedback and
generosity in discussions. Members of PPPC in 2015–2017 were
Caroline Benjamin, Pascal Borry, Angus Clarke, Martina Cornel,
Carla van El, Florence Fellmann, Francesca Forzano, Heidi Carmen
Howard, Hulya Kayserili, Bela Melegh, Alvaro Mendes, Markus
Perola, Dragica Radijkovic, Maria Soller, Emmanuelle Rial-Sebbag
and Guido de Wert. We also thank the anonymous reviewers for their
constructive comments, which have helped to improve the article. Part
of this work has been supported by the Swedish Foundation for
Humanities and Social Science under grant M13-0260:1, and the CHIP
ME COST Action IS1303.

Compliance with ethical standards

Conflict of interest The authors declare that they have no competing
interests.

Open Access This article is licensed under a Creative Commons
Attribution-NonCommercial-NoDerivatives 4.0 International License,
which permits any non-commercial use, sharing, distribution and
reproduction in any medium or format, as long as you give appropriate
credit to the original author(s) and the source, and provide a link to the
Creative Commons license. You do not have permission under this
license to share adapted material derived from this article or parts of it.
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statutory regulation or exceeds the permitted use, you will need to
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of this license, visit http://creativecommons.org/licenses/by-nc-nd/4.0/.

References

1. Dance A. Core concept: CRISPR gene editing. Proc Natl Acad Sci
USA 2015;112:6245–6.

2. Caplan AL, Parent B, Shen M, Plunkett C. No time to waste–the
ethical challenges created by CRISPR. EMBO Rep.
2015;16:1421–6.

3. Maeder ML, Gersbach CA. Genome-editing technologies for gene
and cell therapy. Mol Ther. 2016;24:430–46.

10 H. C. Howard et al.

4. Reardon S. First CRISPR clinical trial gets green light from US
panel. Nature 2016; 22 June 2016.

5. Reardon S. Leukaemia success heralds wave of gene-editing
therapies. Nature 2015;527:146–7.

6. Cyranoski D. CRISPR gene-editing tested in a person for the first
time. Nature 2016;539:479.

7. Liang P, Xu Y, Zhang X, et al. CRISPR/Cas9-mediated gene
editing in human tripronuclear zygotes. Protein Cell
2015;6:363–72.

8. Lanphier E, Urnov F, Haecker SE, Werner M, Smolenski J. Don’t
edit the human germ line. Nature 2015;519:410–1.

9. Baltimore D, Berg P, Botchan M, et al. Biotechnology. A prudent
path forward for genomic engineering and germline gene mod-
ification. Science 2015;348:36–38.

10. Mathews DJ, Chan S, Donovan PJ, et al. CRISPR: a path through
the thicket. Nature 2015;527:159–61.

11. Bosley KS, Botchan M, Bredenoord AL, et al. CRISPR germline
engineering–the community speaks. Nat Biotechnol.
2015;33:478–86.

12. Addison C, Taylor-Alexander S. Gene editing: advising advice.
Science 2015;349:935.

13. Hurlbut JB. Limits of responsibility: genome editing, asilomar,
and the politics of deliberation. Hastings Cent Rep.
2015;45:11–14.

14. Shakespeare T. Gene editing: heed disability views. Nature
2015;527:446.

15. Ledford H. Where in the world could the first CRISPR baby be
born? Nature 2015;526:310–1.

16. Editorial N. After Asilomar. Nature 2015;526:293–4.
17. US National Academy of Sciences, US National Academy of

Medicine, Chinese Academy of Sciences, UK Royal Society. On
human gene editing: International Summit Statement. Washing-
ton, DC: 2015.

18. UNESCO: United Nations Educational Scientific and Cultural
Organization Report of the International Bioethics Committee
(IBC) on Updating its reflection on the Human Genome and
Human Rights. Paris: United Nations Educational, Scientific and
Cultural Organization, UNESCO, 2015.

19. Society for Developmental Biology: Position Statement from the
Society for Developmental Biology on Genomic Editing in
Human Embryos. 2015.

20. National Academy of Sciences, National Academy of Medicine:
Human Genome Editing: Science, Ethics, and Governance. 2017.

21. Baylis F. Human germline genome editing: an impressive sleight
of hand?: Impact Ethicsm Making a DIfference in Bioethics.
Halifax, Canada, 2017, Vol 2017.

22. Hinxton Group: statement on genome editing technologies and
human germline genetic modification, 2015.

23. Howard HC, Swinnen E, Douw K, et al. The ethical introduction
of genome-based information and technologies into public health.
Public Health Genomics 2013;16:100–9.

24. Weiner C, Beeson D, Billings PR, MD. Ethics, Genetic Tech-
nologies and Social Responsibility in the Twenty-first Century.
Berkeley: University of California; 2001.

25. Kevin E, Church G JL: “Gene Drives” And CRISPR Could
Revolutionize Ecosystem Management: Scientific American. https://
blogs.scientificamerican.com/guest-blog/gene-drives-and-crispr-
could-revolutionize-ecosystem-management/, 2014, Vol 2017.

26. Araki M, Ishii T. Providing appropriate risk information on gen-
ome editing for patients. Trends Biotechnol. 2016;34:86–90.

27. Somerville M. Debating the ethics of gene editing; in: Brown J
(ed): The180, 2015

28. Harper JC, Wilton L, Traeger-Synodinos J, et al. The ESHRE
PGD Consortium: 10 years of data collection. Hum Reprod
Update 2012;18:234–47.

29. Walker RL, Morrissey C. Bioethics methods in the ethical, legal,
and social implications of the human genome project literature.
Bioethics 2014;28:481–90.

30. O’Doherty KC, Burgess MM. Engaging the public on biobanks:
outcomes of the BC biobank deliberation. Public Health Geno-
mics 2009;12:203–15.

31. Mohr A, Raman S. Representing the public in public engagement:
the case of the 2008 UK Stem Cell Dialogue. PLoS Biol. 2012;10:
e1001418.

32. National Coordinating Centre for Public Engagement: examples
of successful public engagement: additional evidence submitted to
the Science and Technology Committee (Science Communica-
tion) by the National Coordinating Centre for Public Engagement
(NCCPE). 2016.

33. Resnik DB. Ethics of community engagement in field trials of
genetically modified mosquitoes. Dev World Bioeth 2017.

34. Stirling A. Opening up the politics of knowledge and power in
bioscience. PLoS Biol. 2012;10:e1001233.

35. PE2020 Public Engagement Innovations for Horizon 2020, 2016.
36. Macilwain C. Science’s attitudes must reflect a world in crisis.

Nature 2011;479:447.

ELSI of gene editing 11

© 2018. This work is published under
http://creativecommons.org/licenses/by-nc-nd/4.0/(the “License”).

Notwithstanding the ProQuest Terms and Conditions, you may use this
content in accordance with the terms of the License.

Cambridge Quarterly of Healthcare Ethics (2019), 28, 100–111.
© Cambridge University Press 2018.
doi:10.1017/S0963180118000439100

Articles

Let Us Assume That Gene Editing is Safe—
The Role of Safety Arguments in the
Gene Editing Debate

SØREN HOLM

Abstract: This paper provides an analysis of the statement, made in many papers and
reports on the use of gene editing in humans, that we should only use the technology
when it is safe. It provides an analysis of what the statement means in the context of
nonreproductive and reproductive gene editing and argues that the statement is inconsist-
ent with the philosophical commitments of some of the authors, who put it forward in
relation to reproductive uses of gene editing, specifically their commitment to Parfitian
nonidentity considerations and to a legal principle of reproductive liberty.

But, if that is true it raises a question about why the statement is made. What is its discur-
sive and rhetorical function? Five functions are suggested, some of which are more conten-
tious and problematic than others. It is argued that it is possible, perhaps even likely, that
the “only when it is safe” rider is part of a deliberate obfuscation aimed at hiding the full
implications of the arguments made about the ethics of gene editing and their underlying
philosophical justifications.

Keywords: gene editing; gene modification; gene therapy; harm; nonidentity problem;
principle of procreative beneficence; reproductive liberty; safety; wrong

As noted above, we do not believe that sufficient knowledge is available
to consider the use of genome editing for clinical reproductive purposes
at this time. However, we acknowledge that when all safety, efficacy, and
governance needs are met, there may be morally acceptable uses of this
technology in human reproduction, though further substantial discus-
sion and debate will be required as detailed below.1

Various groups, including ours, agree that numerous technical and safety
issues need to be addressed before genome-editing technologies could
feasibly be used in reproductive clinical applications.2

The clearest ethical concerns regarding current gene editing techniques is
that they are unsafe. The study by Huang and coauthors showed that
current gene editing techniques can lead to a large number of off-target
mutations. This could cause significant defects and disabilities in any
individuals born as the result of the research. While some research sug-
gests there are ways to edit genes that greatly reduce the number of
off-target mutations, . . . it would be highly unethical to bring modified
human embryos to term unless we were very confident that the tech-
nique could be used safely. The risk would simply not be justified by
any potential benefits.3

Translating germline modification into clinical trials and society requires
time, careful research (involving both the science and ethics) and public
deliberation. Broadly, I would propose two conditions for an ethical use

What Is “Safe Gene Editing”?

101

of germline engineering. First, there is a requirement for safety. First-in-
man use for germline modification is ethically challenging by nature,
particularly because the needed evidence to reliably predict risk and
benefit (testing in humans) is missing. This needs careful, long-term,
interdisciplinary research and sufficient evidence to make the leap from
bench to bedside.4

Introduction

In ethical debates about the introduction of new technologies into clinical use
it is often argued that we should not introduce them as long as they are not safe,
but that we should continue basic research and be ready to introduce them
when they are safe. In the meantime we should conduct the ethical analysis of
their various uses on the assumption that they are safe (or at least safe enough)
at the time when they are introduced (let us call this the “‘only when it is safe’
rider”). This discursive move has also been prominent in the debates about the
use of gene editing in humans. It is evidenced by the quotations above; and a
recent review of CRISPR Germline Ethics Statements found that of the 61 ethics
reports and statement identified almost all mentioned safety concerns and the
need for these to be overcome before gene editing can be routinely used in
humans.5

This paper will analyze three issues that are raised by this discursive move. It
will ask:

1) What do we mean by “safe,” i.e., what harms or wrongs are relevant?
2) Does “safe” have a different meaning in relation to technologies with repro-

ductive implications?
3) In the light of 1 & 2, what are the functions of the “only when gene editing is

safe” discursive move?

In the first part of the paper, the analysis of 1 and 2 will initially proceed while
bracketing “reproductive rights” and Parfitian “nonidentity” considerations. The
second part of the paper will reconsider 2 in the light of “reproductive rights” and
“nonidentity” arguments. The third part will analyze the implications of consider-
ations of procreative beneficence. And, the fourth and final part will analyze the
discursive and rhetorical functions of safety arguments in the bioethical debate
about gene modification.

Many official ethics reports concerning gene modification do not contain much
ethical argument and it would be problematic to hold them to a high standard of
argumentative rigor and philosophical understanding. The focus here will there-
fore be on the use of the “only when it is safe” rider in journal articles authored or
coauthored by philosophical bioethicists, where we can legitimately expect more
rigor and knowledge of the philosophical literature, including knowledge of the
authors’ own previous contributions to that literature.

This is not a paper about the ethics of gene editing. Gene editing technologies
enable us to perform a wide range of genetic modifications in humans and ani-
mals. Some of these modifications are ethically contentious, but an analysis of
which modifications are ethically acceptable and which are not is beyond the
scope of this paper.

Søren Holm

102

Safe Gene Modification

What does it mean to say that a particular gene modification is safe or not safe?
What harms, or potentially ethical or legal wrongs are involved?

It is clear that “safe” and “unsafe” should be read in a narrow technical sense
in this discourse. Safety is conceptualized as being about biological harms that
may befall the gene edited organism as a result of the editing, and not about
wider social or ethico-legal harms. This restricted use of “safe” is often not made
explicit, but it is implicit in distinguishing between safety and governance
needs,6 or in focusing on examples of biological lack of safety.7 Such a restrictive
account of safety may in itself be problematic, but a closer analysis of this issue
is outside the scope of this paper where we will accept the narrow biomedical
framing of safety.

If we perform gene editing in an organism, that organism can be harmed in vari-
ous ways (let us call these harm1, harm2, harm3, and harm4):

1) The gene editing may be intended to be harmful
2) The gene editing is technically inefficient and leads to mosaicism
3) The gene editing may be technically efficient (i.e., it makes the desired change

in the genome), but may nevertheless turn out to be harmful either on its
own, in combination with some other part of the organism’s (epi-)genome,
or in combination with some infection or environmental exposure, either
immediately or during the lifetime of the organism

4) Off-target genetic changes may be harmful

In relation to clinical use in humans, including the research leading up to clinical
use, we can discount harm1 and concentrate on harms2-4. No health care profes-
sional would perform gene editing with the express intention to harm. If we per-
form gene modification in humans it will always be in cases where we predict net
benefit from the intervention, but where harm may nevertheless occur.

The likelihood and magnitude of these harms can be estimated from research
evidence, and may be reducible by future research and development. It is, how-
ever, as with all technologies, unlikely that the risks of these harms occurring
can be removed completely. A recent example of a harm falling into category 3
above is the emerging evidence that gene editing is more efficient in cells that are
deficient in p53 function, and that this may lead to the selection of gene-edited
cells with increased risk of tumor formation because p53 is an important tumor
suppressor.8,9 It was only through research that this problem was identified, and
it is only through research that we may find ways to overcome it.

If the gene modification is taking place in vitro—outside of the body in cultured
cells—we may have technical procedures for eliminating the risks of harm2 and
harm4, by modifying cells and then testing and selecting only cells with the correct
gene modification, but in vivo gene modification is likely always to entail some
risk of these harms eventuating.

When is a particular gene modification safe enough to introduce in clinical prac-
tice? The simple answer for adults and children, in terms of harms, seems to be:
when we can be reasonably certain that

− >benefits harms 0,

What Is “Safe Gene Editing”?

103

i.e., when we are reasonably certain of obtaining net benefit from the intervention.
There are many in principle and practical questions about how we estimate the
benefits and harms quantitatively and about what we mean by “reasonably cer-
tain,” but resolving them is not relevant to the current analysis of the function of
the “only when it is safe” rider.

This threshold must be the right one for approval from a regulator for rou-
tine clinical use. A regulator could not allow an intervention to be marketed if
there were still significant concerns about its safety. But what if the condition
that is targeted by the gene modification is very severe, and there are patients
willing to take the gamble before we have sufficient evidence to determine the
risk/benefit ratio? Should such patients be allowed to use the gene modifica-
tion, and should researchers be allowed to offer it to patients as experimental
treatment? This question is not in principle different from similar questions
raised in relation to other therapeutic technologies and methods and discussed
in the literature on “right to try” and expedited/expanded access,10-12 and will
not be analyzed further here. It is, however, important to note that it raises
important questions not only about the limits, if any, of personal autonomy, but
also about the balance between personal liberty to choose and societal interests
in having safety and efficacy questions definitively answered through well-
planned research.

Reproductive Safety

Should we understand safety differently if the gene editing is either performed as
part of the creation of a child by means of a reproductive technology, or affects the
germ cells of the patient?

Bracketing the nonidentity problem and considerations of reproductive liberty,
it seems that there are no relevant differences between the reproductive context
and the nonreproductive context in relation to safety. If gene editing is an intended
part of a reproductive project, “being safe” again simply means that we are rea-
sonably certain that there is net benefit for the child that is the end goal of the
reproductive project.

In the directly reproductive context, in which the entity being edited is either a
gamete or an embryo, the decision of whether or not to use gene editing will be
taken by the people involved in the reproductive project. They choose for some-
one else, i.e., their future child, not for themselves, and the child cannot consent.
But all reproductive decisions by the reproducers are made without the consent of
the children, but if they chose a nonsafe gene modification intervention (i.e., one
where we are not reasonably certain that there is net benefit) they would unjustifi-
ably put the child at risk of harm.

In the indirectly reproductive context, in which germ cells are edited in vivo
either deliberately or as a side effect of some type of in vivo somatic gene editing,
similar considerations about safety and harm apply. We should not introduce
genetic modifications into germ cells unless the modifications are unlikely to lead
to net harm to any future child created using gametes generated by these sperm
cells. Or, if the gene editing is beneficial for the person in whom it is done, but
likely to be harmful to future children, then that harm and the reproductive lack
of safety should be an important consideration in determining whether to bring
the particular type of gene editing into clinical use.

Søren Holm

104

So, the interim conclusion so far is that safety matters both ethically and in rela-
tion to regulation, and it matters because we are at risk of causing significant harm
if we use unsafe gene editing. The “only when it is safe” rider therefore seems
perfectly justified.

Reproductive Safety Reconsidered

The reader who is steeped in the Anglo-American bioethical literature will by
now probably be screaming “but why, oh why have you ignored reproductive/
procreative liberty and the nonidentity problem in the analysis. Because of those
two considerations reproduction is different!”

And, it is true that reproduction is different, so different that there is consid-
erably less agreement about our ethical obligations in the procreative sphere
than there is about our general ethical obligations (see for instance the book-
length exchange between David Benatar and David Wasserman13). So, let us
take reproductive liberty and nonidentity seriously in our analysis of the
meaning of harm in relation to gene editing and imagine a situation where
a set of prospective parents wants to use gene editing as part of the process
leading to the creation of their next child. Do they do something wrong if they
use a gene modification intervention that is unsafe in one of the four ways
outlined above? We will here bracket the issue that this can only be done as
part of technically advanced assisted reproduction and therefore necessarily
involves a number of third parties who may or may not have independent ethical
importance.

The standard interpretation of the nonidentity problem is that if an action (or an
inaction) I is necessary for the coming into being of person P, then P cannot have
been wronged by I being performed, even if I leads to P being in a harmed state as
long as P’s state is not so bad that it constitutes “a life not worth living.”14 P has
not been wronged, because you cannot be wronged if, without the action causing
the putative wrong having been performed, you would not have existed. Some
proponents of nonidentity also claim that P cannot be harmed by I, since harm
should be understood counterfactually and because I is necessary for P coming
into being there is no possible world in which P exists, but in a different unharmed
state. Disentangling the many different possible interpretations of nonidentity
and its implications, and the many and varied highly counterintuitive conse-
quences that flow from it, is far beyond the scope of this paper. What is important
in the present context is that if we accept something like nonidentity, then there is
definitely no one who is wronged, and plausibly no one who is harmed, by the use
of an unsafe gene modification intervention in reproduction, and it is important
that this is accepted by some of those who put forward the “only when it is safe”
rider,15,16 whereas others are more equivocal.17 That there is no wrong done, and
plausibly no harm created, is true of the instant child, and by recursion true of
anyone who will ever carry the modification, given that the first act of modifica-
tion is a necessary condition for their existence. If the parents are doing something
ethically problematic, it is only in relation to the state of the world, if the world
would, counterfactually, have been better without the existence of P.

It might be argued that not all genetic modifications introduced by gene editing
are identity forming or identity changing. Some may, for instance correct a disposi-
tion to a condition with very late onset (e.g., Alzheimer’s disease) without changing

What Is “Safe Gene Editing”?

105

the identity of the embryo that is edited or the person it becomes. And, as a thought
experiment, we can conceive of gene editing of a part of the genome that has no
known functions at all. So, gene editing of an embryo does not necessarily or auto-
matically lead to nonidentity. There are many different conceptions of identity
at play when considering nonidentity. As I have argued in a previous paper in
CQ, nonidentity discussions can involve considerations of numerical, genetic,
phenotypical/physiognomical, psychological, and narrative/social identity, and
it is often important to be clear about what kind of identity we are discussing.18
However, in the present context, almost all conceptions of identity lead to the con-
clusion that the gene editing of embryos affects identity. Not because the actual
modification introduced necessarily affects identity, but because it is very unlikely
that the same unedited embryo would ever have been created, implanted, gestated,
and born.

In a context where gene editing is available to them, prospective parents will,
with their clinicians, plan the IVF + gene editing, and it is very unlikely that this
will happen at exactly the same time and in exactly the same way as it would
have happened if gene editing had not been available, i.e., it is highly unlikely
that exactly the same ova will be retrieved and fertilized by exactly the same
spermatozoon. The resulting child will therefore be numerically, genetically,
phenotypically, and psychologically different from the child which would have
existed (if any) if gene editing had not been available. The only type of identity
that might be preserved is an attenuated form of narrative or social identity. The
child would still have the same number in the birth order and might still have
the same sex, so could still be “the first son born to Jack and Jill”; but because of
the many nonidentities it is likely that its narrative would quite quickly and
quite substantially diverge from the narrative of the child brought into the world
without gene editing being available.

This brings us to reproductive liberty/freedom/autonomy. Reproductive lib-
erty can be understood either as a jurisprudential or an ethical principle or both.
The core of reproductive liberty is the claim that our reproductive choices are
strongly protected from outside interference, and that this strong protection is jus-
tified by the central importance of reproduction to the life plans and personal
identity of people. In his seminal first paper arguing for the importance of procre-
ative liberty, John Robertson writes:

Procreation is a complex activity that develops over time and involves
many disparate behaviors. The importance of procreation as a whole
derives from the genetic, biological, and social experiences that com-
prise it. Reproduction is a basic instinct that supplies societies with the
members who maintain and perpetuate the social order and who pro-
vide services for others. Reproduction also satisfies an individual’s
natural drive for sex and his or her continuity with nature and future
generations. It fulfills cultural norms and individual goals about a good
or fulfilled life, and many consider it the most important thing a person
does with his or her life.
Claims of procreative freedom logically extend to every aspect of repro-
duction: conception, gestation and labor, and childrearing.”19

A strong account of reproductive liberty entails that procreative acts are protected
from outside interference even in cases where the resulting child is harmed and

Søren Holm

106

where the people reproducing are acting in ways that are recognized even by them
as ethically wrong and ordinarily blameworthy.

Taken together, nonidentity and reproductive liberty lead to the conclusion that
almost no uses of genetic modification in reproduction can be deemed as unsafe
(because of nonidentity considerations), and if there are any unsafe uses, they can
nevertheless not be prohibited or interfered with in other ways if the procreating
parents want to use them (because of reproductive liberty). The only exceptions
are gene modifications that are so harmful that they lead to a “life not worth
living,” since creating such a life is a wrong in all circumstances and also a suffi-
cient justification for overriding even a strong right to reproductive liberty.

