Abstract 3550: Dendritic cells pulsed with cancer stem cell antigen showed profound anti-tumoral efficacy in ovarian cancer

Abstract Background: Ovarian cancer serves as the first leading lethally gynecologic cancer, of which around 70% of patients would relapse soon even have undergone standard treatment. Several mechanisms work together attributing to the early relapse. Among these, cancer stem cells (CSCs) act as the root of metastasis and relapse in the early reports. However, limited methods directly targeting these rare populations have been developed so far. Here we develop DC vaccine directly targeting CSCs by loading DCs with enriched cancer stem cell antigen (CSC-Ag) in vitro. Methods: Freshly dissected ovarian tumors are digested into single cell suspension with digestive enzymes. Subsequently, fluorescence-activated cell sorting (FACS) is used to isolate CD44CD133 double-positive cells, previously reported surface markers of CSCs. Basic medium supplied with several cytokines is used to maintain and enrich CSCs. Suspension of enriched stem cell spheres is repeated five freeze-thaw cycles in liquid nitrogen and 37-degree Celsius water bath, and the supernatant after centrifugation is regarded as CSC-Ag. CD14+ cells of patients’ PBMCs are isolated using CD14 microbeads according to manufacturer’s instruction. IL-4 and GM-CSF are added to induce DC. Immature DC are loaded with previously prepared CSC-Ag. Meanwhile, CD14- cells are cultured with the supplement of anti-CD3, anti-CD28 antibodies, and rhIL-2. T cells are soon co-cultured with mature DC for further antigen-presenting. In vitro cytotoxicity against OC cell lines are tested. Additionally, we developed a peritoneal metastatic ovarian cancer cell model by injecting OVCAR8 transfected with luciferase into NSG mice. IVIS system is used later to monitor tumor growth twice a week. Result: Both CD44 and CD133 were highly expressed in ovarian cancers compared with normal tissue in matched TCGA and GTEx datasets. CD44CD133 double-positive cells ranged from 0.00% to 59.10% in our OC cohort (n=21). CSCs were successfully isolated and enriched, confirmed by the expression of stem cell markers, including SOX2, OCT4, and NANOG. CSC-Ag also expressed those markers comparable to freshly cultured CSC. DC-CSC+T showed remarkable efficacy both in vitro and in vivo against ovarian cancer, with more cancer cell apoptosis. Conclusion: Dendritic cells pulsed with cancer-stem cell antigen (DC-CSC+T) could significantly lower the growth rate of OC, and may serve as a potential adjuvant therapy for improving OC disease-free interval. Citation Format: Haixi Liang, Tong Xiang, Lin Zhou, Zhiwen Xie, Mao Ye, Chunyan Lan. Dendritic cells pulsed with cancer stem cell antigen showed profound anti-tumoral efficacy in ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3550.