TGF-Beta Signaling Favors Central Memory Phenotype Expression By Ex-Vivo Stimulated Human T Cells

Abstract

Adoptive immunotherapy using ex vivo differentiated and expanded T cell lines can be remarkably efficient to treat cancer and infections. Unfortunately, the process of ex vivo T cell stimulation can lead to terminal effector differentiation and functional exhaustion thereby limiting the persistence and therapeutic effects of these T cells after transfer. Accumulating evidence suggests that, owing to their proliferative capacity, self-renewal ability and long term persistence in vivo, T cells bearing a central memory (CD45RO+/CD62L+ - Tcm) instead of an effector memory (CD45RO+/CD62L- - Tem) phenotype before adoptive transfer can mediate more significant therapeutic activity. Transforming-growth factor-beta (TGF-β) is a pleiotropic cytokine that influences several aspects of T-cell biology and that is best known for its growth suppression and immunosuppressive activity. We show that TGF-β signaling can have a profound impact on the number of CD4 and CD8 T cells expressing the Tcm and Tem phenotype after anti-CD3e and anti-CD28 stimulation without altering the number of cells recovered at the end of the culture and without inducing regulatory T cells. By enhancing the percentage of the lymph-node homing receptors, L-selectin (CD62L) and CC-chemokine receptor 7 (CCR7) expressing cells, exogenous TGF-β, added to the culture medium to a concentration of 5 ng/ml, favors Tcm over Tem cell accumulation at 7 days (CD4+: 55.80 vs 35.51% (P= 0.0074); CD8+: 56.9 vs 40.3% (P= 0.0063)). Reciprocally, the inhibition of TGF-β signaling with a TGF-β receptor kinase inhibitor (GW788388) accentuated Tem phenotype acquisition. Importantly, these effects of TGF-β on Tcm marker expression were maintained in the presence of exogenous cytokines commonly used in ex-vivo cultures for adoptive immunotherapy (IL-2, IL-7 and IL-15). The manipulation of TGF-β signaling did not increase the expression of exhaustion markers (KRLG-1, CD57) but exogenous TGF-β decreased interferon-gamma (IFN-γ) expression. No effect was noted on TNF-α and IL-2 expression as well as on the percentage of polyfunctional cells generated. We also found that modulating TGF-β signaling during the course of clinically relevant cultures capable of expanding T-cells specific for Epstein-Barr virus LMP2 protein antigens, in the presence of IL-7 and IL-15, could increase the number of CD4 Tcm cells, even 2 weeks after the withdrawal of TGF-β from the culture (38.24 vs 27.82, N=3) without compromising antigen-specific IFN-γ release. In conclusion, the modulation of TGF-β signaling can significantly alter Tcm and Tem phenotype acquisition and may therefore be used to optimize Tcm phenotype expression by ex-vivo pathogen/antigen-specific T cells expanded for adoptive immunotherapy. Disclosures No relevant conflicts of interest to declare.

Functional Consequences of Memory Inflation after Solid Organ Transplantation

“…This finding was surprising given that identification of cells through IFN-g secretion in response to CMV Ag should exclude exhausted or anergic cells that recognize CMV (32,33). TGFb modulates memory T cell differentiation and can promote the formation of less differentiated central memory over effector memory (34). This suggests that TGFB1 in the effector cluster may represent heterogeneity, comprising TGFB1 1 central memory and TGFB1 effectors.…”