Procreative Beneficence and the Ethics of Reproduction

As noted above, the ethics of reproduction is immensely complex and the range of
positions on the duties of prospective parents contemplating reproduction go from a
strong duty not to reproduce (and a fortiori not to reproduce in ways involving gene
editing) to support for the Biblical injunction to be fruitful and multiply. The part of
this wide ranging field that is of relevance here is the question of whether prospective
parents have duties to consider the welfare of the child to be in their reproductive
decision-making. It has been argued that parents have such duties, either a duty not
to bring a child into the world that is in a harmed state,20 or a stronger duty to follow
a principle of procreative beneficence and bring into the world the best child they
can.21 There has been considerable criticism of these posited duties, both in terms of
their justification and in terms of their wider implications.22-24

Some of the writers in the gene editing debate are committed to a trinity or triple
of positions encompassing nonidentity, reproductive (legal) liberty, and procre-
ative beneficence. This commitment to the triple is rarely expressed explicitly, but
accepting two of the positions simultaneously in argument is quite comment.
Julian Savulescu and Guy Kahane, for instance, discuss the interplay between
reproductive liberty and procreative beneficence in their seminal paper on the
principle of procreative beneficence:

Talk about moral obligation can be misunderstood in another way. On
an understanding of obligation that has its roots in Mill, the existence of
an obligation implies the threat of sanction. If this is taken to mean that
there is a conceptual tie between obligation and moral disapproval, then
PB [Procreative Beneficence] is compatible with such a tie. Egregious
procreative choices deserve our disapproval just like other failures to
meet one’s obligations, such as failure to protect the welfare of one’s
children. But although PB claims that parents have a moral reason to
aim to have the most advantaged children, when such a choice is pos-
sible, this is compatible, at the legal level, with enjoyment of a right to
autonomy, including the right to make procreative choices which fore-
seeably and avoidably result in less than the best child. Whether the
public interest ever justifies legal constraints on reproductive choice is
a separate question [reference removed, my emphasis].25

We suspect that most people who support Procreative Autonomy do so
because they fail to distinguish moral and legal principles. PB is a moral
principle. It states what would be morally right or wrong for reproducers

What Is “Safe Gene Editing”?

107

to do. To repeat, PB is not the view that reproducers should be coerced into
selecting the most advantaged child, or punished if they don’t. Liberal political
theory gives strong reasons to grant parents Procreative Autonomy. But this is
compatible with thinking that some legal choices made by parents are
nevertheless deeply wrong [my emphasis].26

Can the “only when it is safe” rider be justified from the triple of positions?
There is a question about whether the triple itself is internally consistent, since
there is a possible conflict between nonidentity and a duty of procreative benefi-
cence, and a possible conflict between reproductive liberty and reproductive
beneficence if reproductive liberty is understood as a moral principle. If the tri-
ple of positions is internally inconsistent, it follows trivially that the rider is
justified, since according to the rules of first order predicate logic, any conclu-
sion can be validly inferred from a set of inconsistent premises. But, this is of
course not real justification but pseudo justification, because soundness is under-
mined by the problem with the premises. In the following, we will therefore
assume that the triple is internally consistent and take reproductive liberty to be
a legal and not a moral principle.

What can be justified is then the claim that prospective reproducers do some-
thing which is seriously morally wrong if they use an unsafe mode of gene editing
(from procreative beneficence), that this wrong is not directly based on any harm
caused to the child (from nonidentity), and that although they are doing some-
thing morally wrong, they should not be legally prevented from doing it (from a
legal understanding of reproductive liberty). This initially looks very much like
the “only when it is safe” rider, but this initial appearance is deceptive because the
“only when it is safe” injunction is now a purely moral injunction with force only
for the prospective reproducers. It is not an injunction aimed at regulators telling
them how to regulate or at health care professionals telling them when to refuse to
assist in a reproductive project involving unsafe gene editing. Health care profes-
sionals can express their disapproval of the reproductive choice (as per Savulescu
and Kahane27) or they can remonstrate, reason, persuade or entreat (as per John
Stuart Mill just after the famous enunciation of the so-called “harm principle” in
On Liberty28), but they cannot refuse to participate unless they give up their neu-
trality as health care professionals.

The Function of Safety Arguments

The argument seems to have arrived at a point of perplexity. Many of the inter-
locutors in the debate who seem to be committed to the “only when it is safe”
rider are also committed to the view that only extreme lack of safety can matter
when regulating the use of gene editing in a reproductive context. The noniden-
tity problem entails that any future child whose coming into existence has
involved gene editing cannot have been wronged by that use of gene editing,
and the invocation of reproductive liberty mean that prospective parents have a
strong right to reproduce in any way they choose, even if the child that is being
brought into existence is in a harmed state. And, even if we commit ourselves to
procreative beneficence this has no bearing on when we should allow gene edit-
ing to be used in reproductive contexts. So what is the real function of the “only
when it is safe” discursive move?

Søren Holm

108

It could of course be inadvertent ignorance of the underlying inconsistency, i.e.,
the ethicists in question are not aware or have not yet realized that they are really
committed to the view that safety does not matter very much, if at all, in reproduc-
tive contexts. Or, that if it matters it matters only morally. I think that is unlikely,
since we are talking about highly intelligent people with excellent philosophical
skills, but cannot prove that it is not the case. I will, however proceed on the
assumption that they know the content of their own prior writings and their stated
philosophical commitments in relation to reproductive ethics sufficiently well to
be able to see, almost immediately, that there is an inconsistency here. So, some-
thing discursively more interesting must be going on. In the following, I will ana-
lyze five possible explanations for the use of the “only when it is safe” rider. These
five options are not mutually exclusive; two or more may be combined in the
explanation of any particular instance of this discursive phenomenon:

1) simplification of argument
2) ritualized hand-waving
3) “consensus building” with nonphilosophers (in casu scientists)
4) blurring of the line between philosophical analysis and policy advice
5) deliberate obfuscation

The simplest and least contentious function of the “only when it is safe” rider is
that it allows the argument about the ethics of eventual uses of gene editing to
proceed, while we are bracketing safety issues. So, we can, for instance, isolate and
analyze questions of gene editing for enhancement purposes without having to
think about any possible interactions between considerations of purpose and con-
siderations about safety. This is standard philosophical technique, tackling the
issues one by one, and is unproblematic, as long as we remember that we have
bracketed safety and that all of our conclusions are therefore qualified by safety.
That is, if we find that X is ethically unproblematic when safety is bracketed, the
conclusion we can draw and state is not that “X is ethically unproblematic,” but
that “X is ethically unproblematic, if gene editing is safe.”

The second function, “ritualized hand waving,” conceptualizes the “only when
it is safe” rider as an almost automatic invocation, on a par with the Muslim’s
“inshallah” finishing of sentences signifying future intentions or “please” in
British English. It has little actual meaning apart from being something that has to
be said every time we discuss a new technology. This can be linked to the observa-
tion that it is one of those propositions which it is very difficult to negate and still
be taken seriously by polite society. Claiming that we should use gene editing in
general, or for a specific purpose “long before it is safe” simply does not sound
like a good idea. We live in a “risk society” where it is a widely shared value that
risks have to be identified and minimised.29 On this account, any inconsistency
with other philosophical commitments is therefore not a real inconsistency,
because uttering or writing “only when it is safe” does not, despite surface
appearances, show any real commitment to the content of the proposition.

The third function of “consensus building with nonphilosophers,” in casu
researchers using gene editing and clinicians wanting to use it, is a plausible
explanation for the frequent occurrence of the “only when it is safe” rider in
reports and papers produced by multidisciplinary groups. As explained above,
negating this rider is pragmatically and discursively difficult, within the general

What Is “Safe Gene Editing”?

109

frame of discourses around new technologies. The precautionary principle and
similar cautionary approaches are part of that general frame, whatever philoso-
phers and regulation theorists might think about them. The philosopher may thus
agree to “only when it is safe” to stay within the standard discursive frame and
seem reasonable and sane, with the tactical goal of, for instance, getting a more
receptive audience in the group for arguments indicating the ethical acceptability
of a wide scope of use of gene editing, e.g., not just restricting it use in humans to
core therapeutic use.

The fourth function of the rider, “blurring of the line between philosophical
analysis and policy advice,” has received some attention in the bioethics literature,
but usually the problem that is discussed is the reverse of the problem we are ana-
lyzing here. That is, the discussion is usually about how philosophers overlook or
elide the difference between putting forward a radical philosophical conclusion
and advocating for a radical public policy change in the direction of that conclu-
sion.30,31 What we have here are philosophers explicitly abjuring themselves of
potential public policy implications of their philosophical conclusions. They could
say “safety does not matter very much” for the regulation of reproductive uses of
gene editing and advocate that we develop our public regulatory policies accord-
ingly, but instead they say that our policy should be only to use the technology
“when it is safe.” Why this reticence to follow the philosophical conclusions where
they lead? Now, it could in an optimistic mode be argued that since gene editing
is a relatively new technology, what has happened is that philosophical bioethi-
cists have read the papers referenced just above and the many similar papers and
books on the gap between philosophical analysis and public policy prescription
and have finally realized the significant difference between reaching a philosophi-
cal conclusion and advocating for that conclusion as directly implementable pub-
lic policy. Having had this “road to Damascus” experience, they have therefore
started to take account of the specific challenges in public policy development and
have stopped immediately transforming philosophical conclusions into policy
advice. I leave it to the reader to decide whether this is a plausible account.

We therefore have to consider the fifth and final possible function of the “only
when it is safe” rider, i.e., deliberate obfuscation. By deliberate obfuscation I mean
that the rider is inserted primarily in order to hide the full implications of the
philosophical positions that underpin the analyses of the ethics of reproductive
gene editing, i.e., it is inserted deliberately in order to placate and mislead the
reader. Here its function is to signal something like “don’t worry too much about
gene editing, or the fairly radical things we say it is OK to do with it; they are all
far in the future and we will only do them when it is safe to do them.” But this
signal elides the difference between biological safety and wider conceptions of
safety, and it obscures the wider role of nonidentity and reproductive freedom
considerations in reaching conclusions on the ethical acceptability of particular
reproductive uses of gene editing.

Conclusion

This paper has analyzed the statement made in many papers and reports on
the use of gene editing in humans that we should only use the technology when
it is safe. It has been argued that the statement is inconsistent with the philosophi-
cal commitments of some of the authors who put it forward in relation to

Søren Holm

110

reproductive uses of gene editing, especially their commitment to Parfitian non-
identity and to legal reproductive liberty.

But, if that is true it raises a question about why the statement is made, what is
its discursive and rhetorical function? Five functions are suggested, some of which
are more contentious and problematic than others, and it is argued that it is pos-
sible, perhaps even likely, that the “only when it is safe” rider is part of a deliberate
obfuscation aimed at hiding the full implications of the arguments made about the
ethics of gene editing and their underlying philosophical justifications.

Notes

1. Chan S, Donovan PJ, Douglas T, Gyngell C, Harris J, Lovell-Badge R, et al. [on behalf of the Hinxton
Group]. Genome editing technologies and human germline genetic modification: The Hinxton
Group Consensus Statement. The American Journal of Bioethics 2015;15(12):42–7.

2. Mathews DJ, Chan S, Donovan PJ, Douglas T, Gyngell C, Harris J, et al. CRISPR: A path through
the thicket. Nature News 2015;527(7577):159–61.

3. Savulescu J, Pugh J, Douglas T, Gyngell C. The moral imperative to continue gene editing research
on human embryos. Protein & cell 2015;6(7):476–9.

4. Bosley KS, Botchan M, Bredenoord AL, Carroll D, Charo RA, Charpentier E, et al. CRISPR germline
engineering—the community speaks. Nature Biotechnology 2015;33(5):478.

5. Brokowski C. Do CRISPR germline ethics statements cut it? The CRISPR Journal 2018;1(2):115–25.
6. See note 1, Chan et al. 2015.
7. See note 3, Savulescu, Pugh, Douglas, Gyngell 2015.
8. Haapaniemi E, Botla S, Persson J, Schmierer B, Taipale J. CRISPR–Cas9 genome editing induces a

p53-mediated DNA damage response. Nature Medicine 2018;24:927–30.
9. Ihry RJ, Worringer KA, Salick MR, Frias E, Ho D, Theriault K, et al. p53 inhibits CRISPR–Cas9

engineering in human pluripotent stem cells. Nature Medicine 2018;24:939–46.
10. Bunnik EM, Aarts N, van de Vathorst S. Little to lose and no other options: Ethical issues in efforts

to facilitate expanded access to investigational drugs. Health Policy 2018;122:977–83.
11. DeTora L. The dangers of magical thinking: Situating right to try laws, patient rights, and the lan-

guage of advocacy. Rhetoric of Health & Medicine 2018;24;1(1-2):37–57.
12. Folkers KM, Bateman-House A. Improving expanded access in the United States: The role of the

institutional review board. Therapeutic Innovation & Regulatory Science 2018;52(3):285–93.
13. Benatar D, Wasserman D. Debating Procreation – Is it wrong to Reproduce? New York: Oxford

University Press; 2015.
14. Parfit D. Reasons and Persons. Oxford: Oxford University Press; 1985.
15. Harris J. One principle and three fallacies of disability studies. Journal of Medical Ethics 2001;

27(6):383–7.
16. Savulescu J. Bioethics: Why philosophy is essential for progress. Journal of Medical Ethics 2015;

41(1):28–33.
17. Bredenoord AL, Dondorp W, Pennings G, De Wert G. Nuclear transfer to prevent mitochondrial

DNA disorders: Revisiting the debate on reproductive cloning. Reproductive Biomedicine Online
2011;22(2):200–7.

18. Holm S. Mitochondrial replacement therapy and identity: A comment on an exchange between
Inmaculada de Melo-Martin and John Harris. Cambridge Quarterly of Healthcare Ethics 2018;
27(3):487–91.

19. Robertson JA. Procreative liberty and the control of conception, pregnancy, and childbirth. Virginia
Law Review 1983;69(3):405–64, at 408.

20. Harris J. Is there a coherent social conception of disability? Journal of Medical Ethics 2000;
26(2):95–100.

21. Savulescu J, Kahane G. The moral obligation to create children with the best chance of the best life.
Bioethics 2009;23(5):274–90.

22. Bennett R. The fallacy of the principle of procreative ceneficence. Bioethics 2009;23(5):265–73.
23. Bennett R. When intuition is not enough: Why the principle of procreative beneficence must work

much harder to justify its eugenic vision. Bioethics 2014;28(9):447–55.
24. Holm S, Bennett R. The proper scope of the principle of procreative beneficence revisited. Monash

Bioethics Review 2014;32(1-2):22–32.

What Is “Safe Gene Editing”?

111

25. See note 21, Savulescu, Kahane 2009, at 278.
26. See note 21, Savulescu, Kahane 2009, at 279.
27. See note 21, Savulescu, Kahane 2009.
28. Mill JS. Introductory. In: Rapaport E, ed. On Liberty. Indianapolis, IN: Hackett Publishers; 1978, at 9.
29. Beck U. Risk society: Towards a new modernity. London: Sage; 1992.
30. Oswald M. Should policy ethics come in two colours: Green or white? Journal of Medical Ethics

2013;39(5):312–5.
31. Holm S. The grand leap of the whale up the Niagara Falls: Converting philosophical conclusions

into policy prescriptions. Cambridge Quarterly of Healthcare Ethics 2015;24(2):195–203.

Copyright © Cambridge University Press 2018

Cambridge Quarterly of Healthcare Ethics (2019), 28, 62–75.
© Cambridge University Press 2018.
doi:10.1017/S096318011800039762

Articles

Can the Thought of Teilhard de Chardin Carry
Us Past Current Contentious Discussions of
Gene Editing Technologies?

MÁRIA ŠULEKOVÁ and KEVIN T. FITZGERALD

Abstract: The advent of CRISPR-Cas9 technology has increased attention, and contention,
regarding the use and regulation of genome editing technologies. Public discussions continue
to give evidence of this debate falling back into the previous polarized positions of techno-
logical enthusiasts versus those who are more cautious in their approach. One response to
this contentious relapse could be to view this promising and problematic new technology
from a radically different perspective that embraces both the excitement of this technologi-
cal advance and the prudence necessary to use it well. The thought of Teilhard de Chardin
provides this desired perspective, and some insights that may help carry forward public
discussions to achieve widely accepted uses and regulations.

Keywords: CRISPR-Cas9; gene editing; genetic engineering; ethics; Pierre Teilhard de
Chardin; evolution; responsibility; biosphere stability; genetic diversity; common good;
transhumanism

CRISPR has greatly increased interest in applying genome editing to plants,
animals, and humans. In addition, it has also increased tensions surrounding
the public debates about how to use this rapidly improving technology. Current
tension between bioenthusiasts and bioconservatives results in significant
gridlock in public discussions. Deliberations about genome editing are falling
into old patterns of polarization and conflict. There is a lack of real, substantive
discussion about the issue. “We can’t get sufficient dialog going,” stated Arthur
Caplan in the June issue of Nature this year, calling for a greater variety of forums
for this discussion.1 There is an increasing need to review the situation and
look at it from a different perspective, one more amenable to substantive dialogue
and a better interchange of ideas and values. It is necessary to gather “informa-
tion from dispersed sources, bringing to the fore perspectives that are often
overlooked and promoting exchange across disciplinary and cultural divides,”2
claim the proposers of a global observatory for gene editing, Sheila Jasanoff
and J. Benjamin Hurlbut. This new institution is being created to foster differ-
ent perspectives and bring them into the larger discussion, where “approaches
currently taken for granted can be tested and recalibrated in the light of alter-
native . . . perspectives.”3

Historically, we have examples of how alternative perspectives to dominant
ethical theories have benefited the overall philosophical discussion of difficult
issues. It is indisputable that one such effort was the “ethics of care.” The ethics of
care is a feminist-oriented philosophical perspective that represents a relational
and context-bound approach to morality. The perspective of the ethics of care con-
trasts with ethical theories that depend on principles or formulas to determine

The research done by Mária Šuleková, PhD, at Georgetown University was supported by the Fulbright
Scholar Program.

Teilhard de Chardin and Genome Editing Technologies

63

moral actions, such as Kantian deontology, utilitarianism, and justice theory.
Also, this perspective does not intend to be absolute and incontrovertible.4,5
The ethics of care has inspired various projects that applied its innovative mindset.
The National Society of Genetic Counselors, for example, used an ethics of care
approach to frame their Code of Ethics.6

When discussing bioethical issues today, those at the forefront of research too
often look for simple and restricted frameworks to understand and evaluate these
issues. This limited approach, however, comes at a cost. One example of this situ-
ation is the case of the reduction of ethical questions regarding germ line genome
editing merely to physical safety, where only the technical assessment of specific
biological endpoints (for instance, off-target effects) is proposed as adequate for an
ethical evaluation. This perspective avoids the fundamental question of how to
care for and value human life as individuals, as a society, and in relation to other
forms of life.7

To address the deficiencies and gridlock in public deliberation described above,
the authors of this article suggest that taking a different approach, that of Teilhard
de Chardin, may help everyone engaged in the discussion about genome editing
to see the current problems from a perspective that helps depolarize the discus-
sion and facilitate substantive dialogue and the interchange of ideas and values.
A comparison of the current two main perspectives regarding CRISPR technology
with the approach of Teilhard will help elucidate the benefits a Teilhardian perspec-
tive could bring to the deliberations regarding the use and regulation of genome
editing technology.

Two Main Approaches Related to Emergent Technologies of Gene Editing

Two main approaches to the use of CRISPR/Cas genome editing can be found in
the literature: enthusiasm-based and caution-based. Enthusiasm generally refers
positively to the application of gene editing to agriculture, animal breeding, and
biomedicine. Cautions are related to a safety-efficacy balance, unforeseen conse-
quences, impact on the environment and biodiversity, and applications in humans.
The use of these emergent techniques in human germ line and embryo research,
and applications in therapeutic and nontherapeutic use, represent particularly
sensitive questions for both groups. Moreover, the use of CRISPR/Cas gene editing
technique for human enhancement purposes raises even greater concerns.

The contention and gridlock found in the larger gene editing discussion is
reflected clearly in the debates surrounding genetically modified foods (GMO).
The two different perspectives can be seen in regulatory frameworks at the
international level. Although Europeans apply a mostly precautionary princi-
ple approach regarding GMOs in their jurisdictions, the approach of the United
States is more permissive: unless there is evidence for harm, use is allowed. In
addition, most European countries ratified the Oviedo Convention on Human
Rights and Biomedicine,8 which prohibits human germ line genome modification.
In contrast, the National Academies of Sciences, Engineering and Medicine hosted
an International Summit on Human Gene Editing in 2015 that concluded that
the clinical use of germ line editing could proceed under regulatory oversight
if safety and efficacy issues are solved and broad societal consensus is obtained.9
Consequently, calls for the reevaluation of the Oviedo Convention’s ban have
intensified.10

Mária Šuleková and Kevin T. FitzGerald

64

In the literature, the enthusiasm-based approach related to CRISPR is well repre-
sented by John Harris,11 who argues that CRISPR should be pursued through research
until it is safe enough for use in humans. He criticizes “panic concerning,” and the
“hostility and suspicion” that the new emerging genetic technologies have recently
encountered. He compares the hostility to the use of CRISPR/Cas9 for editing genes
in in vitro–fertilized zygotes, and mitochondrial replacement therapy, to the fears
associated with in vitro fertilization and other reproductive technologies and cloning.
He considers these fears as baseless since, according to him, the use of both in vitro
fertilization and cloning has proved to be highly beneficial to humanity under effec-
tive regulation and control. Similar to Harris, Julian Savulescu et al.,12 speak about the
moral imperative to continue gene editing research on human embryos. At the same
time, voices against gene editing of the human germ line and human embryos are
raised because “genome editing in human embryos using current technologies could
have unpredictable effects on future generations,” making it dangerous and ethically
unacceptable, according to Edward Lanphier et al.13 Many from this more cautious
perspective are calling for a moratorium on such gene editing research.

In many aspects, the current debate about gene editing is an extension of the dis-
cussion about genetic engineering and human genetic modification in the past.
Current CRISPR supporters and critics recall ideas of Ronald Dworkin on the one
hand, and Francis Fukuyama and Jürgen Habermas on the other. Dworkin argued
that “morality requires society to allow parents to genetically enhance their children
so that they may have broader choices and greater chances of succeeding in life.”14
In contrast, Fukuyama warned that human genetic engineering raises “the ability to
change human nature” and the advancements in this field “challenge dearly held
notions of human equality and the capacity for moral choice.”15 Habermas wrote
that eugenic interventions in the early stage of human development aiming at
enhancement reduce ethical freedom of the person, “barring him from the sponta-
neous self-perception of being the undivided author of his own life.”16

This ongoing dispute points out the “classical” controversy between technologi-
cal conservatives and technological enthusiasts regarding genetic modification.
This conservatives versus enthusiasts debate has been labelled as a controversy
between creatures and creators, or between those who watch the world through
tragic or comic lenses. “Enthusiasts tend to emphasize that we are by nature cre-
ators and that we are true to ourselves when we use technology to transform our
selves. Conservatives, on the other hand, emphasize that we are by nature crea-
tures and that by eschewing technological self-transformation, by affirming the
way we were thrown into the world, we are true to ourselves.”17 As Erik Parens18
interestingly notes when describing these two opposite positions, we should speak
about a “gratitude” stance on the one hand and a “creativity” stance on the other,
instead of using emotionally charged terms. Moreover, he suggests that both groups
could move toward a more binocular thinking about these novel technologies by
claiming that “nobody’s against true enhancement.”19

Extending Parens’ insight, another possible approach to addressing these two
polarized positions is to focus on frameworks that try to identify and valorize
what is worthy in both perspectives. Pierre Teilhard de Chardin’s philosophical-
ethical ideas create a perspective able to avoid uncritical and poorly defined hopes
as well as unbalanced, paralyzing fears, and, hence facilitate the integration of
both the vision of the enthusiasts and the concern of the cautious to promote a
path forward both sides can walk more comfortably together.

