Abstract 6416: Statins inhibit onco-dimerization of 4Ig-B7-H3

Abstract

Abstract B7-H3 (CD276) is a member of the B7 family of immune regulators, but unlike the immune checkpoint targets that have revolutionized cancer therapy, blockade has resulted in mixed outcomes. B7-H3 has 2Ig and 4Ig isoforms, has no known direct ligand, and many of the physiological functions remain elusive. While protein expression of B7-H3 is differentially expressed on many solid tumors and correlates with poor survival, mechanisms of how B7-H3 elicits a pro-tumorigenic phenotype are still poorly understood, significantly hampering therapeutic efforts for targeting. Using a facile, recombinase-enhanced split-luciferase reporter, we visualized 4Ig-B7-H3 homodimerization in live U2OS cells by bioluminescence imaging and confirmed 4Ig-B7-H3 dimerization using a second non-invasive imaging technique: fluorescence lifetime imaging microscopy (FLIM) where 4Ig-B7-H3 was exogenously re-expressed following knockout of CD276 in HeLa cervical cancer cells. Homodimerization of 4Ig-B7-H3 correlated with enhanced clonogenic cell growth in HeLa, SKOv3, MDA-MB-231, and U2OS cells and tumor growth in vivo for several models. This phenotype was reversed following knockout of B7-H3 and rescued upon lentiviral re-expression. By RPPA and phospho-kinase array analysis, increases in PI3K/AKT signaling, Jak/Stat pathway, and modulators of HIF1α and NF-κβ pathways were observed with B7-H3 expression and homodimerization (p<0.05), all which are known to support increased oncogenic functions, such as increased glycolysis, increased survival and proliferation, as well as cytokine modulation and immune evasion. Next, capitalizing on our bioluminescence-based reporter of 4Ig-B7-H3 homodimerization, we performed a high-throughput small molecule screen with a 5,362 Bioactive compound library to identify modulators of 4Ig-B7-H3 dimerization. Notably, our HTS identified several HMG-CoA reductase inhibitors (statins) as significant inhibitors of B7-H3 dimerization (p<0.01). Treatment with 1 μM mevastatin or atorvastatin calcium inhibited clonogenic growth of HeLa cervical, SKOv3 ovarian and MDA-MB-231 breast cancer cells, which all endogenously express B7-H3. In vivo, treatment with 10 mg/kg of atorvastatin calcium reduced tumor growth and increased mouse survival (p=0.035) of SKOv3-ip tumors and produced functional cures (15-30%) of HeLa or MDA-MB-231 xenografts in nude mice in a B7-H3 tumor expression-dependent manner, proven through knockout and rescue experiments in live cells and animals. Thus, in the context of studies to dissect the tumor-intrinsic mechanism(s) of B7-H3 biochemical functions and activation, we identified a novel, dimerization-dependent oncogenic role for 4Ig-B7-H3 that increased oncogenic intracellular signaling activation and was modulated by statin treatment. This may provide a molecular mechanism for the statin-mediated immune modulation and enhanced cancer survival clinically reported for several tumor types. Citation Format: Margie Nicole Sutton, Sarah E. Glazer, Riccardo Muzzioli, Ping Yang, Seth T. Gammon, David Piwnica-Worms. Statins inhibit onco-dimerization of 4Ig-B7-H3 [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 6416.