Abstract 426: Targeting CXCR4 reduced T regulatory cells (Tregs) -mediated cell proliferation suppression in renal cell carcinoma (RCC) patients

Introduction: Renal cell carcinoma (RCC) is an immunogenic cancer with specimens containing tumor infiltrating lymphocytes. T regulatory cells CD4+CD25+Foxp3+ (Tregs) accumulate in RCC and have been implicated in tumor immune escape. Since the chemokine receptor axis CXCR4/CXCL12/CXCR7 plays a crucial role in renal cancer biology and recent evidences demonstrate that targeting CXCR4 modulate inefficient immunotherapy, the aim of the project was to evaluate patients Tregs function from tumor tissue compared to unaffected surrounding tissue and PBL and to study the effect of CXCR4 antagonism on Tregs immuneresponse. Materials and methods: 26 specimens from RCC, unaffected surrounding tissue and relative PBMCs plus heparinized blood from 15 healthy donors (HD) were collected. Single cells suspensions were obtained; Tregs from renal tumor, unaffected surrounding tissues and peripheral blood and circulating T effector (Teff) cells were isolated using a magnetic isolation cell kit. Treg were identified as CD4/CD25/Foxp3 and characterized for surface activation markers (CTLA-4; CXCR-4; PD-1; CD39; ICOS) by flow cytometry. Tregs suppressive activity was evaluated through in vitro co-colture of purified Tregs versus CFSE-labeled autologous Teff cells. Tregs were added at different ratios and co-cultured for 5 days in presence of anti-CD3/CD28 antibody. CXCR4 antagonism on Tregs suppressive function was evaluated through AMD3100, a selective CXCR4 antagonist, and Peptide R/Peptide R29, a new class of CXCR4 antagonists recently developed (1). Results: Tregs were evaluated in tumor tissue, unaffected surrounding tissue and relative PBMCs from 26 RCC patients and 15 healthy donors (HD). Higher number of Tregs was detected in the RCC tumor tissues and PBMCs compared to HD (p