Abstract A192: Role of O(6)-Methylguanine-DNA Methyltransferase in the Proliferation/Invasion Dichotomy and Differential Effect of FAK inhibitor Y11 based on expression of MGMT in Glioblastoma.

Abstract

Abstract Introduction: Glioblastoma multiforme (GBM) is the most frequent and aggressive form of malignant primary brain tumors in adults. Prognosis for patinets diagnosed with GBM remains poor, with 90% of patients recurring within 2 years. Increased proliferation and invasion are the major hallmarks of GBM. Previous studies reported an inverse correlation between proliferation and invasion in GBM, a phenomenon termed the migration/proliferation dichotomy. Our laboratory has established for the first time the role of MGMT The DNA repair protein O6-methylguanine-DNA methyltransferase (MGMT) as a potential negative regulator of GBM angiogenesis and invasion in GBM. SPARC (secreted protein acidic and rich in cysteine) is known for its pro-invasive and anti-proliferative effects in GBM. SPARC interacts with β1 integrin and activates focal adhesion kinase (FAK), a key regulator of migration and proliferation. FAK is overexpressed in patients diagnosed with GBM and has been proposed as a molecular target in GBM. Y11, a novel specific small molecule inhibitor highly selective for Tyr-397, the main autophosphorylation site of FAK has not been tested in GBM. Methods: We used U87/Empty vector (U87/EV) negative for MGMT and its counterpart overexpressing MGMT (U87/MGMT), T98/EV with constitutive expression of MGMT and its knockdown counterpart T98/MGMT-shRNA in addition to GBM cell lines with different levels of MGMT expression (U251, U138, LN18 and A172). The effect of Y11 was investigated using MTT proliferation assay (inhibiting concentration at 50%) and western blotting to assess p-FAK- Tyr-397, Total FAK and SPARC levels. Results: Using isogenic overexpression (U87/EV, U87MGMT) and knockdown (T98/EV, T98/shRNA) models of MGMT in GBM, we showed that expression of MGMT was associated with increased proliferation and decreased invasion. Western blotting (WB) analysis in isogenic cell lines for MGMT and other cell lines with different levels of MGMT expression (U251, U138, LN18 and A172) revealed a striking inverse relationship between expression of MGMT and SPARC. Importantly, we showed that treatment with the small molecule inhibitor Y11, which specifically inhibits FAK Tyr397 induced a dose dependent decrease of pFAK in all cell lines, while total FAK levels were unchanged. This decrease was accompanied by decreased proliferation in all cell lines. Conclusion: Our study reveals that MGMT regulates the proliferation/invasion dichotomy through regulation of SPARC and FAK and highlights the differential effect of Y11 on proliferation and invasion based on expression of MGMT. This will lead to the identification of new biomarkers of invasion and proliferation and prospective therapies targeting molecular effectors involved in invasion and proliferation. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A192. Citation Format: Kenny Chatoor, Siham Sabri, Bassam Abdulkarim. Role of O(6)-Methylguanine-DNA Methyltransferase in the Proliferation/Invasion Dichotomy and Differential Effect of FAK inhibitor Y11 based on expression of MGMT in Glioblastoma. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A192.