Identification and Validation of Proteasome Subunit Alpha Type-7 (PSMA7) as a Therapeutically Vulnerable Target in GBM

Abstract

<h3>Purpose/Objective(s)</h3> A neomorphic gain-of-function mutation of isocitrate dehydrogenase (IDH1/2) is significantly associated with favorable clinical outcome of patients with glioblastoma (GBM; WHO grade IV) compared with ones with wildtype IDH(1/2). The current standard treatment for GBM consists of maximal safe surgical tumor resection followed by radiation treatment (RT) and concomitant and adjuvant temozolomide (TMZ) (Stupp protocol). Despite the standard treatment, the five-year survival rate remains dismal, only 5%. Therefore, there is an urgent need to identify druggable, therapeutically vulnerable targets in GBM. In this study we identified PSMA7, an alpha-type subunit of the 20S proteasome core particle that is involved in the regulated degradation of proteins in cells via the ubiquitin-proteasome system. PSMA7 has been shown to be frequently upregulated in multiple cancer types and blocking PSMA7 inhibits cell proliferation in cervical, colorectal, and gastric cancer. Although the connection between PSMA7 and tumor progression has been studied in several tumor models, little is known about the role of PSMA7 in GBM biology and treatment response where its expression is significantly upregulated. <h3>Materials/Methods</h3> PSMA7 was identified from publicly available mRNA profiling data of primary glioma patients in two patient cohorts from the Chinese Glioma Genome Atlas (CGGA) database. PSMA7 was functionally validated <i>in vitro</i> using RNAi-mediated knockdown approaches. Stable cell lines with shRNA-mediated knockdown of PSMA7 were generated by puromycin selection. Protein expression was quantified by Western blot analysis. mRNA expression from <i>in vitro</i> studies was determined by qRT-PCR. Cell viability and cell proliferation were assessed by MTS or ATP assays. A biological irradiator was used to treat cells with ionizing radiation for evaluating RT response. <h3>Results</h3> Primary glioma patients with high PSMA7 mRNA levels (median split) were found to have worse overall survival than patients with low mRNA levels in two primary glioma cohorts. Moreover, PSMA7 was significantly upregulated in IDH1/2 wildtype (IDHwt) primary gliomas compared to IDH1/2 mutant (IDHmut) primary gliomas in these two cohorts. We confirmed PSMA7 mRNA and protein expression to be significantly higher in U87MG-IDHwt GBM cells compared to U87MG-IDHmut cells <i>in vitro</i>. RNAi-mediated knockdown of PSMA7 in PDX GBM cell lines decreased cell viability and increased sensitivity to ionizing radiation <i>in vitro</i>. We observed that PSMA7 knockdown in a PDX GBM cell line prolonged mouse overall survival in an <i>in vivo</i> orthotopic xenograft mouse model. <h3>Conclusion</h3> Our data indicate that PSMA7 is a potentially vulnerable therapeutic target in GBM as PSMA7 knockdown slowed GBM cell proliferation, sensitized GBM cells to ionizing radiation, and prolonged overall survival in an orthotopic xenograft mouse model.