Frontiers in immunology
Patient-aware single-cell analysis of fibrovascular remodeling in a predominantly colorectal cancer immunotherapy cohort
By Gu Y, Li L, Zhang Y and 3 others.
Abstract
Background
Cancer-associated fibroblasts (CAFs) and vascular mural cells may shape extracellular matrix organization in colorectal cancer. However, single-cell comparisons are susceptible to unequal cell recovery, ambient RNA, doublets, treatment-stage mixing, and repeated sampling within patients. We investigated fibrovascular remodeling using patients as the inferential unit and distinguished reproducible signals from donor-concentrated and context-dependent findings.
Methods
We reanalyzed a published single-cell RNA sequencing cohort of 22 patients, including 20 with colorectal cancer and 2 with duodenal carcinoma. The primary pericyte analysis included 14 pretreatment matched normal-tumor pairs with at least three pericytes per tissue. We assessed ambient contamination and doublets using DecontX and scDblFinder. CAFs were reclustered at six response-blind resolutions, and the stable parent population was tested by patient-blocked pseudobulk analysis after excluding its dominant donor. Independent CRC single-cell and spatial datasets, a cross-cancer immunotherapy cohort, and TCGA were used for external testing.