Abstract C055: Withaferin A offers novel targeted immunotherapy against high-grade serous ovarian carcinoma

Abstract

Abstract Background: Tumor-induced immunosuppression and cell proliferation are the hallmarks of progression of malignant tumors including high-grade serous ovarian carcinoma (HGSOC), a fatal malignancy of women. Natural killer (NK) cell plays critical roles in killing tumor cells in the tumor vicinity as it does not require antigen presentation. Our earlier studies showed the frequency of tumor infiltrating NK cells is significantly low as compared with that of tumor stroma suggesting that tumor-induced factors may be involved in inhibition of NK cell infiltration, and the tumor continue to metastasize. Cytokines including IL-10/IL-15 are used to stimulate NK cells and tumor continuously secretes IL-10 or IL-15 and it is possible that excessive exposure to these cytokines induces cytokine inducible SH2-containing protein (CISH), a marker of NK cell exhaustion. We have shown earlier that root powder of Ashwagandha (ASH), an anti-stress natural herb improved NK cell population and reduced tumor progression. The goal of this study was to determine whether Withaferin A (WFA), an active ingredient of ASH, targets the induction of CISH or cyclin D1 (CCND1), a stimulator cell growth and prevented tumor progression. Materials & methods: Study-1: An exploratory study was conducted with clinical specimens which included normal ovaries (n=5) and ovarian high grade serous carcinoma (HGSC) at both early and late stages (n=5 each) and expression levels of CISH and CCND1 were examined. Study-2: A prospective study using the laying hen model of OVCA supplemented with or without 2% dietary ASH root powder for 90 days (n=7 each;normal, early and late-stage). Normal and tumor ovarian tissues were collected at the end of the study and used for analysis. Both the clinical and hen specimens were used for immunohistochemistry, western blotting and gene expression assays to determine expression levels of CISH and CCND1. In vitro studies were also conducted with WFA on the OVCAR3, an HGSC cell line, where multiple doses were tested, and expression of markers previously mentioned were examined. Data were collected and analyzed by ANOVA and unpaired t-test and considered significant when P<0.05. Results: Frequency of CISH-expressing cells in both clinical and hen specimens in normal, early and late-stages were localized. Compared to normal, increased expression of both CISH and CCND1 was seen in early-stages and increased further through late stages. Similarly, strong expression of both CISH and CCND1 was seen in proteomic and genomic studies. ASH supplementation is hens compared to untreated showed a significant decrease (P<0.001) in CISH expression as well as in CCND1 gene expression studies. In vitro studies WFA treatment significantly decreased expression of both markers (P<0.05). Conclusion: The results of this study suggest that both CISH and CCND1 are involved in progression of OVCA and dietary supplementation with ASH or treatment with WFA may be an effective bi-targeted immunotherapy against HGSOC. Support: Ovarian Cancer Spore, Roswell Park Citation Format: Jasmin C. Acosta, Animesh Barua. Withaferin A offers novel targeted immunotherapy against high-grade serous ovarian carcinoma [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr C055.