Teilhard de Chardin and Genome Editing Technologies

65

Pierre Teilhard de Chardin’s Thought as a Path for Pursuing Gene Editing
Technology

Pierre Teilhard de Chardin never worked out a comprehensive synthesis of his
ethical thought. However, as his commentator Joseph A. Grau concluded, Teilhard
was concerned in his early writings about morality and “his moral concern contin-
ued to deepen, and, particularly during the last two decades of his life, came to
focus with particular emphasis on the critical problems of human unification on a
global scale, which he saw generating so much turmoil and confusion about him.”20
Thus, it is possible to retrace his reflections on morality by bringing together his
different insights about moral matters in conjunction with some of the fundamen-
tal concepts of his general theory of creation. Going further, Teilhard reflects how
science and technological progress can play a transformative role in human and
planetary life. Indeed, he saw that transformations of humankind brought about
by technological progress profoundly changed human action, and opened up new
responsibilities21 for employing those technologies.

Teilhard’s Evolutionary Perspective

Teilhard understood evolution in a different way than Charles Darwin and
thought that different mechanisms operate in evolution.22 The key mechanisms
could only be discovered working on a macro scale. Mechanisms that are focused
on the level of populations are not helpful or adequate to explain evolution at this
larger scale. Instead, one needs to employ a global approach to evolution to
observe the aspects that are indiscernible at the level of populations. Teilhard’s
definition of biology was the science of the infinitely complex. Hence, Teilhard
took the difference between micro and macro evolution and connected it to the
issue of complexity in biology.

Teilhard considered evolution as a moving toward complexity based on his scien-
tific research (e.g., the evolution of Siphnaeidae). The evolution of the universe,
matter, and life is described by a moving towards complexity, and in animals
towards cerebralization in different branches. We can only understand his process
of moving towards by using a holistic or systemic approach. From this perspective,
harmonious or coordinate evolution is the sufficiency and preservation of natural
equilibria and stability at the level of ecosystems, and it is because of connections
between the ecosystems and their different species that these natural equilibria are
preserved and are stable. Hence, for Teilhard, continuous evolution and stability
are intrinsically interrelated.

Moving Towards

From the perspective of Teilhard de Chardin, the evolutionary design of the Earth
unfolds from the prelife or inorganic world to the organic world, that in its devel-
opment constitutes the biosphere. Today, the concept of biosphere is widely used
in environmental and ecological discussions. Teilhard de Chardin was one of the
first advocates of the concept. The term was first defined by the Austrian scientist
Eduard Suess in 1875. Inspired by Suess’s definition, Teilhard de Chardin noted
that a “frail but superactive film of highly complex, self-reproducing matter spread
around the world.”23

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66

The biosphere gave rise to the noosphere. This was caused by an ongoing pro-
cess of the movement toward a complexity. The noosphere is defined by Teilhard as
“the psychically reflexive human surface.”24 The noosphere is related to the
human “self-conscious” type of consciousness.25 Teilhard recognized the immen-
sity of both the earth and its human inhabitants, as well as their shared origin
and destiny. He proposed that the biological, societal, and cultural dimensions
of evolution are interwoven as implied by his idea of the organic evolution of life
that unfolds into the reflective effort of human thinking.26

The term noosphere comes from a Greek word nous, commonly translated as
“mind” or “intellect.” For Teilhard, the noosphere represents an integrating reality
and points to the “layer of mind, thought and spirit within the layer of life cover-
ing the earth.”27 The noosphere is profoundly connected to the organic layers of
the biosphere and represents its further development. It cannot be understood
merely as a sphere of knowledge or inventiveness. Teilhard sees it as a sphere of
human thought, will, love, action, and interaction and considers all these as closely
interconnected. When Teilhard talks about the origin of the noosphere, he describes
it as the “biological interpretation of human history.”28 Furthermore, Teilhard pro-
posed the idea that humans are the ultimate outcomes of the general laws of bio-
sphere evolution, but with the condition that the general laws of the biosphere are
linked to the preservation of equilibria.29

Teilhard de Chardin often voiced his concern for building the Earth and culti-
vating the spirit of one Earth that emphasizes seeing the whole world and all its
people as one. He expressed this perspective of the Earth as “the passionate
sense of common destiny that draws the thinking fraction of life ever further
forward.”30 Teilhard further expounded about “the evolution of a greater con-
sciousness,” by which human thought “introduces a new era in the history of
nature.”31 He sees the Earth as a New Earth that needs to be constructed.32 The
construction, according to Teilhard, does not imply our rule over nature, but
denotes the responsible construction of a future for humanity. Teilhard was not
formally an ethicist, but according to Grau, his way of understanding moral
dimensions can be labeled as a morality of movement.33

Role of Responsibility

Movement towards a New Earth requires a new future-oriented ethics. Teilhard’s
ethics for the future34 is based on a hopeful and positive image of the world, and
needed to be developed if there is to be a future for humankind in this world. Its
main characteristic is the essential role of responsibility.35 Humankind should be
responsible for its own future, and this future should be developed considering all
forms of life and all of nature.36 Teilhard claims that with humans evolution
“becomes free to dispose to itself—it can give itself or refuse itself. Not only do we
read in our slightest acts the secret of its proceedings; but for an elementary part
we hold it in our hands, responsible for its past to its future.”37

For Teilhard, freedom is a continually expanding condition that correlates with
growing consciousness. Through reflection, humanity expands its freedom. At the
same time, with the growing consciousness comes responsibility to take part in
the expanded vision. Hence, humans need to align their actions to a purpose and
goal.38 If humankind wants to achieve the “higher plane of humanity,” it’s neces-
sary to profoundly change people’s “fundamental way of valuation and action.”39

Teilhard de Chardin and Genome Editing Technologies

67

The required internal conditions are related to the authentic exercise of freedom,
that is “a know-how to do” to evade traps and dead ends, and, most importantly,
“a will to do,” so as not to be deterred by fears or adversity.40

Teilhard’s vision of the Earth is to build it as mutual home for all living beings,
both human and nonhuman. This goal requires responsibility to preserve this
home. The preservation of this common home means the survival of humankind
as well.41 Teilhard understood the world as evolutionary and becoming aware
of its responsibility to direct the development of humankind toward complete
fulfillment. Moreover, he claims that responsibility cannot be developed in human
beings without allowing, to some extent, the development of other beings around
them.42 Responsibility is deeply connected to the unity of the biosphere and to
universal human solidarity.

Common Aspiration—Advance Human Unity

Teilhard was worried about the future of humankind and of all life as he perceived
a transformation within humankind on planet earth. He wrote that “the whole
future of the Earth […] seems to me to depend on the awakening of our faith in the
future.”43 Faith in the future means faith in the potential further development of
human beings, faith in peace, and faith in the greater unity and collaboration
among the people on a global level. Teilhard readily declared his own faith in the
intellectual, moral, and spiritual development of humankind.44

Teilhard raised the question that “a profound common aspiration arising out of
the very shape of the modern world—is not this specifically what is most to be
desired, what we most need to offset the growing forces of dissolution and disper-
sal at work among us?”45 Indeed, he saw the hopes and desires of people and the
need for the unity of humankind as steadily growing, as well as the need for mutual
help and encouragement. Teilhard described “the well-ordered integration” of the
individual “with the unified group in which Mankind must eventually culminate,
both organically and spiritually,” and of the “two processes of collectivization and
personalization” as interdependent.46

Teilhard de Chardin talks about a new threshold in the development of human
consciousness and organization. He suggests that human beings should make
every effort to create a higher form of life represented by a more unified humanity
instead of trying to live longer or just surviving.47 “The more scientifically I regard
the world, the less can I see any possible biological future for it except the active
consciousness of its unity.”48

Teilhard sought the “miracle of common soul.” He saw it as a convergence and
union of the diverse elements of humanity. It is impossible to accomplish this
union without love and compassion, and without developing the Spirit of Earth,
that might be understood as the openness to the presence of the Spirit. The Spirit of
Earth and human unity look currently more like dreams than reality, but Teilhard
felt that they were “in process of formation.”49 It is “the irresistible pressure which
unites people at a given moment in a passion they share.”50 This pressure generates
a progress towards human convergence and union through a new form of love
exercised by “interlinking.” Teilhard thinks that “a superabundance of love” may
be produced by the active forward movement of the noosphere.51

Teilhard emphasized the necessity of solidarity among peoples. “At no moment
in history has man been found, as he is today, so bound, actively and passively,

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68

in the depth of his being, to the value and perfection of everyone around him.”52
The person cannot be perfected without an authentic encounter with others.
The human future lies in the direction of a deliberately, individually chosen,
communal life.53

Facing Technology

Teilhard included a role for technology in the construction of the New Earth.
For him, technology is an instrument that would be used in the progress toward
the future of humankind. He had a fundamentally positive, optimistic, and enthu-
siastic attitude to technological developments.54 Spearheading the growth of
the noosphere is “a systematic organization and exploration of our universe.”
For Teilhard, research is “the highest of human functions,” and not an accessory,
an eccentricity, or a danger.55

This assertion can appear to be overly optimistic and careless, considering
concerns about the potential applications of modern scientific research.56 However,
Teilhard recognized both this potential crisis and the burden that is connected
with acting in a technological age. The urgency to act grows proportionately
with the increase of knowledge and power, because now people can see with
more clarity what the consequences of acting or not acting are. With the power
in their hands people understand that they cannot blame God, chance, or fate
if this power is not directed at human flourishing.57 Teilhard lamented, “Our plan
as to build a big house, larger but similar in design to our good old dwelling places.
And now we have been led by the higher logic of progress which is in us, to
collect components that are too big for the use we intended to make of them.”58
Overall, Teilhard interpreted these various issues as a “crisis of birth,” a pro-
cess of moving the structure of life to a new stage.59 In recalling the analogy of
“building,” Teilhard stated that research should be directed responsibly toward
building the earth and enriching human life.60

Using Emerging Technologies in Biosphere

Teilhard does not interpret progress as an absolute value. According to him, prog-
ress is the movement toward the future and preservation of life, and requires certain
responsibilities.61 Ludovico Galleni shows that Teilhard demands the examination
of the laws of the biosphere, and its fundamental mechanisms, in order that bio-
sphere stability may be achieved.62 To continue moving towards the future, we do
not need to discourage technological progress, but we should obtain the knowledge
of the processes related to biosphere stability and use technology so that this
stability is preserved. Moving toward represents openness to the future shaped,
at least in part, by technological progress, but it also represents the protection
of the existing stability of the biosphere.63

It appears self-evident that not all technologies are suitable to be used in the
construction of the New Earth. According to Anto Čartolovni, a suitable technol-
ogy for Teilhard must fulfill the following criteria: irreversibility, proportionality,
and foreseeability.64 However, a question arises concerning the issue: must every
irreversible change be considered inherently bad? Teilhard’s understanding of the
importance of the stability of the biosphere provides some clarity and helps in
determining which changes would be desirable. According to Teilhard, only the

Teilhard de Chardin and Genome Editing Technologies

69

irreversible changes that damage the stability of biosphere are inherently bad.
Since, as mentioned previously, stability and evolution towards complexity are
interrelated, one can conclude that an application of a technology could be consid-
ered unsuitable if it reduced overall biosphere or noosphere complexity or pre-
vented its continued development. This assessment would also have to include
the social conditions of humankind and, hence, the level of care for all human
beings and the environment. From this perspective, Čartolovni65 concludes that in
the case of using CRISPR to create gene drives to remove certain species, such as
malaria-spreading mosquitos, humankind would first have to obtain sufficient
understanding of the potential consequences of gene drive technology before
being able to decide whether or not to use it, and at the moment, we do not have
that level of understanding.

Using Emerging Technologies in Noosphere

Humankind continuously expands its presence over the earth, and through culture
it intensifies that presence. Socialization strengthens the personal growth of indi-
viduals and acts as an incubus for the formation of a global awareness and ability
to act. Globalization has expanded the capacities of human awareness and inter-
relatedness in the domain of culture, and has also increased the power of human
action. We have obtained complex knowledge, and with it a growing power to
affect the process of evolution. The discourse is no longer about how evolution has
formed us over the ages, but about what we can now do to transform the biology
that took millions of years to evolve to this point in time and space. As Teilhard
realized, evolution is nowadays not about where we have come from, but about
where we’re heading.66

Teilhard and the Transhumanist Movement

According to Eric Steinhart,67 Teilhard is one of the first to articulate transhuman-
ist themes, and his thought has influenced several important transhumanists.
Teilhard argues for the ethical use of technology “in order to advance humanity
beyond the limitations of natural biology,” including the use of both biotechnolo-
gies and intelligence technologies. In addition, he formed preliminary thoughts
about other themes often found in transhumanist writings, e.g., related to the con-
cept of a singularity in which human intelligence will become superintelligence.68
However, Ilia Delio69 argues that Teilhard did not seek to transcend biological
limits through technology, and that he was not a forerunner to the currently promoted
ideas of transhumanism. He was a scientist and visionary who saw technology as
a positive step in the whole evolutionary process. However, his concept of noo-
sphere, the next step in evolution, was perceived as a level of global consciousness
that leads not to transhumanism, but to an ultrahumanism, that was instead “a
deepening of human life through technologically-mediated collective conscious-
ness.”70 Moreover, “the evolution of humanity is not only an evolution of con-
sciousness; it is also a new phase of life in the universe moving toward unification
of mind by which cosmic evolution progresses toward greater unity.”71 Teilhard
indicated that ultimate knowing is love that draws together and unites in such a
way that a new complexified being transcends mere individual being. Hence, inte-
gral to the noosphere is the necessity of love of others, and living for others, instead

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70

of only for oneself. In this sense, one can easily conclude that Teilhards’s thought
is far different from the oft purported transhumanist goals of individual perfec-
tion and the creation of a separate posthuman techno sapiens species.

Novel Technologies, Responsibility, and the Goal of Human Unity

In his writings, Teilhard focused on convergence within the noosphere and on
interconnection between thinking beings. That is why some see him as the first
internet and social networking visionary. It is also true that in Teilhard we do not
encounter any paralyzing fear of technology. On the contrary, technology is part of
his “moving toward,” and is a critical part of the evolutionary process we are now
holding in our hands through that technology. It is also true that Teilhard can be
seen as a member of the group of “enthusiasts” or “creators,” rather than belong-
ing to the group of “cautious” or “grateful,” regarding the new methods of genetic
engineering. For Teilhard, the rapid development of technology does not trigger
a sense of fear or anxiety for the future. On the contrary, he perceives the advance
of technology as the opportunity to realize the story of the Earth, that began to be
written billions of years ago, and the continuation of that story depends on our
understanding of the deeper meaning of the purpose of the story.

However, for Teilhard, enthusiasm and courage in the face of technological
advance is intrinsically linked to human responsibility for all human action.
Moreover, evolution itself is now put into the hands of human freedom, and this
makes all of humankind responsible for the future of evolutionary processes.
Hence, this responsibility is for all nonliving, living, and human entities. Humans
are not only essential to the development of the New Creation—humankind is
responsible for it also. Thus, Teilhard can also be seen as a member of the group of
“cautious” or “grateful,” who consider themselves to be creatures rather than cre-
ators, pointing to the importance of restraint and careful discernment in the assess-
ment of the new technologies of gene engineering.

For Teilhard, according to his commentators, this responsibility manifests itself
in the protection of biosphere stability—and in the assessment of the proportional-
ity, predictability, and irreversibility of applying genome editing technologies to
organisms in the environment. In relation to the noosphere the issue can be more
complicated, but responsibility still obliges us to carefully delineate the goals, and
the means, that we pursue in using genetic editing technology on humans. The
common aspiration of humankind for Teilhard is to “enhance” the unity of human-
kind in love and care, not the technological “enhancing” of individuals in pursuit
of a singular superhuman.

Respecting Human Genetic Diversity

The common aspiration of gradual human unification does not mean that the value
of individuality and diversity is denied or diminished. Teilhard thought that even in
a global project, diversity must be protected. He talks about diversity in terms of
culture: its evolution and value. When the noosphere began to grow and develop,
a new kind of evolution, where cultural attributes were passed on, was introduced
by the increase and diffusion of thinking creatures. The result was a new and special
form of evolution that was identified by the establishment of different cultures. This
development of cultures highlights the biological unity of humankind.72

Teilhard de Chardin and Genome Editing Technologies

71

We can find a connection between the biosphere and the noosphere here. The
significance of biodiversity is one of the main points of the theory of evolution of
the biosphere. Stability is another key feature, according to James Lovelock’s theory,73
in which stability is sustained by biodiversity. This relationship means that global
stability is connected to local diversity. Hence, if there are feedback links between
the global and local levels that sustain overall stability, then when local diversity
is rich there will be a larger number of those links and the global stability is
increased. Similarly, in the noosphere, diversity produces stability. The noosphere
embodies cultural diversity that needs to be preserved in order to support and
increase stability.74

This preservation of cultural diversity can serve as a source of insight for the
evaluation of CRISPR genetic editing applications in humans. We are to value
individual diversity. This goal cannot be achieved if gene editing techniques are
used to reduce the genetic diversification of the human family, either by intent
or inadvertently. This reduction in diversity could happen, for instance, if there is
a preference for instilling certain genetic traits in a society, or strong pressures on
parental choices in the field of reproductive medicine. Obviously, genetic identity
does not exhaust personal identity, but since it is fundamental to it and to both
social and species health, genetic diversity should be respected in individuals and
considered a source of enrichment for the society.

Considering Common Good

What does it mean to follow Teilhard’s goal of common unity in gene editing
research and application? We showed how this does not mean a pursuit of “super-
humanity” in the transhumanist sense. At the same time, Teilhard presents an
enthusiasm for research and technological progress. The rhetoric of the common
good as a common goal is often used in the context of genetic research.75 The promo-
tion of this goal refers to research progress and public health benefits. What does
it mean to follow the common good and link it with an authentic Teilhardian prog-
ress in gene editing applications?

Building upon the comments of Ludovico Galleni and Francesco Scalfari,76
Teilhard foresees maintaining noosphere stability as a common effort of all human-
kind. In one of Teilhard’s last writings, collected in The Future of Man, he clearly
posed the problem of the stability of the noosphere: “If a real power of love does
not indeed arise at the earth of evolution, stronger than all individual egotisms
and passions, how can the noosphere even be stabilized?”77 Galleni and Scalfari
try to find a parallel between the biosphere and the noosphere by exploring which
ethical parameters correspond to the actual physical parameters that, at the level
of the biosphere, allow for the survival of life. They conclude that these cultural
and ethical parameters are in accord with the rights described in the Declaration
of Human Rights of the United Nations Charter.78 When these rights are fully
recognized, we will then have the necessary protections for a diversity which pro-
motes and preserves global stability. “Although difficult, this is the only way to
preserve cultural diversity and its advantages for the survival of the noosphere.”79
Human rights are a fundamental result of the “real power of love” as articulated
by Teilhard. Hence, we can conclude that human rights are pragmatic examples of
the movement toward human unity, and these rights represent a common ground
for pursuing both individual and common goods.

Mária Šuleková and Kevin T. FitzGerald

72

As presented above, if we want to stabilize the world of the noosphere, soli-
darity and care must be practiced and promoted. This goal requires a special
care for marginalized people and people in need. When CRISPR technology is
used, the questions of social justice will need to be addressed. Who will have
access to gene editing techniques? Who will decide who has the access? How
can these techniques be used for the good of all people, not only for a select few?
Moreover, the question of solidarity cannot be reduced simply to the issue of
access, control, and distribution. All peoples must be integrated into the decision-
making processes surrounding the development and applications of genome
editing technologies. Teilhard points to the importance of love and care for the
maintenance of human community. Today, the concept of love is widely sim-
plified and corrupted, despite its historically rich philosophical tradition. The
challenge today is for people to recover a more traditional concept of love and
see how it can be applied in a normative sense for the evaluation the use of
gene editing technologies by all peoples, in particular the most vulnerable and
in need.

Obtaining Knowledge of Complex Reality

Teilhard encourages us to move toward the construction of the New Earth
through the discovery and application of new technologies. At the same time,
the criterion of responsibility must be respected and developed further. As pre-
viously presented, the maintenance of biosphere stability serves as a fundamen-
tal criterion of assessment regarding the application of novel technologies in the
environment, such as the gene drive release. In referring to the global human
community, Teilhard is concerned about noosphere survival and development
that is indissolubly linked to the maintenance of biosphere stability and devel-
opment. This linkage is a concern that he shares with Hans Jonas,80 and it is
about the environment as a requirement for the survival of future generations.
We argued above that it is necessary to obtain a reasonable understanding of
processes related to biosphere stability in order to evaluate well any genetic edit-
ing applications in the environment. It is the same situation regarding the case
of noosphere stability. The problem is that the noosphere is characterized by
complexity: Teilhard speaks of the progressive growth toward higher states of
complexity. According to Lovelock, stability is attained as a result of diversifica-
tion and the increase of complexity.81 Hence, obtaining adequate understanding
before genome editing applications in humans should be seen in the broader
sense of understanding these complexities at the anthropological, ethical, and
social levels, as well as genetic and physiological levels. Thus, a condition of suf-
ficient understanding of CRISPR phenomena for use in humans will require the
integration of knowledge from various disciplines, including philosophical, legal,
social, and religious perspectives. The human reality is characterized by complex-
ity, and we cannot afford to ignore that complexity by reducing our deliberations
to scientific dimensions only.

Conclusion: Searching for the Dynamic-Stability Approach

Maintenance of biosphere and noosphere stability cannot be understood in a static
sense. There is a continuous moving forward within the process of biosphere and

Teilhard de Chardin and Genome Editing Technologies

73

noosphere preservation. Thus, the permanent process of movement toward is essen-
tial for the preservation of the stability of both the sphere of all living beings and
sphere of human beings—and, at the same time, this stability is necessary for the
continuous movement toward.

