Abstract 2731: Cell-type deconvolution of African American breast tumors reveals spatial heterogeneity of the immune microenvironment
Authors: Reynier Jean-Baptiste , Yu Jovian , Biernacka Anna , Khramtsova Galina , Yoshimatsu Toshio , Niu Qun , Woodard Anna , Zheng Yonglan , Mueller Jeffrey , Chen Mengjie , Olopade Olufunmilayo I. 
Published: July 2021

Abstract
Background: As immunotherapy emerges as a possible treatment for triple negative breast cancer (TNBC), it is crucial to understand heterogeneity and the complexities of the tumor microenvironment. GeoMx, a new platform developed by NanoString, allows for gene expression analysis within localized segments of the tumor, and is therefore ideal for determining the spatial distribution of immune cell-types. To test the feasibility of applying spatial transcriptomic to breast tumors from diverse populations, we selected ten paraffin embedded blocks from a cohort of self-reported African American patients with TNBC in the Chicago Multi-Ethnic Breast Cancer Study (ChiMEC).
Methods: Tissue sections were immunostained with four antibodies (PanCK, CD3E, CD20 and DAPI) and regions of interest (ROIs) were manually selected based on specific morphology, resulting in 10 to 39 ROIs per slide (with a total of 234 ROIs across the ten slides). Using the GeoMx Cancer Transcriptome Atlas assay, expression levels of 1825 genes were measured within each ROI. For this cell-type deconvolution analysis, we used the Spatial Deconvolution algorithm, developed specifically for GeoMx data. We first normalized read counts across all ROIs of a slide and removed the background expression levels. In order to account for cancer cell types in the deconvolution process, we selected ROIs with high tumor content to predict the gene expression profiles of these tumor cells. We then performed deconvolution using both cancer and immune expression profiles, and clustered ROIs into immune signature groups.
Results: As expected, we saw a higher abundance of immune cells in the stroma compared with the tumor, which resulted in lower immune cell-type diversity within the tumor segments. Tumor segments were CD8+ T-cell and neutrophil enriched: 20.3% of immune cells in tumor segments were neutrophils, and 23.7% were CD8+ T-cells, whereas in the stroma, only 3.6% and 13.8% were neutrophils and CD8+ T-cells respectively. The proportion of naïve versus memory cytotoxic T-cells differed between the tumor and the stroma, with 84.1% of CD8+ T-cells in the stroma identified as memory versus only 23.9% for tumor segments. We were also able to compare two slides from the same patient and found similar immune patterns across the two sections of the tumor. Clustering of immune cell-type distribution revealed three major immune signatures within both sections of this tumor: macrophage-rich, B-cell rich, and CD8+ T-cell rich ROIs.
Conclusion: Spatial transcriptomics allows for granular analysis of the tumor microenvironment. This level of detail is crucial to understanding the cancer-immune relationship, as our analysis revealed high heterogeneity of the immune landscape within tumors. Future work will compare the tumor microenvironment within and across racial groups.
Citation Format: Jean-Baptiste Reynier, Jovian Yu, Anna Biernacka, Galina Khramtsova, Toshio Yoshimatsu, Qun Niu, Anna Woodard, Yonglan Zheng, Jeffrey Mueller, Mengjie Chen, Olufunmilayo I. Olopade. Cell-type deconvolution of African American breast tumors reveals spatial heterogeneity of the immune microenvironment [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 2731.