Abstract Background Despite intensive effort, currently no effective anti-Ras therapies have successfully reached clinical application. Previous studies suggest that the histone deacetylatse (HDAC) inhibitor romidepsin, which is currently in clinical trials for the treatment of multiple malignancies, can block Ras-dependent signaling and growth transformation. These studies suggest that mutational activation of Ras may be a useful biomarker for sensitivity to romidepsin and that the anti-tumor activity of this HDAC inhibitor may involve inhibition of Ras effector-mediated signaling. Results To rigorously assess romidepsin as an antagonist of Ras, we utilized two well-characterized cell models for Ras transformation. We found that romidepsin blocked the anchorage-dependent and -independent growth of NIH 3T3 fibroblasts and RIE-1 epithelial cells transformed by all three Ras isoforms. However, romidepsin treatment also blocked growth transformation caused by other oncoproteins (B-Raf and ErbB2/Neu), suggesting that romidepsin is not selective for Ras. We also observed striking differences in romidepsin-mediated growth inhibition between transformed NIH 3T3 fibroblasts compared to RIE-1 epithelial cells, suggesting that the mechanism by which romidepsin blocks transformation is dependent on cellular context. Finally, we found that romidepsin did not inhibit Ras activation of the ERK and AKT effector pathways in NIH 3T3 and RIE-1 cells, suggesting that romidepsin does not directly antagonize Ras. Conclusion Taken together, our results suggest that romidepsin is not selective for Ras-transformed cells and that the anti-tumor activity of romidepsin is not due to direct inhibition of Ras function.