# Single‐Cell RNA Sequencing and Bulk RNA Sequencing Revealed the Interplay Between Intratumoral Heterogeneity and the Tumor Microenvironment in Breast Cancer

## TL;DR

This study explores how different cell types interact in breast cancer subtypes, revealing key pathways that could influence treatment and prognosis.

## Contribution

The study identifies novel cell communication pathways and their roles in different breast cancer subtypes using single-cell and bulk RNA sequencing.

## Key findings

- ANGPTL pathway is critical in estrogen receptor-positive breast cancer.
- PTN pathway plays a key role in both ER+BC and HER2+BC, while GAS pathway is linked to poor prognosis in TNBC.
- SPP1 and GRN pathways in macrophage-fibroblast interactions influence tumor progression in ER+BC.

## Abstract

The study is to investigate differential signaling pathways within the tumor microenvironment across molecular subtypes of breast cancer (BC).

Single‐cell RNA (scRNA‐seq) sequencing data of BC samples were obtained from the Gene Expression Omnibus database. Cell types were identified using the SingleR package, in conjunction with the analysis of marker genes. Subsequently, Monocle was used for pseudotime analysis of epithelial cells, fibroblasts, and macrophages to characterize their differentiation states. CellChat was employed to study the ligand‐receptor interactions among various cell types across different BC molecular subtypes. In addition, we used common bulk RNA sequencing data from The Cancer Genome Atlas to investigate the correlation between key signaling pathway factors identified by scRNA‐seq and clinical outcomes.

Inference of copy number variation analysis using T cells revealed significantly elevated copy number variation scores in epithelial cells and fibroblasts. In the communication between epithelial cells and fibroblasts, the ANGPTL pathway is critical in estrogen receptor‐positive breast cancer (ER+BC), while the PTN pathway plays a key role in both ER+BC and human epidermal growth factor receptor 2‐positive breast cancer (HER2+BC), and the GAS pathway is associated with poor prognosis in triple‐negative breast cancer (TNBC). In the interaction between fibroblasts and macrophages, the macrophage subpopulation supporting tumor angiogenesis exhibits significant activity in ER+BC, with the associated SPP1 and GRN pathways strongly influencing tumor progression. The SEMA3 pathway mainly acts through dividing tumor‐associated fibroblast clusters across all BC subtypes. When exploring the role of lymphocytes, the PTN pathway also plays a role in HER2+BC, while in TNBC, CXCL and CD70 pathways are significantly involved in immune response modulation.

Our comprehensive analysis of cell–cell communication networks among epithelial cells, fibroblasts, macrophages, and lymphocytes across BC subtypes focuses on ligand‐receptor interactions. This study revealed that certain molecules within these networks exhibit significant prognostic value and therapeutic promise.

The figure shows single‐cell datasets of breast cancer collected from the GEO database, including estrogen receptor‐positive breast cancer, HER2‐positive breast cancer, and triple‐negative breast cancer. These datasets are used to analyze differences among breast cancer subtypes by studying the interactions among epithelial cells, fibroblasts, macrophages, and lymphocytes.