Abstract 4811: Oxygen tension - dependent differences in cancer cell kinome
Authors: Adebayo Adedeji K. , Kumar Brijesh , Davis Christopher , Angus Steven P. , Nakshatri Harikrishna 
Published: April 2023

Abstract
Current approaches to preclinical cancer research often fail to consider the impact of ambient oxygen (O2; ~21%) on cancer cells. This is also true for hypoxia studies that typically involves cancer cells previously grown in ambient O2 before subsequent transfer to hypoxia. However, the tumor microenvironment is characterized by significantly lower O2 levels. We have previously demonstrated the impact of ambient O2 on stem cell populations, signaling pathways and resistance to therapy. We developed an experimental approach that allows us to collect and process tumor tissues from transgenic mammary tumor mouse models and ovarian cancer patients under physioxia (3% O2), such that they are never exposed to ambient O2. In this study, our goal was to explore oxygen-dependent signaling pathway alterations and to determine how these pathways are influenced by targeted drugs in the context of physioxia or ambient air (AA). Our studies revealed increased basal phosphorylation levels of EGFR (Y1068) in the tumor cells in AA, relative to physioxia. However, downstream signaling effectors AKT and ERK showed higher phosphorylation levels under physioxia, compared to AA, suggesting that their activation is independent of EGFR signaling. These findings correlate with the decreased sensitivity of the tumor cells under physioxia to target drugs lapatinib and alpelisib. We then sought to examine basal and target drug induced kinome changes in tumor cells under physioxia and AA via Multiplexed Inhibitor Beads (MIBs) kinome assay. This assay revealed significant differences in the kinome of the tumor cells under physioxia compared to AA. Although direct comparison between vehicle and lapatinib treated cells in physioxia and ambient air showed very minimal changes, pairwise comparison between lapatinib treated physioxia cells and vehicle treated AA cells revealed an increase in the activity of PDGFRB in lapatinib treated physioxia cells. Similarly, a receptor tyrosine kinase (RTK) array and western blotting showed increased basal and lapatinib induced phosphorylation of PDGFRB (Y751) under physioxia. Next, we determined the potential role of PDGFRB in downstream signaling pathway activation of AKT and ERK and resistance to lapatinib. We found that sunitinib, a multitarget RTK inhibitor with high affinity for PDGFR effectively decreased PGDFRB activity under physioxia, with a concurrent decrease in the phosphorylation of AKT. Moreover, tumor cells under physioxia were more sensitive to sunitinib treatment, relative to ambient air. Furthermore, a combination of lapatinib and sunitinib rendered tumor cells under physioxia more sensitive to treatment than with lapatinib alone. These findings suggest that ambient and physioxic oxygen tensions differentially impact cancer relevant signaling pathways. Therefore, it may be necessary to carry out preclinical cancer studies in the context of physiologically relevant oxygen tensions to aid translatability.
Citation Format: Adedeji K. Adebayo, Brijesh Kumar, Christopher Davis, Steven P. Angus, Harikrishna Nakshatri. Oxygen tension - dependent differences in cancer cell kinome. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4811.