# Despite medical and ethical complications, is xenotransplantation a potential solution to human organ shortages?

### ABSTRACT

Xenotransplantation is a medical procedure, whereby animal organs are transplanted into humans. The organ crisis is an ever prominent issue, as globally the demand for healthy, well-functioning organs significantly exceeds the supply. The process of xenotransplantation has the potential to curb this issue, as well as allowing ethnic minorities to access transplanted organs more easily, and potentially limit the number of sales on the black market. However, there are a multitude of ethical and medical considerations which have to be accounted for. The rejection of a foreign organ remains an impending issue, as well as the contraction of Porcine Endogenous Retrovirus, which could have unknown effects on humans. There are many ethical arguments which oppose xenotransplantation, such as the role and treatment of transgenic animals, and the righteousness of exploiting animals for human benefit. These benefits and risks will be evaluated to determine whether xenotransplantation is a potential solution to human organ shortages.

### INTRODUCTION

Xenotransplantation is the scientific fusion of species by transplanting organs or grafts from one genus to another, such as animals to humans (US Food and Drug Association, 2019). The largest hurdle in transplantation is that demand for human organs worldwide exceeds supply drastically, generating a need for alternative sources of organs (Ekser, Cooper and Tector, 2015). As a consequence of this, xenotransplantation has gained support from an increasing number of medical professionals. However, there are numerous medical and ethical implications which have to be assessed in order to determine the plausibility of xenotransplantation in a clinical environment.

Organ xenotransplantation was first hypothesized by Professor Keith Reemtsma, in the late 1960’s. He claimed that chimpanzee kidneys could function as human kidneys due to the close evolutionary relationship between the two species, and carried out the first organ xenotransplantation (Cooper, Ekser and Tector, 2015). Between 1963 and 1964, 13 kidney xenotransplantations were carried out, with the majority of the patients dying weeks after the surgery. The cause of death within these patients was either organ rejection or infection, which highlighted the complexity of the human immune system (Berki, 2018).

Figure 1: Complex mechanism of hyperacute rejection in humans (Upadhyay, 2019)

Shortly after this, the first immunosuppressive drug, cyclosporine, had been invented. Immunosuppressive drugs reduce the strength of the immune system, and hence provide doctors with some hope, as potentially, if the immune response to the foreign tissues could be hindered, then the organ may not be rejected. Unfortunately, this proved to be unsuccessful as well, as the immunosuppression was not enough to prevent organ rejection (Berki, 2018). Dr. Thomas E. Starzl attempted liver transplants between humans and nonhuman primates, which resulted in only one patient living for a further 70 days after surgery (Cooper, Ekser and Tector, 2015). The failure of this experiment meant the study was unable to proceed to a clinical trial, due to the high risk. In 1997, a ban was initially placed on xenotransplantation research with concerns growing about the contraction of Porcine Endogenous Retrovirus (PERV) (Science Learning Hub, 2011). The ban was set in place in order to establish a full legal framework which highlighted all the issues presented by animal-to-human transplants (Science Magazine, 1997). However, some countries including the United Kingdom, United States and New Zealand now allow xenotransplantation research to occur, but only on an individual basis (Science Learning Hub, 2011).

### MAIN BODY

#### Potential for xenotransplantation

Xenotransplantation has clear life saving benefits. Surgeons believe that xenotransplantation could be used in babies born with congenital heart defects since that mortality rate exceeds 50%. A pig’s heart could keep a baby alive, as a replacement for the failing heart until a human heart donor is found (Weintraub, 2019). As 40,000 babies are born with a congenital heart defect in the United States each year, the success of transplanting animal organs could potentially help save many lives (Centers For Disease Control and Prevention, 2019).

Additionally, xenotransplantation could be particularly beneficial for those of ethnic minorities, where locating an organ of similar tissue type is a heightened problem (Morgan et al., 2016). For example, the Asian and Black populations in the UK are considered minorities, and, in 2013, patients had to wait one year longer than the general population for a kidney-only transplant (Morgan et al., 2016). This problem is then heightened due to the fact that such ethnic minorities constitute a significantly larger proportion of the organ donation waitlist, as they are three to four times more likely to develop end-stage renal failure and therefore require a kidney transplant. Furthermore, to reduce the risk of rejection, donors and recipients must have matching blood group and antigen tissue type, which is generally hard to coordinate with an ethnic minority group. All of these factors place tremendous stress on the supply of organs, so xenotransplantation could help alleviate this issue.

