Selective Expansion of Human T Regulatory Cell Subsets and T-cell Depletion: Role of Rapamycin (Sirolimus)

Christoph Bergmann; Laura Strauss; Stephan Lang; Magis Mandapathil; Theresa L. Whiteside
Objective: The immunosuppressive drug rapamycin promotes the expansion of regulatory T-cell subsets, CD4+CD25high Foxp3+ and interleukin (IL)-10+ Tr1 cells. The mechanisms still remain unknown. Here, we studied expansion, survival and apoptotic pathways of human regulatory T cells (nTreg) and Tr1 cells in response to rapamycin. Methods: CD4+CD25+ and CD4+CD25- T cells were sorted from peripheral blood mononuclear cells of normal controls (n=15) using AutoMACS. CD4+CD25+ T cells were expanded in the presence of anti-CD3/CD28 Abs and 1000 IU/mL IL-2 for 3 weeks (nTreg). Sorted CD4+CD25- T cells were co-cultured with autologous immature dendritic cells and tumor cells of HNSCC cell line in presence of low doses of IL-2, IL-10, and IL-15 for 10 days (Tr1 cells). Rapamycin (1 nM) was added to half of the cultures. After harvest, phenotype expression of survival proteins or rapamycin-induced apoptosis (AnnexinV) was determined. Regulatory function was tested in co-cultures with autologous CFSE-labeled CD4+CD25- or CD8+CD25- T responders. Results: Expansion of CD4+CD25high Foxp3+ nTreg cells was significantly promoted after expansion of CD4+CD25+ T cells in presence of rapamycin (P<0.001) and cells showed high suppressor activity (>82%). Furthermore, rapamycin promoted up-regulation of antiapoptotic and down-regulation of proapoptotic proteins on nTreg, opposed to inverse effect on naive T cells. In vitro induction of Tr1 cells with suppressor activity in CD4+CD25- T cells was significantly augmented in presence of rapamycin, as was level of expressions of apoptotic proteins and cytotoxins on these cells. Both nTreg and Tr1 cells were resistant to rapamycin-mediated apoptosis. Conclusions: Rapamycin models the sensitivity of effector and suppressor T cells to apoptosis favoring the survival of Treg subsets, nTreg and Tr1 cells. Thus, this drug is applicable decedent studies of these cells and may be beneficial in immunotherapy in T-cell mediated diseases.