Bcl-2/Bcl-xL inhibition predominantly synergistically enhances the anti-neoplastic activity of a low-dose CUSP9 repurposed drug regime against glioblastoma.

Drug repurposing represents a promising approach to safely accelerate the clinical application of therapeutics with anti-cancer activity. In this study, we examined whether inhibition of the anti-apoptotic Bcl-2 family proteins Bcl-2 and Bcl-xL enhances the biological effects of the repurposed CUSP9 regimen in an in vitro setting of glioblastoma. We applied 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assays to assess cellular proliferation. Annexin V/propidium iodide and tetramethylrhodamine, ethyl ester staining were used to examine apoptosis. Western blotting, RT-PCR, and specific knockdown experiments using siRNA were employed to examine molecular mechanisms of action. Bcl-2/Bcl-xL inhibition exerted synergistic anti-proliferative effects across established, primary cultured, and stem-like glioblastoma cells when combined with CUSP9 which had been reduced to only one tenth of its proposed original concentration (CUSP9-LD). The combination treatment also led to enhanced apoptosis with loss of mitochondrial membrane potential and activation of caspases. On the molecular level, CUSP9-LD counteracted ABT263-mediated up-regulation of Mcl-1. Silencing of Mcl-1 enhanced ABT263-mediated apoptosis which indicates that down-regulation of Mcl-1 is crucial for the induction of cell death by the combination treatment. These data suggest that Bcl-2/Bcl-xL inhibition enhances the susceptibility of glioblastoma cells towards CUSP9, allowing dramatic dose reduction and potentially decreased toxicity when applied clinically. A clinical trial involving the original CUSP doses (CUSP9v3) is currently ongoing in our institution (NCT02770378). The Bcl-2/Bcl-xL inhibitor ABT263 is in clinical trials and might represent a valuable adjunct to the original CUSP.

Halatsch ME ，Kast RE ，Dwucet A ，Hlavac M ，Heiland T ，Westhoff MA ，Debatin KM ，Wirtz CR ，Siegelin MD ，Karpel-Massler G ... -  《-》

TIC10/ONC201 synergizes with Bcl-2/Bcl-xL inhibition in glioblastoma by suppression of Mcl-1 and its binding partners in vitro and in vivo.

Karpel-Massler G ，Bâ M ，Shu C ，Halatsch ME ，Westhoff MA ，Bruce JN ，Canoll P ，Siegelin MD ... -  《Oncotarget》

Combined inhibition of RAC1 and Bcl-2/Bcl-xL synergistically induces glioblastoma cell death through down-regulation of the Usp9X/Mcl-1 axis.

Hlavac M ，Dwucet A ，Kast RE ，Engelke J ，Westhoff MA ，Siegelin MD ，Debatin KM ，Wirtz CR ，Halatsch ME ，Karpel-Massler G ... -  《-》

Dual Inhibition of Bcl-2/Bcl-xL and XPO1 is synthetically lethal in glioblastoma model systems.

Shang E ，Zhang Y ，Shu C ，Ishida CT ，Bianchetti E ，Westhoff MA ，Karpel-Massler G ，Siegelin MD ... -  《Scientific Reports》

Inhibition of Bcl-2/Bcl-xL and c-MET causes synthetic lethality in model systems of glioblastoma.

Zhang Y ，Ishida CT ，Shu C ，Kleiner G ，Sanchez-Quintero MJ ，Bianchetti E ，Quinzii CM ，Westhoff MA ，Karpel-Massler G ，Siegelin MD ... -  《Scientific Reports》