In developing countries, citizens with a relatively low-income status may decide to engage in the black market organ trade, by selling organs such as kidneys and livers, on the black market. This is done in order to afford other goods, needed for survival. The payment which donors receive for the organ varies greatly between countries and the organ itself, however it is approximately $1000 and$5000 (US dollars). However, the amount paid to the organ donors is insignificant compared to the money taken by the agents who facilitate the organ sale, who enjoy drastically larger profits by selling these organs, in the United States, for $100,000 to$200,000 (US dollars), therefore presenting organ trade on the black market as a highly futile endeavor, even for those of low-income status (Nullis-Knapp, 2004).

Those receiving organs also place their own health at risk, which could impose a cost on them later in life. Purchasing from untrustworthy sources carries a multitude of risks, including that there are usually no regulations with no real oversight and there is a possibility that the organ may be diseased, causing the transplant to fail (Taylor, 2006). With this in mind, using tissue and organs from primates, rather than those donated by humans using the black market, could increase the number of successful transplants. Despite this, organ trade on the black market may still continue; not due to a need for the supply of organs, but a need for the money. The reason why the majority of sales on the black market occur is primarily financially related, and therefore, even if xenotransplantation reduced the dependency on organ trafficking, the trade would most likely still continue (Maher, 2016).

Xenotransplantation would help to lessen the organ crisis, where there is a sufficient shortage of transplantable organs. As of March 2020, there are over 112,000 people on the organ waiting list in the United States, with another person being added every 9-10 minutes. Approximately 20 people a day die due to their place on the waiting list (U.S. Health Resources and Services Administration, 2020). If organs were taken from animals instead of relying on human organs, which have to be matched specifically to its recipient, this could help significantly reduce the number of people on the waiting list.

For xenotransplantation, pigs, who are part of the Suidae family, are the preferable animal for organ donation due to the anatomical similarity of their size, shape and weight to humans (Griesemer, Yamada and Sykes, 2014). Additionally, they are rapidly reproducing animals, and offspring quantities are relatively large, so there will always be an abundant supply of the animal.  Since many pigs are slaughtered annually for human consumption, there would be less of an ethical objection to using their organs for treating diseases, with exceptions to vegetarians or vegans whose reason for the diet is due to preventing animal deaths. Nevertheless, it can be argued that the benefits of saving lives is far greater than the discomfort and dispute caused amongst animal rights activists, and therefore the use of xenotransplantation should be accepted.

Furthermore, it is clear from past studies that pigs are viable for genetic engineering. In 2018, researchers from the University of Edinburgh successfully bred pigs that were completely immune to porcine reproductive and respiratory syndrome (PRRS) (Devlin, 2018). By removing the CD163 protein (the cellular receptor for PRRS) or the SRCR5 region (interaction site for the virus) on the genome, the pigs no longer showed symptoms of PRRS, such as reduced litter size and breathing problems (Yang et al., 2018). Therefore, if the animals needed to have their genome edited in order to produce offspring which had more favourable or useful organs in terms of xenotransplantation, it could be done. However, this does inevitably result in ethical questions surrounding the righteousness of this action and the destroying of their natural state in order for humans to benefit from.

Xenotransplantation from non-human donors also eliminates the dependency on grieving families, avoiding the pressure to consult the relatives of the deceased. Although, human organ donations are still preferable over other species and therefore consulting relatives should still be encouraged. With xenotransplantation, nurses and carers may be able to prepare for the surgery beforehand, which may save in administrative costs, as the process is quickened and becomes easier to coordinate. Furthermore, the legal and operational problems that correspond with the use of organs from recently deceased donors will be reduced, which would be of particular importance in countries where  organ donation from the deceased is complicated due to a network of cultural taboos. For example, some South Asia Muslim Ulemas oppose donation from living and deceased human donors as they view the human body asa trusteeship from God thatmust not be desecrated following death (Bruzzone, 2008). However, they do encourage xenotransplantation research. This does stimulate another ethical dilemma, which is if a certain ethnic group refuses xenotransplantation on religious or cultural grounds, and are in need of an organ, some may argue they should be prioritized for allotransplantation – the transplantation of human organs. Since, this would maximise the benefit of the majority of parties. If xenotransplantation was to become a reality, the allocation of human organs and animal organs would have to be thoroughly evaluated. Systematic and fair approaches would have to be implemented to ensure the maximum number of people are receiving organs, and allocation is based on clinical need.

