THE ROLE OF SERPINB3 ON GLIOBLASTOMA CANCER STEM CELL PROLIFERATION - Use of Patient Derived Glioblastoma Cell Lines to Show the Effect of SerpinB3 on

Prognosis

Glioblastoma (GBM) is the most common primary human malignant brain tumor; the median overall survival of those affected is 14 to 16 months. Poor prognosis can be attributed to the high recurrence rate of the disease, which is due, in part, to the presence of cancer stem cells (CSCs). This research is focused on studying the interaction between SerpinB3 and junction adhesion molecule A (JAM-A) in GBM CSCs. Previous studies provide evidence that JAM-A has an intrinsic, pro-tumorigenic role in regulating the CSC phenotype. None of these studies, however, focused specifically on JAM-A's intracellular downstream signaling. A pulldown of His-tagged JAMA was done to identify binding partners and resulted in the identification of the serine protease inhibitor SerpinB3. Endogenous JAMA binding to SerpinB3 was confirmed through immunoprecipitation of SerpinB3. To investigate the CSC-specific role of SerpinB3, SerpinB3 was knocked-down in a human GBM xenograft model (T4121) utilizing 2 non-overlapping short-hairpin RNA constructs. This generated 2 knockdown conditions (each a variation of expressed SerpinB3) and a nontarget control. A CellTiter-Glo® assay was used to measure CSC growth dynamics and a limiting dilution assay was done to determine the stem cell frequency. These in vitro studies demonstrated reduced function of the CSC when SerpinB3 was knocked down. GBM CSCs were orthotopically implanted into NSG mice via an intracranial injection (n=30). Median survival for non-target control mice was 28 days while neither knockdown condition reached median survival at day 50 (log-rank test, P< 0.001). This finding suggests that SerpinB3 disrupts the CSC state in GBM.