Abstract MIP-071: TARGETING A CHEMORESISTANT OVARIAN CANCER CELL POPULATION VIA THE CARBOHYDRATE ANTIGEN SIALYL TN
Authors: Rueda B.R. , Starbuck K. , Eavarone D. , Prendergast J. , Stein J. , Foster R. , Behrens J. 
Published: June 2017

Key words: Sialyl Tn, drug resistance, antibody-drug conjugates, cancer stem cells

OBJECTIVES: A successful therapeutic strategy for ovarian cancer will require direct targeting of inherently chemoresistant tumor cells which are comprised in part of cancer stem cells (CSCs) that survive current cytotoxic treatment regimes and drive tumor resurgence. The sialyl-Tn (STn) antigen is a carbohydrate moiety present on tumor cells but rarely seen in normal adult tissue. Importantly, STn has been shown to be present on CSCs in pancreatic, colon, and gastric malignancies. Our objective was to assess the expression of STn and the known CSC marker CD133 in human ovarian cancer (OvCa) cell lines and primary serous carcinomas, and evaluate the ability of STn+ and STn- cells to both grow in an anchorage independent manner and survive standard-of-care cytotoxic therapy. Furthermore, we sought to assess the effect of murine and humanized α -STn antibody-drug conjugates (ADCs) on OvCa cells in vitro and tumor viability in vivo.

METHODS: STn and CD133 expression in established OvCa cell lines was analyzed by flow cytometry. STn-CD133-, STn+CD133-, STn-CD133+ and STn+CD133+ cells were purified from OVCAR3 and OVCAR4 by FACS, plated in soft agar, and incubated for 21 days. Colony forming efficiency of each sub-population was calculated. Unsorted cells were treated in vitro with either murine α -STn-monomethyl auristatin E (MMAE) ADC or vehicle control and cell viability was assessed by MTT assay. Subsequently, cells were treated in vitro with α -STn-MMAE, paclitaxel and carboplatin, or appropriate controls, and the profile of cells surviving 72 hours post-treatment was determined by flow cytometric analysis. Finally, OVCAR3-derived mouse xenografts were treated with murine and humanized α-STn-MMAE, unconjugated mAbs alone, and vehicle control. Mice were assessed regularly for tumor growth and cytotoxic effects.

RESULTS: In the OvCa cell lines OV90, OVCAR3 and OVCAR4, when grown in traditional 2D culture, STn+ cells comprised 98.4%, 40.0%, and 26.4% of the total cell population, respectively. In each of these cell lines, we readily detected STn+CD133+ sub-populations suggesting that STn is expressed on CD133+ ovarian CSCs. Colony formation assays analyzing FACS-purified STn-CD133-, STn+CD133-, STn-CD133+ and STn+ CD133+ sub-populations suggest that STn expression correlates with anchorage independent growth, a characteristic of cell stemness. Paclitaxel and carboplatin treatment in vitro significantly increased the proportion of STn+ and CD133+ cells, demonstrating the chemoresistant characteristics of these cells. Treatment with the murine α-STn-MMAE ADCs reduced the viability of OvCa cell lines in vitro in a dose-dependent manner. Treatment with murine and humanized α -STn- MMAE antibodies in vivo reduced tumor volumes, whereas vehicle treatment did not impede tumor growth. Interestingly, the unconjugated antibody also had a modest negative impact on tumor volume.

CONCLUSION: A novel, highly specific STn antibody identifies the STn antigen in OvCa cell lines and patient samples. STn+ and CD133+ cells demonstrate stem-like characteristics such as anchorage-independent growth and chemoresistance. STn ADCs decreased cell viability in vitro and reduced tumor volumes in vivo, suggesting that specific therapeutic targeting of STn in ovarian tumors may be an effective clinical strategy to eliminate quiescent CSCs.

Citation Format: B.R. Rueda, K. Starbuck, D. Eavarone, J. Prendergast, J. Stein, R. Foster, J. Behrens. TARGETING A CHEMORESISTANT OVARIAN CANCER CELL POPULATION VIA THE CARBOHYDRATE ANTIGEN SIALYL TN [abstract]. In: Proceedings of the 11th Biennial Ovarian Cancer Research Symposium; Sep 12-13, 2016; Seattle, WA. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(11 Suppl):Abstract nr MIP-071.