A recent study pinpoints the precise molecular targets of organ transplant rejection. This discovery prompted the development of tools to identify and characterise donor-reactive T cells in the laboratory. If the findings of this preclinical study can be translated to the human setting, significant advances in post-transplant immune monitoring and other benefits may result.
by A/Prof Alexandra Sharland, Dr Nicole Mifsud, and Eric Son
Organ transplants save thousands of lives each year. Graft rejection is an ever-present threat, and the maintenance of transplant function relies on the use of powerful drugs to suppress the host immune system. Transplanted organs are recognised as foreign (and therefore rejected), because of differences between the donor and recipient in a class of cell-surface proteins called MHC (or major histocompatibility complex) molecules. However, there is more to this story than MHC mismatches alone. MHC molecules carry a diverse cargo of small protein fragments called peptides. By altering the composition of this peptide cargo, we were able to show that the peptides partner with the MHC to form the molecular target recognised by the host T cells that cause graft rejection1 (Figure 1). Working in a mouse model, we screened a large starting pool and identified the individual peptides that T cells recognise. We subsequently used this knowledge to develop molecular tools to follow donor-reactive T cells during transplant immune responses.
MHC molecules loaded with specified peptides can be produced in a soluble form and combined in multimeric complexes or “multimers” (such as tetramers of peptide-MHC with streptavidin, or dextramers where peptide-MHC is added to a dextran backbone, Figure 2). Using a panel of peptide-MHC multimers, we were able to detect and characterise donor-reactive T cells before, during and after transplant rejection1 (Figure 3). We are now aiming to translate these findings to human MHC molecules (called Human Leucocyte Antigen or HLA) and subsequently to a human clinical transplant setting.
Transplant rejection is suspected whenever a decline in graft function is noted and is confirmed through tissue biopsies. Biopsies are invasive and associated with risks of bleeding and other complications. Moreover, by the time that rejection is detected, graft damage has already occurred and may not be completely reversible, eventually leading to graft loss. While there have been some promising advances towards the early non-invasive diagnosis of rejection through measurement of cell-free donor DNA in the recipient’s circulation,2,3 and in estimating the diversity and number of T cells that respond to organ transplants,4 no tests currently available permit the detection of donor-reactive T cells directly from the blood or tissues of transplant recipients.
Peptide-MHC multimer panels would enable measurement of the frequency of donor-reactive T cells in blood samples, along with their gene expression and functional state, significantly advancing immune monitoring in the clinic. Such panels are compatible with techniques and instrumentation that are widely available in diagnostic laboratories, and suitable for automation. The initial use of these multimer panels would likely be in the immune monitoring arms of clinical trials of novel therapies (such as infusion of regulatory cells), while they could later be adopted more widely in risk stratification of transplant recipients and to fine-tune levels of conventional immunosuppressive drugs. If changes in the number or characteristics of donor-reactive cells precede clinical evidence of acute rejection, it may be possible to avoid graft damage by taking pre-emptive action.
Powerful new techniques to analyse gene expression within transplanted organs and recipient T cells are being used in attempts to develop biomarkers to diagnose acute rejection,5,6,7 predict the induction of immunological tolerance to the donor,8,10 or estimate the likelihood of graft loss in the longer-term.11,12
Throughout this intense effort, one important deficiency has been our lack of ability to confidently identify the donor-reactive T cells and distinguish them from the large number of irrelevant or bystander cells within a heterogeneous cell population. The result is that signals from the donor-reactive T cells can be obscured by noise generated from cells of irrelevant specificity, potentially leading to inconclusive study findings. Incorporating peptide-MHC multimer panels into biomarker discovery strategies would allow these signals to emerge.
In the sphere of basic biomedical discovery, peptide-MHC multimers can be used to isolate individual donor-reactive T cells for multi-OMIC single-cell biology experiments, where the T cell receptor sequence, cell-surface proteins, and gene expression can all be determined at the same time as donor antigen-specificity,9 enabling greater understanding of the processes underpinning transplant rejection or tolerance induction.
Knowledge of both the peptide-MHC ligand and the T cell receptor sequences makes it possible to interrogate their interactions in detail at a biophysical and biochemical level. In future, this knowledge will enable the development of new technologies for visualising donor-specific cells in situ in tissues, and may lead to the development of antigen-specific therapies, where peptide-MHC multimers are used for the deletion, depletion, or immunomodulation of donor-reactive T cells.