In the CRISPR debate, there is currently a call for constructive dialogue and
alternative approaches. Employing different types of frameworks capable of con-
sidering both enthusiastic and cautious approaches regarding the use of gene edit-
ing is required. We believe that Teilhard de Chardin’s perspective represents a
valuable contribution to the effort to find inclusive frameworks. The possibility of
gene editing in the living world and in humans can be seen as another step of
evolution—a sign of continuous moving forward. Moreover, in the Teilhardian
school of thought, a permanent movement forward is considered necessary for
maintenance of the world and, indeed, the entire universe. From this perspective,
technology enthusiasts can feel fully engaged in the discussion. At the same time,
the process of moving toward is an intrinsic part of the natural equilibrium and
stability of the Earth and the universe. Application of CRISPR technologies should
respect and foster this stability. This perspective will encourage technology-cautious
people to contribute much to the discussion. In the biosphere, the concept of sta-
bility can serve as a fulcrum balancing these often opposing perspectives of enthu-
siasm and caution. In the noosphere, the movement forward is put in the hands of
humans through their freedom and responsibility for both humans and all other
living forms. It is on this level of reflection that the movement forward becomes
the movement toward, because humans act intentionally. In the context of the CRISPR
debate, the crucial task will be to decide what goal(s) we are willing to pursue as
individuals, as societies, and as a species when we speak about the use of gene
editing. Teilhard does not provide that specific answer, not having had knowledge
of this genetic technology, but he can provide a better way to think about how we
might find that answer together.

Notes

1. Smalley E. As CRISPR–Cas adoption soars, summit calls for genome editing oversight. Nature
Biotechnology 2018;36(6):486.

2. Jasanoff S, Hurlbut JB. A global observatory for gene editing. Nature 2018;555(7697):435–7.
3. See note 2, Jasanoff, Hurlbut 2018:435–7.
4. Burton B, Dunn CP. Ethics of care. Encyclopædia Britannica Inc; 2017 Jun 19; available at https://

www.britannica.com/topic/ethics-of-care (last accessed 26 Jul 2018).
5. For more about relationality and dependency in ethics of care, see also Šuleková M. Vzťahovosť,

závislosť a cnosť v etike starostlivosti. In: Rajský A, Wiesenganger M, eds. Pomoc druhému na ceste
cnosti. Trnava, Slovak Republic: Typi Universitatis Tyrnaviensis; 2018:124–151.

6. National Society of Genetic Counselors, Inc. NSGC Code of Ethics. Adopted 1/92, revised 12/04,
1/06, 4/17; available at https://www.nsgc.org/p/cm/ld/fid=12 (last accessed 26 Jul 2018).

7. See note 2, Jasanoff, Hurlbut 2018:435–7.
8. Council of Europe. Convention for the Protection of Human Rights and Dignity of the Human

Being with regard to the Application of Biology and Medicine: Convention on Human Rights and
Biomedicine; 1997; available at https://www.coe.int/en/web/conventions/full-list/-/conventions/
treaty/164 (last accessed 26 Jul 2018).

9. On Human Gene Editing: International Summit Statement; 2015; available at http://www8.
nationalacademies.org/onpinews/newsitem.aspx?RecordID=12032015a (last accessed 26 Jul
2018).

10. Sykora P, Caplan A. The Council of Europe should not reaffirm the ban on germline genome edit-
ing in humans. EMBO Reports 2017;18(11):1871–2.

11. Harris J. Germline manipulation and our future. The American Journal of Bioethics 2015;15(2):30–4.

Mária Šuleková and Kevin T. FitzGerald

74

12. Savulescu J, Pugh J, Douglas T, Gyngell C. The moral imperative to continue gene editing research
on human embryos. Protein & Cell 2015;6(7):476–9.

13. Lanphier E, Urnov F, Haecker SE, Werner M, Smolenski J. Don’t edit the human germ line. Nature
2015;519(7544):410–1.

14. Nordberg A, Minssen T, Holm S, Horst M, Mortensen K, Lindberg Moller B. Cutting edges and
weaving threads in the gene editing (ᴙ)evolution: reconciling scientific progress with legal, ethical,
and social concerns. Journal of Law and Biosciences 2018;5(1):35–83.

15. Fukuyama F. Our Posthuman Future: Consequences of the Biology Revolution. New York: Farrar, Straus
and Giroux; 2002.

16. Habermas J. The Future of Human Nature. Cambridge: Polity Press; 2003.
17. Parens E. Shaping Our Selves: On Technology, Flourishing, and a Habit of Thinking. New York: Oxford

University Press; 2015.
18. See note 17, Parens 2015.
19. See note 17, Parens 2015.
20. Grau JA. Morality and the Human Future in the Thought of Teilhard de Chardin. New Jersey-London:

Associated University Presses, Inc.; 1976.
21. Gibellini R. Teilhard de Chardin: L’opera e le interpretazioni. Brescia, Italy: Editrice Queriniana; 1981.
22. Galleni L. Teilhard de Chardin and the Latin school of evolution: Complexity, moving towards and

equilibriums of nature. Pensamiento 2011;67(254):689–708.
23. Teilhard de Chardin 1971 qtd. in Galleni L, Scalfari F. Teilhard de Chardin’s engagement with the

relationship between science and theology in the light of discussions about environmental ethics.
In: Deane-Drummond C, ed. Pierre Teilhard De Chardin on People and Planet. London: Equinox;
2006:196–214.

24. Teilhard de Chardin 1971 qtd. in See note 23, Galleni, Scalfari 2006:196–214.
25. See note 23, Galleni, Scalfari 2006:196–214.
26. King U. Feeding the zest for life: Spiritual energy resources for the future of humanity. In: Meynard

T, ed. Teilhard and the Future of Humanity. New York: Fordham University Press; 2006:3–19.
27. See note 26, King 2006:3–19.
28. Teilhard de Chardin 1964 qtd. in See note 26, King 2006:3–19.
29. See note 22, Galleni 2011:689–708.
30. Teilhard de Chardin 1969 qtd. in See note 26, King 2006:3–19.
31. Teilhard de Chardin 1969 qtd. in See note 26, King 2006:3–19.
32. Čartolovni A. Teilhard de Chardin’s oeuvre within an ongoing discussion of a gene drive release

for public health reasons. Life Sciences, Society and Policy 2017;13(1):1–18.
33. See note 20, Grau 1976.
34. Jeličić A. Intelektualna i duhovna baština Pierrea Teilharda de Chardina iz perspektive suvremenih

bioetičkih problema. Filozofska istraživanja 2015;35(2):289–300.
35. See note 32, Čartolovni 2017:1–18.
36. See note 26, King 2006:3–19.
37. Teilhard de Chardin P. The Phenomenon of Man, 5th ed. London-Glasgow: Collins Clear-Type Press;

1974.
38. Grumett 2005 qtd. in Ayotte DJ. An introduction to the thought of Teilhard de Chardin as a catalyst

for interreligious dialogue. Religious Inquiries 2012;1(2):65–82.
39. See note 26, King 2006:3–19.
40. See note 26, King 2006:3–19.
41. See note 32, Čartolovni 2017:1–18.
42. See note 20, Grau 1976.
43. Teilhard de Chardin 1964 qtd. in See note 26, King 2006:3–19.
44. See note 26, King 2006:3–19.
45. Teilhard de Chardin 1999 qtd. in See note 26, King 2006:3–19.
46. See note 26, King 2006:3–19.
47. See note 26, King 2006:3–19.
48. Teilhard de Chardin 1962 qtd. in See note 26, King 2006:3–19.
49. See note 26, King 2006:3–19.
50. Teilhard de Chardin 1962 qtd. in See note 26, King 2006:3–19.
51. See note 26, King 2006:3–19.
52. Teilhard de Chardin 1970 qtd. in See note 20, Grau 1976.
53. Ligneul A. Teilhard and Personalism. New York: Paulist Press Deus Books; 1968.

Teilhard de Chardin and Genome Editing Technologies

75

54. See note 20, Grau 1976.
55. See note 26, King 2006:3–19.
56. See note 26, King 2006:3–19.
57. See note 20, Grau 1976.
58. Teilhard de Chardin 1962 qtd. in See note 26, King 2006:3–19.
59. See note 26, King 2006:3–19.
60. See note 20, Grau 1976.
61. Jeličić 2015 qtd. in See note 32, Čartolovni 2017:1–18.
62. Galleni 2016 qtd. in See note 32, Čartolovni 2017:1–18.
63. See note 32, Čartolovni 2017:1–18.
64. See note 32, Čartolovni 2017:1–18.
65. See note 32, Čartolovni 2017:1–18.
66. See note 38, Ayotte 2012:65–82.
67. Steinhart E. Teilhard de Chardin and Transhumanism. Journal of Evolution and Technology

2008;20(1):1–22
68. See note 67, Steinhart 2008:1–22.
69. Delio I. Transhumanism or ultrahumanism: Teilhard de Chardin on technology, religion and

evolution. Theology and Science 2012;10(2):153–66.
70. See note 69, Delio 2012:153–66.
71. See note 69, Delio 2012:153–66.
72. See note 23, Galleni, Scalfari 2006:196–214.
73. Lovelock 1998 qtd. in See note 23, Galleni, Scalfari 2006:196–214.
74. See note 23, Galleni, Scalfari 2006:196–214.
75. About communitarian prospective in genetic research see also Šuleková M. Le biobanche di ricerca:

Problematiche etico-sociali tra diritti dell´individuo e prospettiva comunitaria. Medicina e Morale
2013;3:477–510.

76. See note 23, Galleni, Scalfari 2006:196–214.
77. Teilhard de Chardin 1973 qtd. in See note 23, Galleni, Scalfari 2006:196–214.
78. United Nations. Universal Declaration of Human Rights; 1948; available at http://www.un.org/

en/universal-declaration-human-rights/ (last accessed 26 Jul 2018).
79. See note 23, Galleni, Scalfari 2006:196–214.
80. Jonas H. The Imperative of Responsibility: In Search of an Ethics for the Technological Age. Chicago-

London: The University of Chicago Press; 1984.
81. Lovelock 1991 qtd. in Galleni L. Is Biosphere doing theology? Zygon 2001;1:33–7.

Copyright © Cambridge University Press 2018

Xu Cell Biosci (2020) 10:48
https://doi.org/10.1186/s13578-020-00410-6

R E S E A R C H H I G H L I G H T

CCR5-Δ32 biology, gene editing,
and warnings for the future of CRISPR-Cas9
as a human and humane gene editing tool
MengMeng Xu*

Abstract
Background: Biomedical technologies have not just improved human health but also assisted in the creation of
human life. Since the first birth of a healthy baby by in vitro fertilization (IVF) 40 years ago, IVF has been the mainstay
treatment for couples struggling with infertility. This technology, in addition to increasingly accessible genetic testing,
has made it possible for countless couples to have children. Since CRISPR-Cas9 gene editing was described in 2015,
its potential for targeting genetic diseases has been much anticipated. However, the potential of using CRISPR-Cas9
for human germline modification has led to many fears of “designer babies” and widespread concerns for the impact
of this technology on human evolution and its implications in Social Darwinism. In addition to these ethical/moral
concerns, there remain many unknowns about CRISPR-Cas9 technology and endless unanticipated consequence to
gene editing.

Methods: In this paper, we analyze the current progresses of CRISPR-Cas9 technology and discuss the theoretical
advantages of certain allelic variances in the C-C chemokine receptor 5 gene (CCR5) in the setting of recent ethical/
moral concerns regarding gene editing using the CRISPR-Cas9 system.

Results: These uncertainties have been highlighted recently by the birth of Chinese twins whose C-C chemokine
receptor 5 (CCR5) gene had been inactivated via CRISPR-Cas9 to be theoretically protective against HIV infection. CCR5
signaling is critical for the successful infection of human immunodeficiency virus (HIV ) and people with homozy-
gous inactivating CCR5-Δ32 mutations have been shown to be protected against HIV infection. Those with the
CCR5-Δ32/Δ32 mutation also have greater neuroplasticity, allowing for improved recovery from neurological trauma,
and decreased Chagas cardiomyopathy. However, the CCR5-Δ32/Δ32 mutation has also been associated with earlier
clinical manifestations for West Nile infection, ambiguous effects on osteoclast function, and a four-fold increased
mortality from influenza infection. These detrimental health impacts, in addition to the confounding factor that these
CRISPR babies do not carry this exact CCR5-Δ32/Δ32 mutation, lead to many questions regarding the children’s future
health and the moral conundrum of their birth. The creation and birth of these babies was not completed with any
scientific, ethical, or governmental oversight, which has spurned the acceleration of talks regarding global regulations
for human genetic editing.

Conclusions: Although we can try to regulate for ethical, health-related only use of this technology, moral and
governmental oversights need to be supplemented by technical regulations. For instance, whole genome sequenc-
ing needs to be used to eliminate off-target mutations that could affect the health and safety of infants born to this

© The Author(s) 2020. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing,
adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and
the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material
in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material
is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the
permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creat iveco
mmons .org/licen ses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creat iveco mmons .org/publi cdoma in/
zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

Open Access

Cell & Bioscience

*Correspondence: [email protected]
Department of Pediatrics, Morgan Stanley Children’s Hospital, Columbia
University, 3959 Broadway, New York, NY 10032, USA

Page 2 of 6Xu Cell Biosci (2020) 10:48

As the most efficient and precise genome editing tool
available, CRISPR-Cas9 technology presents a powerful
and lost-cost method of genetic editing that has never
been available before. The availability of this technique
has radically changed the biomedical field and has the
potential to radically alter human healthcare [1–3]. It
has made in  vitro modeling of human mutations possi-
ble, increased the speed of genetically engineered animal
models, and made the treatment of genetic diseases more
than a pipedream. In fact, a pilot clinical trial in sickle
cell anemia just reported promising preliminary results
in the first patient ever treated with CRISPR-Cas9 gene-
therapy [4] and there are multiple other ongoing trials
assessing gene therapy in hematologic disease [5]. The
power of CRISPR-Cas9 technology is not limited to the
correction of disease-causing genetic mutations, but also
being considered as a method for taking advantage of
genetic traits inherent in some populations. For instance,
the C-C chemokine receptor 5 (CCR5) Δ32 mutation
found in ~ 11% of northern Europeans is known to pro-
tect against HIV infection. Last year, twin Chinese girls
were engineered by CRISPR-Cas9 to carry a CCR5 gene
with similar properties to CCR5-Δ32, specifically to be
resistant to HIV. The announcement of these unexpected
births has highlighted the fear of a new era of eugenics
brought on by CRISPR-Cas9. Here we discuss the protec-
tive and detrimental effects of this mutation and contrib-
ute to the ongoing moral, philosophical, and regulational
conversation with considerations regarding the technical
safety of CRISPR-Cas9 technique in humans.

The CCR5 gene was first identified in 1977 [6] but did
not become a subject of great public interest until 2009,
when an HIV positive individual transplanted with bone
marrow from a donor with a homozygous CCR5-Δ32
mutation, became HIV negative despite stopping anti-
retroviral (ARV) therapy [7]. This seminal clinical case
study was founded on decades of work showing CCR5’s
role as a co-stimulator in T-cell function, activation, and
the production of antigen specific T-cells [8]. These stud-
ies showed the CCR5-Δ32 mutation to cause deletion of
32-base pairs in CCR5, leading to non-functional expres-
sion of this gene that does not localize to the cell surface.
These mechanistic findings along with the discovery of
CCR5 as a necessary co-receptor for entry of macrophage
tropic HIV strains [9, 10] led to increased interest in this
gene as a target for HIV treatment and other immuno-
logical processes.

CCR5 deletions have also been shown to provide pro-
tection against other pathogens, including smallpox and
flaviviruses such as dengue, Zika, and West Nile virus
[11]. In fact, smallpox endemics in Europe are believed
to be the selective pressure that led to an increased pres-
ence of the allele in European populations [11]. CCR5
deletion was also found to be protective against non-viral
infections. Early reports have found the CCR5-Δ32 dele-
tion to be protective against inflammatory cardiomyo-
pathy in patients with chronic Chagas’ disease [12]. This
result was recently disputed in a polymorphism analysis
between wild-type, heterozygous, and homozygous Cha-
ga’s disease patients [13]. However, a Brazilian genetic
polymorphism study of CCR1, CCR5, and their ligands
CCL2 and CCL5, respectively, found CCL5-CCR1 to be
the target for immune-stimulation from Trypanosoma
cruzi infection. Certain variants of CCL5-CCR1  were
subsequently found to be significantly protective against
Chagas’s disease [14]. Outside of the infectious disease
realm, CCR5 has also been found to be involved in neu-
ronal recovery from stroke and traumatic brain injury
(TBI) through upregulation of  CREB (cAMP response
element-binding protein) and DLK (Delta-like protein 1)
signaling [15]. Joy et  al. first identified the expression of
CCR5 in cortical neurons after stroke and later discov-
ered neuronal knockdown of CCR5 to result in enhanced
cortical projections during regeneration and preservation
of dendritic spines [15]. These in vitro findings were sub-
sequently confirmed as clinically significant in an analysis
of 1,563 stroke patients (300 CCR5-Δ32 carriers vs 1265
non-carriers) in the Tel Aviv Brain Acute Stroke Cohort
(TABASCO). Patients with Δ32/Δ32 loss-of-function
mutation CCR5 recovered significantly faster from stroke
with improved measures of memory, verbal function, and
attention- indicating improved neuronal plasticity [15].
While CCR5 is clinically relevant in this wide variety of
diseases, its importance in HIV infection has been the
most studied in the clinical setting.

As a cell membrane integrated protein with seven
transmembrane segments and an eighth α-helix parallel
to the plasma membrane, CCR5 presents on the cell sur-
face and functions in tandem with CD4-recptors as the
initial co-docking site for the HIV PG120-PG41 surface
protein. This initial association between the HIV PG120-
PG41, CCR5, and CD4-receptors allows for the initial
viral invasion and subsequent infection and replication
(Fig.  1a). The essential binding site on CCR5 for HIV

process. Like Pandora’s Box, we cannot pretend to forget CRISPR-Cas9 technology, all we can do is ensure a safe,
moral, and equitable used of this technology.

Keywords: CCR5-Δ32, Human genome editing, HIV infection, CRISPR-Cas

Page 3 of 6Xu Cell Biosci (2020) 10:48

PG120-PG41 is known as 2D7. It is located on the third
extracellular element (second loop) of the membrane
integrated CCR5 and works in tandem with the PA12
binding site and the G protein linkage domains found on
the first extra-cellular element of CCR5. The CCR5-Δ32
mutation, describes a 32 base pair deletion just before
the 2D7 structural loop. This results in the creation of a
premature stop codon, and thus, the absence of the 2D7
loop necessary for HIV viral binding, but preserves  the
PA12 binding site (Fig.  2). This mutation hampers HIV
binding two-fold: by removing the necessary 2D7 binding
domain and by rendering the protein cytosolic. Around
10% of the European population have paired missense
mutations C20S and C178R or C101X and FS299, collec-
tively known as CCR5-Δ32, which protects against HIV
infection by inhibiting the initial viral docking process
(Fig. 1b) [16, 17].

Ever since the theoretical protection of CCR5-Δ32/
Δ32 against HIV was clinically supported by the cure of
a HIV-positive patient transplanted with bone marrow
from a homozygous CCR5-Δ32 donor [7], the potential
for CCR5-Δ32 as a curative therapy for HIV has been
greatly debated and anticipated [8, 17, 18]. However,
most controlled and regulated studies are still in the pre-
clinical phase using human stem cells or mouse models.
The Deng group established a CRISPR/Cas9 gene edit-
ing system in human CD34+ hematopoietic stem cells

(HSPCs) which allowed for long-term CCR5 ablation.
Mice transplanted with these CCR5-deleted HSPCs
exhibited lasting HIV-1 resistance in  vivo [19]. Another
study found editing of co-receptors CCR5 and CXCR4
by CRISPR-Cas9 to protect CD4+ T cells from HIV-1
infection in  vitro [20]. Although another group was able
to successfully transplant and achieve long-term engraft-
ment of CRISPR-edited HSPCs into a patient, they were
only able to disrupt 5% of CCR5 function. This unex-
pected result hinted at unanticipated factors in in  vivo
editing, thus halting the study for fear of harm to patient
health [21, 22]. Despite the lack of complete understand-
ing of the CCR5 gene and incomplete pre-clinical test-
ing proving CCR5 gene manipulation to be benign, some
have already jumped ahead to human genome manipu-
lation. Last year, Jiankui He, a researcher at the South-
ern University of Science and Technology in Guandong,
China announced the birth of twins whose genomes he
had manipulated by CRISPR-Cas9 to have non-func-
tional CCR5. This editing was made in an effort to pro-
tect the infants against HIV infection. This unregulated
experiment immediately generated massive concern over
the moral impact of this human experiment and earned
universal condemnation for advancing to human experi-
mentation without adequate safety precautions and
assessments.

Fig. 1 The HIV infection process (a): The HIV GP-120 first associates with both the CD4 and CCR5 on the surface of a cell, which is the first step in
viral invasion and further viral replication. Molecular mechanism of CCR5 in HIV infection and the protective effect of cytoplasmic CCR5-Δ32 against
HIV-1 infection (b)

Page 4 of 6Xu Cell Biosci (2020) 10:48

While the use of CRISPR-Cas9 technology as a eugenics
tool is morally confounding and difficult to justify given
the human health, evolution, and social equality impli-
cations; it is naïve to say that CRISPR-Cas9 will not be
used by futures parents and scientist to give an advanta-
geous foundation to their children. Thus, the best course
of action that global summits on genome editing can pro-
duce are exact allowances and restrictions for genome
editing and specific punishments for both the researcher
and the local/federal governments responsible for enforc-
ing regulations. Inherited disease caused by specific point
mutations may be the most realistic targets for germline
alternation. For instance, correcting the point mutation
causing the glutamine to valine mutation in sickle cell
disease could free future generation from the constant
threat of pain crises and eliminate the risk of acute chest
and stroke that often claim these patients’ lives. However,
even in these clear-cut cases we still need further data on
the exact time period during which germline alteration is
safe for the embryo. However, to ensure at least the meth-
odological safety of using CRISPR-Cas9 in humans, two
technical aspects must be met: total understanding of the
gene being altered and complete control over off target
effects of CRISPR-Cas9 editing. Editing of CCR5 does
not fit the first requirement as those homozygous for the
CCR5-Δ32 mutation have  unexpected negative effects
such as earlier clinical manifestations for West Nile infec-
tion [23], four-fold likelihood of mortality from influenza

infections [24], and disadvantageous osteoclast function
[25]. In addition, multiple publications have reported
unexpected off-target mutations generated by CRISPR-
Cas9. Although one retracted publication demonstrated
few unexpected mutational events [26], one study found
rare but notable  mutations [27], several others found
large deletions [28, 29], while another found unexplain-
able complex deletions and insertions in mice generated
by CRISPR-Cas9 [30]. As such, the CCR5 twins need to
be monitored both for possible known effects, such as
an increased susceptibility to influenza infection, abnor-
mal  bone growth and other immunological conditions,
and also require close monitoring of their general growth
and development for unanticipated effects.

