Progress towards Clinical Pig Organ Xenotransplantation

The early years of research

Research into pig organ xenotransplantation has made great progress, but it has been a long journey fraught with setbacks. For example, I began to investigate whether pig organs could be transplanted successfully into non-human primates (NHPs) almost 40 years ago¹ and found that pig kidneys and hearts were rejected hyperacutely (i.e. within minutes or hours).

The initial problem was the presence in humans and NHPs of ‘preformed’ anti-pig antibodies. All humans develop anti-pig antibodies in infancy, and this is not because they are exposed to pigs or eat pork. In infancy, when their gastrointestinal tract is colonized by microflora, they develop protective antibodies against many bacteria and viruses. Unfortunately, some of the microbes express the same carbohydrate antigens as pigs and so, when a pig organ is transplanted, the recipient is ‘primed’ to immediately reject it.

Indeed, multiple barriers needed to be overcome. For many years, it was said that xenotransplantation research was like peeling an onion—you peeled one layer off, only to be faced by another. The analogy went further in that peeling the xenotransplantation onion was inevitably accompanied by tears.

What has led to progress?

Nevertheless, my own group (in which several Indian postdoctoral surgical research fellows, such as Murali Basker, Goutham Kumar Mehta, Vikas Satyananda, Santosh Nagaraju and Abhijit Jagdale, made important contributions) and a handful of other groups slowly made progress.² This resulted from a combination of two advances: (i) the ability to genetically engineer pigs to reduce the injury caused by the human innate immune response and (ii) the introduction of novel immunosuppressive agents that inhibit the adaptive (T cell) immune response to the pig organ graft.²

Innovations in gene-editing of pigs

Those in the field of gene editing have developed the ability to (i) delete expression (knockout) of the pig antigens against which humans have ‘preformed’ antibodies that, when they bind to the pig organ, activate the complement cascade and (ii) introduce species-specific human ‘protective’ genes that provide further protection to the pig tissues from the human immune response.

The first step taken in this respect was by David White in Cambridge in the UK who produced pigs expressing a human complement-regulatory protein (CD55) that provided significant protection to the pig organ from human complement injury,³ extending pig kidney transplantation in NHPs to a maximum of 3 months. These were among the first experiments that demonstrated the potential of gene editing in pig organ xenotransplantation.

Since then, 3 pig antigens have been identified, all of which are carbohydrates, the most important of which is galactose-α1,3-galactose (Gal).⁴ At the time it was identified (1991), a technique to ‘knockout’ the gene in pigs had not yet been developed⁵ and so it was a further decade before a Gal-KO pig was produced.⁶ The transplantation of a heterotopic (non-life-supporting) Gal-KO pig heart into an immunosuppressed NHP extended graft survival to a maximum of 6 months⁷ and of a life-supporting pig kidney to 3 months. Two other pig xenoantigens have since been identified and have been deleted from the pigs available today. The combination of triple gene knockout and insertion of protective human transgenes extended graft survival further.

Novel immunosuppressive therapy

In 2000, a young Swiss research fellow in my group, Leo Bühler, demonstrated that conventional immunosuppressive therapy (i.e. cyclosporine or tacrolimus-based) administered to NHPs with pig organ xenografts was not efficient in inhibiting the adaptive immune response.⁸ However, a regimen based on a new immunosuppressive agent that inhibited the CD40/CD154 T cell co-stimulation pathway was much more successful. Since then, one of these agents has formed the basis of almost all of the immunosuppressive regimens used in xenotransplantation research.

Today, we have pigs available with 10 or more gene edits aimed at protecting the organs from the human immune response.^9,10 Life-supporting pig kidneys have now functioned for periods of >2 years¹⁰ and life-supporting pig hearts for >16 months (Cleveland JD, personal communication). For a number of complex reasons (only some of which we understand), survival of pig livers and lungs in NHP recipients has been much less successful, generally being measured only in days.

Moving into the clinic: Which patients should be offered a pig organ?

I had always anticipated that the US Food and Drug Administration (FDA) would approve a pig kidney transplant before a pig heart transplant. If a pig kidney failed, the possibility of returning the patient to dialysis may be life-saving whereas there may be no such back-up after pig heart transplantation. However, this proved not to be the case (see below).

