Abstract LB078: Hexyl-(cuban-1-yl-methyl)-biguanide (HCB) inhibits hormone therapy resistant breast cancer cells, in part by Inhibiting CYP3A4 arachidonic acid epoxygenase activity

Abstract

Abstract Introduction: Small molecule therapeutics of estrogen receptor-positive/HER2-negative breast cancer remains an area of active investigation where novel agents are greatly needed for treatment of hormone therapy resistant metastatic disease. The biguanide hexyl-benzyl-biguanide (HBB) is a potent inhibitor of CYP3A4 arachidonic acid (AA) epoxygenase activity and inhibits breast cancer cell proliferation and MCF-7 breast cancer tumor growth in nude mice. To explore the impact of bioisosteric substitution of the benzyl moiety of HBB with a cubane moiety, we synthesized hexyl-(cuban-1-yl-methyl)-biguanide (HCB) and tested its potency for the inhibition of the cognate CYP3A4 target AA epoxygenase activity as well as breast cancer cell proliferation of hormone therapy sensitive and resistant cell lines. Results: HCB selectively inhibited CYP3A4-mediated biosynthesis of (±)-14,15-EET with an IC50 of 4.7±0.2 uM vs. 64.8±6.5 uM for 8,9-EET and 26.5±1.9 uM for 11,12-EET. At 24 hours, HCB inhibited proliferation of MCF-7 (ER+HER2-), BT474 (ER+HER2+) and MDA-MB-231 (ER-HER2-) cells at IC50 of 8.4±1.2, 11±1.3 and 15±0.9 uM, respectively. At 48 hours, HCB inhibited proliferation of aromatase inhibitor and fulvestrant resistant (LR,FR), and cyclin dependent kinase inhibitor (CDKi) palbociclib resistant (LR,FR,PR) MCF-7 cell lines; LR,FR MCF-7AC1 (IC50 =1.34±0.1 uM) and LR,FR,PR MCF-7AC1 (IC50 =1.64±0.2 uM). Addition of 14,15-EET (1 uM) partially rescues MCF-7 cells from HCB-mediated inhibition of proliferation. OXPHOS is promoted, in part, by EETs. HCB is a potent OXPHOS inhibitor and rapidly inhibits O2 consumption of the MCF-7 and ZR75 (ER+HER2-) cells in a dose-dependent fashion (P<0.05). HCB treatment (10 uM) reduces mitochondrial membrane potential to 57.4±15.3% (P<0.001) of vehicle control in MCF-7 cells. Treatment with HCB at 20 uM for 0.5 hour also causes mitochondrial swelling in MCF-7 cells. HCB (10 uM) activates AMPK within 0.5 hour and increases the level of phosphorylation from 2.4±0.3 to 25.1±6.0 folds in a time dependent fashion in MCF-7 cells from 0.5-24 hours. Conclusion: These results show that HCB inhibits proliferation of ER+HER2- breast cancer cells, in part through inhibition of OXPHOS and suppression of the CYP product 14,15-EET. This inhibition is highly active in hormonal therapy and CDKi resistant ER+HER2- breast cancer cells. These results suggest that HCB is a novel and potent biguanide that has potential to be developed for inhibition of hormone therapy resistant and CDKi resistant breast cancer. Citation Format: Zhijun Guo, Jianxun Lei, Kwon Ho Hong, Beverly Norris, Craig M. Flory, Swaathi Jayaraman, Connor McDermott, Elizabeth Ambrose, Irina Sevrioukova, Tom Poulos, Ilia Denisov, Stephen Sliga, Robert J. Schumacher, Gunda I. Georg, John R. Hawse, Matthew P. Goetz, David A. Potter. Hexyl-(cuban-1-yl-methyl)-biguanide (HCB) inhibits hormone therapy resistant breast cancer cells, in part by Inhibiting CYP3A4 arachidonic acid epoxygenase activity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB078.