Even should these unknowns be overcome, there may
still be small deletions or insertions that cause deleteri-
ous frame-shift mutations, or rarer effects we have yet
to identify. As such, the only way to ensure the coding
fidelity of edited cells is by sequencing the full genome
of each edited cell in comparison to parents’ genomes.
This safety check itself will require further technologi-
cal development allowing for rapid, inexpensive whole-
genome sequencing and analysis while in the narrow
window of implantable embryos. Even these precau-
tions would not account for the epigenetic factors that
may impact growth and development. Should complica-
tions from these identified elements be resolved, there
are still a myriad of unknown factors in CRISPR-Cas9

Fig. 2 The structure of membrane integrated CCR5. The elements important in HIV binding and structure (PA12 binding site and 2D7 binding
site, and sites of tyrosine sulfonation and G-protein linkage) are highlighted. The CCR5-Δ32 deletion site is denoted with a triangle and found just
before the 2D7 binding site. Mutation at this site results in a premature stop codon, and thus the deletion of all protein structures after this location,
resulting in the loss of the 2D7 binding site and a cytosolic CCR5

Page 5 of 6Xu Cell Biosci (2020) 10:48

technology that should present an independent techno-
logical precaution against human genetic editing regard-
less of the moral/ethical conundrum (Fig. 3). We suggest
that there needs to be a more vigorous and annual global
debate to established the specific mutations on which
human gene-editing research should be allowed, and that
these genes be limited to those what would solve clear
clinical problems (i.e. sickle cell disease, other diseases
with known mutational causes). Ideally, such a body of
experts would also be able to advise a multinational con-
sortium such  as the United Nations on the appropriate
punitive and incentive actions necessary to dissuade indi-
viduals and institutions from supporting unsanctioned
human genome editing.

Abbreviations
ARV: Anti-retroviral; CCR5: C-C chemokine receptor 5; CCL2: C-C motif
chemokine ligand 2; CREB: cAMP response element-binding protein; CRISPR:
Clustered regularly interspaced short palindromic repeats; Cas9: CRISPR-
associated protein-9 nuclease; CXCR4: C-X-C chemokine receptor type 4; DLK1:
Delta-like protein 1; GP120: Glycoprotein 120, viral envelope glycoprotein 120;
GP41: Glycoprotein 41, viral envelope glycoprotein 41; HSPC: Hematopoi-
etic stem cell; HIV: Human immunodeficiency virus; IVF: In vitro fertilization;
TABASCO: Tel Aviv Brain Acute Stroke Cohort; TBI: Traumatic brain injury.

Authors’ contributions
MX conceived and prepared the manuscript. The author read and approved
the final manuscript.

Funding and acknowledgements
There is no funding support for this study. The collection of references for this
article is supported by the Medical Scientist Training Program (MSPT ), NIH.

Availability of data and materials
Not applicable.

Ethics approval and consent to participate
All applicable international, national, and/or institutional guidelines for the
care and use of animals were followed in the discussed studies.

Consent for publication
Not applicable.

Competing interests
The authors declare that they have no competing interests.

Received: 13 January 2020 Accepted: 14 March 2020

References
1. Mali P, Yang L, Esvelt KM, Aach J, Guell M, DiCarlo JE, Norville JE, Church

GM. RNA-guided human genome engineering via Cas9. Science.
2013;339(6121):823–6. https ://doi.org/10.1126/scien ce.12320 33.

2. Cong L, Ran FA, Cox D, Lin S, Barretto R, Habib N, Hsu PD, Wu X, Jiang W,
Marraffini LA, Zhang F. Multiplex genome engineering using CRISPR/Cas

Fig. 3 Advantages and disadvantages of CCR5-Δ32/Δ32 (a) and CRISPR-Cas9 (b) genome editing

Page 6 of 6Xu Cell Biosci (2020) 10:48

systems. Science. 2013;339(6121):819–23. https ://doi.org/10.1126/scien
ce.12311 43.

3. Fonfara I, Richter H, Bratovič M, Le Rhun A, Charpentier E. The CRISPR-
associated DNA-cleaving enzyme Cpf1 also processes precursor CRISPR
RNA. Nature. 2016;532(7600):517–21. https ://doi.org/10.1038/natur e1794
5.

4. Stein R. Gene-edited ’Supercells’ make progress In fight Against Sickle
Cell Disease. National Public Radio. https ://www.npr.org/secti ons/healt
h-shots /2019/11/19/78051 0277/gene-edite d-super cells -make-progr ess-
in-fight -again st-sickl e-cell-disea se. Accessed 19 Nov 2019.

5. Demirci S, Uchida N, Tisdale JF. Gene therapy for sickle cell disease: an
update. Cytotherapy. 2018;20(7):899–910. https ://doi.org/10.1016/j.
jcyt.2018.04.003.

6. Walz DA, Wu VY, de Lamo R, Dene H, McCoy LE. Primary structure
of human platelet factor 4. Thromb Res. 1977;11:893–8. https ://doi.
org/10.1016/0049-3848(77)90117 -7.

7. Hütter G, Nowak D, Mossner M, Ganepola S, Müßig A, Allers K. Long-term
control of HIV by CCR5 Delta32/Delta32 stem-cell transplantation. N Engl
J Med. 2009;360:692–8. https ://doi.org/10.1056/NEJMo a0802 905.

8. Barmania F, Pepper MS. C-C chemokine receptor type five (CCR5): An
emerging target for the control of HIV infection. Appl Transl Genom.
2013;2:3–16. https ://doi.org/10.1016/j.atg.2013.05.004.

9. Dragic T, Litwin V, Allaway GP, Martin SR, Huang Y, Nagashima KA. HIV-1
entry into CD4+ cells is mediated by the chemokine receptor CC-CKR-5.
Nature. 1996;381:667–73. https ://doi.org/10.1038/38166 7a0.

10. Deng H, Liu R, Ellmeier W, Choe S, Unutmaz D, Burkhart M. Identifi-
cation of a major co-receptor for primary isolates of HIV-1. Nature.
1996;381:661–6. https ://doi.org/10.1038/38166 1a0.

11. Galvani A, Slatkin M. Evaluating plague and smallpox as histori-
cal selective pressures for the CCR5-Δ32 HIV-resistance allele. PNAS.
2003;100(25):15276–9. https ://doi.org/10.1073/pnas.24350 85100 .

12. Flórez O, Martín J, González CI. Genetic variants in the chemokines and
chemokine receptors in Chagas disease. Hum Immunol. 2012;73(8):852–
8. https ://doi.org/10.1016/j.humim m.2012.04.

13. Oliveira AP, Bernardo CR, Camargo AV, Villafanha DF, Cavasini CE, de Mat-
tos CC, de Godoy MF, Bestetti RB, de Mattos LC. CCR5 chemokine receptor
gene variants in chronic Chagas’ disease. Int J Cardiol. 2014;176(2):520–2.
https ://doi.org/10.1016/j.ijcar d.2014.07.043.

14. Batista AM, Alvarado-Arnez LE, Alves SM, Melo G, Pereira IR, Ruivo LAS, da
Silva AA, Gibaldi D, da Silva TDESP, de Lorena VMB, de Melo AS, de Araújo
Soares AK, Barros MDS, Costa VMA, Cardoso CC, Pacheco AG, Carrazzone
C, Oliveira W Jr, Moraes MO, Lannes-Vieira J. Genetic polymorphism at
CCL5 is associated with protection in Chagas’ heart disease: antagonistic
participation of CCR1+ and CCR5+ cells in chronic Chagasic cardio-
myopathy. Front Immunol. 2018;9:615. https ://doi.org/10.3389/fimmu
.2018.00615 .

15. Joy MT, Ben Assayag E, Shabashov-Stone D, Liraz-Zaltsman S, Mazzitelli
J, Arenas M, Abduljawad N, Kliper E, Korczyn AD, Thareja NS, Kesner EL,
Zhou M, Huang S, Silva TK, Katz N, Bornstein NM, Silva AJ, Shohami E, Car-
michael ST. CCR5 is a therapeutic target for recovery after stroke and trau-
matic brain injury. Cell. 2019;176(5):1143–57. https ://doi.org/10.1016/j.
cell.2019.01.044.

16. Parmentier M. CCR5 and HIV infection, a view from Brussels. Front Immu-
nol. 2015;6:295. https ://doi.org/10.3389/fimmu .2015.00295 .

17. Allers K, Schneider T. CCR5Δ32 mutation and HIV infection: basis for cura-
tive HIV therapy. Curr Opin Virol. 2015;14:24–9. https ://doi.org/10.1016/j.
covir o.2015.06.007.

18. Hütter G, Bodor J, Ledger S, Boyd M, Millington M, Tsie M, Symonds G.
CCR5 targeted cell therapy for HIV and prevention of viral escape. Viruses.
2015;7(8):4186–203. https ://doi.org/10.3390/v7082 816.

19. Xu L, Yang H, Gao Y, Chen Z, Xie L, Liu Y, Liu Y, Wang X, Li H, Lai W, He Y,
Yao A, Ma L, Shao Y, Zhang B, Wang C, Chen H, Deng H. CRISPR/Cas9-
mediated CCR5 ablation in human hematopoietic stem/progenitor cells
confers HIV-1 resistance in vivo. Mol Ther. 2017;25(8):1782–9. https ://doi.
org/10.1016/j.ymthe .2017.04.027.

20. Liu Z, Chen S, Jin X, Wang Q, Yang K, Li C, Xiao Q, Hou P, Liu S, Wu S, Hou
W, Xiong Y, Kong C, Zhao X, Wu L, Li C, Sun G, Guo D. Genome editing of
the HIV co-receptors CCR5 and CXCR4 by CRISPR-Cas9 protects CD4+ T
cells from HIV-1 infection. Cell Biosci. 2017;7:47. https ://doi.org/10.1186/
s1357 8-017-0174-2.

21. Xu L, Wang J, Liu Y, Xie L, Su B, Mou D, Wang L, Liu T, Wang X, Zhang B,
Zhao L, Hu L, Ning H, Zhang Y, Deng K, Liu L, Lu X, Zhang T, Xu J, Li C, Wu
H, Deng H, Chen H. CRISPR-edited stem cells in a patient with HIV and
acute lymphocytic leukemia. N Engl J Med. 2019;381(13):1240–7. https ://
doi.org/10.1056/NEJMo a1817 426.

22. Wei X, Nielsen R. CCR5-Δ32 is deleterious in the homozygous state in
humans. Nat Med. 2019;25(6):909–10. https ://doi.org/10.1038/s4159
1-019-0459-6.

23. Lim JK, McDermott DH, Lisco A, Foster GA, Krysztof D, Follmann D,
Stramer SL, Murphy PM. CCR5 deficiency is a risk factor for early clinical
manifestations of West Nile virus infection but not for viral transmission. J
Infect Dis. 2010;201(2):178–85. https ://doi.org/10.1086/64942 6.

24. Falcon A, Cuevas MT, Rodriguez-Frandsen A, Reyes N, Pozo F, Moreno
S, Ledesma J, Martínez-Alarcón J, Nieto A, Casas I. CCR5 deficiency
predisposes to fatal outcome in influenza virus infection. J Gen Virol.
2015;96:2074–8. https ://doi.org/10.1099/vir.0.00016 5.

25. Xie Y, Zhan S, Ge W, Tang P. The potential risks of C-C chemokine receptor
5-edited babies in bone development. Bone Res. 2019;7:1–4. https ://doi.
org/10.1038/s4141 3-019-0044-0.

26. Schaefer KA, Wu WH, Colgan DF, Tsang SH, Bassuk AG, Mahajan VB.
Unexpected mutations after CRISPR–Cas9 editing in vivo. Nat Methods.
2017;14(6):547–8. https ://doi.org/10.1038/nmeth .4293(Retracted).

27. Iyer V, Shen B, Zhang W, Hodgkins A, Keane T, Huang X, Skarnes WC.
Off-target mutations are rare in Cas9-modified mice. Nat Methods.
2015;12(6):479. https ://doi.org/10.1038/nmeth .34088 .

28. Adikusuma F, Piltz S, Corbett MA, Turvey M, McColl SR, Helbig KJ, Beard
MR, Hughes J, Pomerantz RT, Thomas PQ. Large deletions induced by
Cas9 cleavage. Nature. 2018;560:E8–E9. https ://doi.org/10.1038/s4158
6-018-0380-z.

29. Ma H, Marti-Gutierrez N, Park SW, Wu J, Lee Y, Suzuki K, Koski A, Ji D, Hay-
ama T, Ahmed R, Darby H, Van Dyken C, Li Y, Kang E, Park AR, Kim D, Kim
ST, Gong J, Gu Y, Xu X, Battaglia D, Krieg SA, Lee DM, Wu DH, Wolf DP, Heit-
ner SB, Belmonte JCI, Amato P, Kim JS, Kaul S, Mitalipov S. Correction of a
pathogenic gene mutation in human embryos. Nature. 2017;548:413–9.
https ://doi.org/10.1038/natur e2330 5.

30. Shin HY, Wang C, Lee HK, Yoo KH, Zeng X, Kuhns T, Yang CM, Mohr T,
Liu C, Hennighausen L. CRISPR/Cas9 targeting events cause complex
deletions and insertions at 17 sites in the mouse genome. Nat Commun.
2017;31(8):15464. https ://doi.org/10.1038/ncomm s1546 4.

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Review

Genome-Editing Technologies: Concept, Pros, and
Cons of Various Genome-Editing Techniques and
Bioethical Concerns for Clinical Application
Sikandar Hayat Khan1

1Department of Pathology, PNS HAFEEZ Hospital, Pathology E-8, Islamabad, Islamabad 44400, Pakistan

The traditional healthcare system is at the doorstep for entering
into the arena of molecular medicine. The enormous knowl-
edge and ongoing research have now been able to demonstrate
methodologies that can alter DNA coding. The techniques used
to edit or change the genome evolved from the earlier attempts
like nuclease technologies, homing endonucleases, and certain
chemical methods. Molecular techniques like meganuclease,
transcription activator-like effector nucleases (TALENs), and
zinc-finger nucleases (ZFNs) initially emerged as genome-edit-
ing technologies. These initial technologies suffer from lower
specificity due to their off-targets side effects. Moreover, from
biotechnology’s perspective, the main obstacle was to develop
simple but effective delivery methods for host cell entry. Later,
small RNAs, including microRNA (miRNA) and small inter-
fering RNA (siRNA), have been widely adopted in the research
laboratories to replace lab animals and cell lines. The latest dis-
covery of CRISPR/Cas9 technology seems more encouraging by
providing better efficiency, feasibility, and multi-role clinical
application. This later biotechnology seem to take genome-
engineering techniques to the next level of molecular engineer-
ing. This review generally discusses the various gene-editing
technologies in terms of the mechanisms of action, advantages,
and side effects.

https://doi.org/10.1016/j.omtn.2019.02.027.

Correspondence: Sikandar Hayat Khan, Department of Pathology, PNS HAFEEZ
Hospital, Pathology E-8, Islamabad, Islamabad 44400, Pakistan.
E-mail: [email protected]

Over the last half century after post-DNA helical structure discovery,
the world has seen a continuous staircase outburst of various molec-
ular technologies, which are now heading forward toward transla-
tion into clinical and laboratory practice.1 Given the availability of
sequencing platforms, acquired wisdom about the micro-mechanics
at work within the genetic apparatus, and the introduction of user-
friendly nanotechnologies, it was possible for next-generation scien-
tists to manipulate the genetic codes at various levels.2 Over the last
two decades we saw a plethora of molecular techniques, which al-
lowed us to edit genes or their alter pathways, allowing humans for
the first time to micro-edit the DNA codes and further to alter the
mRNA fate through post-transcriptional modifications.3

Principally, genome-wide editing techniques can be interpreted as
methods where DNA sequences are changed by deletions, mRNA
processing, and post-transcriptional modifications to result in altered
gene expression, leading to functional behavior of proteins.4,5 Com-

326 Molecular Therapy: Nucleic Acids Vol. 16 June 2019 ª 2019 The A
This is an open access article under the CC BY-NC-ND license (http

mon to these methods are three basic steps, including mechanisms
for genetic tool entry into the cell and later nucleus; altering gene tran-
scription and onward processing function; and, finally, the end-
output in the shape of a suppressed, overexpressed, or simply an
altered protein product.6,7 From a holistic point of view, the tech-
niques involve an apparently simplistic concept involving multiple re-
ceptor-ligand interactions; varying cell entry modes like lipofection,
sonification, and transfection; and further downstream pathway ef-
fects. Furthermore, these technologies are variable in terms of their
specificity and sensitivity, off-target effects, finances, and technique
expertise. The body’s immune response to accept the foreign genetic
elements within the cells can lead to the rejection of foreign tissues.

Moreover, molecular knowledge, in terms of methodology differences,
defining targetable diseases, innovative nanotechnology tools for gene
editing, and ethical aspects, also needs to be understood. The plat-
forms for these technologies are improving every day, with a plethora
of new data appearing due to technology miniaturization and automa-
tion and newer discoveries to improve the yield and specificity of
an edited product. Alongside the developmental improvement in
genome-wide engineering the regulatory work-up, standardization
protocols need to be devised to reduce inter and intra-method impre-
cision, defining the indications and contraindications of every tech-
nique to help improve the concept of personalized medicine.

This review briefly explains the available technologies, provides com-
parison and contrast between different genome-editing methods,
and identifies some newer versions of genome editing with possible
bioethical concerns.

Review Methodology

PubMed searches with the keywords genome-editing techniques or
gene-editing techniques in the last 10 and 5 years yielded a total of
4,466 and 4,054 references, except some historical and related refer-
ences. Specific searches for articles dealing with specific genome-edit-
ing methods included conventional genome-editing systems (n = 100),

uthor(s).
://creativecommons.org/licenses/by-nc-nd/4.0/).

Figure 1. A Consolidated Overview of Genome-Editing Techniques

www.moleculartherapy.org

Review

chemical methods (n = 252), meganucleases (n = 83), zinc-finger
nucleases (ZFNs) (n = 890), transcription activator-like effector nucle-
ases (TALENs) (n = 1,136), homing endonucleases (n = 265), and
CRISPR (n = 11,421). The search was therefore limited to reviews
showing conceptual information of common techniques and compar-
ative information about ZFNs, TALENSs, and CRISPR technologies.
Finally, the literature was searched for learning newer and advanced
gene-editing methods and bioethicalconcerns associated with genome
biotechnologies.

Genome-Editing Techniques

The recent expansion and advancements in the field of biotechnology
provided us with information and insight into the biochemical and
molecular mechanisms to edit DNA and, thus, modify downstream
pathways. To date, multiple biotechnologies have shown promise
for clinical use, but the field of genome-editing technologies is rapidly
evolving and improving. The new techniques seem promising, but the
earlier ones have also been updated and improved. For simplicity and
consolidation, an overview of genome-editing techniques is presented
in Figure 1.

Representative genome-editing techniques are discussed below.

(1) Conventional genome-editing technique. In the true sense, the
technique may not relate with evolving genome-editing tech-
niques. As highlighted in Figure 1, it includes homologous
recombination related with gene intervention. While not much

in vogue or lab use today, the technique is based on physiological
processes involving a double-stranded repair system. However,
some recent data have shown RAD52 protein to be important
in mediating homologous recombination, and this protein there-
fore has been considered as a therapy target in certain cancers like
BRCA 1 and 2 repair pathways.6,7 However, the technique as of
now could not gain widespread introduction due to the emer-
gence of newer techniques.

(2) Chemical modalities of genome editing. Komiyama8 utilized
non-restriction enzyme methodology termed artificial restriction
DNA cutter (ARCUT). This method uses pseudo-complemen-
tary peptide nucleic acid (pcPNA), whose job is to specify the
cleavage site within the chromosome or the telomeric region.
Once pcPNA specifies the site, excision here is carried out by
cerium (CE) and EDTA (chemical mixture), which performs
the splicing function.8 Furthermore, the technology uses a
DNA ligase that can later attach any desirable DNA within the
spliced site. The advantage of this particular technique is that it
can be used in high salt concentrations. Upon initial introduc-
tion, the technique looked quite appealing to the clinical market;
however, later issues like increased turnaround time and specif-
ically the manufacturing of site-specific pcPNA became huge
hurdles (Figure 2).8,9

(3) Homing endonuclease systems. Homing endocucleases (HEs)
with this word “homing” practically is interpreted as lateral trans-
mission of a genome DNA sequence. The general concept involves
a DNA segment where a site is removed by the endocnulceases,

Molecular Therapy: Nucleic Acids Vol. 16 June 2019 327

Figure 2. Excision of Selective Site of dsDNA by Utilizing Artificial Restriction DNA Cutter

www.moleculartherapy.org

Review

which thus results in the formation of 2 segments of DNA frag-
ment.10 So what are these HEs? They are nucleases that occur
naturally, with a size almost equivalent to 14 bp, and they are
capable of splicing slightly larger DNA sequences.11 Recently,
the introduction of recombinant adeno-associated viruses
(rAAVs) have allowed them as efficient vehicles for transporting
genetic tools of genome engineering into the cell, as depicted in
Figure 3.12 Issues pertaining to this technology include engineer-
ing difficulties in the preparation of these nucleases as well as
developing vectors for their entry into cells.13 Another issue
with rAAV, though improving with better biotechnology, was

328 Molecular Therapy: Nucleic Acids Vol. 16 June 2019

off-target effects like reducing site specificity, less DNA integra-
tion, and possible host genome mutations.14

(4) Protein-based nuclease systems. These systems incorporate
nuclease proteins for DNA sequence editing. The common tech-
niques are described below.
Meganucleases. Also termed molecular DNA scissors, these are
large base pair structures that are sometimes found in the genome.
Their potential to excise large pieces of DNA sequences was
recently recognized as a genetic tool to modify DNA. This genetic
potential has been manipulated in labs by modifying the recogni-
tion sites to create nicks, as required for DNA sequence change.

Figure 3. Schematic Showing rAAV Entry,

Movement within Cytoplasm, Attachment with

DNA, and Integration with DNA Segment for

Possible Genome Modification

The steps include the following: (1) entry of rAAV into cell,

(2) uptake by exosome and transport within cytoplasm, (3)

release of rAAV for entry into nucleus, (4) rAAV delivery of

homing endocnulease (HE) and desirable DNA segment,

(5) HE cut of the non-desirable DNA code, and (6) rAAV-

delivered desirable DNA code replacement of the DNA.