- Son, E. T., Faridi, P., Paul-Heng, M., Leong, M. L., English, K., Ramarathinam, S. H., . . . Sharland, A. F. (2021). The self-peptide repertoire plays a critical role in transplant tolerance induction. J Clin Invest, 131(21). doi:10.1172/JCI146771
- Fearon, W. F., & Valantine, H. A. (2021). Can We Predict Rejection Early After Heart Transplantation? Circulation, 144(18), 1473-1475. doi:10.1161/CIRCULATIONAHA.121.056808
- Snyder, T. M., Khush, K. K., Valantine, H. A., & Quake, S. R. (2011). Universal noninvasive detection of solid organ transplant rejection. Proc Natl Acad Sci U S A, 108(15), 6229-6234. doi:10.1073/pnas.1013924108
- DeWolf, S., Shen, Y., & Sykes, M. (2016). A New Window into the Human Alloresponse. Transplantation, 100(8), 1639-1649. doi:10.1097/TP.0000000000001064
- Madill-Thomsen, K., Abouljoud, M., Bhati, C., Ciszek, M., Durlik, M., Feng, S., . . . Halloran, P. F. (2020). The molecular diagnosis of rejection in liver transplant biopsies: First results of the INTERLIVER study. Am J Transplant, 20(8), 2156-2172. doi:10.1111/ajt.15828
- Salem, F., Perin, L., Sedrakyan, S., Angeletti, A., Ghiggeri, G. M., Coccia, M. C., . . . Cravedi, P. (2021). The spatially resolved transcriptional profile of acute T cell-mediated rejection in a kidney allograft. Kidney Int. doi:10.1016/j.kint.2021.09.004
- Verma, A., Muthukumar, T., Yang, H., Lubetzky, M., Cassidy, M. F., Lee, J. R., . . . Suthanthiran, M. (2020). Urinary cell transcriptomics and acute rejection in human kidney allografts. JCI Insight, 5(4). doi:10.1172/jci.insight.131552
- Baron, D., Ramstein, G., Chesneau, M., Echasseriau, Y., Pallier, A., Paul, C., . . . Brouard, S. (2015). A common gene signature across multiple studies relate biomarkers and functional regulation in tolerance to renal allograft. Kidney Int, 87(5), 984-995. doi:10.1038/ki.2014.395
- Ma, K. Y., Schonnesen, A. A., He, C., Xia, A. Y., Sun, E., Chen, E., . . . Jiang, N. (2021). High-throughput and high-dimensional single-cell analysis of antigen-specific CD8(+) T cells. Nat Immunol, 22(12), 1590-1598. doi:10.1038/s41590-021-01073-2
- Vionnet, J., & Sanchez-Fueyo, A. (2018). Biomarkers of immune tolerance in liver transplantation. Hum Immunol, 79(5), 388-394. doi:10.1016/j.humimm.2018.02.010.
- Halloran, P. F., Venner, J. M., Madill-Thomsen, K. S., Einecke, G., Parkes, M. D., Hidalgo, L. G., & Famulski, K. S. (2018). Review: The transcripts associated with organ allograft rejection. Am J Transplant, 18(4), 785-795. doi:10.1111/ajt.14600
- O’Connell, P. J., Zhang, W., Menon, M. C., Yi, Z., Schroppel, B., Gallon, L., . . . Murphy, B. (2016). Biopsy transcriptome expression profiling to identify kidney transplants at risk of chronic injury: a multicentre, prospective study. Lancet, 388(10048), 983-993. doi:10.1016/S0140-6736(16)30826-1
About the Authors
Alexandra Sharland, MBBS, PhD, FRACP. Associate Professor of Transplantation Immunobiology, Faculty of Medicine and Health, The University of Sydney. Transplant Immunologist with a particular interest in understanding the factors which determine the fate of alloreactive T cells.
Nicole Mifsud, PhD. Group Leader of the Clinical Immunology Laboratory, Faculty of Medicine, Nursing and Health Sciences, Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute, Monash University, Clayton, VIC, Australia. T cell immunologist with particular focus on understanding the contribution of both alloreactive and cross-reactive T cells in transplant rejection.
Eric Taeyoung Son, MD-PhD candidate at the University of Sydney. Eric is conducting transplantation immunology research under the supervision of A/Prof Alexandra Sharland.