Identification of approved and investigational drugs that inhibit hypoxia-inducible factor-1 signaling

// Chia-Wen Hsu 1 , Ruili Huang 1 , Thai Khuc 1 , David Shou 1 , Joshua Bullock 2 , Suzanne Grooby 2 , Sue Griffin 2 , Chaozhong Zou 3 , Annette Little 2 , Holly Astley 2 , Menghang Xia 1 1 Division of Pre-Clinical Innovation, National Center for Advancing Translational Sciences, National Institutes of Health, Bethesda, MD, USA 2 Horizon Discovery Ltd., Waterbeach, Cambridge, UK 3 American Type Culture Collection, Gaithersburg, MD, USA Correspondence to: Menghang Xia, e-mail: mxia@mail.nih.gov Keywords: hypoxia inducible factor, cancer, genome editing, drug, high-throughput screening Received: September 10, 2015 Accepted: January 05, 2016 Published: January 23, 2016 ABSTRACT One of the requirements for tumor development is blood supply, most often driven by hypoxia-induced angiogenesis. Hypoxia induces the stabilization of hypoxia-inducible factor-1 alpha (HIF-1α), which induces expression of an angiogenic factor, vascular endothelial growth factor (VEGF). The purpose of this study is to validate a new screening platform combined with orthogonal assays to rapidly identify HIF-1 inhibitors and to evaluate the effectiveness of approved drugs on modulating HIF-1 signaling. We generated an endogenous HIF-1α–NanoLuc luciferase reporter allele in the human HCT116 colon cancer cell line using genome editing and screened a panel of small interfering RNAs (siRNAs) to 960 druggable targets and approximately 2,500 drugs on a quantitative high-throughput screening (qHTS) platform. Selected compounds were further investigated with secondary assays to confirm their anti-HIF activity and to study their mode of action. The qHTS assay identified over 300 drugs that inhibited HIF-1α-NanoLuc expression. The siRNA screening results supported the effectiveness of several target-specific inhibitors. Moreover, the identified HIF-1 inhibitors, such as mycophenolate mofetil, niclosamide, and trametinib, were able to suppress cancer cell proliferation and angiogenesis. Our study indicates that blocking the mitogen-activated protein kinase (MAPK) and phosphoinositol 3-kinase (PI3K) pathways effectively inhibits hypoxia-induced HIF-1α accumulation and HIF-1α transactivation and that proteasome inhibitors induce accumulation and decrease transcriptional activity of HIF-1α. These findings underline the importance of developing a battery of robust assay platforms and confirmation studies that focus on endogenous protein targets so that only relevant and reliable data will be taken into pre-clinical and clinical studies.