Abstract 5483: Macrospatial ecotype heterogeneity in early-stage breast cancer identified by spatial nucleus barcoding

Abstract Ductal carcinoma in situ (DCIS), the predominant form of early-stage breast cancer and a precursor to invasive breast cancer, remains inadequately understood in terms of spatial microenvironment reprogramming. In this study, we developed a Spatial Nucleus Barcoding (SNuBar) approach for single-nucleus RNA sequencing (SNuBar-RNA) that utilizes a single oligonucleotide adaptor to preserve spatial information. The accuracy and scalability of SNuBar was validated using cell line mixture experiments. We then applied SNuBar-RNA to investigate the macrospatial ecotype heterogeneity in early-stage breast cancer. By analyzing single cells from different macrodissected spatial regions in normal breast tissues, matched normal and DCIS tissues, we identified 13 major cell types and 43 distinct cell states, co-localizing in diverse combinations across four primary topographic areas (adipose-enriched, normal epithelial, activated stroma and tumor). Spatial analysis revealed the reprogramming of vascular and myoepithelial cells in localized zones near premalignant cells, while fibroblasts, T-cells, and myeloid cells exhibited reprogramming across broader fields. Validation using the MERSCOPE assay confirmed stromal heterogeneity between tumor and normal areas. Collectively, our data shows that many diverse cell types are reprogrammed in spatially-defined areas in the early-stage breast cancer microenvironment, relative to the normal breast tissue regions. Citation Format: Kaile Wang, Zhenna Xiao, Yiyun Lin, Runmin Wei, Shanshan Bai, Jie Ye, Rui Ye, Min Hu, Aatish Thennavan, Emi Sei, Alastair Thompson, Savitri Krishnamurthy, Nicholas Navin. Macrospatial ecotype heterogeneity in early-stage breast cancer identified by spatial nucleus barcoding [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5483.