Integrated scRNA-Seq and Spatial Transcriptomics Reveal CHPF and GPC1 as Markers of Cancer-Associated Fibroblast-Dominant Domains in Colorectal Cancer

Spatial heterogeneity in the colorectal cancer (CRC) tumor microenvironment (TME) is closely related to stromal organization and may influence treatment-related features. In this study, we integrated single-cell RNA sequencing and spatial transcriptomic data from CRC tissues to examine cancer-associated fibroblast (CAF)-dominant spatial domains. GPC1 and CHPF were enriched in stromal regions and showed higher expression in CRC tissues than in normal tissues. In the analyzed cohorts, higher GPC1 and CHPF expression was associated with advanced clinicopathological features and poorer survival outcomes in univariate analyses. Single-cell analyses showed that GPC1 and CHPF were mainly expressed in fibroblasts and were enriched in an ADAM12-associated fibroblast subset with myofibroblast-like features, extracellular matrix-related genes, and hypoxia/glycolysis-related signatures. Cell-cell communication analysis suggested that this fibroblast subset was linked to higher inferred WNT signaling. Drug-response and immunotherapy-related analyses further showed that GPC1 and CHPF expression was associated with transcriptomic patterns related to treatment response. In mucinous adenocarcinoma (MA), the ADAM12-associated fibroblast program showed stronger inferred MIF-related signaling and was associated with macrophage M2-like features. Spatial mapping and multiplex immunofluorescence supported partial colocalization of GPC1, CHPF, and ADAM12 in CAF-rich stromal regions. In CAFs, GPC1 or CHPF knockdown reduced ADAM12, CAF activation markers, and WNT/MIF-related proteins, while ADAM12 knockdown weakened macrophage M2-like marker expression in a co-culture model. These findings suggest that GPC1 and CHPF mark an ADAM12-associated CAF program linked to stromal remodeling and immune-related features in CRC, especially in MA, but further studies are needed to define its functional role.