High-resolution spatial transcriptomics identifies that insufficient radiofrequency ablation induces hepatocellular carcinoma progression via the CEBPD/CXCL2 axis

BACKGROUND AND AIMS: Radiofrequency ablation (RFA) is becoming a standard treatment for early-stage hepatocellular carcinoma (HCC). However, high rates of postoperative recurrence, particularly following insufficient RFA (iRFA), remains a clinical obstacle. iRFA contributes to the formation of an immunosuppressive tumor microenvironment and facilitates tumor progression, though the molecular mechanisms driving these processes are not fully elucidated. This study seeks to comprehensively characterize the spatial architecture and cellular interactions within iRFA HCC, and to identify key transcriptional mechanisms through which sublethal heat stress promotes immune evasion and recurrence. APPROACH AND RESULTS: We integrated high-definition spatial transcriptomics (Visium HD) with single-cell RNA sequencing data to map the cellular heterogeneity and communication networks in samples from 6 HCC patients who underwent surgical resection after iRFA. Functional validation was performed using in vitro heat stress models, co-cultured assays, and orthotopic mouse models. We provided a high-resolution spatial atlas of iRFA HCC and the ablation zone, identifying a subpopulation of tumor cells exhibiting activation of the transcription factor CEBPD in response to sublethal heat stress. CEBPD directly bound to the promoter of CXCL2 and drove its expression. Upregulation of CXCL2 diminished immune checkpoint inhibitors' response in iRFA HCC through promoting SPP1+ TAM accumulation via the CXCL2/STAT3/SPP1 axis, and enhanced tumor cell invasion through autocrine signaling. CONCLUSIONS: Our study provides a comprehensive spatial characterization of HCC following iRFA, identifies the heat stress-CEBPD-CXCL2 axis as a key driver of tumor progression post-RFA, and demonstrates the therapeutic potential of CXCR2 inhibition for preventing tumor recurrence following RFA.