Inhibition of B-Raf/MEK/ERK signaling is an effective therapeutic strategy against certain types of cancers such as melanoma and thyroid cancer. While demonstrated to be effective anticancer agents, B-Raf or MEK inhibitors have also been associated with early tumor progression and development of secondary neoplasms. The ligation of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) with its receptor, death receptor 5 (DR5), leading to induction of apoptosis, offers a promising anticancer strategy. Importantly, this is also a natural immunosurveillance mechanism against cancer development. We previously demonstrated that activated B-Raf/MEK/ERK signaling positively regulates DR5 expression. Hence, our current work sought to address whether B-Raf/MEK/ERK inhibition and the consequent suppression of DR5 expression impede cancer cell response to DR5 activation-induced apoptosis and activated immune cell-induced killing. We found that both B-Raf (for example, PLX4032) and MEK inhibitors (for example, AZD6244 and PD0325901) effectively inhibited ERK1/2 phosphorylation and reduced DR5 levels in both human thyroid cancer and melanoma cells. Similar to the observed effect of genetic knockdown of the B-Raf gene, pre-treatment of cancer cell lines with either B-Raf or MEK inhibitors attenuated or abolished cellular apoptotic response induced by TRAIL or the DR5 agonistic antibody AMG655 or cell killing by activated T cells. Our findings clearly show that inhibition of B-Raf/MEK/ERK signaling suppresses DR5 expression and impairs DR5 activation-induced apoptosis and T cell-mediated killing of cancer cells. These findings suggest a potential negative impact of B-Raf or MEK inhibition on TRAIL- or DR5-mediated anticancer therapy and on TRAIL/DR5-mediated immune-clearance of cancer cells.