Abstract 2746: 3D genome structural alterations in biological pathway genes drive the progression of cells from normal epithelium to metastatic cancer

Abstract Large scale structural variation in the genome is known to be linked to cancer pathways. Breast cancer progression from normal epithelium to localized, and metastatic cancer involves genomic perturbations such as mutations, deletions, and duplications. Emerging research has shown that the structure of the 3D genome, specifically how chromosomes and genes are folded and organized in the nucleus, directly impacts gene expression and thus, may contribute to disease progression. In fact, local folding around critical genes, especially those involved in important biological pathways, may have direct impact on cancer initiation and progression. Limits in technology has restricted a more complete understanding linking 3D genome to disease. Biochemical methods require many cells and are unable to preserve single cell and sub-population level information. Previous imaging methods are limited by the number of analyzable targets, reducing the generalizability of results across other genomic loci and limited a comprehensive genome-wide understanding. Here, we use jebFISHTM and the PaintScapeTM platform to resolve 3D structural features of genomic loci at high fidelity. With our Biological Pathways Panel, we focus on genes that are known to play critical roles in DNA repair, cell cycle, apoptosis, chromatin modifications, and transcriptional regulation. Using breast cancer progression as the model, we apply jebFISHTM to cell lines from normal epithelium, localized cancer, and metastatic cancer. We observe dramatic changes in 3D genome at both single cell and population levels. At each stage of breast cancer progression, we identify specific alterations to 3D genome features including changes in structure and switching of A/B chromatin compartments. Structural alterations are also present at whole chromosome level. We see disruption and repositioning of chromosome territories as well as interchromosomal interactions. jebFISHTM single-cell, single-homolog resolution enables identification of cellular sub-populations at each stage of breast cancer progression. We observe changes in the frequency of sub-populations with disease progression. Our results suggest that genome organization, at the local gene as well as chromosome level is modulating the expression of genes important in biological pathways. The presence of sub-populations suggests that the heterogeneous nature of 3D genome organization within the population is an important feature, impacting our understanding and treatment of diseases. While this study focuses on breast cancer, we envisage that these results are generalizable and further the understanding of genome organization and gene expression. Citation Format: Huy Nguyen, Shyamtanu Chattoraj, Jude Dunne. 3D genome structural alterations in biological pathway genes drive the progression of cells from normal epithelium to metastatic cancer [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 2746.