Rapid and profound potentiation of Apo2L/TRAIL-mediated cytotoxicity and apoptosis in thoracic cancer cells by the histone deacetylase inhibitor Trichostatin A: the essential role of the mitochondria-mediated caspase activation cascade

Abstract

Apo2L/TRAIL is actively investigated as a novel targeted agent to directly induce apoptosis of susceptible cancer cells. Apo2L/TRAIL-refractory cells can be sensitized to the cytotoxic effect of this ligand by cytotoxic chemotherapeutics. The aim of this study was to evaluate the in vitro tumoricidal activity of the Apo2L/TRAIL + Trichostatin A in cultured thoracic cancer cells and to elucidate the molecular basis of the synergistic cytotoxicity of this combination. Concurrent exposure of cultured cancer cells to sublethal concentrations of Apo2L/TRAIL and Trichostatin A resulted in profound enhancement of Apo2L/TRAIL-mediated cytotoxicity in all cell lines regardless of their intrinsic susceptibility to this ligand. This combination was not toxic to primary normal cells. While Apo2L/TRAIL alone or Trichostatin A alone mediated < 20% cell death, 60 to 90% of cancer cells were apoptotic following treatment with TSA + Apo2L/TRAIL combinations. Complete translocation of Bax from the cytosol to the mitochondria compartment was mainly observed in combination-treated cells and this was correlated with robust elevation of caspase 9 proteolytic activity indicative of activation of the mitochondria apoptogenic effect. Profound TSA + Apo2L/TRAIL-mediated cytotoxicity and apoptosis were completely abrogated by either Bcl2 over-expression or by the selective caspase 9 inhibitor, highlighting the essential role of mitochondria-dependent apoptosis signaling cascade in this process. Moreover, increased caspase 8 activity observed in cells treated with the TSA + Apo2L/TRAIL combination was completely suppressed by Bcl-2 over-expression or by the selective caspase 9 inhibitor indicating that the elevated caspase 8 activity in combination-treated cells was secondary to a mitochondria-mediated amplification feedback loop of caspase activation. These finding form the basis for further development of HDAC inhibitors + Apo2L/TRAIL combination as novel targeted therapy for thoracic malignancies.

Cisplatin enhances the antitumor effect of tumor necrosis factor-related apoptosis-inducing ligand gene therapy via recruitment of the mitochondria-dependent death signaling pathway

“…[41][42][43] We have previously observed that cultured thoracic cancer cells, while expressing adequate levels of DR4/ DR5, are generally refractory to the cytotoxic effect of the soluble recombinant death ligand Apo2L/TRAIL. [21][22][23] Infecting these cells with AdVgTRAIL caused at most moderate reduction of cell viability at an MOI of 10 PFU per cell. Higher cytotoxic effect could be achieved in some cell lines at higher MOIs of 20 or 50 PFU per cell (data not shown) but this might not be clinically practicable or achievable.…”

“…We elected not to focus our effort on dissecting out the effect of CDDP on modulating the phenotypic expression of pro-and antiapoptotic proteins or members of the deathinducing signaling complex that play crucial roles in transducing death signals through the extrinsic and intrinsic pathways as previous works in our laboratory found this approach to be nonproductive. [21][22][23] We, instead, concentrated on functionally evaluating the role of the mitochondria-dependent intrinsic pathway and the positive caspase activation feedback loop on mediating the enhanced cytotoxicity observed in combinationtreated cells. The observation that profound cytotoxicity by the CDDP/AdVgTRAIL combinations was totally abrogated either by the selective caspase 9 inhibitor or siRNA-mediated knockdown of caspase 9 expression or by overexpression of Bcl2 provided irrefutable evidence that cell death in this setting was mediated exclusively by type II receptor-mediated apoptosis.…”

“…19 Despite expressing adequate levels of DR4/DR5 functional receptors for TRAIL, a significant proportion of cancer cells exhibit resistance to the cytotoxic effect of this ligand. [20][21][22] This can be overcome by combining Apo2L/TRAIL or anti-DR4 antibody with cytotoxic chemotherapeutics such as cisplatin or paclitaxel. 20,[23][24][25][26][27][28][29][30][31][32] While the underlying mechanisms responsible for the synergistic interactions between chemotherapeutics and TRAIL receptor agonists to mediate profound induction of apoptosis are complex, 5,24,25,[33][34][35][36] it appears that recruitment/activation of the intrinsic death pathway (mitochondria-mediated) in combination-treated cells plays the crucial role.…”

Mammalian Target of Rapamycin Contributes to the Acquired Apoptotic Resistance of Human Mesothelioma Multicellular Spheroids

“…Final concentrations of inhibitors were chosen based on prior work (10,22). Concentrations of apoptotic agents were derived from the literature (23)(24)(25). Anisomycin was used at a low, subtoxic concentration (25 ng/ml) that we have previously shown does not inhibit the proliferation of our cells (23).…”

Comprehensive identification of genes driven by ERV9-LTRs reveals TNFRSF10B as a re-activatable mediator of testicular cancer cell death

“…Treatment of testicular cancer cells with HDACi not only results in the induction of ERV9-LTR promoter activity, but also in rapid apoptosis. 3,25,26 We therefore attempted to identify ERV9-LTR-driven genes that may contribute to programmed cell death. Strikingly, TNFRSF10B, encoding the death receptor 5, was among the HDACi-responsive and ERV9-LTR-downstream genes control 6h TSA 12hTSA 18h TSA Treatment with the solvent DMSO served as control.…”