#### Concerns with Xenotransplantation

The largest ethical complication with xenotransplantation is possible animal mistreatment. It is argued that it is a violation of nature to exploit animals for their organs. The welfare of pigs is the primary concern for many  animal right activists, as they would be subject to a series of trials and experiments even before xenotransplantation could become commercially available (Manesh, 2014). Primates and pigs are exceptionally sensitive and socially complex animals and the conditions for mass breeding may place tremendous emotional stress on them (Marino and Colvin, 2016). Additionally, it is completely unethical to keep animals in battery farm environments. Some animal advocates fear that if xenotransplants become a clinical reality, there is the prospect of “biological factories” in which animals are bred purely as a primary source of organs for humans, which is fundamentally exploiting the animals and may cause an uproar of opinion in the public.

Another key ethical argument surrounds whether it is right to exploit another species for the benefit of humans. Those who oppose anthropocentric views (that humans are the most important beings) may be disturbed by the concept of placing the survival of humans above that of another species, and would greatly disagree with such violation of animal rights. This is undoubtedly an area of great debate.

There are also several religious concerns surrounding the procedure. In some cultures, and ethnicities’, such a Judaism and Islam, pigs are considered unclean animals, and to receive an organ for transplant from this animal could be considered sinful. The acceptance of xenotransplantation depends on the culture, as the chairman of the Sharia council states that, under Islamic law, while eating pig is forbidden, other uses are to be allowed, such as xenotransplantation. Jewish communities have a similar mindset. However, the Union of Muslim Organisations believe xenotransplantation could be allowed as long as pigs or any other prohibited animal is not used (Nuffield Council on Bioethics, 1997).

Pigs are social animals, who require others around them to feel at ease (Landsberg and Denenberg, 2014). The necessary conditions required for mass breeding (isolation and caesarean section births) would place tremendous emotional stress on them. Some may have a utilitarian approach, whereby actions are considered ethical if they reduce the harm as much as possible. Thus, the stress placed on the animals would be warranted if it resulted in a solution to combating the organ crisis.

While the organs may be relatively close in size and shape, there are numerous other factors which provide great disparity. Physiological differences between each species need to be properly accounted for. The life span of a pig is approximately 15 years (Dooldeniya and Warrens, 2003). Therefore, the use of the pig’s heart will only provide the patient with up to 15 more years before another transplant is needed? Furthermore, some proteins specific to porcine species are molecularly incompatible for humans, which could cause malfunction of important regulatory processes (Candinas, 2000). Lastly, porcine organs operate under different hydrostatic pressures than humans as well as having an elevated body temperature of 39°C, which would be considered dangerous for humans.

Since the organ is from another species, there would undoubtedly be risks which are unknown and not accounted for, which the patient must be fully aware of when agreeing to receive a transplant. Consent is key in regards to xenotransplantation. However, when a patient is in need of an organ, questions may be raised about their emotional stability and capacity to make an informed decision (NYU School of Medicine, 2013). Receiving an organ can be highly long-awaited, and therefore patients may be desperate and not fully aware of the implications.

Porcine organs are considered to be the most favourable for xenotransplants as they are similar in size and the animals can be bred on mass. However, the clinical use of these organs has been hindered, due to the prominent risk of xenozoonosis (the contraction of a harmful disease through transplantation of an animal tissue into a human). All recipients of porcine organs will be exposed to the risk of contracting Porcine Endogenous Retrovirus (PERV), a pig virus, from the same family as HIV, which splices into human DNA (Blusch, Patience and Martin, 2002). PERV-A and PERV-B are polytropic, meaning they are to infect a range of host species including humans, whereas PERV-C is ecotropic and restricted to infecting pigs.

