Abstract 3529: Immunotherapy targeting folate receptor induces autophagy in ovarian cancer

Abstract

Abstract Cancer cells are highly dependent on folate metabolism mediated by the folate receptor (FRa). Gene encoding FRa (FOLR1) is overexpressed in endometrial and ovarian cancers. We aim to investigate the functional mechanisms of blocking FRa with MORAb-003 antibody, a humanized monoclonal antibody directly against FRa. We first analyzed clinical data from the Cancer Genome Atlas (TCGA) for correlation between copy numbers of FOLR1 gene and disease-free survival in patients with high-grade serous ovarian carcinoma (HGSC). Next, we determined the surface−expression of FRa on HGSC cells using flow cytometry (FACS). We then used xenograft mouse models to test tumor inhibition by MORAb-003 antibody-induced FRa blockade. We performed reverse-phase protein array (RPPA) analysis in FRa-positive and FRa-negative HGSC tumors to compare their responses to MORAb-003 treatment. To recapitulate the in vivo environment, we cultured FRa-positive and -negative epithelial ovarian cancer cells in three-dimensional (3D) condition and targeted the specific factors identified by RPPA using shRNAs knockdown. MORAb-003 monotherapy significantly reduced tumor weights and numbers of tumor nodules in both FRa-positive IGROV1 and SKOV3 models; combination of MORAb-003 and docetaxel significantly reduced both tumor weights and numbers of tumor nodules compared to docetaxel monotherapy. As expected, we did not observe these effects in the A2780 model, which has low FRa expression. Immunohistochemistry using PCNA and TUNEL staining in resulting tumors revealed that MORAb-003 reduced proliferation, but had no significant effect on apoptosis. Systematic analysis from RPPA identified a number of autophagy factors and cell-death initiators as being significantly upregulated by MORAb-003 treatment in FRa-positive tumors including Beclin-1 and PEA-15 (a factor involved in activation of autophagic cell death). cDNA microarrays further showed that MORAb-003 treatment up-regulated the expression of BECN1, ATG8, ATG3 and LC3B in FRa-positive tumors. Using FACS analysis with acridine orange (AO) staining, we observed significantly more AO positive population in 3D cultured SKOV3 cells treated with MORAb-003 alone (16.9%) or MORAb-003+docetaxel (21.2%) than in cells treated with control IgG (5.24%). Such differences were not observed in A2780 cells. We also observed significant increase of autophagy flux in SKOV3 cells but not in A2780 cells. We also found that blocking autophagy with hydroxychloroquine or bafilomycin A1, or knockdown Beclin-1 or PEA-15 with shRNAs reversed the growth inhibition induced by MORAB-003 in 3D cultured SKOV3 cells. We reported autophagy-associated cell death is a previously unrecognized mechanism for targeting of FRa in addition to the known mechanisms including antibody-dependent cellular cytotoxicity. Our studies also suggested that Beclin-1 and PEA-15 could be further explored as prognostic markers for FRa-targeted therapy. Citation Format: Yun-Fei Wen, Anil Sood. Immunotherapy targeting folate receptor induces autophagy in ovarian cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3529.