Abstract 5565: Dysfunction in ATP-binding cassette 1 (ABCA1): a driver of intracellular cholesterol accumulation and potential therapeutic vulnerability of bowel cancers

Background: Cholesterol is an integral sterol lipid that influences the homeostatic maintenance of cellular structure, hormone synthesis, and metabolic regulation within the cell. Accumulation of intracellular cholesterol has shown to drive the progression of multiple human cancers. Here we report that mutation or transcriptional repression of the cholesterol exporter, ABCA1, is a common phenomenon conserved in bowel cancers with novel mechanistic and therapeutic implications. Methods: Whole-exome sequence analysis was conducted on 24 appendiceal cancers (AC) from consented patients. ABCA1 coding mutations and expression levels were analyzed in tumors from The Cancer Genome Atlas (TCGA) and Broad DepMap databases. One AC cell line (Naxorini) was created in house and four commercial CRC cell lines were used in ABCA1 functional studies. Combination treatment of 22-(R)-hydroxycholesterol (OHC) and 9-cis-retinoic acid (CRA) was used to induce ABCA1. ABCA1 knockdown (KD), measures of cellular viability, cell death, apoptosis, and free cholesterol were quantified. Results: Protein coding mutations in the ABCA1 gene were identified in 7/24 (29%) AC specimens. Mutational analysis of other gastrointestinal cancers shows that ABCA1 is preferentially mutated in mucinous CRCs and mucinous stomach adenocarcinomas compared to their non-mucinous counterparts (X2 p-value <0.001 and 0.047). ABCA1 mRNA expression levels were significantly lower in CRC and gastric tumors as compared to 28 different tumor types (ANOVA p<0.001), and significantly lower in bowel cancer cell lines as compared to 34 different cancer lineages (ANOVA p<0.001). 10uM OHC+CRA treatment robustly induced ABCA1 protein expression in ABCA1 wildtype (WT) cell lines (Naxorini, MDST8, COLO201), but not in ABCA1 mutant lines (HCT15, SNU81), significantly induced cell death in ABCA1 WT cell lines (all, p<0.001), and decreased free cholesterol accumulation (MDST8, p=0.010; Naxorini, p=0.007). Importantly, KD of ABCA1 rescued the viability of OHC+CRA treated cells (MDST8, p<0.001) demonstrating that OHC+CRA cytotoxicity is dependent on the induction of ABCA1. Furthermore, the KD of ABCA1 in untreated MDST8 cells significantly enhanced cell viability (p=0.004) consistent with a growth advantage owing to reduced activity of ABCA1. Analysis of ABCA1-dependent, OHC+CRA induced cell death found alterations in protein levels of Bad, Bcl-2 and HtrA2/Omi (all, p<0.05), associating ABCA1 induction with mitochondrial apoptosis. Conclusions: Dysregulation of cellular cholesterol due to dysfunctional ABCA1, whether by somatic mutation or transcriptional downregulation, may represent an oncogenic addiction conserved in bowel cancers. These findings warrant further investigation into the therapeutic potential of ABCA1 and cholesterol targeting strategies for bowel cancers. Citation Format: Daniel J. Gironda, Calvin J. Wagner, Ashok K. Pullikuth, Lance D. Miller. Dysfunction in ATP-binding cassette 1 (ABCA1): a driver of intracellular cholesterol accumulation and potential therapeutic vulnerability of bowel cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5565.