Role of CD28 Signaling on T cell Phenotype following Lymphodepletion and Homeostatic Reconstitution

Abstract

Calcineurin inhibitors, the current standard of care in immunosuppression for transplant patients, leads to nephrotoxicity and loss of graft function over time. Preclinical studies have shown selective CD28 blockade (CD28 dAb, Lulizumab) to be superior to calcineurin inhibitors in reducing toxicity and prolonging allograft survival. Emory University will soon commence a Phase 2 clinical trial testing the efficacy of a Lulizumab‐based immunosuppression regimen in kidney transplantation. Patients in the trial will be T cell depleted (TCD), rendering them lymphopenic at the time of transplant and will therefore undergo homeostatic reconstitution in the weeks following transplantation. Lulizumab prevents T cell activation by selectively targeting CD28 signaling, but it is presently unclear what effect blocking this pathway will have on T cell phenotypes following homeostatic reconstitution (HR) in these patients. Previous studies have shown that HR leads to increased differentiation of naïve T cells into memory cells and to premature senescence. We hypothesized that blocking CD28 signaling in T cells during this process could prevent memory differentiation and senescence. We investigated this question using a murine model of TCD and skin transplantation. Four groups of mice were established: no treatment control, TCD alone, CD28 dAb alone, and both TCD and CD28 dAb (n = 4). Mice were sacrificed at 6 weeks post‐depletion and transplantation, and blood and tissues were collected for flow cytometric analysis. Using both traditional flow cytometry analysis and an unsupervised machine‐learning approach, we found that TCD CD4+ T cells have significantly elevated expression of the memory markers CD27, CD103, and CD69 when compared to no treatment control CD4+ T cells, indicating HR‐induced differentiation into memory T cells. In addition, TCD CD4+ T cells also expressed significantly higher levels of PD‐1, TIGIT, and CD226 (DNAM‐1) when compared to no treatment control CD4+ T cells, indicating a higher degree of senescence. Finally, we found that treating TCD mice with CD28 dAb restored expression of both memory markers and costimulatory markers to baseline levels. As such, we believe CD28 dAb presents a viable method to control HR‐induced differentiation and senescence of CD4+ T cells. These results were consistent across the blood, lung, and spleen. Our future directions include using murine models to determine whether the selective CD28 blockade‐mediated rescue of the HR‐induced phenotypic changes significantly affects protective immunity to infection.Support or Funding InformationResearch reported here was supported by the National Institutes of Health under award number R25 DK 101390 ‐ Title: Summer Undergraduate Program in Renal Research (SUPERR).