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BRAF(V600E)-inhibitors (BRAFi; e.g., vemurafenib) and modern immune-based therapies such as PD-1/PD-L1 and CTLA-4 checkpoints blockade and adoptive cell transfer (ACT) have significantly improved the care of melanoma patients. Having these two effective (BRAFi and immunotherapy) therapies raises the question whether there is a rational biological basis for using them in combination. We developed an in vitro model to determine whether tumor resistance mechanisms to a small molecule inhibitor of a driver oncogene, and to cytotoxic T lymphocyte (CTL)- and natural killer (NK) cell-delivered apoptotic death signals were exclusive or intersecting. We generated melanoma sublines resistant to BRAFi vemurafenib and to CTL recognizing the MART-1 melanoma antigen. Vemurafenib-resistant (VemR) sublines were cross-resistant to MART CTL and NK cells indicating that a common apoptotic pathway governing tumor response to both modalities was disrupted. Pretreatment of VemR melanomas with a histone deacetylase inhibitor (HDACi) restored sensitivity to MART CTL and NK apoptosis by skewing the apoptotic gene programs towards a proapoptotic phenotype. Our in vitro findings suggest that during the course of acquisition of BRAFi resistance, melanomas develop cross-resistance to CTL- and NK-killing. Further, aberrant apoptotic pathways, amenable by an FDA-approved chromatin remodeling drug, regulate tumor resistance mechanisms to immune effector cells. These results may provide rational molecular basis for further investigations to combine these therapies clinically.

Protein degradation by proteasome is essential to the regulation of important cellular functions including cell cycle progression, proliferation, differentiation and apoptosis. Abnormal proteasomal degradation of key regulatory proteins perturbs the normal dynamics of these cellular processes culminating in uncontrolled cell cycle progression and decreased apoptosis leading to the characteristic cancer cell phenotype. Proteasome inhibitors are a novel group of therapeutic agents designed to oppose the increased proteasomal degradation observed in various cancers while restoring key cellular functions such as apoptosis, cell cycle progression, and the inhibition of angiogenesis. Several proteasome inhibitors have been evaluated in pre- and clinical studies for their potential usage in clinical oncology. Bortezomib (Velcade, PS-341) is the first Food and Drug Administration-approved proteasome inhibitor for the treatment of multiple myeloma and mantle cell lymphoma. Bortezomib's ability to preferentially induce toxicity and cell death in tumor cells while rendering healthy cells unaffected makes it a powerful therapeutic agent and has extended its use in other types of malignancies. The ability of bortezomib and other proteasome inhibitors to synergize with conventional therapies in killing tumors in various in vitro and in vivo models makes this class of drugs a powerful tool in overcoming acquired and inherent resistance observed in many cancers. This is achieved through modulation of aberrant cellular survival signal transduction pathways and their downstream anti-apoptotic gene products. This review will discuss the anti-neoplastic effects of various proteasome inhibitors in a variety of cancers with a special emphasis on bortezomib, its mechanism of action and role in cancer therapy. We further discuss the potential use of bortezomib in the treatment of metastatic melanoma.

TRAIL promotes apoptotic tumor cell death; however, TRAIL-resistant tumors need to be sensitized to reverse resistance. Proteasome inhibitors potentiate TRAIL apoptosis in vitro and in vivo and correlate with up-regulation of death receptor 5 (DR5) via an unknown mechanism. We hypothesized that the proteasome inhibitor NPI-0052 inhibits the transcription repressor Yin Yang 1 (YY1) which regulates TRAIL resistance and negatively regulates DR5 transcription. Treatment of PC-3 and Ramos cells with NPI-0052 (/=2.5 nM) and TRAIL sensitizes the tumor cells to TRAIL-induced apoptosis. By comparison to bortezomib, a 400-fold less concentration of NPI-0052 was used. NPI-0052 up-regulated DR5 reporter activity and both surface and total DR5 protein expression. NPI-0052-induced inhibition of NF-kappaB activity was involved in TRAIL sensitization as corroborated by the use of the NF-kappaB inhibitor dehydroxymethylepoxyquinomicin. NPI-0052 inhibited YY1 promoter activity as well as both YY1 mRNA and protein expression. The direct role of NPI-0052-induced inhibition of YY1 and up-regulation of DR5 in the regulation of TRAIL sensitivity was demonstrated by the use of YY1 small interfering RNA. The NPI-0052-induced sensitization to TRAIL involved activation of the intrinsic apoptotic pathway and dysregulation of genes that regulate apoptosis. The NPI-0052 concentrations used for TRAIL sensitization were not toxic to human hematopoetic stem cells. The present findings demonstrate, for the first time, the potential mechanism by which a proteasome inhibitor, like NPI-0052, inhibits the transcription repressor YY1 involved in TRAIL resistance and DR5 regulation. The findings also suggest the therapeutic application of subtoxic NPI-0052 concentrations in combination with TRAIL/agonist DR4/DR5 mAbs in the treatment of TRAIL-resistant tumors.