Background Cancer-associated fibroblasts (CAFs) are highly heterogeneous in the progression of pancreatic ductal adenocarcinoma (PDAC) and vasculogenic mimicry (VM) refers to a phenomenon in which cancer cells adopt endothelial-like characteristics.

Objective To identify a novel protumoural CAF subtype undertaking VM.

Design We used single-cell RNA sequencing and mIHC to identify FAPα⁺CD144⁺ endothelial-like CAFs (endoCAFs) and combined prospective and retrospective analyses to assess its clinical outcomes. Tube formation, proliferation and invasion assay were conducted on cell lines, organoids, the orthotopic tumour model and LSL-Kras^G12D/+, LSL-Trp53^R172H/+ and Pdx1-Cre (KPC) mouse model. Mechanically, we performed cytokine array assays, RNA-sequencing, IP-mass spectrometry, ChIP and luciferase analyses. Importantly, an siRNA delivery nanosystem was developed to precisely target FAPα⁺CD144⁺endoCAFs in vivo.

Results FAPα⁺CD144⁺endoCAFs were present in the tumour microenvironment of PDAC, and patients with a higher CD144⁺CAFs proportion displayed poor prognosis of PDAC. FAPα⁺CD144⁺endoCAFs not only acquired a VM phenotype to provide metastatic conduits but also promoted the proliferation and invasion of tumour cells in situ through paracrine signalling, thereby actively facilitating the metastasis of tumour cells. The CD144-β-catenin-STAT3 signalling axis was activated, and CD144 and downstream secreted cytokines were transcriptionally upregulated to maintain the dual roles of endoCAFs. A CAF-targeting siRNA delivery nanosystem, via loading FAPα and siCD144, was administered to precisely target FAPα⁺CD144⁺ endoCAFs, which substantially inhibited their protumoural roles in vivo.

Conclusion FAPα⁺CD144⁺endoCAFs can promote metastasis of PDAC via undertaking VM and paracrine through activation of the CD144-β-catenin-STAT3 signalling axis. CAF-targeting siRNA delivery nanosystem can inhibit tumour progression by precisely targeting FAPα⁺CD144⁺endoCAFs.