Figure 4. Schematic Showing Step by Step Zinc-Finger Nuclease-Induced Genome Editing

The mechanisms include the following: (1) ZFNs containing FokI endonucleases and protein-binding domains are introduced into the cell, (2) FokI and protein-binding

domains are released to enter the nucleus, (3) protein-binding domains attach with DNA fragment to be removed, (3) FokI cuts out the identified DNA segment by creating

double-stranded DNA break, and (4) the desirable DNA segment is inserted and integrated into the DNA sequence.

www.moleculartherapy.org

Review

These meganucleases are sometimes joined by proteins to create
large variants like DmoCre and E-Drel, which can further provide
nucleotide site-specific cleavage.15 The technique revolves around
two basic steps: first is the recognition of a cleavage site, and then
endonucleases splice out the region.16 The positive aspect related
to meganucleases is less toxicity, as they are naturally occurring
and provide very specific site cleavage. However, there are newer
techniques now in the clinical arena that have not allowed them to
flourish more.

ZFNs. ZFNs are purely artificial structures generated by a combi-
natorial approach where restriction endonucleases are joined with
zinc-finger-binding domain protein. Figure 4 explains their mech-
anism of action in detail, where a binding protein domain iden-
tifies after reaching the desirable splice site, which is then cut at
a specific codon by special restriction endonucleases called FokI.
The biotechnology is restricted in terms of attachment with 3 co-
dons on either side of the DNA chain. The technique in recent
years has gained widespread popularity due to its simplicity and
specificity, and it is being employed in clinical usage for certain
diseases.17,18

TALENs. TALENS almost resemble ZFNs in terms of
manufacturing and mode of action. They are made by a similar
principle where a restriction nuclease is bound to a DNA-binding
protein domain called TAL effector.19 The difference between
TALENs and ZFNs is that the former can target 3 nt in one go
and the latter can only address 1 nt, thus making TALENs slightly

more site specific with fewer off-target effects.20 However, the
techniques share many similarities (Figure 5).

(5) RNA DNA systems. These systems primarily include the
different types of CRISPR methods. The concept of CRISPR is
primitive and has been derived from an ancient immunity sys-
tem, adopted in nature by some prokaryotic cells like Archea
and probably some bacteria.21 CRISPR in itself has two compo-
nents, including SPR termed sometimes as spacers, which are
hallmarked by varying and differing nucleotide sequences, and
probably each one of them represents a past exposure to foreign
antigen. The CRI may represent the genetic memory for a bacte-
rium and can be re-activated once encountered with a similar
foreign antigen. CRI has similar nucleotides (repeats) represent-
ing like separators between different CRIs.22 Figure 6 attempts to
provide a basic overview of the CRISPR/Cas9 concept.
Cas especially Cas9 as depicted in Figure 6 has a nuclease function.
Whenever CRISPR RNA (crRNA; also termed guide RNA
[gRNA]) guides the Cas9 protein regarding a possible antigenic
threat, like a bacteriophage, it with the help of gRNA creates dou-
ble-stranded DNA (dsDNA) nicks at the guided selected sites,
causing a site-specific cleavage and, thus, destruction of the anti-
gen.23 Moreover, the memory from the antigen is stored as spacer
within CRISPR.24

This physiological role of Cas9/CRISPR as explained above had
recently been extensively utilized for multiple clinical condi-
tions.25–27 At the time of writing this review, the news broke about

Molecular Therapy: Nucleic Acids Vol. 16 June 2019 329

Figure 5. Diagram Showing Mechanisms of Transcription Activator-like Effector Nucleases

The steps of gene editing include the following: (1) TALENs containing FokI endonucleases and TALE domains are introduced into the cell, (2) FokI and TALE domains are

released to enter the nucleus, (3) TALE recognizes the non-desirable DNA segments and attaches with them, (4) FokI cleaves the non-desirable DNA segments, and (5) after

the non-desirable DNA segments are cleaved, the desirable segment of DNA is incorporated into the DNA.

330

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Review

Lulu and Nana being claimed to be the first genetically modified
babies, where the human genome was edited to create resistance
against HIV infection.28 Specific crRNA/gRNA has been engi-
neered, which can be introduced into cell nuclei and later Cas9
where the non-desirable dsDNA is associated with the Cas9 after
guidance provided by the specific crRNA/gRNA. This comple-
mentary binding between gRNA and the non-desirable segment
allows Cas9 to destroy the DNA fragment. In clinical and research
practice, the created nick can be specifically filled by inserting the
sequence of choice to change the non-desirable sequence of nucle-
otides.29

Over the last few years, CRISPR/Cas9 technology has gained wide-
spread popularity on account of its simplicity and specificity, with
different versions of the original now under research. Multiple ex-
perimentations and biotechnologies have been re-defining the
CRISPR/Cas technologies into 3 distinct types of CRISPR-Cas
types, based on crRNA processing and further action, including
the following:

Type 1 CRISPR/Cas system. This version utilized Cas5 or Cas6 for
pre-processing of crRNA; further cleavage function needs Cas3,
Cascade, and crRNA for interference.

Type 2 CRISPR/Cas system. Though Cas9 typically functions un-
der the guidance of crRNA to target DNA, RNase III, trans acti-
vating RNA (tracrRNA), and a yet-to-be-identified protein factor
are involved in trimming at the 50 end.

Molecular Therapy: Nucleic Acids Vol. 16 June 2019

Type 3 CRISPR/Cas system. Like the type 1 system, this category
uses Cas6 for processing crRNA 30 end trimming. The uniqueness
of this technique is its targeting of RNA, which is done by a specific
complex called type III Csm/Cmr complex.30

Apart from the aforementioned conventional style classification of
CRISPR/Cas classification, the data review provided multiple
other biotechnologies now being utilized. Some examples include
photo-activating CRISPR system,31 Intein-inducible split Cas9,32

and modifications like hybrid crRNA-tracrRNA.33

(6) Gene-silencingtechniques. These methodsmaynotfalltrulyunder
genome editing, but they still are capable of modifying the DNA
sequence. These technologies include RNAi, CRISPR interference
(CRISPRi), and morpholino oligonucleotide techniques.34–36

Comparative Analysis

The above provides a gist of the various commonly used genome-edit-
ing techniques. Though there is enormous development, innovation,
and design of newer ways to edit the genome, we focus our further dis-
cussion on the comparison of common techniques, including ZFN,
TALEN, and CRISPR methods. Tables 1, 2, and 3 provide a compara-
tive assessment among these methods.

Advancements in Genome Engineering

The biotechnology is booming with a lot of newer modalities to edit
the genome. Oligodeoxyribonucleotide (ODN) can be utilized with

Figure 6. Schematic Demonstrating the Concept of CRISPR/Cas9 Interactions Leading to the Destruction of Viral Genome at the Selected Splice Site by the

crRNA/gRNA

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Review

double-stranded transcription factor decoy (TFD) to act as a thera-
peutic target for multiple diseases, which can affect the transcription
factor and thus bring in the requisite change in transcription and
further downstream protein actions.37 Papaioannou et al.38 have uti-
lized single-stranded ODNs to precisely cut genomes for repairing
very small point mutations, giving a footprint-free genome-editing
modality. This concept involves a drug (doxycycline)-induced Cs9
transgene, which is carried into the cell by a specific transposon,
providing us with very specific and efficient Cas9-mediated editing
of the genome. This technique does not need the conventional donor

Table 1. Biotechnology Differences among Prototype Genome-Editing Techniq

Serial No. Parameter ZFN TALEN

1 design simplicity
moderate (ZFNs need customized
protein for every DNA sequence)

slightly
multip
of engi

2 engineering feasibility low higher

3 multiplex genome editing few models few mo

4 large-scale library preparation
not much progress (need
individual gene tailoring)

not mu
gene ta

5 specificity low higher

6 efficiency normala norma

7 cost low high

aSome new versions are more efficient24,48 but CRISPR science is evolving more.
bCpf1 protein addition will probably improve cell delivery methods.51,52

template, and, thus, it is termed footprint-free genome editing.38 The
technique seems to have minimal off-target effects and is considered
to be a safer version.

Other novel modalities of genome editing are also appearing in the
literature, with slight modifications of existing techniques. Martínez-
Gálvez et al.39 used single-stranded DNA (ssDNA) and argonautes
in gene editing and helped improved gene editing. Some researchers
have utilized certain enzymes like integrases and in the future may
obviate the need for nucleases.40

ues

CRISPER/Cas Reference

complex (identical repeats are
le, which creates technical issues
neering and delivery into cells)

simpler (available versions for
crRNA can be easily designed)

48

highest 24,49

dels
high-yield multiplexing available
(no need for obtaining embryonic
stem cells)

48,50

ch progress (need individual
iloring)

progress demonstrated (CRISPR
only requires plasmid containing
small oligonucleotides)

51

highest 24

lb high 24,48,52

low 53

Molecular Therapy: Nucleic Acids Vol. 16 June 2019 331

Table 2. Side Effect Profiles for Genome-Editing Methods

Serial No. Parameter ZFN TALEN CRISPER/Cas Reference

1
off-target effect
incidence

– – – 54

a
homologous
recombination
rate frequency

+ + + –

b
non-homologous
end joining (NHEJ)
mutation rates

+ +
++ (only with
earlier versions)

55,56

c
immune reaction
susceptibility

less less more 57,58

d

RNA-guided
endonuclease
(RGEN)-induced
off-target
mutatagenesis

� � ++ 59

2 cytotoxicity chances ++ + + –

Table 3. Clinical and Research Applications across Important Genome-

Editing Techniques

Serial No. Parameter ZFN TALEN CRISPER/Cas Reference

1 diagnostic utility + + +++ 60

2 clinical trial use ++ + +++ 61

3
utility as epigenetic
marker

++ +++ ++++ 62

4
making gene-knockout
models for research

no no yes (CRISPRi) 63

5
capacity for modification
of mitochondrial DNA

no no probable 64

6
genetic editing in
human babies

no no yes 65

7 RNA editing no no yes 66

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Review

The most interesting part of the genome-editing technique, which
may be the game changer in genome editing, is the whole genome
engineering by synthesis that in fact would re-create the genome
from scratch as per the given designed DNA code. This probably
will become the synthetic genomics of the future.41 Though research
work in this domain stands preliminary, over time it is anticipated
that this technology may overtake the concept of genome editing.

Bioethical Issues and Genome-Editing Techniques

Genome-editing tools are powerful in terms of their potential to not
only bring biotechnological revolution in the field of crop develop-
ment and human pathology but also, in the wrong hands, lead to
abuse and misuse in multiple ways, including manipulation of germ-
line genetics. Genuine bioethical concerns have been raised by many
experts.42 While time will be the actual judge of these technologies as
boon or bane, still the methods can impact the human race probably
in the most nuclear ways, and our incoming human race may be
victimized in ways we do not yet understand.43 Principal concerns
apart from illegal germline mutation include the morality, the eu-
genics helping the fittest to survive, ongoing clinical debates about
informed consent, religious debate, the possible rise of clones,
designer babies, and possibly superhumans.44–46 Moreover, the cur-
rent literature also rules in the possibility of genome editing as a future
weapon of war.47

While the quest for a healthy baby and right of best possible treatment
choice have been acknowledged in many societies, the approaching
biotechnological revolution seems imminent and undeniable. The
pressing need, therefore demands a harmonious and regulated
translation of needed aspects of genome-editing-related technologies
for molecular medicine and other non-clinical crop and food indus-
tries. This will need consensus in public opinion, debates among
experts, involvement of biotechnologists, opinions of bioethical ex-
perts, regulatory frameworks within legislatures, and final guidelines
and oversight for the finally allowed limited application.

332 Molecular Therapy: Nucleic Acids Vol. 16 June 2019

Conclusions

This review discussed multiples aspects of genome-editing technolo-
gies, including a classification; some basic explanatory concepts on
mechanisms;andcomparisonbetween methods,newer advancements,
and bioethical concerns. It seems that CRISPR/Cas technologies are
probably superseding ZFNs and TALENS. However, the CRIPSR/
Cas methods are also being improvised, and newer additions have
further enhanced its functional capabilities with reduced off-target
effects. Furthermore, the process of engineering better gene modifica-
tion technologies is evolving and can one day replace even CRISPR/
Cas, possibly shifting to synthetic genomics. Among all these revolu-
tionary developments, bioethical concerns need serious attention.

REFERENCES
1. Friedrich, M. (1871). Ueber die chemische Zusammensetzung der Eiterzellen [On the

chemical composition of pus cells]. Medicinisch-chemische Untersuchungen 4,
441–460.

2. Watson, J.D., and Crick, F.H.C. (1953). Molecular structure of nucleic acids; a struc-
ture for deoxyribose nucleic acid. Nature 171, 737–738.

3. Sanger, F., and Coulson, A.R. (1975). A rapid method for determining sequences in
DNA by primed synthesis with DNA polymerase. J. Mol. Biol. 94, 441–448.

4. Mullis, K.B. (2016). https://www.karymullis.com/.

5. Pavletich, N.P., and Pabo, C.O. (1991). Zinc finger-DNA recognition: crystal struc-
ture of a Zif268-DNA complex at 2.1 A. Science 252, 809–817.

6. Claussin, C., and Chang, M. (2016). Multiple Rad52-Mediated Homology-Directed
Repair Mechanisms Are Required to Prevent Telomere Attrition-Induced
Senescence in Saccharomyces cerevisiae. PLoS Genet. 12, e1006176.

7. Arai, N., and Kagawa, W. (2014). [Molecular mechanisms of homologous recombi-
nation promoted by budding yeast Rad52]. Seikagaku 86, 693–697.

8. Komiyama, M. (2014). Chemical modifications of artificial restriction DNA cutter
(ARCUT) to promote its in vivo and in vitro applications. Artif. DNA PNA XNA
5, e1112457.

9. Ito, K., and Komiyama, M. (2014). Site-selective scission of human genome using
PNA-based artificial restriction DNA cutter. Methods Mol. Biol. 1050, 111–120.

10. Schultz, B.R., and Chamberlain, J.S. (2008). Recombinant adeno-associated virus
transduction and integration. Mol. Ther. 16, 1189–1199.

11. Smith, J., Grizot, S., Arnould, S., Duclert, A., Epinat, J.C., Chames, P., Prieto, J.,
Redondo, P., Blanco, F.J., Bravo, J., et al. (2006). A combinatorial approach to create
artificial homing endonucleases cleaving chosen sequences. Nucleic Acids Res. 34,
e149.

www.moleculartherapy.org

Review

12. Yoon, Y., Wang, D., Tai, P.W.L., Riley, J., Gao, G., and Rivera-Pérez, J.A. (2018).
Streamlined ex vivo and in vivo genome editing in mouse embryos using recombinant
adeno-associated viruses. Nat. Commun. 9, 412.

13. Rey-Rico, A., and Cucchiarini, M. (2016). Controlled release strategies for rAAV-
mediated gene delivery. Acta Biomater. 29, 1–10.

14. Jin, L., Lange, W., Kempmann, A., Maybeck, V., Günther, A., Gruteser, N., Baumann,
A., and Offenhäusser, A. (2016). High-efficiency transduction and specific expression
of ChR2opt for optogenetic manipulation of primary cortical neurons mediated by
recombinant adeno-associated viruses. J. Biotechnol. 233, 171–180.

15. Zaslavskiy, M., Bertonati, C., Duchateau, P., Duclert, A., and Silva, G.H. (2014).
Efficient design of meganucleases using a machine learning approach. BMC
Bioinformatics 15, 191.

16. McMurrough, T.A., Brown, C.M., Zhang, K., Hausner, G., Junop, M.S., Gloor, G.B.,
and Edgell, D.R. (2018). Active site residue identity regulates cleavage preference of
LAGLIDADG homing endonucleases. Nucleic Acids Res. 46, 11990–12007.

17. Ochiai, H., and Yamamoto, T. (2017). Construction and Evaluation of Zinc Finger
Nucleases. Methods Mol. Biol. 1630, 1–24.

18. Ji, H., Lu, P., Liu, B., Qu, X., Wang, Y., Jiang, Z., Yang, X., Zhong, Y., Yang, H., Pan, H.,
et al. (2018). Zinc-Finger Nucleases Induced by HIV-1 Tat Excise HIV-1 from the
Host Genome in Infected and Latently Infected Cells. Mol. Ther. Nucleic Acids 12,
67–74.

19. Hensel, G., and Kumlehn, J. (2019). Genome Engineering Using TALENs. Methods
Mol. Biol. 1900, 195–215.

20. Chandrasegaran, S., and Carroll, D. (2016). Origins of Programmable Nucleases for
Genome Engineering. J. Mol. Biol. 428 (5 Pt B), 963–989.

21. Fischer, S., Maier, L.K., Stoll, B., Brendel, J., Fischer, E., Pfeiffer, F., Dyall-Smith, M.,
and Marchfelder, A. (2012). An archaeal immune system can detect multiple proto-
spacer adjacent motifs (PAMs) to target invader DNA. J. Biol. Chem. 287, 33351–
33363.

22. Eid, A., and Mahfouz, M.M. (2016). Genome editing: the road of CRISPR/Cas9 from
bench to clinic. Exp. Mol. Med. 48, e265.

23. Zhang, F., Wen, Y., and Guo, X. (2014). CRISPR/Cas9 for genome editing: progress,
implications and challenges. Hum. Mol. Genet. 23 (R1), R40–R46.

24. Zhang, J.H., Adikaram, P., Pandey, M., Genis, A., and Simonds, W.F. (2016).
Optimization of genome editing through CRISPR-Cas9 engineering. Bioengineered
7, 166–174.

25. Hussain, W., Mahmood, T., Hussain, J., Ali, N., Shah, T., Qayyum, S., and Khan, I.
(2019). CRISPR/Cas system: A game changing genome editing technology, to treat
human genetic diseases. Gene 685, 70–75.

26. Wang, L., Zheng, W., Liu, S., Li, B., and Jiang, X. (2019). Delivery of CRISPR/Cas9 by
novel strategies for gene therapy. ChemBioChem 20, 634–643.

27. Cai, B., Sun, S., Li, Z., Zhang, X., Ke, Y., Yang, J., and Li, X. (2018). Application of
CRISPR/Cas9 technologies combined with iPSCs in the study and treatment of retinal
degenerative diseases. Hum. Genet. 137, 679–688.

28. Regalado, A. (2018). Exclusive: Chinese scientists are creating CRISPR babies. A
daring effort is under way to create the first children whose DNA has been tailored
using gene editing. MIT Technology Review. https://www.technologyreview.com/s/
612458/exclusive-chinese-scientists-are-creating-crispr-babies/.

29. Damian, M., and Porteus, M.H. (2013). A crisper look at genome editing: RNA-
guided genome modification. Mol. Ther. 21, 720–722.

30. Rath, D., Amlinger, L., Rath, A., and Lundgren, M. (2015). The CRISPR-Cas immune
system: biology, mechanisms and applications. Biochimie 117, 119–128.

31. Polstein, L.R., and Gersbach, C.A. (2015). A light-inducible CRISPR-Cas9 system for
control of endogenous gene activation. Nat. Chem. Biol. 11, 198–200.

32. Zetsche, B., Volz, S.E., and Zhang, F. (2015). A split-Cas9 architecture for inducible
genome editing and transcription modulation. Nat. Biotechnol. 33, 139–142.

33. Mali, P., Aach, J., Stranges, P.B., Esvelt, K.M., Moosburner, M., Kosuri, S., Yang, L.,
and Church, G.M. (2013). CAS9 transcriptional activators for target specificity
screening and paired nickases for cooperative genome engineering. Nat. Biotechnol.
31, 833–838.

34. Fischer, S.E. (2015). RNA Interference and MicroRNA-Mediated Silencing. Curr.
Protoc. Mol. Biol. 112, 26.1.1–5.

35. Yoshida, M., Yokota, E., Sakuma, T., Yamatsuji, T., Takigawa, N., Ushijima, T.,
Yamamoto, T., Fukazawa, T., and Naomoto, Y. (2018). Development of an integrated
CRISPRi targeting DNp63 for treatment of squamous cell carcinoma. Oncotarget 9,
29220–29232.

36. Paul, S., and Caruthers, M.H. (2016). Synthesis of Phosphorodiamidate Morpholino
Oligonucleotides and Their Chimeras Using Phosphoramidite Chemistry. J. Am.
Chem. Soc. 138, 15663–15672.

37. Hecker, M., and Wagner, A.H. (2017). Transcription factor decoy technology: A ther-
apeutic update. Biochem. Pharmacol. 144, 29–34.

38. Papaioannou, I., Simons, J.P., and Owen, J.S. (2012). Oligonucleotide-directed gene-
editing technology: mechanisms and future prospects. Expert Opin. Biol. Ther. 12,
329–342.

39. Martínez-Gálvez, G., Ata, H., Campbell, J.M., and Ekker, S.C. (2016). ssDNA and the
Argonautes: The Quest for the Next Golden Editor. Hum. Gene Ther. 27, 419–422.

40. Ledford, H. (2016). Beyond CRISPR: A guide to the many other ways to edit a
genome. Nature 536, 136–137.

41. Luo, Z., Yang, Q., Geng, B., Jiang, S., Yang, S., Li, X., Cai, Y., and Dai, J. (2018). Whole
genome engineering by synthesis. Sci. China Life Sci. 61, 1515–1527.

42. Krishan, K., Kanchan, T., and Singh, B. (2016). Human Genome Editing and Ethical
Considerations. Sci. Eng. Ethics 22, 597–599.

43. Krishan, K., Kanchan, T., Singh, B., Baryah, N., and Puri, S. (2018). Germline Editing:
Editors Cautionary. Clin. Ter. 169, e58–e59.

44. Shinwari, Z.K., Tanveer, F., and Khalil, A.T. (2018). Ethical Issues Regarding CRISPR
Mediated Genome Editing. Curr. Issues Mol. Biol. 26, 103–110.

45. Liao, S.M. (2019). Designing humans: A human rights approach. Bioethics 33,
98–104.

46. Hofmann, B. (2018). The gene-editing of super-ego. Med. Health Care Philos. 21,
295–302.

47. Fraser, C.M., and Dando, M.R. (2001). Genomics and future biological weapons: the
need for preventive action by the biomedical community. Nat. Genet. 29, 253–256.

48. Wang, H., La Russa, M., and Qi, L.S. (2016). CRISPR/Cas9 in Genome Editing and
Beyond. Annu. Rev. Biochem. 85, 227–264.

