Monday September 14, 2020 - 14:30 to 15:15
TCR repertoire sequencing of total T cells and circulating follicular helper T cells in kidney transplant recipients
Patricia Almendro Vázquez3, Rocío Laguna Goya1,3, Alberto Utrero Rico3, Patricia Suárez Férnandez3, Cecila González Cuadrado3, Marta Chivite Lacaba3, Jara Caro Espado 2, Esther González Montes2, Esther Mancebo Sierra1,3, Estela Paz Artal1,3.
1Immunology, Hospital 12 de Octubre, Madrid, Spain; 2Nephrology, Hospital 12 de Octubre, Madrid, Spain; 3Immunodeficiencies and Transplant Immunology Group , imas12, Madrid, Spain
Introduction: In renal transplantation (RT), allorecognition by immune cells has deleterious effects on the graft. In particular, follicular helper T cells (Tfh) have been associated with anti-HLA antibody development and rejection. All T cells recognize antigens through their receptor (TCR) and the set of TCR sequences of an individual is called ‘TCR repertoire’. Early identification of alloreactive T cell clones (or allo-Tfh clones) by TCR repertoire sequencing could be useful to better understand the immune response against the allograft, to monitor graft evolution and potentially to develop targeted therapies in RT rejection.
Materials and Methods: Peripheral blood was obtained pre-transplant (pre-Tx) and at different times post-transplant (post-Tx) in a prospective cohort of 86 RT recipients (RTR). Genomic DNA (gDNA) was obtained from PBMCs (i.e., total T cells). To date, the TCR repertoire of 9 RTR has been sequenced pre-Tx and post-Tx (6 patients with biopsy-proven acute rejection (BPAR) and 3 long-term stable recipients). In parallel, circulating Tfh (cTfh) from 3 healthy controls (HC) and 15 RTR (6 acute antibody mediated rejection, 6 chronic rejection and 3 long-term stable recipients) were isolated by cell sorting and gDNA of cTfh was extracted. In addition, the repertoire was sequenced in DNA isolated from a patient's rejection biopsy. In all cases, a survey resolution analysis was performed using the Adaptive Biotechnologies immunoSEQ human T-cell receptor beta (hsTCRB) kit. Raw data was analysed using the immunoSEQ software.
Results and Discussion: The TCR repertoire of total T cells was sequenced, with no significant differences observed between the repertoires of HC and patients with chronic kidney disease pre-Tx, nor between RTR with stable graft and rejection. A decrease in the number of TCR rearrangements post-Tx compared to pre-Tx was observed in RTR (p<0.01), possibly due to the effect of immunosuppressive therapy. Regarding cTfh, their TCR repertoire was less clonal and therefore much more diverse than that of total T lymphocytes, both in HC and RTR (p=0.001). In addition, there was an increase in cTfh clonality in patients with acute antibody mediated rejection during the first year post-Tx compared with HC (p<0.05) and with patients with chronic rejection after 5 years post-Tx (p=0.01), which could be explained as a consequence of either the alloresponse or the immunosuppressive therapy. Of interest, in the patient whose biopsy was included, the same expanded clone was found in his biopsy and in peripheral blood.
Conclusion: The study of total T cells and cTfh TCR repertoire in RTR can provide early and accurate information about alloresponses against the graft. cTfh repertoire seems more diverse than that of total T cells. Further analysis to evaluate TCR repertoire is in progress.
Government of Spain PI-18/969. FEDER (European Funds for Regional Development).