Abstract 4948: The role of zafirlukast and isoquercetin in thiol isomerase inhibition in relation to cancer and its complications

Abstract

Abstract High levels of thiol isomerases in cancer patients have been correlated with increased tumor growth and an increased risk of cancer-induced thrombosis. Inhibition of major extracellular thiol isomerases, which include the enzymes PDI, ERp57, ERp72 and ERp5 in both cancer and thrombosis, have strong therapeutic potential to treat cancer while also inhibiting cancer-induced thrombosis. In this study, we explored the effects of two thiol isomerase inhibitors, isoquercetin (IsoQ) and zafirlukast (Zafi) against OVCAR8 ovarian cancer cells and a xenograft mouse model created from those cells. To explore the potential of these drugs to inhibit tumor growth and cancer-induced thrombosis, multiple in vitro and in vivo experiments were performed. Alterations of cellular OVCAR8 thiol isomerase activity were measured using a DI-E-GSSG assay where, after drug treatment separately with both drugs, DTT and the PDI specific fluorescent probe were added to OVCAR8 cells and then monitored kinetically at 520nm/550nm. Further, the ability of the drugs to inhibit the coagulative potential of OVCAR8 cells was measured by monitoring Factor Xa generation as a marker for tissue factor release. Cells were seeded at a concentration of 100,000 cells, then monitored for Factor Xa generation after addition of a fluorescent probe that is a Factor Xa substrate, Factor VIIa, calcium and Factor X. To explore the effects of IsoQ and Zafi in vivo, an OVCAR8 xenograft mouse model was created. Tumors were allowed to grow to an average of 30mm2 and mice were then treated with 10 mg/kg IsoQ, 30 mg/kg IsoQ or 30 mg/kg Zafi for 46 days via oral gavage. Blood was collected on Day 46, plasma was isolated and examined for alterations in p-selectin expression via ELISA and alterations in thiol isomerase activity by the DI-E-GSSG assay. Cell surface thiol isomerase activity on OVCAR8 cells was inhibited by 32% after with Zafi and by 35% after treatment with IsoQ. OVCAR8 cell induced Factor Xa generation was significantly reduced by 79% after Zafi treatment and by 35% after IsoQ treatment. In the xenografts, 10mg/kg of IsoQ, 30 mg/kg of IsoQ, and 30 mg/kg of Zafi were all separately able to suppress tumor growth by approximately 85% compared to controls. P-selectin expression decreased by approximately 50% and thiol isomerase activity was decreased by approximately 40% under all 3 treatment conditions. These results demonstrate the potential of thiol isomerase inhibitors to not only treat cancer-induced thrombosis but also the cancer itself, as cancer induced thrombosis markers and tumor size were both significantly reduced in these experiments. Citation Format: Justine A. Gelzinis, Daniel R. Kennedy. The role of zafirlukast and isoquercetin in thiol isomerase inhibition in relation to cancer and its complications. [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 4948.