Abstract 2048: Mutations in FOXA1 alter chromatin remodeling and cell fate in prostate organoids

Abstract Genomic analysis of targeted patient tumor sequencing identified frequent mutations, 41% in prostate cancer (Li, et al., 2020) in the gene FOXA1, a developmentally important pioneer transcription factor (TF) in mammary and prostate tissues. Previous work by our group and others has shown that these FOXA1 mutations alter global chromatin accessibility and promote growth in prostate cells (Adams, 2019), but the underlying molecular details, including the identity of partner TFs, remain unclear. To address this topic, we generated mouse prostate organoids expressing Foxa1 alleles harboring three distinct classes of mutations: (i) overexpression of WT Foxa1 (reflecting focal gene amplification seen in tumors), (ii) a series of mutants within the Wing2region of the forkhead binding domain (FHBD) and (iii) a mutant bearing a stop codon after the FHBD to represent a series of C-terminal truncation mutants. We performed single nucleus multiome sequencing to obtain gene expression (snRNA-seq) and chromatin accessibility (snATAC-seq) readouts from the same individual nuclei. Whereas each Foxa1 mutant has distinct, often mutant-specific features, several themes emerge. These include alterations in the relative proportion of stem-like (L2) luminal cells vs secretory (L1) luminal cells as well as changes in luminal or basal gene signatures, increased androgen receptor signaling output, and enrichment for motifs of distinct classes of partner TFs. For example, cells expressing the truncation mutant show gain in the accessibility of Gata3 and Pou2f1 TF binding motifs, as well as enhanced numbers of L1-like luminal cells. Functional studies demonstrate that Pou2f1 is specifically required for the pro-luminal phenotype in cells expressing the truncation mutant whereas Gata3 plays a more general pro-luminal role. Correlations in motif accessibility and transcription factor expression across single cells further revealed a composite androgen receptor (AR)-FOXA1 motif enriched in the pro-luminal truncation mutant, while the canonical AR motif was enriched in pro-basal cell mutants. Finally, Foxa1 mutants cooperative with Trp53 and Pten loss in orthotopic prostate tumorigenicity assays, most strikingly manifest by reversion of the basal-like features characteristic of Trp53/Pten loss tumors to Ck8+ luminal adenocarcinoma histology, mirroring that seen in Foxa1-mutant human prostate cancer tumors in mice. Thus, mutant Foxa1 alleles cooperate with canonical prostate cancer tumor suppressors and alter the histologic phenotype of prostate cancers in mice through the activation of basal or luminal lineage differentiation programs. Citation Format: Erik Ladewig, Abbas Nazir, Christina Leslie, Charles Sawyers. Mutations in FOXA1 alter chromatin remodeling and cell fate in prostate organoids [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2048.