Pig organs make their way into humans. Credit: Laoise Mac Gabhann

On 16 March 2024, doctors at Massachusetts General Hospital (Mass General Hospital) transplanted a genetically modified pig kidney into a human patient for the first time. The patient, 62-year-old Richard ‘Rick’ Slayman, had end-stage kidney disease and dialysis had stopped working. This radical new approach offered hope for patients with organ failure who could not receive a human organ. However, transplanting pig organs into humans had only been attempted twice before in living patients — two men with heart failure — and both recipients died within two months. Slayman’s doctors saw this transplant as a last-ditch effort to save his life.

Two weeks later, Slayman walked out of Mass General Hospital in high spirits, and there were no signs of organ rejection. The results offered promising news for 90,000 people in the United States who were on a waitlist for a kidney transplant in 2023, of whom only 28,000 received a kidney in that time frame. However, several weeks later, Mr Slayman sadly died, his fate mirroring those who came before him. His doctors said his death was unrelated to organ rejection, but the details have not yet emerged.

The field of xenotransplantation, once relegated to the fringes of medicine, has grown by leaps and bounds over the past few years. Several companies in the United States, China, New Zealand and elsewhere are racing to breed and genetically engineer pigs so that their organs are better suited for human transplantation. Their aim — to fill the desperate need for organ transplants — is especially urgent when considering that in the United States alone, 17 people die each day waiting for an organ donation.

Since the 1990s, scientists have used pigs as an alternative organ source because they are readily available and physiologically similar to humans, and thanks to gene-editing tools like CRISPR, they can be engineered to resemble humans more closely. So far, their investments have paid off. Since 2021, two pig hearts, two pig kidneys and at least one pig liver have made it into living human patients (Table 1). While they have not functioned long term in human trials, none was immediately rejected, and in preclinical studies using nonhuman primates, pig organs functioned for up to three years1,2.

Table 1 A brief history of pig-to-human transplantation
Full size table

According to Muhammad Mohiuddin, a xenotransplantation surgeon at the University of Maryland, these successes have breathed enthusiasm into the field and made “believers out of non-believers.” But there are still considerable hurdles that remain. For each of the pig-to-human transplants, the organs either failed or needed to be removed within two months. “We will have a long way to go before we are confident that we can offer this to patients and give them ten more years of life,” says Mohiuddin. “But, it is a viable option,” he adds.

History of xenotransplantation: piggybacking on past experiments

Xenotransplantation has a surprisingly long history. In the 1900s, surgeons experimented with animal-to-human transplantation by grafting organs and tissues from goats, pigs and even rabbits onto human recipients. Without any modifications made to the animal or the human immune system, these early attempts failed spectacularly. “It is a very daunting task,” said Mohiuddin. When he started working with baboons he found “an unmodified pig heart will be rejected in minutes right before your eyes.” The difficulty of the task deterred people from entering the field for decades.

Things changed, however, in the 1960s with the development of immunosuppressant drugs like azathioprine, which revolutionized organ transplantation in general and later xenotransplantation. During this time period, researchers relied on organs from chimpanzees, baboons and macaques, hoping their closer biological kinship to humans would reduce rejection. These primate-to-human attempts yielded mixed results. The most successful experiment was in a patient who lived for nine months with a chimpanzee kidney before dying from pneumonia3.

Meanwhile, the burgeoning field of human-to-human transplants became an accepted therapy for organ failure in the 1970s and 1980s thanks to advances in immunosuppression. But still the availability of organs for donation could not keep up with demand, and so the dream of animal-to-human transplants remained alive.

Enter the pig. Owing to concerns over viral transmission from primates to humans, the FDA put a moratorium on primate organ donation in 1999 and these readily available animals have emerged as the frontrunners for xenotransplantation. They are physiological similar to humans and offer rapid reproduction, large litters and lower breeding costs. Additionally, using pigs instead of nonhuman primates sidesteps some ethical concerns. But pigs were not a perfect solution, due to their considerable immune incompatibility. “Xenograft rejection is predominantly an antibody-mediated rejection,” says Mohiuddin. “Our initial goal was to find those antigens and remove them, and before CRISPR–Cas9 time this was done a single gene at a time,” he adds.

The path to porcine perfection or porcine progress

Pigs present several challenges for xenotransplantation. “When you put a porcine organ into humans, all the alarms are going to be sounded,” says Wenning Qin, a bioengineer at eGenesis. “They’re gonna be highly inflammatory.” Perhaps the most significant issue is the presence of cell-surface sugars that humans and primates do not have. One sugar, called galactose-α-1,3-galactose (α-gal), was particularly challenging because most people carry preformed antibodies to it stemming from exposure through food.

