Abstract 131: Remodeling of stromal-epithelial interactions in breast cancer progression as inferred from regional and single-cell gene expression analysis
Authors: Officer Adam , Dempsey Andre , Hasteh Farnaz , Slyper Michal , Segerstolpe Asa , Klughammer Joanna , Jane-Valbuena Judit , Rozenblatt-Rosen Orit , Regev Aviv , Yau Christina , Harismendy Olivier 
Published: July 2021

Abstract
Background: The transition from ductal carcinoma in situ (DCIS) to invasive ductal carcinoma (IDC) is defined by the invasion of malignant cells into surrounding tissues, but associated changes in the cellular composition and interactions in the tumor microenvironment are poorly understood. Systematic analysis of gene expression differences between DCIS and IDC are confounded by cellular heterogeneity and few studies have separated stromal and epithelial contributions or distinguished pure DCIS from those synchronous with IDC.
Methods: The gene expression of 192 microdissected regions from 140 cases was assembled from 5 studies and included DCIS (83 epithelial, 22 stromal) and IDC (67 epithelial, 20 stromal) cases. Differences in infiltration of 10 immune cell types were measured using expression signatures. Cell-cell interactions between stromal and epithelial regions were measured using the co-expression of 1175 curated pairs of ligand-receptor genes and compared between DCIS and IDC using a permutation test. The corresponding interacting cell types were identified using RNA sequencing of 5444 single nuclei extracted from fine needle aspirates of two excised DCIS specimens.
Results: Stromal regions of Her2+ cases had the highest level of immune infiltration compared to IDC and Her2- cases. Of the measured immune cell types, B cells showed a progressive depletion from normal to DCIS to IDC. The expression-based measurement of stromal-epithelial interactions identified 99 and 115 ligand-receptor interactions stronger in DCIS and IDC, respectively (FDR&lt;0.1). To precisely identify the corresponding interacting cell types, the expression profile of cells within the DCIS microenvironment was determined using single-nucleus RNA sequencing from two DCIS patients. The analysis identified 9 cell types, including luminal, basal, macrophage, adipocyte and endothelial, and 54% of the candidate cell-cell interactions could be mapped to at least one cell type pair. Based on these cell type mappings, interactions between luminal cells and fibroblasts were gained in IDC while those involving the vascular endothelium were lost, including interactions between CD24 and SELP, an interaction involved in leukocyte recruitment. Increased macrophage autocrine interactions were identified in both IDC stroma and epithelium through PLAU-PLAUR co-expression, an interaction previously associated with a transition to the M2 phenotype.
Conclusion: The analysis of published gene expression studies with novel computational methods, augmented with a single-cell landscape representative of the disease, can help compensate for the difficulty in studying pure DCIS specimens. The microenvironment changes in cellular dynamics involved both immune and non-immune cell types suggesting mechanisms other than direct immune escape contribute to progression.
Citation Format: Adam Officer, Andre Dempsey, Farnaz Hasteh, Michal Slyper, Asa Segerstolpe, Joanna Klughammer, Judit Jane-Valbuena, Orit Rozenblatt-Rosen, Aviv Regev, Christina Yau, Olivier Harismendy. Remodeling of stromal-epithelial interactions in breast cancer progression as inferred from regional and single-cell gene expression analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 131.