Clinically, the greatest complication to overcome in xenotransplantation is organ rejection. Humans have a complex immune system, so the response to foreign, newly transplanted organs is immediate. This results in complete destruction. of the organ and has been termed hyper-acute rejection (Dorling et al., 1997). This form of rejection occurs when the antigens are completely unmatched and will begin within minutes of transplantation. Consequently, The tissue must be removed immediately so the recipient does not die. Hyperacute rejection involves ischemia (starving the tissue of blood) and necrosis (death of the cells in the tissue) of the organ, which it thought to be the result of cytotoxic antibodies in the recipient (Vitak, 2014). Clearly, if the organ dies, it will no longer function properly which can be lethal for patients, as all organs are needed to maintain bodily functions. This poses serious complications to the clinical reality of the procedure.

#### REALITY OF XENOTRANSPLANTATION

Since organ rejection is the principal medical complication of xenotransplantation, many scientists are exploring new methods of eliminating this risk, the first being the use of transgenic pigs. This is where a gene from one species, such as pigs, is transferred into another species genome, such as humans. This allows the transgenic pigs, bred for xenotransplantation, to contain a human gene which produces a complement regulating protein (CD55 and CD59). Kidneys from pigs which were expressing the gene coding for CD55 showed increased protection against hyperacute rejection, even in the absence of immunosuppression, hence reducing the immune response to transplanted organs (Ménoret et al., 2004).

One biomedical research firm looking to help solve the problems of organ rejection and PERV contraction is eGenesis Bio, founded in 2015 by CRISPR and genomics pioneers, Dr. Luhan Yang and Dr. George Church. eGenesis aims to create transgenic pigs through the use of CRISPR-Cas 9 technology (eGenesis Bio, 2020). CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) is a gene-editing tool, which allows scientists to modify and selectively delete, or correct, a disease-causing abnormality in a specific DNA segment (Kararoudi et al., 2018). Two RNA strands bind to form a complex with a protein called Cas-9. This natural process was first discovered in the immune response of bacteria, which form Cas-9 protein if invaded by a virus. This was first experimentally proven in 2007, when researchers used Streptococcus thermophilus (Ishino, Krupovic and Forterre, 2018). When the matching sequence known as the guide RNA, finds its target within the viral genome, the Cas-9 cuts the target DNA, disabling the gene. Hence, the Cas-9 protein can be viewed as molecular scissors. Scientists have used this technology by altering the guide RNA sequences to cut and modify the genome of other species.

Figure 2: The use of CRISPR for xenotransplantation (eGenesis, 2017)

The contraction of PERV is a large hurdle in assessing the reality of xenotransplantation. Although so far no humans have contracted PERV in a clinical setting, it has been proven that transmission is possible, which could lead to immunodeficiency and tumorigenesis, as per other similar retroviruses (Niu et al, 2017). In a study conducted in 2017, scientists successfully created PERV-inactivated primary porcine cells, through the combined use of CRISPR, apoptosis inhibitors and growth factors. They did this by using modified fibroblasts (a cell which produces collagen or other fibrous tissues) to produce embryos through somatic cell nuclear transfer (SCNT), which were then implanted into surrogate sows (Niu et al, 2017). The offspring of these pigs were completely PERV-free.

Another possibility which has recently been explored is the use of pigs without the gene coding for α1,3-galactosyltransferase. These pigs are known as α1,3-galactosyltransferase gene-knockout pigs (GTKO) (Ekser, Cooper and Tector, 2015). The use of such porcine tissues have proven to be extremely successful in a study conducted in 2005, where the elimination of the galactose-alpha 1,3-galactose gene prevented hyperacute rejection and extended survival of pig hearts in baboons for 2-6 months (Tseng et al., 2005). Furthermore, after hyperacute rejection is prevented, it would be expected that another form of rejection occurs – acute humoral xenograft rejection (AHXR) (Ekser, Cooper and Tector, 2015). Similarly to hyperacute rejection, it is the result of endothelium and innate immune cell activation which collectively destroy the graft. However, GTKO pigs expressed prevention of this form of rejection as well (Ekser, Cooper and Tector, 2015).

Regardless of all of this progress, graft vasculopathy has been noted in some attempts of pig heart xenotransplantations (Ekser, Cooper and Tector, 2015). This is also seen in allotransplantation and is categorized by a thickening of the innermost membrane (initima) of major heart arteries (Ramzy et al., 2005).