49. Salsman, J., and Dellaire, G. (2017). Precision genome editing in the CRISPR era.
Biochem. Cell Biol. 95, 187–201.

50. Sakuma, T., Nishikawa, A., Kume, S., Chayama, K., and Yamamoto, T. (2014).
Multiplex genome engineering in human cells using all-in-one CRISPR/Cas9 vector
system. Sci. Rep. 4, 5400.

51. Poirier, J.T. (2017). CRISPR Libraries and Screening. Prog. Mol. Biol. Transl. Sci. 152,
69–82.

52. Sun, B., Yang, J., Yang, S., Ye, R.D., Chen, D., and Jiang, Y. (2018). A CRISPR-Cpf1-
Assisted Non-Homologous End Joining Genome Editing System of Mycobacterium
smegmatis. Biotechnol. J. 13, e1700588.

53. Qiu, X.Y., Zhu, L.Y., Zhu, C.S., Ma, J.X., Hou, T., Wu, X.M., Xie, S.S., Min, L., Tan,
D.A., Zhang, D.Y., and Zhu, L. (2018). Highly Effective and Low-Cost MicroRNA
Detection with CRISPR-Cas9. ACS Synth. Biol. 7, 807–813.

54. Koo, T., Lee, J., and Kim, J.S. (2015). Measuring and Reducing Off-Target Activities of
Programmable Nucleases Including CRISPR-Cas9. Mol. Cells 38, 475–481.

55. Symington, L.S., and Gautier, J. (2011). Double-strand break end resection and repair
pathway choice. Annu. Rev. Genet. 45, 247–271.

56. Yin, H., Xue, W., Chen, S., Bogorad, R.L., Benedetti, E., Grompe, M., Koteliansky, V.,
Sharp, P.A., Jacks, T., and Anderson, D.G. (2014). Genome editing with Cas9 in adult
mice corrects a disease mutation and phenotype. Nat. Biotechnol. 32, 551–553.

57. Jordan, B. (2016). Les débuts de CRISPR en thérapie génique – Chroniques
génomiques [First use of CRISPR for gene therapy]. Med. Sci. (Paris) 32, 1035–1037.

58. Chew, W.L. (2018). Immunity to CRISPR Cas9 and Cas12a therapeutics. Wiley
Interdiscip. Rev. Syst. Biol. Med. 10, https://doi.org/10.1002/wsbm.1408.

Molecular Therapy: Nucleic Acids Vol. 16 June 2019 333

www.moleculartherapy.org

Review

59. Cho, S.W., Kim, S., Kim, Y., Kweon, J., Kim, H.S., Bae, S., and Kim, J.S. (2014).
Analysis of off-target effects of CRISPR/Cas-derived RNA-guided endonucleases
and nickases. Genome Res. 24, 132–141.

60. Shang, W., Wang, F., Fan, G., and Wang, H. (2017). Key elements for designing and
performing a CRISPR/Cas9-based genetic screen. J. Genet. Genomics 44, 439–449.

61. Wassef, M., Luscan, A., Battistella, A., Le Corre, S., Li, H., Wallace, M.R., Vidaud, M.,
and Margueron, R. (2017). Versatile and precise gene-targeting strategies for func-
tional studies in mammalian cell lines. Methods 121-122, 45–54.

62. Vojta, A., Dobrini�c, P., Tadi�c, V., Bo�ckor, L., Kora�c, P., Julg, B., Klasi�c, M., and Zoldo�s,
V. (2016). Repurposing the CRISPR-Cas9 system for targeted DNA methylation.
Nucleic Acids Res. 44, 5615–5628.

334 Molecular Therapy: Nucleic Acids Vol. 16 June 2019

63. Zhang, B., Yang, Q., Chen, J., Wu, L., Yao, T., Wu, Y., Xu, H., Zhang, L., Xia, Q., and
Zhou, D. (2016). CRISPRi-Manipulation of Genetic Code Expansion via RF1 for
Reassignment of Amber Codon in Bacteria. Sci. Rep. 6, 20000.

64. Fogleman, S., Santana, C., Bishop, C., Miller, A., and Capco, D.G. (2016). CRISPR/
Cas9 and mitochondrial gene replacement therapy: promising techniques and ethical
considerations. Am. J. Stem Cells 5, 39–52.

65. Callaway, E. (2016). Gene-editing research in human embryos gains momentum.
Nature 532, 289–290.

66. Zimmer, C. (2016). Scientists FindFormofCrisprGene EditingWith New Capabilities.
The New York Times, June 3, 2016. https://www.nytimes.com/2016/06/04/science/
rna-c2c2-gene-editing-dna-crispr.html.

© The Author(s) 2019. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved.
For permissions, please e-mail: [email protected]

Human Reproduction, Vol.34, No.11, pp. 2104–2111, 2019
Advance Access Publication on November 14, 2019 doi:10.1093/humrep/dez162

INVITED COMMENTARY

The technical risks of human gene
editing
Benjamin Davies*
Welcome Centre for Human Genetics, University of Oxford, Oxford OX3 7BN, UK.

*Correspondence. Email: [email protected]

Submitted on May 29, 2019; resubmitted on July 2, 2019; editorial decision on July 15, 2019

ABSTRACT: A recent report from Dr He Jiankui concerning the birth of twin girls harbouring mutations engineered by CRISPR/Cas nucleases
has been met with international condemnation. Beside the serious ethical concerns, there are known technical risks associated with CRISPR/Cas
gene editing which further raise questions about how these events could have been allowed to occur. Numerous studies have reported
unexpected genomic mutation and mosaicism following the use of CRISPR/Cas nucleases, and it is currently unclear how prevalent these
disadvantageous events are and how robust and sensitive the strategies to detect these unwanted events may be. Although Dr Jiankui’s study
appears to have involved certain checks to ascertain these risks, the decision to implant the manipulated embryos, given these unknowns, must
nonetheless be considered reckless. Here I review the technical concerns surrounding genome editing and consider the available data from Dr
Jiankui in this context. Although the data remains unpublished, preventing a thorough assessment of what was performed, it seems clear that
the rationale behind the undertaking was seriously flawed; the procedures involved substantial technical risks which, when added to the serious
ethical concerns, fully justify the widespread criticism that the events have received.

Key words: gene editing / CRISPR / nuclease / mutagenesis / Cas9

Introduction
The development of site-specific nucleases over the last decade now
makes it possible to introduce precise changes into the DNA sequence
of our cells (Carroll, 2017). In particular, RNA-guided CRISPR/Cas
nucleases are very easy to design against specific genomic target
sequences and high efficiencies of mutagenesis can be achieved (Sander
and Joung, 2014). These qualities are making the therapeutic applica-
tion of CRISPR/Cas nucleases to tackle genetic disease feasible, and
there has been a diverse range of success stories published in pre-
clinical models (Porteus, 2019). For example, CRISPR/Cas9 nucleases
have been designed to ablate the mutation responsible for muscular
dystrophy, restoring normal gene expression (Long, et al., 2014). In
both small (Nelson, et al., 2016) and large (Amoasii, et al., 2018) animal
models, viral delivery of these nucleases into the diseased muscle was
shown to restore muscle condition and strength. Such studies typify the
exciting new field of therapeutic gene editing and highlight its potential
in tackling genetic disease.

The target cell for therapeutic gene editing needs to be carefully
considered to maximise the therapeutic potential and its longevity.
Clearly, editing a stem cell population would have clear advantages for
a prolonged therapeutic effect. Indeed, CRISPR/Cas nucleases have
been introduced into haematopoietic stem cells to correct the under-
lying mutations responsible for sickle-cell anaemia (Vakulskas, et al.,

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2018) and there is thus considerable interest in this approach for long-
lasting treatments for genetic diseases of the blood.

The application of gene editing tools in somatic stem cell therapies
has raised the possibility that they could be applied in the ultimate
stem cell, the one-cell embryo, allowing the genetic correction to be
permanent and thus present in all cells of the resulting individual.
Furthermore, a successful genetic manipulation of the one-cell embryo
would lead to the presence of the genetic change in the germ cells of
the manipulated individual, and thus the inheritance of these genome
edits would be achieved. Early work introducing CRISPR/Cas nucle-
ases into the one-cell mouse embryo by microinjection demonstrated
that genetic modification of the whole organism could be achieved
efficiently and confirmed the inheritance of the genetic change in
subsequent generations (Wang et al., 2013).

CRISPR/Cas nucleases in human embryos
Reports soon emerged that CRISPR/Cas could indeed be used to
manipulate the human one-cell embryo. The first published stud-
ies used discarded tripronuclear zygotes and achieved mutagenesis
efficiencies of up to 50%, with specific gene editing (i.e. the incor-
poration of information from a co-injected repair template) occur-
ring at around 15% (Kang et al., 2016; Liang et al., 2015). This was
quickly followed by a further report of efficient mutagenesis in healthy

The technical risks of human gene editing 2105

Figure 1 Disadvantageous outcomes of CRISPR/Cas mutagenesis within the 1-cell embryo. (A) Off-target mutagenesis. In addition to
the correct mutagenesis event, similar sequences elsewhere in the genome are also mutated leading to unpredictable effects. (B) Mosaicism. Prolonged
activity of the nuclease within the developing embryo can lead to different mutations in different parts of the resulting individual. (C) Large deletions. The
left panel shows the intended mutagenesis event where a target gene is inactivated. The right panel shows a potential consequence of a large deletion
event, where a neighbouring gene is also inactivated. (D) On-site damage. The top panel shows the intended mutagenesis event, with a CRISPR/Cas
nuclease specifically recognising only the mutant allele, leading to its inactivation. The bottom panel shows the biallelic mutation that could occur if the
CRISPR/Cas nuclease is not able to discriminate the mutant sequence from the normal sequence.

two-pronuclear embryos (Tang, et al., 2017). Two more substantial
articles followed, both published in Nature (Ma et al., 2017; Fogarty
et al., 2017). The first demonstrated efficient correction of a dominant
pathogenic mutation at the MYBPC3 gene, encoding a cardiac myosin-
binding protein, in heterozygous human embryos, and proposed a
mechanism of inter-homologue repair using the wild-type allele as
a repair template (Ma et al., 2017). The second study focussed on
the use of the technology to explore the role of genes involved in
preimplantation human development and achieved targeted mutation
of the gene encoding the transcription factor OCT4 (POU5F1) in 71% of
manipulated embryos (Fogarty et al., 2017). More recent studies have
successfully achieved gene editing using exogenous repair templates to
repair pathogenic mutations (Tang et al., 2018) or to introduce specific
reporter sequences (Yao et al., 2018).

An alternative strategy for achieving site-specific change within the
genome has been reported: base editing, which relies upon the fusion
of enzymatic domains to the CRISPR/Cas machinery, capable of
chemically converting one nucleotide base to another (Rees and Liu
2018). Base editing has also been successfully applied within the human
one-cell embryo (Li et al., 2017; Liang et al., 2018), and pathogenic
mutations have been successfully corrected using this technology (Liang
et al., 2017; Zeng et al., 2018).

These studies demonstrate that it is now feasible to achieve muta-
tions and edits in human embryos at manageable frequencies and
suggest that the tools for therapeutic germline editing are now avail-
able. There are numerous ethical concerns surrounding this technology
which have been widely discussed and reviewed elsewhere (van Dijke
et al., 2018), but a major requirement before germline editing can be

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considered is to assess the safety of the manipulations. Investigations
of CRISPR/Cas9 mutagenesis both in cell culture experiments and in
embryos have highlighted a number of disadvantageous consequences,
where research is needed to assess and mitigate the risk and optimise
methods permitting the reliable detection of such events. There thus
remains an appreciable amount of technology development and opti-
misation to be done before therapeutic editing can be considered.

Off-target mutagenesis
Soon after the demonstration that CRISPR/Cas9 could be used for
targeted manipulation of the mammalian genome, reports emerged
that its use carries a risk of unintended mutagenesis at closely matched
genomic sequences (Fu et al., 2013). This so-called off-target muta-
genesis is also more pronounced than initially expected as the com-
monly used Cas9 enzyme can tolerate certain mismatches within its
targeting sequence (Fig. 1A). Many of the studies addressing off-target
mutagenesis have been performed in cell culture experiments where
the CRISPR/Cas enzymes are transfected into millions of cells, the
genomic DNA of which is then deep-sequenced to ascertain levels
of accuracy. These types of experiment may overestimate the risk of
off-target mutagenesis occurring when the CRISPR/Cas nucleases are
applied in a single cell, i.e. the one-cell embryo. Indeed, one carefully
controlled study in mouse used whole genome sequencing on a trio
(sequencing both parental and offspring DNA) to address off-target
mutagenesis resulting from a one-cell embryo microinjection experi-
ment and the authors were unable to detect any events in the founder
mice analysed (Iyer et al., 2018). In contrast, a larger study investigated

2106 Davies

founder rodent lines generated with multiple CRISPR/Cas9 enzymes
addressing a number of different target sequences and found that
almost 30% of the mutant lines harboured putative off-target mutations
(Anderson et al., 2018). Interestingly, base editors designed to convert
cytidine to thymidine residues were also found to have substantial
off-target effects when applied within the mouse one-cell embryo
(Zuo et al., 2019). Some of the human studies have also analysed
the resulting embryos for off-target effects. Candidate off-target sites,
localised using bioinformatic approaches, have been analysed by either
Sanger or next generation sequencing. One of the studies confirmed
an off-target mutation in two of the resulting embryos (Liang et al.,
2015). All of the other studies found no evidence for significant levels
of off-target mutation (Fogarty et al., 2017; Kang et al., 2016; Ma et al.,
2017; Tang et al., 2017). In contrast with the results from the cytidine
to thymidine base editors in mouse (Zuo et al., 2019), only low or
entirely absent levels of off-target mutagenesis were detected when
these reagents were applied in human embryos (Li et al., 2017; Liang
et al., 2017; Zeng et al., 2018).

The risk of off-target mutagenesis is thus clearly dependent upon
the target sequence and can be reduced by designing CRISPR/Cas
nucleases against truly unique genomic sequences; a number of online
algorithms are available to facilitate this improved design (Haeussler
et al., 2016; Hodgkins et al., 2015). These bioinformatic assessments
of off-target risk can be somewhat flawed however, as many of the
available tools do not take into consideration human genetic sequence
variation. A true off-target profile and thus risk assessment for a
selected CRISPR/Cas nuclease can only really be accomplished by
establishing a personalised genome. Indeed, studies have suggested that
naturally occurring human SNPs can alter the off-target landscape of
site-specific nucleases substantially (Lessard et al., 2017).

It has also been shown that the concentration and persistence of
the nuclease can increase the chance of off-target cleavage (Kim et al.,
2014; Zuris et al., 2015), and subsequently, a number of approaches
aimed at limiting the activity of the nuclease have been shown to reduce
the level of off-target mutation (Chen et al., 2016; Shen et al., 2018a).
Structural investigations and molecular evolution of the Cas9 nuclease
have enabled the design of variant sequences which show increased
levels of accuracy (Kleinstiver et al., 2016; Slaymaker et al., 2016) and
reduced risk of off-target mutagenesis in rodent models (Anderson
et al., 2018). Furthermore, orthologues of Cas nucleases from alter-
native bacterial species have been shown to have increased levels
of accuracy (Kim et al., 2016; Teng et al., 2018). A recent study
also improved the accuracy of CRISPR/Cas effectors by altering the
structure of its cofactor guide-RNA (Kocak et al., 2019), highlighting a
different approach to addressing this problem.

Taken together, there is a measurable risk of off-target mutagenesis
when applying CRISPR/Cas nucleases in cells and embryos, but there
has been significant technology refinement and bioinformatics tool
development to reduce these risks substantially. Nonetheless, more
research is needed in this area to assess the risk, improve the accuracy
of the enzymes and explore methods for detecting these off-target
events.

Mosaicism
The microinjection of CRISPR/Cas nucleases into mouse zygotes
soon revealed that the CRISPR/Cas nucleases frequently retain activ-

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ity after the first embryo cleavage event. Consequently, there is an
appreciable risk that individual cells of the two-cell or even four-
cell embryo harbour different combinations of wild-type and mutant
alleles. The resulting organism is thus frequently a genetic mosaic, with
different combinations of mutations in different parts of the animal
(Fig. 1B). This phenomenon is easily demonstrated by microinjection
experiments in mouse embryos which use CRISPR/Cas nucleases
designed against coat colour genes. Addressing the tyrosinase gene,
the loss of function of which leads to the albino phenotype, clear
somatic mosaicism was evident as the majority of founder offspring
show a speckled, patchy coat colour, rather than complete albinism
(Yen et al., 2014). Clear evidence of mosaicism was also found in the
human studies (Fogarty et al., 2017; Ma et al., 2017; Tang et al., 2017;
Yao et al., 2018).

Mosaicism may be tackled by altering the timing of the nuclease
activity within the one-cell embryo, effectively restricting its activity
to the one-cell stage. Reducing the half-life of the nuclease through
the use of a destabilised version of Cas9 was shown to reduce
mosaicism whilst editing non-human primate embryos (Tu et al., 2017).
Another improvement was achieved by delivering the nuclease to in
vitro fertilised embryos by electroporation, permitting the delivery of
the CRISPR/Cas machinery at a very early developmental stage, even
before pronuclei have formed (Hashimoto et al., 2016). One study
in human embryos was able to effectively eliminate mosaicism by
introducing the CRISPR/Cas reagents at the same time as performing
the fertilisation by intracytoplasmic sperm injection (Ma et al., 2017).

Large deletions and rearrangements
The mutagenesis occurring following the application of CRISPR/Cas
nucleases relies upon the innate DNA repair machinery of the tar-
get cells. Most frequently, the induced double-strand break (DSB) is
repaired by non-homologous end joining, which can lead to the intro-
duction of small deletions and insertions. The mutations are frequently
small in size, the most common being a single-nucleotide insertion or
deletion (Chakrabarti et al., 2018; Taheri-Ghahfarokhi et al., 2018).
Nonetheless, larger deletions do occur and there is evidence to suggest
that, on occasion, the repair event can result in large kilobase-scale
deletions. In one in vivo study, introducing CRISPR/Cas nucleases as
a virus to correct a mutation in the Otc gene, an appreciable rate
(6.5%) of disruptive large deletions was found (Yang et al., 2016). More
recently, various studies have observed large deletions occurring at
significant levels following the use of CRISPR/Cas9 in vitro (Kosicki
et al., 2018) and when applied within the one-cell embryo (Parikh
et al., 2015; Shin et al., 2017). These unexpectedly large deletions have
the capacity to delete whole genes or cause misregulation of nearby
expressed sequences (Fig. 1C).

In addition to large deletions, a mouse study has revealed that
complex rearrangements can occur following the application of nucle-
ases in the one-cell embryo (Boroviak et al., 2017). These events
seem particularly prevalent when using multiple CRISPR/Cas nucleases
which cleave in cis. The repair of the resulting two or more DSBs can
result in the deletion of the intervening sequence (which is often the
aim of the experiment), but also the inversion of the sequences can
occur, as well as many surprising duplications and insertion events.

Interestingly, the prevalence of these deletions and rearrangements
may have been underestimated since these complex events are often

The technical risks of human gene editing 2107

invisible to the molecular assays used to genotype the resulting muta-
tions. Simple PCR-based genotyping strategies can be compromised
by the deletion encompassing the primer binding sites. Short-read next
generation sequencing technologies are not well equipped to detect
and assess genomic inversions and duplications. Genomic technolo-
gies based on long-reads or more traditional assessment of target
locus integrity by Southern blotting or fluorescent in situ hybridisa-
tion analysis may help detection, but these methods are difficult to
apply and may not very applicable for genotyping embryo biopsy
material.

The difficulty in detecting large deletions has fuelled controversy
surrounding the inter-homologue repair mechanism proposed when
CRISPR/Cas9 nucleases are used to selectively ablate a pathogenic
mutation present heterozygously (Ma et al., 2017). It has been sug-
gested that the inability to detect a rearranged or damaged mutant
allele could lead to the misinterpretation that the allele has been
repaired from the intact wild-type allele (Adikusuma et al., 2018;
Egli et al., 2018), although follow-up analysis of the original study
provided evidence arguing against this explanation (Ma et al., 2018).
At the moment, we know too little about the dominant DNA repair
machinery active within the early preimplantation embryos and these
discussions highlight the requirement for further research to fully
establish what repair events are likely, how they can be harnessed
for therapeutic effect and how disadvantageous large deletions and
rearrangements of the target locus can be detected.

On-site damage and biallelic modification
The high efficiency of CRISPR/Cas nucleases frequently leads to the
mutagenesis of both autosomal copies of a target gene. Where a com-
plete loss of function is therapeutic, this is, of course, advantageous;
however, there are frequent situations where only one copy of a gene
needs to be addressed, in particular when trying to correct or ablate
dominant heterozygous mutations. Although CRISPR/Cas nucleases
can be designed against the mutated copy of a gene, the tolerance of
Cas9 for small mismatches may make it challenging to design nucleases
that can discriminate between a mutant copy and a normal copy of
a gene (Fig. 1D). Interestingly, in the human study which successfully
applied CRISPR/Cas nucleases to correct a dominant heterozygous
mutation in the MYBPC3 gene (Ma et al., 2017), the mutation chosen
for this proof-of-concept study was a 4-bp deletion. This relatively
large mutation allowed the nuclease to be designed specifically against
this mutant allele, thus eliminating the risk of mutating the wild-type
allele. The majority of disease-associated mutations, however, are
single-nucleotide changes where discrimination may be challenging, and
mutagenesis of the normal copy of the gene or even reprocessing
and subsequent mutagenesis of the correctly repaired mutation would
be expected to occur at appreciable frequencies. Development of
enzymes with a higher level of discrimination may help the selective
correction of the mutant alleles. The developed Cas enzymes and
orthologues with reported higher accuracy may be very useful in this
context (Kim et al., 2016; Kleinstiver et al., 2016; Slaymaker et al., 2016;
Teng et al., 2018).

Another approach to help improve the predictability of gene editing
outcomes is emerging from the analysis of large numbers of mutage-
nesis events. It has become clear that, for a specific target sequence,
certain mutational outcomes can be quite common. Part of the expla-

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nation for this lies in regions of microhomology that lie upstream and
downstream of the target site (Bae et al., 2014), but recent studies have
identified other key attributes of the underlying primary sequence that
can be used to predict the predominant pattern of repair (Allen et al.,
2018; Chakrabarti et al., 2018; Shen et al., 2018b; Taheri-Ghahfarokhi
et al., 2018). These studies demonstrate the power of using machine
learning tools and large data set analysis to help make CRISPR/Cas
mutagenesis more predictive. With the help of these new tools, it
could be possible to achieve the desired repair of a mutation simply by
design considerations alone. Indeed, one of the recent papers in this
area has already confirmed the feasibility of this approach by using the
predictive outcome of DSB processing to repair a pathogenic mutation
(Shen et al., 2018b).

