Abstract B17: Targeting MK2 to improve temozolomide efficacy in glioblastoma

Abstract

Abstract Glioblastoma is the most prevalent and aggressive form of brain cancer. The heterogeneous nature and survival mechanisms that glioblastoma tumors develop impede chemotherapy efficacy of temozolomide (TMZ) and result in tumor relapse. The p38 MAPK and its downstream kinase MK2 have been well established in regulating signaling pathways influencing inflammation, cell division and differentiation, as well as cell motility in response to a wide range of extracellular stimuli. Recent studies have identified p38 MAPK-MK2 pathway as an effector kinase complex that is activated following DNA damage and results in cell cycle arrest, so that cells have the capacity to repair their DNA and continue to proliferate. Importantly, cells with p53 mutation/deficiency were shown to rely on MK2 pathway for G2/M arrest and survival after exposure to chemotherapy and thus highlighting MK2 as a potential therapeutic target. We sought to determine if inhibition of MK2 in glioblastoma cells would result in increased efficacy of TMZ and to delineate the underlying mechanisms. Using genetic knockdown and pharmacological inhibitors we examined whether MK2 inhibition improves temzolomide efficacy in glioblastoma cell lines and patient-derived tumor cells of different subtypes (classical, mesenchymal, proneural and neural). Viability and clonogenic assays were used to determine survival of tumor cells to combination treatment. Apoptosis was determined using Annexin-V staining, cell cycle analysis using flow cytometry, mitotic entry by p-HH3 staining, extent of DNA damage using γH2AX staining and senescence using β-galactosidase staining. Western blotting analysis was performed to study changes in protein expression. We show that MK2 inhibition significantly improved TMZ anti-proliferative efficacy in A172 glioblastoma cell line and primary cells of classical subtype. Furthermore, MK2 inhibition reduced ability of TMZ treated cells to form colonies in clonogenic assay. Mechanistic studies revealed that MK2 knockdown reduced G2/M arrest induced by TMZ and promoted G0/G1 arrest. These changes were associated with increase in TMZ-induced apoptosis and senescence. We show that MK2 knockdown does not promote premature mitosis and mitotic catastrophe due to accumulation of DNA damage, as evidenced by lack of p-HH3 and γH2AX staining. In summary, our results demonstrate that MK2 inhibition is a novel strategy to sensitize glioblastoma cells to chemotherapy. We propose that in glioblastoma cells, MK2 signaling does not contribute to G2/M arrest induced by TMZ and instead MK2 plays a crucial role in regulating G0/G1 arrest in response to DNA damaging agents. Citation Format: Fadi Gurgis, Bryan Day, Brett Stringer, Terrance G. Johns, Lenka Munoz. Targeting MK2 to improve temozolomide efficacy in glioblastoma. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Cancer Cell Cycle - Tumor Progression and Therapeutic Response; Feb 28-Mar 2, 2016; Orlando, FL. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(11_Suppl):Abstract nr B17.