Kidney. Two types of patients can be considered. The first are those at ‘high risk’ with no life-saving alternative, e.g. those who are no longer able to be supported by dialysis (because vascular access is no longer possible), or have comorbid conditions that exclude them from allotransplantation. In these patients, kidney xenotransplantation is a clinical experiment rather than a formal clinical trial. The second would include ‘low risk’ patients who are acceptable candidates for allotransplantation but who are unlikely ever to receive a deceased human donor kidney (Fig 1).¹¹ These patients can largely be predicted when placed on the waiting list. They are older (e.g. 60–70 years), of blood groups O or B, and with diabetes as the cause of their nephropathy, as these patients are more likely to die before being allocated a deceased human kidney in the USA.

Heart. As ventricular assist devices are readily available in the USA for adult patients with end-stage cardiac disease, it is perhaps more difficult to identify patients for whom it would be ethical to offer a pig heart transplant. However, the situation is quite different in the case of infants born with complex congenital heart disease. These babies do extremely well if allotransplantation can be carried out, but this is frequently not possible because they sadly die before a suitable human donor heart can be found for them. The results of palliative surgical procedures are less than ideal, and support with a ventricular assist device is rarely fully successful in this age group.

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FIG 1.

Survival of patients in the USA (i) with a living donor kidney (top), (ii) a deceased donor kidney (second from top), (iii) on the waitlist while on dialysis (second from bottom) and (iv) while on dialysis but not waitlisted for a kidney transplant (bottom). Approximately 45% of patients receiving chronic dialysis while on the waitlist for a deceased human donor kidney either die or are removed from the waitlist (as no longer acceptable candidates for the procedure) within 5 years (reproduced with permission from Jagdale et al. Transplantation 2021;105:1904–8, based on data from two sources (i) USRDS. Annual data report 2017 [cited 2019 4/08/2019], available from www.usrds. org/adr.aspx. and (ii) Orandi et al. New Engl J Med 2016;374:940–50).

Export to PPT

In these patients, a strong case can be made for inserting a pig heart soon after birth to ‘bridge’ them until a deceased human donor heart becomes available,¹² which takes a mean of 4 months in the USA. Although potentially life-saving, this does not commit the patient to a pig heart for the remainder of his/her life (but does enable the medical team to obtain valuable experience of clinical xenotransplantation in an ethical manner).

The first clinical experiments

On the basis of the encouraging laboratory results summarized above, surgeons have taken the first tentative steps to apply xenotransplantation to patients with end-stage heart or kidney disease (a handful of studies in which pig kidneys or hearts were transplanted into brain-dead human subjects [decedents] confirmed that hyperacute rejection did not occur but provided few new data¹³).

The first clinical attempts were to transplant pig hearts into patients for whom any alternative therapy, e.g. allotransplantation, mechanical device support, had been excluded. In retrospect, the first patient (in 2022) was too debilitated to recover from the procedure despite excellent function of the pig heart for >6 weeks.¹⁴ For example, in the 8 weeks of his survival, he was strong enough to sit out of bed on only a single occasion. Ultimately, he died from a combination of graft rejection, pig cytomegalovirus in the heart graft and extreme debility. The second patient died from rejection, possibly associated with inadequate immunosuppressive therapy.

The first pig kidney transplant in a living patient was carried out by my colleagues at the Massachusetts General Hospital (MGH) in 2024. The pig kidney functioned well, but the patient, who was known to have ischaemic heart disease, died of a sudden cardiac dysrhythmia after 52 days.¹⁵ Since then, two kidney transplants have been carried out by the group at New York University, one of whom remains alive and well after approximately 3 months, and a second patient at MGH continues to do well after 1 month.

In view of these moderately encouraging results, the US FDA has approved a small number of further pig kidney transplants in patients who are acceptable candidates for allotransplantation but are unlikely to survive long enough on the waiting list to obtain a deceased human kidney.¹⁶

Nevertheless, potential problems remain. For example, although classical rejection of a pig kidney graft in an NHP is relatively rarely problematic today, there is a significant incidence of the development of proteinuria that can be severe. However, in view of the several advantages of caring for a patient in a hospital setting (when compared to managing an NHP in a laboratory environment), I am optimistic that problems like this may be prevented or controlled.

I suggest that xenotransplantation is an example of what Tom Starzl, a great pioneer in allotransplantation, had in mind when he wrote:

‘History tells us that procedures that were inconceivable yesterday, and are barely achievable today, often become routine tomorrow’.