The reported birth of gene-edited twins
Despite technical improvements addressing the shortcomings, there
remains uncertainty about the prevalence, extent and detection of
genomic damage and mosaicism. Considerable research and technical
development are needed to quantify and address the issues before
therapeutic gene editing can be considered. Given these unresolved
safety concerns, it was alarming to hear the reports emerging late
last year from the Southern University of Science and Technology,
Shenzhen, China, which suggested that human embryos had been
manipulated by CRISPR/Cas9, reimplanted into the mother and car-
ried to term. The principal scientist involved in this study, Dr He Jiankui,
reported his results at the Second International Summit on Human
Genome Editing in Hong Kong (National Academies of Sciences,
2019) and described the birth of twin girls, Lula and Nana, who both
carried CRISPR/Cas9-engineered mutations. Dr Jiankui attempted
to introduce loss-of-function mutations into the gene encoding the
CCR5 chemokine receptor, a co-receptor for certain subtypes of
HIV virus.

A naturally occurring CCR5 variant involving a deletion of 32 bp
introduces a premature STOP codon into the gene, resulting in the
expression of a truncated protein which is not able to act as an
HIV co-receptor. Subsequently, individuals homozygous for this so
called �32 mutation are resistant to infection by certain HIV subtypes.
There has been wide interest in this mutation, since an HIV-infected
patient has been effectively cured of viral infection by an allogeneic
stem-cell transplantation with haematopoietic stem cells from a donor
homozygous for this �32 CCR5 variant (Hutter et al., 2009). The aim
of Dr Jiankui’s study was to engineer loss-of-function mutations within
the CCR5 gene in human embryos generated by IVF from parents where
the father was infected with HIV. In doing so, the goal was to protect
the resulting embryos from HIV infection.

Flawed scientific rationale and experimental
design
The scientific rationale behind the study is questionable for a number
of reasons. Firstly, there are established protocols involving semen
washing which can be used to reduce the risk of infection when using
HIV-infected semen in assisted reproductive therapy (Zafer et al.,
2016). There appears no need to invoke genome editing for this
purpose. Secondly, CCR5 is a co-receptor for one subtype of HIV;
a different chemokine receptor, CXCR4, can also act as a co-receptor

2108 Davies

for different classes of HIV. In patients with CCR5 mutations, CXCR4-
tropic HIV subtypes can, albeit inefficiently, enter the cells through this
alternative receptor which would continue to be expressed (Agrawal
et al., 2004). Potentially the edited offspring would thus still be sus-
ceptible to HIV infection, despite engineered mutations in their CCR5
receptor gene.

With respect to the experimental design, the genome engineering
strategy adopted did not involve the incorporation of the naturally
occurring �32 mutation, despite the fact that a previous study in
human embryos showed successfully that the �32 mutation could be
incorporated at the CCR5 gene with a repair template (Kang et al.,
2016). Instead, a random mutagenesis approach was adopted, albeit at
the same position within the gene, which would be expected to lead to
the incorporation of de novo mutations within the gene. The biological
consequences of these novel mutations are impossible to predict and
could lead to a global CCR5 knockout by affecting mRNA or protein
stability.

Importantly, it has been suggested that stable expression of the CCR5
�32 variant may be important for HIV resistance (Agrawal et al.,
2007); thus, mutations that cause a global knockout might not lead
to the immunity which was the primary goal of the study. More
worrying, results from Ccr5 knockout mouse models are revealing
that there may be other consequences of CCR5 loss of function,
besides HIV entry. Ccr5 loss of function led to an increased severity
following infection of influenza virus (Falcon et al., 2015) and West
Nile virus (Durrant et al., 2015) and have implicated CCR5 function
in neuronal plasticity (Zhou et al., 2016) and recovery after brain injury
(Joy et al., 2019). Another recent study explored human UK Biobank
data to assess the impact of the CCR5 �32 variant on longevity
and reported an estimated 21% increase in mortality for individuals
homozygous for this mutation (Wei & Nielsen, 2019). Mutations at
CCR5, especially those with uncharacterised consequences on protein
stability and expression, might thus be expected to have unpredictable
and disadvantageous consequences.

Genotyping data from Dr Jiankui’s study
The results of Dr Jiankui’s study remain unpublished, and thus, the
primary data has not been peer-reviewed, making it difficult to assess
the study. However, from his presentation at the Second International
Summit on Human Genome Editing, we can learn some of the geno-
typing approaches he adopted in an attempt to mitigate the known
problems of CRISPR/Cas nucleases outlined above.

Following CRISPR/Cas9 injection, the embryos were cultured to the
blastocyst stage in vitro and the CCR5 genotype was then assessed by
preimplantation genetic diagnosis (PGD) using whole genome sequenc-
ing on trophectoderm biopsies. Two embryos were selected for trans-
plantation, the first of which (Lulu) harboured +1-bp and −4-bp
frameshift alleles, predicted to encode truncation mutations that are
similar but different from the natural occurring �32 mutation. As
discussed above, the biological activity of these mutations is completely
unclear.

The second embryo (Nana) harboured a 15-bp deletion on one
allele, whilst the other allele remained unedited. This 15-bp deletion
results in an in-frame deletion that would effectively remove five amino
acids from the mature CCR5 peptide chain. Dr Jiankui hypothesised
that this allele might destabilise the CCR5 structure near the HIV-

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binding site, but no experimental validation of this assumption was
presented. The in-frame deletion would almost certainly result in a
mature protein of completely unknown function; one could envisage
a dominant effect which might compromise the function of organ
systems in which CCR5 is known to play a role.

The fact that an embryo with an in-frame deletion and with a
remaining intact copy of the CCR5 gene was selected for implantation is
highly questionable. The unedited CCR5 allele would still be expressed,
and thus, a normal version of this chemokine receptor would remain
on the surface of T-cells, thus rendering the cells infectable by HIV, at
odds with the primary goal of the study.

Dr Jiankui highlighted in his presentation that the parents made
the decision concerning whether this embryo should be implanted,
although it is entirely unclear how the parents were advised or coun-
selled. Indeed, in the question-and-answer session that followed his
presentation, it became clear that the scientists directly involved in
the study may have performed the genetic counselling of the parents,
rather than correctly qualified and trained genetic counsellors.

Known problems of CRISPR/Cas
mutagenesis were addressed
Concerning off-target site mutagenesis, theoretical off-target sites were
bioinformatically assessed using the parental genomes to allow the
consequence of SNP variation to be considered. As mentioned above,
naturally occurring genome variation has the potential to alter the off-
target landscape substantially (Lessard, et al., 2017) and it is interesting
that this study did indeed ascertain a personalised genome for this pur-
pose. These theoretical off-targets were combined with experimentally
reported off-target sites from published in vitro experiments which
used the same target site to establish a panel of risk sites within the
genome. PGD using whole genome sequencing on biopsied cells from
the embryos revealed the presence of a single intergenic off-target
site within Lulu’s genome. A decision was thus made to knowingly
implant an embryo harbouring a CRISPR/Cas-induced mutation at
an off-target site; although localised to an intergenic region of the
genome, the functional consequences of this off-target mutation are
unclear. Intriguingly, this off-target was no longer detected in fetal
DNA analysis obtained from maternal blood during gestation and in
the cord and placental samples obtained at birth, indicating that the
initial PGD result might have been an artefact of whole genome ampli-
fication or reflect a low-level mosaicism in the trophectoderm cells
biopsied.

Concerning mosaicism, the CRISPR/Cas9 reagents were applied
during the ICSI fertilisation procedure using the same approach as
adopted previously (Ma et al., 2017). Similarly, despite what appears to
be mosaic sequence traces in the PGD Sanger sequencing, the results
of the whole genome sequences revealed equal proportions of two
alleles in each of the implanted embryos, suggesting that the embryos
were not genetic mosaics.

Concerning large deletion analysis, the presence of large deletions
was investigated by searching for chimeric sequencing reads arising
from two regions of the genome in cis. Interestingly, in one edited
embryo, not selected for implantation, a 6-kb deletion at the CCR5
target site was indeed found, confirming the prevalence of this kind
of repair outcome. The embryos chosen for implantation revealed no
evidence of large deletions.

The technical risks of human gene editing 2109

Despite these relatively thorough sequencing experiments which,
to a degree, seek to mitigate the known problems of CRISPR/Cas
mutagenesis outlined above, it is unclear how thorough and complete
the analysis was. Without an in-depth assessment of the primary data,
it is impossible to know whether the investigations were sufficient to
completely rule out non-specific mutagenesis events, large deletions or
genomic rearrangements.

Conclusions
Given the above technical and scientific issues, combined with the grave
ethical concerns (Krimsky, 2019), it is not surprising that Dr Jiankui’s
study has been widely condemned as being a reckless and premature
use of the technology. As a direct result, experts in the field have called
for a moratorium on germline editing (Lander et al., 2019), which has
been widely supported by the scientific community. It is important to
recognise however that this moratorium concerns the implantation of
edited embryos. Indeed, many scientists, clinicians, patient groups and
ethicists support that research is needed to understand and address
the risks involved. There is thus an understanding that this research
may necessitate the use of human embryos, and the argument has been
made that intentionally refraining from engaging in life-saving research is
not morally defensible (Savulescu et al., 2015). However, at this point in
time, at the very beginning of this emerging field with many unknowns,
the implantation of edited embryos cannot be justified.

Research is needed to fully understand the repair outcomes occur-
ring following the action of CRISPR/Cas9 nucleases within the early
embryo, especially those involving repair templates. Further improve-
ments in the accuracy of nucleases by either mutagenesis or molec-
ular evolution, or by mining the bacterial and archeal kingdoms for
alternative more accurate enzymes, would be advantageous. Improving
methodologies for the detection of off-target mutation and large
deletion and rearrangement events is also an area where continued
research would be beneficial. Furthermore, a more thorough exam-
ination of the effects of base editing technology would also be of
considerable interest.

PGD provides one alternative strategy for combatting genetic dis-
ease. However, there are concerns that, with reproductive age increas-
ing in the Western world and given that PGD is known to impact
reproductive success (Steffann et al., 2018), the number of viable
embryos from which healthy individuals can be selected may frequently
be too low for PGD selection to provide an effective solution. There
are, of course, also situations where PGD cannot provide a solution,
for example, where one patient is homozygous for a mutation. It is
thus not too far-fetched to imagine a growing necessity to consider
human germline editing in the future. It is thus clear that, in parallel with
research addressing the safety concerns, the debate exploring ethical
aspects of human germline editing must continue.

Funding
Wellcome Trust Core Award Grant (203141/Z/16/Z).

Conflict of interest
The author has nothing to declare.

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References
Adikusuma F, Piltz S, Corbett MA, Turvey M, McColl SR, Helbig KJ,

Beard MR, Hughes J, Pomerantz RT, Thomas PQ. Large deletions
induced by Cas9 cleavage. Nature 2018;560:E8–E9.

Agrawal L, Jin Q, Altenburg J, Meyer L, Tubiana R, Theodorou I,
Alkhatib G. CCR5Delta32 protein expression and stability are critical
for resistance to human immunodeficiency virus type 1 in vivo. J Virol
2007;81:8041–8049.

Agrawal L, Lu X, Qingwen J, VanHorn-Ali Z, Nicolescu IV, McDermott
DH, Murphy PM, Alkhatib G. Role for CCR5Delta32 protein in
resistance to R5, R5X4, and X4 human immunodeficiency virus type
1 in primary CD4+ cells. J Virol 2004;78:2277–2287.

Allen F, Crepaldi L, Alsinet C, Strong AJ, Kleshchevnikov V, De Angeli
P, Palenikova P, Khodak A, Kiselev V, Kosicki M et al. Predicting
the mutations generated by repair of Cas9-induced double-strand
breaks. Nat Biotechnol 2018.

Amoasii L, Hildyard JCW, Li H, Sanchez-Ortiz E, Mireault A, Caballero
D, Harron R, Stathopoulou TR, Massey C, Shelton JM et al.
Gene editing restores dystrophin expression in a canine model of
Duchenne muscular dystrophy. Science 2018;362:86–91.

Anderson KR, Haeussler M, Watanabe C, Janakiraman V, Lund J,
Modrusan Z, Stinson J, Bei Q, Buechler A, Yu C et al. CRISPR off-
target analysis in genetically engineered rats and mice. Nat Methods
2018;15:512–514.

Bae S, Kweon J, Kim HS, Kim JS. Microhomology-based choice of Cas9
nuclease target sites. Nat Methods 2014;11:705–706.

Boroviak K, Fu B, Yang F, Doe B, Bradley A. Revealing hidden com-
plexities of genomic rearrangements generated with Cas9. Sci Rep
2017;7:12867.

Carroll D. Genome Editing: Past, Present, and Future. Yale J Biol Med
2017;90:653–659.

Chakrabarti AM, Henser-Brownhill T, Monserrat J, Poetsch AR,
Luscombe NM, Scaffidi P. Target-specific precision of CRISPR-
mediated genome editing. Mol Cell 2018.

Chen Y, Liu X, Zhang Y, Wang H, Ying H, Liu M, Li D, Lui KO, Ding
Q. A self-restricted CRISPR system to reduce off-target effects. Mol
Ther 2016;24:1508–1510.

Durrant DM, Daniels BP, Pasieka T, Dorsey D, Klein RS. CCR5 limits
cortical viral loads during West Nile virus infection of the central
nervous system. J Neuroinflammation 2015;12:233.

Egli D, Zuccaro MV, Kosicki M, Church GM, Bradley A, Jasin M. Inter-
homologue repair in fertilized human eggs? Nature 2018;560:E5–E7.

Falcon A, Cuevas MT, Rodriguez-Frandsen A, Reyes N, Pozo F,
Moreno S, Ledesma J, Martinez-Alarcon J, Nieto A, Casas I. CCR5
deficiency predisposes to fatal outcome in influenza virus infection.
J Gen Virol 2015;96:2074–2078.

Fogarty NME, McCarthy A, Snijders KE, Powell BE, Kubikova N,
Blakeley P, Lea R, Elder K, Wamaitha SE, Kim D et al. Genome
editing reveals a role for OCT4 in human embryogenesis. Nature
2017;550:67–73.

Fu Y, Foden JA, Khayter C, Maeder ML, Reyon D, Joung JK. High-
frequency off-target mutagenesis induced by CRISPR-Cas nucleases
in human cells. Nat Biotechnol 2013;31.

Haeussler M, Schonig K, Eckert H, Eschstruth A, Mianne J, Renaud
JB, Schneider-Maunoury S, Shkumatava A, Teboul L, Kent J et al.
Evaluation of off-target and on-target scoring algorithms and

2110 Davies

integration into the guide RNA selection tool CRISPOR. Genome Biol
2016;17:148.

Hashimoto M, Yamashita Y, Takemoto T. Electroporation of Cas9
protein/sgRNA into early pronuclear zygotes generates non-mosaic
mutants in the mouse. Dev Biol 2016;418:1–9.

Hodgkins A, Farne A, Perera S, Grego T, Parry-Smith DJ, Skarnes
WC, Iyer V. WGE: a CRISPR database for genome engineering.
Bioinformatics 2015;31:3078–3080.

Hutter G, Nowak D, Mossner M, Ganepola S, Mussig A, Allers K,
Schneider T, Hofmann J, Kucherer C, Blau O et al. Long-term control
of HIV by CCR5 Delta32/Delta32 stem-cell transplantation. N Engl
J Med 2009;360:692–698.

Iyer V, Boroviak K, Thomas M, Doe B, Riva L, Ryder E, Adams DJ.
No unexpected CRISPR-Cas9 off-target activity revealed by trio
sequencing of gene-edited mice. PLoS Genet 2018;14:e1007503.

Joy MT, Ben Assayag E, Shabashov-Stone D, Liraz-Zaltsman S, Mazzitelli
J, Arenas M, Abduljawad N, Kliper E, Korczyn AD, Thareja NS et al.
CCR5 is a therapeutic target for recovery after stroke and traumatic
brain injury. Cell 2019;176:1143–1157.e1113.

Kang X, He W, Huang Y, Yu Q, Chen Y, Gao X, Sun X, Fan Y.
Introducing precise genetic modifications into human 3PN embryos
by CRISPR/Cas-mediated genome editing. J Assist Reprod Genet
2016;33:581–588.

Kim D, Kim J, Hur JK, Been KW, Yoon SH, Kim JS. Genome-wide
analysis reveals specificities of Cpf1 endonucleases in human cells.
Nat Biotechnol 2016;34:863–868.

Kim S, Kim D, Cho SW, Kim J, Kim JS. Highly efficient RNA-guided
genome editing in human cells via delivery of purified Cas9 ribonu-
cleoproteins. Genome Res 2014;24:1012–1019.

Kleinstiver BP, Pattanayak V, Prew MS, Tsai SQ, Nguyen NT,
Zheng Z, Joung JK. High-fidelity CRISPR–Cas9 nucleases with
no detectable genome-wide off-target effects. Nature 2016;529:
490–495.

Kocak DD, Josephs EA, Bhandarkar V, Adkar SS, Kwon JB, Gersbach
CA. Increasing the specificity of CRISPR systems with engineered
RNA secondary structures. Nat Biotechnol 2019.

Kosicki M, Tomberg K, Bradley A. Repair of double-strand breaks
induced by CRISPR-Cas9 leads to large deletions and complex
rearrangements. Nat Biotechnol 2018;36:765–771.

Krimsky S. Ten ways in which He Jiankui violated ethics. Nat Biotechnol
2019;37:19–20.

Lander ES, Baylis F, Zhang F, Charpentier E, Berg P, Bourgain C,
Friedrich B, Joung JK, Li J, Liu D et al. Adopt a moratorium on
heritable genome editing. Nature 2019;567:165–168.

Lessard S, Francioli L, Alfoldi J, Tardif JC, Ellinor PT, MacArthur
DG, Lettre G, Orkin SH, Canver MC. Human genetic variation
alters CRISPR-Cas9 on- and off-targeting specificity at therapeutically
implicated loci. Proc Natl Acad Sci U S A 2017;114:E11257–e11266.

Li G, Liu Y, Zeng Y, Li J, Wang L, Yang G, Chen D, Shang X, Chen J,
Huang X et al. Highly efficient and precise base editing in discarded
human tripronuclear embryos. Protein Cell 2017;8:776–779.

Liang P, Ding C, Sun H, Xie X, Xu Y, Zhang X, Sun Y, Xiong Y, Ma W,
Liu Y et al. Correction of beta-thalassemia mutant by base editor in
human embryos. Protein Cell 2017;8:811–822.

Liang P, Sun H, Zhang X, Xie X, Zhang J, Bai Y, Ouyang X, Zhi S, Xiong
Y, Ma W et al. Effective and precise adenine base editing in mouse
zygotes. Protein Cell 2018;9:808–813.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

.

Liang P, Xu Y, Zhang X, Ding C, Huang R, Zhang Z, Lv J, Xie X,
Chen Y, Li Y et al. CRISPR/Cas9-mediated gene editing in human
tripronuclear zygotes. Protein Cell 2015;6:363–372.

Long C, McAnally JR, Shelton JM, Mireault AA, Bassel-Duby R,
Olson EN. Prevention of muscular dystrophy in mice by CRISPR/
Cas9-mediated editing of germline DNA. Science 2014;345:
1184–1188.

Ma H, Marti-Gutierrez N, Park S-W, Wu J, Hayama T, Darby H,
Van Dyken C, Li Y, Koski A, Liang D et al. Ma et al. reply. Nature
2018;560:E10–E23.

Ma H, Marti-Gutierrez N, Park SW, Wu J, Lee Y, Suzuki K, Koski A,
Ji D, Hayama T, Ahmed R et al. Correction of a pathogenic gene
mutation in human embryos. Nature 2017;548:413–419.

National Academies of Sciences, Engineering, and Medicine, Policy and
Global Affairs. The National Academies Collection: Reports funded
by National Institutes of Health. In: Second International Summit on
Human Genome Editing: Continuing the Global Discussion: Proceedings
of a Workshop-in Brief . US: National Academies Press, 2019

Nelson CE, Hakim CH, Ousterout DG, Thakore PI, Moreb EA,
Castellanos Rivera RM, Madhavan S, Pan X, Ran FA, Yan WX et al.
In vivo genome editing improves muscle function in a mouse model
of Duchenne muscular dystrophy. Science 2016;351:403–407.

Parikh BA, Beckman DL, Patel SJ, White JM, Yokoyama WM.
Detailed phenotypic and molecular analyses of genetically modi-
fied mice generated by CRISPR-Cas9-mediated editing. PLoS One
2015;10:e0116484.

Porteus MH. A new class of medicines through DNA editing. N Engl J
Med 2019;380:947–959.

Rees HA, Liu DR. Base editing: precision chemistry on the genome and
transcriptome of living cells. Nat Rev Genet 2018;19:770–788.

Sander JD, Joung JK. CRISPR-Cas systems for editing, regulating and
targeting genomes. Nat Biotechnol 2014;32:347–355.

Savulescu J, Pugh J, Douglas T, Gyngell C. The moral imperative
to continue gene editing research on human embryos. Protein Cell
2015;6:476–479.

Shen C-C, Hsu M-N, Chang C-W, Lin M-W, Hwu J-R, Tu Y, Hu
Y-C. Synthetic switch to minimize CRISPR off-target effects by
self-restricting Cas9 transcription and translation. Nucleic Acids Res
2018a.

Shen MW, Arbab M, Hsu JY, Worstell D, Culbertson SJ, Krabbe
O, Cassa CA, Liu DR, Gifford DK, Sherwood RI. Predictable and
precise template-free CRISPR editing of pathogenic variants. Nature
2018b;563:646–651.

Shin HY, Wang C, Lee HK, Yoo KH, Zeng X, Kuhns T, Yang CM, Mohr
T, Liu C, Hennighausen L. CRISPR/Cas9 targeting events cause
complex deletions and insertions at 17 sites in the mouse genome.
Nat Commun 2017;8:15464.

Slaymaker IM, Gao L, Zetsche B, Scott DA, Yan WX, Zhang F. Ratio-
nally engineered Cas9 nucleases with improved specificity. Science
2016;351:84–88.

Steffann J, Jouannet P, Bonnefont JP, Chneiweiss H, Frydman N. Could
failure in preimplantation genetic diagnosis justify editing the human
embryo genome? Cell Stem Cell 2018;22:481–482.

Taheri-Ghahfarokhi A, Taylor BJM, Nitsch R, Lundin A, Cavallo AL,
Madeyski-Bengtson K, Karlsson F, Clausen M, Hicks R, Mayr LM et al.
Decoding non-random mutational signatures at Cas9 targeted sites.
Nucleic Acids Res 2018;46:8417–8434.