β-Catenin-regulated ALDH1A1 is a target in ovarian cancer spheroids

Cancer cells form three-dimensional (3D) multicellular aggregates (or spheroids) under non-adherent culture conditions. In ovarian cancer (OC), spheroids serve as a vehicle for cancer cell dissemination in the peritoneal cavity, protecting cells from environmental stress-induced anoikis. To identify new targetable molecules in OC spheroids, we investigated gene expression profiles and networks upregulated in 3D vs traditional monolayer culture conditions. We identified ALDH1A1, a cancer stem cell marker as being overexpressed in OC spheroids and directly connected to key elements of the beta-catenin pathway. beta-Catenin function and ALDH1A1 expression were increased in OC spheroids vs monolayers and in successive spheroid generations, suggesting that 3D aggregates are enriched in cells with stem cell characteristics. beta-Catenin knockdown decreased ALDH1A1 expression levels and beta-catenin co-immunoprecipitated with the ALDH1A1 promoter, suggesting that ALDH1A1 is a direct beta-catenin target. Both short interfering RNA-mediated beta-catenin knockdown and A37 ((ethyl-2-((4-oxo-3-(3-(pryrrolidin-1-yl) propyl)-3,4-dihydrobenzo [4,5]thioeno [3,2-d] pyrimidin-2-yl) thio) acetate)), a novel ALDH1A1 small-molecule enzymatic inhibitor described here for the first time, disrupted OC spheroid formation and cell viability (P<0.001). beta-Catenin knockdown blocked tumor growth and peritoneal metastasis in an OC xenograft model. These data strongly support the role of beta-catenin-regulated ALDH1A1 in the maintenance of OC spheroids and propose new ALDH1A1 inhibitors targeting this cell population.