Xenotransplantation: A potential solution to human organ shortages

Highlights:

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• Xenotransplantation does not only refer to organs but also tissues and cells
• Genetically-engineered pigs represent a good potential, cost-effective source of organs
• CRISPR gene editing technique is making the process of creating genetically modified pigs an easier, cost-effective feat
• Immunological reactions to the xenograft involve elements from the innate and adaptive immune systems
• Economic, ethical, societal, and religious barriers are other hurdles facing xenotransplantation
• Xenotransplantation research faces difficulties, as some governments place restrictions on this type of research
• The three main types of xenograft rejection are hyperacute, acute humoral, and acute cellular xenograft rejection

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Introduction

Xenotransplantation is a term meaning cross-species transfer of living cells, tissues, or organs. In other words, it describes any procedure that involves transplanting, implanting, or infusing live cells, tissues, or organs between different species (like nonhuman-to-human transfer). The transplanted cells, tissues, or organs are called xenografts or xenotransplants. The word “Xenos" originates from Greek, meaning "foreign". Xenotransplantation is not a new concept and has been first mentioned over 350 years ago in the context of xenotransfusion of blood from lambs to humans. The twentieth century has seen several xenotransplantation trials, like transplanting a rabbit kidney to a human in 1905. In January 2022, a 57-year-old man received a heart transplant from a genetically modified pig. Two months later, he passed away, not due to rejection but rather due to a porcine virus that infected the heart before the transplantation procedure.

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Xenotransplantation: A promising technique to fill a gap

Organ transplants carry many risks, but they could be lifesaving for people on many occasions, given the availability of organs. Current data from the literature suggests the presence of organ donor shortage, negatively impacting people with terminal organ failure (1). Research shows that the number of human organs available for transplantation is as low as 5% of the required number (2). Data from the World Health Organization reported similar findings, indicating that the number of annual organ transplants is <10% of global needs (3). In the United States, thirty patients each day either die or are removed from the organ transplant “waiting list” because their sickness makes them too weak or sick to undergo the procedure. The shortages in available organs impact the lifespan and longevity of individuals who need them. For example, evidence indicates that kidney transplant patients live twice as long as dialysis patients, including having a better quality of life (1). The latter was true regardless of age.

In addition to the above, having readily available organs for patients would save significant costs (1, 3). For example, the literature shows that a patient with a transplanted kidney would save around $1 million in lifetime costs compared to dialysis patients (1). In the United States, costs associated with end-stage organ failures mount up to almost 60% of Medicare’s budget (3). This highlights the potential of xenotransplantation in covering this gap.

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Potential sources for xenografts

Since the 17^th century, the use of various animals to provide xenografts has been explored. From that period to the 20^th century, primates like chimpanzees and baboons and animals like rabbits, frogs, sheep, chickens, and cats were tested as potential donors (4).

In the 21^st century, advances in research and genetics allowed the creation of genetically modified animals (4). The pig is an example of a genetically-engineered organ donor animal (3). Pigs are an excellent organ-source candidate for numerous reasons, like anatomical and physiologicalsimilarities to human organs, ease ofbreeding, avoidance of potential, life-threatening complications to human donors, and elimination of illegal trafficking. In addition to the above, genetically-engineered pigs offer advantages over primates. Some advantages over the chosen primate (baboon) are covered in the table below (1).

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In the early 2000s, xenotransplantation research was stimulated by the introduction of Gal-knockout pigs, which improved survival in heart and kidney xenotransplantations (5). Galactose-α1,3-galactose (Gal) antigen knockout and identifying other xenoantigens presented opportunities for further advancement in the field. In one case of heart xenograft transplantation from a genetically modified pig to a nonhuman primate (NPH), the NPH survived for over 900 days (5). The effect was sustained as the NPH was given a regimen based on a chimeric anti-CD40 monoclonal antibody, preventing humoral rejection and coagulation.

Genetic modification of pigs usually involves two key processes, including the deletion of one or more of the three key pig antigens and the insertion of a human transgene to protect against human complement and/or coagulation pathways (1). The outcome of the process is the reduced capacity of the human immune system to react to the xenograft. The use of technologies like Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) makes the genetic modification process easier, cheaper, and faster.

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Barriers to xenotransplantation

Immunological rejection is the first barrier when considering transplantation procedures. Also, it constitutes the most substantial obstacle to xenotransplantation (6). The literature indicates that immunological reactions to xenografts involve adaptive immunity and innate immunity (7). During these reactions, natural antibodies and various immune system components elicit a response against the transplanted organ.

Innate immunity barriers to xenotransplantation consist of phagocytic cells (like monocytes), natural killer cells, cells producing inflammatory mediators (like basophils), and complement proteins (6). It poses a more significant barrier to xenografts than allografts (tissue transfer between humans) due to molecular discrepancies between the host and the donor. Innate immunity triggering depends on the balance between activation and inhibition signals. For example, macrophages can mediate a strong rejection reaction to xenografts due to overwhelming activation signals and ineffective inhibitory ones (6). The cluster of differentiation 47 (CD47) is a glycoprotein expressed in tissues and serves as a “self-identifying” marker when interacting with inhibitor receptor SIRPα in macrophages. When xenografts fail to be identified by macrophages, a strong immune response is triggered (6). In addition to the previous, activated macrophages play a role in modulating adaptive immunity. In return, adaptive immune reaction helps make the innate immune response more efficient through cytokine production and antibody interaction.

