Background: Immunosuppressive factors such as regulatory T cells (Tregs) limit the efficacy of immunotherapies. Histone deacetylase (HDAC) inhibitors have been shown to have anti-tumor activity in different malignancies and to induce immuno-modulatory effects. We have previously reported that a class I selective HDAC inhibitor, entinostat, has synergistic anti-tumor effects in combination with high dose interleukin-2 (IL-2) in a renal cell carcinoma model (Kato Y, et al. Clinical Cancer Res 2007). Our group has also recently shown that low dose entinostat induces STAT3 acetylation, down-regulates Foxp3 expression in Tregs, and blocks Tregs suppressive function without affecting T effector cells (Shen Li, et al. PLoSONE 2012). Methods: Based on these preclinical evidences, we have initiated a Phase I/II clinical study with entinostat and high dose IL-2 in patients (pts) with metastatic renal cell carcinoma. The primary objective of the phase I portion was to evaluate the safety and tolerability of this combination strategy. The main eligibility criteria were clear cell histology, no prior treatments, and being fit to receive high dose IL-2. Results: The phase I portion consisted of two dose levels of entinostat (3 and 5 mg) and a fixed standard dose of IL-2 (600,000 units/kg every 8 hrs) according to a 3+3 design. To date, dose level one and two have been completed without DLTs. No severe adverse events have been observed during the first 45 days of treatment. The most common transient grade 3/4 toxicities were hypophosphatemia and thrombocytopenia (6 pts), neutropenia and lymphopenia (2 pts). The 5 mg dose level has been expanded. To date we have enrolled eight eligible pts and six have completed one cycle (84 days) treatment with four achieving objective responses by RECIST criteria. Decreased Tregs and increased CD8 numbers have been observed in tumor biopsies following treatment. Conclusions: The results from the phase I portion suggest that entinostat can be given safely in combination with high dose IL-2. Preliminary results also show encouraging biological and efficacy data (R21CA137649; U0-1CA70095). Clinical trial information: NCT01038778.