Single-Cell Analysis of Chemotherapy-induced Remodeling Reveals CD276-driven Basal-like Chemoresistance in Pancreatic Cancer

BACKGROUND & AIMS: Unresectable advanced pancreatic ductal adenocarcinoma (PDAC) typically requires systematic chemotherapy, but it remains unclear how this treatment remodels tumor cell plasticity and the tumor microenvironment (TME) to influence clinical outcomes. METHODS: We conducted single-cell RNA sequencing on paired pre- and posttreatment tumor biopsies and peripheral blood mononuclear cells from 28 patients with PDAC receiving abraxane plus gemcitabine chemotherapy. To validate the chemoresistant niche, we employed multiplex immunofluorescence and spatial transcriptomics, ranging from in situ sequencing to 10X Visium HD at 2-μm resolution. In addition, we performed functional validation experiments, including CRISPR-Cas9 knockout and tumor-killing assays in vitro, and in vivo studies using the KPC mouse model and xenograft tumors in nude mice, with a focus on CD276/B7-H3 as the key regulator identified in our study. RESULTS: tumor-associated macrophages, and exhausted T cells, which progressively dominated the TME during treatment in nonresponders. Importantly, we identified CD276/B7-H3 as a dual-function immune checkpoint: it promotes tumor transition to a chemoresistant basal-like state, induces T cell exhaustion, and enhances the angiogenesis signature of tumor-associated macrophages. CONCLUSIONS: Our work uncovers the plasticity of PDAC tumor cell states and interactions with the TME that are modified during chemotherapy of unresectable advanced PDAC and pinpoints CD276/B7-H3 as a critical regulator and a promising therapeutic target for overcoming chemotherapy resistance.