RAPAMYCIN EXPANDS AND CONFERS RESISTANCE TO APOPTOSIS OF HUMAN INDUCIBLE REGULATORY T CELLS (TR1)

The immunosuppressive drug Rap amycin (RAPA) has been shown to promote expansion of CD4(+) IL-10(+) natural human regulatory T cells (nTreg) in vitro and in vivo. RAPA effects on inducible Treg (Tr1) are unknown, and this study explores in vitro responses of Tr1 to this drug. CD4(+)CD25(neg) T cells isolated from PBMC of normal donors were used to generate Tr1 cells. Expanded Tr1 cells were tested for surface markers, expression of survival proteins, resistance to apoptosis and the ability to suppress proliferation of autologous CD4(+)CD25(neg) responder T cells (RC) in functional assays. PAPA was found to promote the generation of human Tr1 cells from autologous CD4(+)CD25(neg) precursors in peripheral blood. Tr1 cells + PAPA mediated higher suppression (p<0.01) of RC proliferation than Tr1 cells cultured without RAPA. Tr1 cells + RAPA also expressed higher levels of FasL and granzyme B (p<0.002), produced more IL-10 and TGF-beta 1 and were more resistant to activation-induced cell death (p<0.02). PAPA up-regulated expression of the Bcl-2 family anti-apoptotic proteins in Tr1. In addition, stimulation of Tr1 by LPS + PAPA resulted in increased proliferation and resistance to apoptosis. PAPA favors in vitro generation of inducible human Treg (Tr1) from CD4(+)CD25(neg) precursor cells and significantly enhances their survival and suppressor functions.