Intrinsic resistance of prostate cancer to the BH3 mimetic ABT-737 is related to expression of the anti-apoptotic Mcl-1 and additional survival and apoptosis regulating pathways

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Background: ABT-737 is a novel and potent oral small molecule inhibitor of the anti-apoptotic Bcl-2 and Bcl-xL proteins. ABT-737 has substantial single agent or synergistic activity in myeloid/lymphoid malignancies and small cell lung cancer. Overexpression of Bcl-2/Bcl-xL is linked to prostate cancer progression, hence rendering Bcl-2 and Bcl-xL attractive therapeutic targets. The in vitro activities of ABT-737 in prostate cancer models and potential resistance mechanisms were investigated.
 Methods: Cytotoxicity of ABT-737 and potential synergism with apoptosis inducing stimuli was investigated in PC3, DU145, C42, LNCaP cell using cell viability assay. Expression of anti-apoptotic Bcl-2 members in PC cell lines was assessed by immunoblotting (IB) and related to cytotoxic effect of ABT-737. The effect of serum deprivation (SD) on ABT-737 cytotoxicity was assessed and explorative studies of potential ABT-737 resistance mechanisms under serum deprived growing conditions were conducted. The impact of Mcl-1 down regulation following transfection with specific antisense oligodeoxynuleotide (ODN) on ABT-737 induced cytotoxicity was evaluated.
 Results: ABT-737 had no to minor cytotoxic effect in tested PC lines (IC₅₀ > 10μM, 72hours). Combination of ABT-737 with androgen-deprivation or cyctotoxic agents (paclitaxel, CDDP) caused no or mild additive effect (20-40%) in some but not other cell lines. In contrast, SD (72hours) exhibited pronounced synergistic activity with ABT-737 (IC₅₀ 0.5μM PC3/DU145/C42, 2.5μM LNCaP) but not the inactive enantiomer while no impact on cell viability by SD alone was noted. Expression signature of anti-apoptotic Bcl-2 proteins demonstrated that Mcl-1 was expressed in all PC lines on IB. Mcl-1 down regulation following antisense ODN markedly restored sensitivity to ABT-737 (IC50 2-3 μM) but not enantiomer in all but DU145 (IC₅₀ 5-10μM). Expression dynamics of pro-/anti-apoptotic Bcl-2 proteins, clusterin, PI3K, MAPK under normal and SD growing conditions indicated several potential mechanisms associated with ABT-737 resistance in PC.
 Conclusion: ABT-737 did not demonstrate single agent activity or meaningful additive/synergistic activity with commonly used therapeutic maneuvers in preclinical prostate cancer models. Sensitivity to ABT-737 appeared to be restored upon serum withdrawal in PC model. Multiple and cell line specific expression and alterations in signaling and apoptosis regulating proteins were associated with serum withdrawal. Our results identify Mcl-1 as an important determinant of ABT-737 resistance and validate Mcl-1 as potential therapeutic target in prostate cancer.