Abstract 3376: Molecular Drivers of Endothelial Hypoxic Adaption During Angiogenesis Deciphered Using the Berg Interrogative Biology® Platform

Angiogenesis is a key feature of tumor progression providing oxygen and nutrients required for tumor cell growth, hypoxia being a major driver of this phenomenon. A systems biology approach using the Berg Interrogative Biology® platform was implemented to identify drivers of the endothelial cell (EC) angiogenic response to hypoxia. To determine the proteomic profile of proliferating ECs and non-proliferating confluent ECs exposed to normoxia or hypoxia, a functional proteomic approach employing activity-based probes and phosphorylation analysis was utilized. Kinases and ATPases were labeled with ATP-binding domain enrichment probes and titanium dioxide enrichment of phosphopeptides was employed for capture of protein phosphorylation events. Phenotypic assays including proliferation rates, apoptosis, mitochondrial superoxide and ROS/NO signaling were included as a measure of EC phenotype. Comparative proteomics, kinase activity, phosphoproteomics and assay data were integrated using an AI based Bayesian Network Inference approach to investigate causal signaling networks in order to elucidate the complexity and dynamics of angiogenesis and more specifically, the role of hypoxia in driving intracellular signaling in response to changes in oxygen tension. High confidence causal networks identified novel proteins that modulate the EC hypoxic response, validation of which are in process. Previously characterized proteins that are responsive to hypoxia were also identified in the hypoxic, but not normoxic signaling networks, namely Aldoc, Rac1, mTor, Cav-1 and Bax. Endothelial proliferation rate is closely related to both hydrogen peroxide and nitric oxide signaling, as has previously been reported. Novel regulatory networks that determine these interactions were identified. Using the Berg Interrogative Biology® platform, we are deciphering both the effects of the hypoxic microenvironment, and the unique characteristics of proliferating ECs by applying integrated functional proteomic assays and a systems approach to determine global changes in intracellular signaling in response to hypoxia. Citation Format: Tony E. Walshe, Justin Bourdelais, Viatcheslav R. Akmaev, Leonardo O. Rodrigues, Socheata Lao, Stephane Gesta, Michael A. Kiebish, Vivek K. Vishnudas, Rangaprasad Sarangarajan, Niven R. Narain. Molecular drivers of endothelial hypoxic adaption during angiogenesis deciphered using the Berg Interrogative Biology® platform. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3376.