Elements of adaptive immunity involved in xenograft rejection include B and T cells, and are engaged at later stages than innate immunity (6). B cells mediate humoral response — they are secreting antibodies and presenting antigens to T cells, which release cytokines that modulate immune response towards xenografts. In addition, T cells can directly attack xenotransplants and modulate B and natural killer cell responses (6). In other words, T-cell response is triggered by direct and indirect recognition. In the former, T cell receptors identify and interact with Major Histocompatibility Complex (MHC are genes that code for proteins expressed on the cellular surface that allow the immune system to identify them) molecules. Indirect recognition occurs through antigen-presenting cells, like macrophages and B cells (6). Genetic modification of the xenograft provider is a technique to overcome T-cell mediated immune response. For example, research shows that genetically-modified pigs expressing anti-CD2 monoclonal antibodies have demonstrated promising results. Anti-CD2 antibodies are antibodies that have T-cell depleting and co-stimulatory blocking properties. In addition, silencing some genes, like MHC, has been shown to lower T-cell response (6). For best outcomes, combined techniques should be implemented.

Other barriers include economic, ethical, regulatory, and policy considerations. Currently, it is difficult to estimate the economic value associated with xenotransplantation (8). This is because costs associated with creating a xenograft and medication use when a xenograft is transplanted cannot be accurately estimated at this stage. For example, if a xenotransplant only reduced the cost of anti-rejection drugs, it might not be deemed as cost-effective (8). However, if it also reduced side effects from traditional transplant surgery, like that of the heart, it would have significant financial and health value. The other aforementioned issues relate to regulations ensuring animal rights, health, societal acceptance, and controlling cross-species zoonotic infection transmission (9). Also, some governments, like the United Kingdom, have discouraged and banned the use of nonhuman primates for transplantation for reasons such as the increased risk of disease spread due to genetic relatedness.

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Types of xenograft rejection

According to the literature, three main types of xenograft rejection outcomes can occur successively (7). They are hyperacute, acute humoral, and acute cellular xenograft rejection (6, 7). They are discussed below:

• Hyperacute xenograft rejection (HAR): In HAR, the immune reaction usually destroys the transplanted organ within minutes to hours (7). This occurs when a wild-type pig graft is transplanted into humans. The process is usually mediated by a complement system, which activates the coagulation cascade, leading to rapid xenograft rejection (6). Plasmapheresis is a technique to avoid HAR, which depletes antipig antibodies or inhibits complement activation (7). 
• Acute humoral xenograft rejection: The abovementioned procedure only delays HAR for hours to weeks because once the host-antibodies titer returns, the xenograft is attacked again. This is known as acute humoral xenograft rejection, also referred to as acute vascular rejection or delayed xenograft rejection (6, 7).
• Acute cellular xenograft rejection: In this type, the immune response is mediated by innate and adaptive immunities. Organ rejection occurs days to weeks after organ transplantation (6, 7). Examples of ways to overcome this kind of organ rejection are immunosuppressive treatments, which prevent T-cell and B-cell infiltration. However, they have significant side effects that could negatively influence health and quality of life.

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Tying up loose ends

Xenotransplantation offers a solution to a growing problem, which is organ shortages. Despite its potential, it still faces many acceptance and implementation barriers, like those related to public acceptance, funding, and religious beliefs. In addition, we still have not perfected the process of creating xenografts that patients could live with for years. This limits the implementation across multiple diseases and organs. Xenografts could potentially influence longevity by providing replacement organs for people with end-stage life diseases, whose lives could be cut short without such an intervention. This does not mean that xenotransplantation is the only approach to address organ shortages, research exploring techniques based on stem cell use and cloning is taking place with promising results. Eventually, the utilization of these approaches will depend on the time they become available to patients, the costs they will save, and the quality-of-life enhancements they will offer.

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References

1. Cooper DKC, Gaston R, Eckhoff D, Ladowski J, Yamamoto T, Wang L, et al. Xenotransplantation-the current status and prospects. Br Med Bull. 2018;125(1):5-14.
2. Cascalho M, Platt JL. Challenges and potentials of xenotransplantation. Clinical Immunology: Copyright © 2008 Elsevier Ltd. All rights reserved.; 2008. p. 1215-22.
3. Ekser B, Cooper DKC, Tector AJ. The need for xenotransplantation as a source of organs and cells for clinical transplantation. Int J Surg. 2015;23(Pt B):199-204.
4. Cooper DKC, Ekser B, Tector AJ. A brief history of clinical xenotransplantation. Int J Surg. 2015;23(Pt B):205-10.
5. Ekser B, Li P, Cooper DKC. Xenotransplantation: past, present, and future. Curr Opin Organ Transplant. 2017;22(6):513-21.
6. Carvalho-Oliveira M, Valdivia E, Blasczyk R, Figueiredo C. Immunogenetics of xenotransplantation. International Journal of Immunogenetics. 2021;48(2):120-34.
7. Lu T, Yang B, Wang R, Qin C. Xenotransplantation: Current Status in Preclinical Research. Front Immunol. 2019;10:3060.
8. Loike JD, Kadish A. Ethical rejections of xenotransplantation? The potential and challenges of using human‐pig chimeras to create organs for transplantation. EMBO reports. 2018;19(8):e46337.
9. Rollin BE. Ethical and societal issues occasioned by xenotransplantation. Animals. 2020;10(9):1695.

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