Abstract 2422: The lysine demethylase KDM1A inhibition attenuates hypoxic responses in glioblastoma

Abstract

Abstract Background: Glioblastoma (GBM) is associated with poor survival (1 year-34.6% and 5 year-4.75%) and affects approximately 13,000 patients per year. Standard treatment consists of surgical resection, external beam radiation therapy, adjuvant chemotherapy with temozolomide and tumor treating fields. Nonetheless, despite a heavy investment in therapy, all patients eventually succumb to their disease. Hypoxic regions are common in GBM and are implicated in maintenance of glioma stem cells (GSCs), promoting angiogenesis and therapeutic resistance. Recently, we and others have shown that lysine-specific histone demethylase 1A (KDM1A) is overexpressed in GBM and GSCs. However, the role of KDM1A and its functions in hypoxic response remains unknown. The objective of this study is to elucidate the functional role of KDM1A in hypoxia and test the efficacy of novel KDM1A inhibitors on hypoxia-mediated functions in GBM. Methods: The expression of KDM1A following hypoxia induction was determined in vitro by Western blotting and qRT-PCR. The expression of KDM1A in GBM tissue was examined by immunohistochemistry (IHC). KDM1A knockout cells were generated using CRISPR/Cas9 system and knockdown cells were generated using KDM1A shRNA lentiviral particles. The effect of KDM1A knockdown or treatment with KDM1A inhibitors NCL-1 and NCD-38 on stemness and self-renewal of primary patient-derived GBM cells was examined using neurosphere formation and extreme limiting dilution assays. Interaction of KDM1A with hypoxia inducible factor 1α (HIF-1α) was examined by immunoprecipitation (IP). The role of KDM1A on HIF target genes was determined using qRT-PCR and HRE-reporter activity. Mouse orthotopic xenografts models were used for preclinical evaluation of KDM1A inhibitors. Results: Western blot analysis revealed that KDM1A expression is induced in primary GBM cells following hypoxia. IHC analysis demonstrated elevated expression of KDM1A in hypoxic regions in human GBM tissues as well as mouse GBM xenografts. IP analysis demonstrated that KDM1A interacts with HIF-1α. Cell viability assays revealed that NCD-38 is more potent in reducing the cell viability of primary GBM cells compared to other KDM1A inhibitors. KDM1A knockdown or KDM1A inhibitor treatment significantly reduced the HIF- target gene expression including VEGF, PGK, CAIX, and HRE-Luc reporter activity in GBM cells. Further, hypoxia mediated cell proliferation and self-renewal ability of GSCs was compromised in KDM1A knockdown and KDM1A inhibitor-treated cells. Further, knockdown of KDM1A or treatment with KDM1A inhibitor significantly reduced the in vivo tumor growth and improved survival in orthotopic models. Conclusions: Our results demonstrate that KDM1A plays a critical role in hypoxia response and inhibition of KDM1A is a novel therapeutic approach for the treatment of GBM. Citation Format: Gangadhara R. Sareddy, Suryavathi Viswanadhapalli, Mei Zhou, Yiliao Luo, Xiaonan Li, Jinyou Liu, Aleksandra Gruslova, Takayoshi Suzuki, Andrew Brenner. The lysine demethylase KDM1A inhibition attenuates hypoxic responses in glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2422.