Despite this, a study in 2011 is providing some hope on this matter. GTKO pig organs were transplanted into baboons and given immunosuppression as well. This resulted in a survival of 236 days for the cardiac xenograft (Mohiuddin et al., 2011). However, in both of these case studies, the cause of death of the baboons was due to thrombotic microangiopathy and coagulation dysregulation, which seems to be hindering the baboons potential longer life. Coagulation dysregulation, which can result in thrombotic microangiopathy (TAM) is when there is thrombosis (a blood clot in the blood vessels) (Ekser, Cooper and Tector, 2015). TAM in the graft may be observed as fibrin deposition or platelet aggregation which will cause the blood vessel to clot. Eventually, this will block the vessels, leading to ischemic damage where tissues do not receive a sufficient blood supply and can result in a necrotic organ (Ezzelarab et al., 2009).

Additionally, coagulation in xenotransplantation presents another hurdle. When pancreatic islet tissues are transplanted, a response known as the instant blood-mediated inflammatory reaction (IBMIR) occurs which can lead to thrombosis amongst other complications (Goto et al., 2008). Depending on the tissue, the coagulation response to xenotransplantation can differ, with renal xenografts much more likely to develop coagulopathy (when the blood’s ability to clot is impaired) than cardiac tissues (Cowan, Robson and dʼApice, 2011). Although this occurs in all forms of transplantation, the problem is heightened in xenotransplantation due to antibodies which are present in humans and the clear differences in species. Attempts to overcome this issue have involved the use of common anticoagulants including Heparin, but have proved unsuccessful (Byrne et al., 2005). Therefore, resolving this issue may therefore lie in the hands of genetic engineering, however would require more attention and research.

There are multiple concerns surrounding the use of transgenic pigs for xenotransplantation. Firstly, some may argue that moving or altering the genes destroys the integrity of species as natural organisms, creating atypical hybrids, which cannot be considered as animals (Lamb, 1997). Furthermore, some may argue that altering the genome of another species without complete knowledge of possible side effects, which may cause significant harm to the animal is completely unethical. Genetically modified animals can also be patented by humans (Dresser, 1988). Deeming transgenic pigs as “human inventions” strips away their identity as a species and intellectual beings, and reinforces the idea that animals are a property of humans. Although the majority of the public shows acceptance of this idea, in the event it will benefit humans, it may spark huge debate amongst opinionated animal rights advocates, and management of such groups would have to be considered (Lesser, 2006).

CRISPR could be the future of biomedicine, but it is unreasonable to assume this tool could instantly be used in xenotransplantation. Many years of research would have to be conducted, with an abundance of support and funding from government agencies, which could prove to be quite challenging. For example, in Canada, a moratorium was held which prevented the transplantation of porcine organs from entering into a clinical trial (Cheng, 2015). This was to continue until the public was assured that the potential disease transfer of PERV (Porcine Endogenous Retrovirus) from pigs to humans could be satisfactorily managed. Canada has now imposed regulation whereby those wishing to pursue a clinical trial involving xenotransplantation must seek approval by Health Canada, however still, nearly 20 years later, no trials have been approved (Health Canada, 2010). In the UK, a portion of tax-payers money would have to be allocated to conduct the trials, which could spark a dispute between ethical and religious groups who are unhappy with the allocation of tax revenue. Religions such as Jainism do not believe in xenotransplantation. However, when it comes to life or death, some ethical obligations may have to be reconsidered. As xenotransplantation pioneer David Cooper quotes, “When it hits you personally and you are going to die, I think your attitude changes” (Weintraub, 2019).

### CONCLUSION

Xenotransplantation could be a revolutionary procedure. It involves transplanting porcine or primate organs into humans in need of a new organ, with the potential to solve the NHS organ crisis, resolve the black-market trade of organs and save thousands of lives worldwide. Despite being surrounded by medical issues, several solutions have arisen in recent years, including using transgenic pigs, though CRISPR technology. These solutions still require an abundance of thorough research and funding for clinical trials. Therefore, the main complications associated with the procedure are the ethical controversies, from the role animals would play in the process to religious limitations. If society can acquiesce with the moral and ethical obligations that are linked to xenotransplantation, it undoubtedly has the potential to develop into a legitimate solution to organ defects.

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