According to Paul Tan, the founder and CEO of NZeno, a New Zealand-based xenotransplant company, the best way to think about these genes is like the human ABO blood types. “They are important because they cause immune rejection in the same way that if you have type A blood, you have antibodies against type B blood,” he explains.

David Ayares, president and chief scientific officer of Revivicor, a Virginia-based xenotransplantation biotech, says the presence of this sugar was the first major hurdle to overcome. When he started, the longest a pig heart could last in a baboon before being attacked by the immune system was mere hours to days, but after the genetic removal of α-gal, survival times jumped to weeks or months4. Later, scientists removed two other cell-surface sugars called N-glycolylneuraminic acid (Neu5Gc) and SDa. Experiments using human blood show significantly less antibody binding in pigs with all three genetic changes to remove the cell-surface sugars4.

But more work was needed. Primates that received genetically modified pig hearts still showed latent forms of immune rejection like microscopic blood clotting and T cell-mediated rejection. And so, scientists had the idea to add a spate of human genes to the pig genome. First, they added genes in the human complement cascade, CD46 and CD55. These genes act like ‘off switches’ that dampen the immune attack. Then, they added two more genes in the blood clotting cascade that prevent the formation of blood clots.

Finally, they added two more human genes, the heme oxygenase 1 (HO1) gene (HMOX1) and CD47, to the pig’s genome. HO1 offers anti-inflammatory benefits and guards against ischemia–reperfusion injury, which occurs when blood flow is interrupted and then restored in an organ. The influx of oxygenated blood can release reactive oxygen species that damage cells, leading to cell death and organ failure. CD47, on the other hand, provides protection from macrophages, a key type of innate immune cell that identify, attack and ‘eat’ dead, foreign or threatening invader cells. The CD47 gene is termed the ‘don’t eat me gene’.

With ten edits, including the three sugar knockouts, the six human gene ‘knock-ins’ and one gene to reduce the size of the pig organs, the first pig organ was ready for human transplantation. In January 2022, this ten-edit pig heart was transplanted into a patient for the first time, marking a milestone for the field.

In this photo provided by the University of Maryland School of Medicine, members of the surgical team show the pig heart for transplant into a patient that was performed in Baltimore on Friday 7 January 2022. Credit: ZUMA Press, Inc. / Alamy Stock Photo

Meanwhile, in other labs, scientists were tinkering with different combinations. Revivicor, the creator of the ten-edit pig, also created a pig with just one gene edit — the removal of the α-gal sugar. This pig, called the GalSafe pig, earned FDA approval in December 2020 for use in food and therapeutics. In 2024, a GalSafe kidney was used in a kidney transplant for Lisa Pisano, a 54-year-old woman from New Jersey. It was the second pig kidney to be transplanted into a human (after Rick Slayman), but this version combined the α-gal knockout with a pig thymus, which they surgically connected to the kidney in the hope it would help to educate human T cells. A Chinese company, Clonorgan Biotechnology, transplanted a pig liver into a human from a six-edit Bama miniature pig5. And Boston-area eGenesis has created a pig from another breed, called the Yucatan miniature pig, with a staggering 69 edits6.

While immune rejection remains a critical hurdle, different approaches to pig selection raise additional concerns such as size, growth management and viruses. Companies like Revivicor use larger pigs with faster growth rates, reaching 800 pounds. They must genetically modify these pigs to inhibit their natural growth patterns. eGenesis, on the other hand, opts for smaller Yucatan minipigs, naturally reaching a maximum of 150 pounds and offering a better size match for human recipients. However, these pigs carry a different risk: their genome contains endogenous retroviruses.

Porcine endogenous retroviruses (PERVs) are unusual viruses that integrate into the genome and cannot be removed like most infectious viruses. The worry is that once they get into humans, they will activate and cause infections or integrate into the human genome, causing genetic mutations. There is no evidence that PERVs have infected host cells in xenotransplantation studies, but lab experiments have shown some risks.

“The fear is that we would have a zoonotic event very similar to what happened with HIV and the retrovirus will cross over into the human genome,” says Mike Curtis, CEO of eGenesis. He acknowledges the scant evidence that PERVs can infect human cells, but he also cautions that xenotransplantation is a new practice, and we should be wary of its safety record. “We’ve never had a retroviral transmission from a pig to a person,” but he worries that transplanting pig organs into humans who are on an immunosuppression regime to prevent organ rejection is risky. “What is the risk of retroviral transmission from a pig to a person? We don’t really know,” he says.

