Cyclin D1-CDK4 activity drives sensitivity to bortezomib in mantle cell lymphoma by blocking autophagy-mediated proteolysis of NOXA

Abstract

BackgroundMantle cell lymphoma (MCL) is an aggressive B-non-Hodgkin lymphoma with generally poor outcome. MCL is characterized by an aberrantly high cyclin D1-driven CDK4 activity. New molecular targeted therapies such as inhibitors of the ubiquitin-proteasome system (UPS) have shown promising results in preclinical studies and MCL patients. Our previous research revealed stabilization of the short-lived pro-apoptotic NOXA as a critical determinant for sensitivity to these inhibitors. It is currently unclear how cyclin D1 overexpression and aberrant CDK4 activity affect NOXA stabilization and treatment efficacy of UPS inhibitors in MCL.MethodsThe effect of cyclin D1-driven CDK4 activity on response of MCL cell lines and primary cells to proteasome inhibitor treatment was investigated using survival assays (Flow cytometry, AnnexinV/PI) and Western blot analysis of NOXA protein. Half-life of NOXA protein was determined by cycloheximide treatment and subsequent Western blot analysis. The role of autophagy was analyzed by LC3-II protein expression and autophagolysosome detection. Furthermore, silencing of autophagy-related genes was performed using siRNA and MCL cells were treated with autophagy inhibitors in combination with proteasome and CDK4 inhibition.ResultsIn this study, we show that proteasome inhibitor-mediated cell death in MCL depends on cyclin D1-driven CDK4 activity. Inhibition of cyclin D1/CDK4 activity significantly reduced proteasome inhibitor-mediated stabilization of NOXA protein, mainly driven by an autophagy-mediated proteolysis. Bortezomib-induced cell death was significantly potentiated by compounds that interfere with autophagosomal function. Combined treatment with bortezomib and autophagy inhibitors enhanced NOXA stability leading to super-induction of NOXA protein. In addition to established autophagy modulators, we identified the fatty acid synthase inhibitor orlistat to be an efficient autophagy inhibitor when used in combination with bortezomib. Accordingly, this combination synergistically induced apoptosis both in MCL cell lines and in patient samples.ConclusionOur data demonstrate that CDK4 activity in MCL is critical for NOXA stabilization upon treatment with UPS inhibitors allowing preferential induction of cell death in cyclin D transformed cells. Under UPS blocked conditions, autophagy appears as the critical regulator of NOXA induction. Therefore, inhibitors of autophagy are promising candidates to increase the activity of proteasome inhibitors in MCL.Electronic supplementary materialThe online version of this article (10.1186/s13045-018-0657-6) contains supplementary material, which is available to authorized users.

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“…Blocking autophagy could increase cancer cells sensitivity to chemotherapy. For instance, bortezomib-induced MCL cell death was significantly potentiated by compounds that interfered with autophagosomal function [164]. HMGB1-dependent autophagy promotes chemotherapy resistance in three ways: nuclear HMGB1 upregulates the expression of HSP27, cytoplasmic HMGB1 activates the Beclin-1/PI3K-III complex, and extracellular HMGB1 binds to RAGE [45,165].…”

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“…Interestingly, CDK4/6 inhibitors might display influences on the tumor microenvironment. For instance, inhibition of CDK4 activity would significantly reduce the stabilization of autophagy-mediated proteasome NOXA and induce cell apoptosis in mantle cell lymphoma 78. Also, studies demonstrated that cyclin D-CDK4 kinase destabilized PD-L1 via Cullin3 SPOP to control cancer immune surveillance and revealed the potential combination of CDK4/6 inhibitors and PD-1/PD-L1 blockade to enhance therapeutic efficacy for human cancers 79.…”

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“…By interacting with MCL-1, NOXA allows the release of the pro-apoptotic effector, BAK, leading to mitochondrial depolarization and initiation of the apoptotic cascade [101]. Interestingly, the inhibition of cyclin D1/CDK4 activity in MCL cells reduces the stabilization of NOXA, directing the protein through degradation by an autophagy mechanism [110].…”