Abstract 3817: Suicidal autophagy induced by immunotherapy targeting folate receptor in ovarian cancer
Authors: Wen Yun-Fei , Sood Anil K. 
Published: August 2015

Abstract
Objective Cancer cells depend on folate metabolism for DNA replication, making folate receptor alpha (FRα) an important therapeutic target. Blocking FRα with a humanized anti-FRα monoclonal antibody, MORAB-003 offers a potent clinical opportunity; however, the mechanisms underlying its efficacy were not well understood. We investigated the functional mechanisms underlying MORAB-003 induced tumor inhibition in ovarian cancer. Methods Using FACS analysis and real-time PCR, FRα expression was determined in an array of human ovarian cancer cells. Next, we examined the effects of MORAB-003 monotherapy and combination with docetaxel in tumor growth using the xenograft mouse models. In vitro, we analyzed ovarian cancer cells with 3-dimensional (3-D) spheroid-like architecture. We next performed systems-based analysis including reverse-phase protein and genomic microarrays, and discovered an array of factors/genes involved in autophagy was upregulated by MORAB-003. Functional assays including FACS with AO staining, tandem mRFP/mGFP-LC3 fluorescence microscopy, transmission electron microscopy (TEM) and siRNAs knockdown were performed to further investigate MORAB-003-induced autophagy in tumor inhibition. Results IGROV1 and SKOV3ip1 cells express high-level FRα and A2780 cells express low-level FRα. In vivo, MORAB-003 monotherapy or a combination of MORAB-003 and docetaxel led to significantly decreased tumor growth in IGROV1 and SKOV3ip1 models, but not in A2780 model. Immunohistochemistry analysis on tumor sections indicated that MORAB-003 significantly reduced proliferative cancer cells, but had minimum effects on tumor cell apoptosis or angiogenesis. Network analysis on protein and gene expression profiles in IGROV1 and SKOV3ip1 tumors revealed that MORAB-003 significantly upregulated a number of factors/genes involved in autophagy, including LC3II, ATG3, ATG4B, S116-PEA-15 and BECN1. Furthermore, S116-PEA-15 and Beclin-1 were required for caspase-3-activated cell death in 3-D IGROV1 and SKOV3ip1 spheroids treated with MORAB-003. Results from autophagic flux assays indicated that MORAB-003 treatment led to an abundant accumulation of late-stage autophagic vacuoles. Blocking the autophagic vacuolization or fusion using bafilomycin A1 or hydroxychloroquine reversed the MORAB-003 induced reduction of cell viability in 3-D IGROV1 and SKOV3ip1 spheroids. Furthermore, queries from the Cancer Genome Atlas data (TCGA) revealed that copy-number gains of the FOLR1 gene significantly correlated with shorter disease-free survival in patients with the immunoreactive subtype of ovarian serous carcinoma and uterine carcinoma. Conclusion MORAB-003 inhibits tumor growth via a non-canonical autophagy associated with programmed cell death. This finding has substantial implications for clinical development of anti-FRα therapies, especially with regard to novel combinations and predictive biomarkers.
Citation Format: Yun-Fei Wen, Anil K. Sood. Suicidal autophagy induced by immunotherapy targeting folate receptor in ovarian cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3817. doi:10.1158/1538-7445.AM2015-3817