To mitigate this risk, eGenesis used CRISPR–Cas9 gene editing technology to inactivate all the PERVs in a pig genome. The PERV deactivation added 59 edits to the 10-edit pig, putting the total at 69 genetic changes. In March 2024, surgeons transplanted a 69-edit pig kidney into a human, Rick Slayman, for the first time.

In addition to endogenous viruses, there is also the threat of conventional infectious viruses. Following the death of the recipient of the first pig-to-human transplant in March 2022, scientists detected the presence of a pig herpes virus called cytomegalovirus (CMV) in the pig’s heart after it was removed from the patient7. The presence of the virus raises concerns about the limitations of current screening protocols.

Despite repeated tests for CMV before the transplant, the virus remained undetected. While the link between CMV and the patient’s death remains unclear, it emphasizes the importance of closely monitoring viral activity in pig organs intended for human recipients.

Looking ahead

The race to make pig organs viable for human transplant is rapidly accelerating, with different companies and research groups testing various approaches. Paul Tan compares the growth to the development of COVID-19 vaccines during the pandemic, “it’s almost like the different types of vaccines for COVID on the market,” he says.

The addition of human data from pig-to-human transplants and experiments in deceased patients in recent years is allowing scientists to generate a wealth of information on how the body reacts.

In a recently published paper8, researchers analyzed blood taken from two deceased recipients of the 10-edited pig heart. This analysis used advanced techniques, including single-cell transcriptomics, lipidomics, proteomics and metabolomics, providing a detailed view of the body’s response to the transplanted organ. It has also generated a groundswell of data from which to learn about the molecular details of rejection.

According to the study’s author, Brendan Keating, surgery professor at NYU Grossman School of Medicine, they are pinpointing which pig proteins are shedding from the donated organs and generating antibody responses. “We are at the point where we are digging down into what is going on,” he says. In his dataset, he and his team analyzed over 8,000 proteins from the immune response of a deceased patient who received a pig heart over 61 days. Signs of rejection began on day 33, long before any clinical signs appeared. “Because we have all of these daily time points, we can see a concerted immune response across that whole 61-day procedure,” he says. “Now, using all of these systems biology approaches, we can try and come up with an approach for knocking out immune pathways or using additional immunosuppression to target those pathways,” he adds.

Overall, the study offers valuable insights into xenotransplantation challenges and possibly paves the way for more targeted immunosuppressive strategies and potentially personalized transplants.

Another way of personalizing the transplants is through cross-matching donors with recipients. This approach is commonplace in human-to-human transplants when doctors match patients to donors based on their histocompatibility or tissue type. According to Angelika Schnieke, Professor Emerita at the Technical University of Munich, who spent her career researching xenotransplantation, this is something we could start doing with xenotransplantation as well. She says these immunohistocompatibility complexes could potentially be edited out of gene-edited pigs to reduce incompatibility9.

Major histocompatibility complex (MHC) class I-deficient pigs have been generated and could be used to study the relevance of porcine MHC I molecules in xenotransplantion. Adapted from ref. 9, Springer Nature Limited.

Some scientists have taken a cue from cancer research and are experimenting with adding PDL1, a protein that acts as a ‘brake’ to keep the body’s immune responses under control10. PDL1 is expressed on the surface of many cancers, which allows them to grow undetected by the immune system, and new powerful immunotherapies are targeted at blocking this protein.

According to Curtis, Ayares and Tan, the end goal is to engineer the pigs enough so that no immunosuppressants are needed in the future, but they still have a long road ahead. Schnieke says that pig organs could serve a more temporary purpose as researchers iron out the issues with immune rejection. She suggests using them as a stop gap for those who are suffering from liver toxicity or waiting for a human organ. “If you are already on dialysis, you could get a kidney and it might work well, but if it doesn’t you could take it out again, go back onto the dialysis,” she says. She also says that the human liver is unique because it can regenerate and says people are working on developing pig livers that can be used temporarily until their livers heal.

In the meantime, scientists will continue gathering data and using it to improve the pigs, the surgical protocols and more. “I consider this as a stepwise process,” says Mohiuddin. “We are entering into a big world of the unknown, but we have confidence from our nonhuman primate experiments that this is achievable.”