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  3. Integrating single cell- and spatial- resolved transcriptomics unravels the inter-tumor heterogeneity and immunosuppressive landscape in HBV- and Clonorchis sinensis-associated hepatocellular carcinoma

Integrating single cell- and spatial- resolved transcriptomics unravels the inter-tumor heterogeneity and immunosuppressive landscape in HBV- and Clonorchis sinensis-associated hepatocellular carcinoma

  • Mol Cancer. 2026 Jan 5;25(1):3. doi: 10.1186/s12943-025-02381-z.
Jiayun Chen # 1 2 3,  Wenmin Lu # 1,  Yanni Lou # 4,  Jing Liu # 3,  Xiwen Liao # 5,  Yunmeng Bai 2,  Guangqing Cheng 2,  Guangzhi Zhu 5,  Ji Feng 1 6,  Junqi Liu 5,  Zhaoji Liu 7,  Liqun Jia 8,  Jing Zhou 9,  Tao Peng 10,  Guo-Dong Lu 11 12,  Jigang Wang 13 14 15
Affiliations

Affiliations

  • 1 Department of Toxicology, School of Public Health, Guangxi Medical University, Nanning, Guangxi, 530021, China.
  • 2 State Key Laboratory for Quality Ensurance and Sustainable Use of Dao-di Herbs, Artemisinin Research Center, Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing, 100700, China.
  • 3 Center for Drug Research and Development, Guangdong Provincial Key Laboratory for Research and Evaluation of Pharmaceutical Preparations, Guangdong Pharmaceutical University, Guangzhou, Guangdong, 510006, China.
  • 4 Department of Integrative Oncology, China-Japan Friendship Hospital, No. 2 East Yinghua Road, Chaoyang District, Beijing, 100029, China.
  • 5 Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, 6 Shuangyong Road, Nanning, Guangxi, 530021, China.
  • 6 School of Public Health, Fudan University, 130 Dong-An Road, Shanghai, 200032, China.
  • 7 Department of Physiology, School of Preclinical Medicine, Guangxi Medical University, 22 Shuangyong Road, Nanning, Guangxi, 530021, China.
  • 8 Department of Integrative Oncology, China-Japan Friendship Hospital, No. 2 East Yinghua Road, Chaoyang District, Beijing, 100029, China. [email protected].
  • 9 Department of Physiology, School of Preclinical Medicine, Guangxi Medical University, 22 Shuangyong Road, Nanning, Guangxi, 530021, China. [email protected].
  • 10 Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, 6 Shuangyong Road, Nanning, Guangxi, 530021, China. [email protected].
  • 11 Department of Toxicology, School of Public Health, Guangxi Medical University, Nanning, Guangxi, 530021, China. [email protected].
  • 12 School of Public Health, Fudan University, 130 Dong-An Road, Shanghai, 200032, China. [email protected].
  • 13 Department of Toxicology, School of Public Health, Guangxi Medical University, Nanning, Guangxi, 530021, China. [email protected].
  • 14 State Key Laboratory for Quality Ensurance and Sustainable Use of Dao-di Herbs, Artemisinin Research Center, Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing, 100700, China. [email protected].
  • 15 Center for Drug Research and Development, Guangdong Provincial Key Laboratory for Research and Evaluation of Pharmaceutical Preparations, Guangdong Pharmaceutical University, Guangzhou, Guangdong, 510006, China. [email protected].
  • # Contributed equally.
PMID: 41491521 DOI: 10.1186/s12943-025-02381-z
Abstract

Background: Hepatocellular carcinoma (HCC) is the most common primary liver carcinoma with high lethality. Both of hepatitis B virus (HBV) and Clonorchis sinensis (C. sinensis) are critical infectious contributors to HCC development. However, the inter-tumor heterogeneity and tumor microenvironment (TME) of HCC patients with different infectious background remain largely unknown.

Methods: We compiled a cohort of 269 primary HCC patients to assess the clinical impact of C. sinensis and HBV infections on patient prognosis. Single-cell RNA Sequencing (scRNA-seq) and spatial transcriptomic (ST-seq) analyses were performed on tumor and adjacent normal samples from C. sinensis-associated HCC (CP), and double-infection HCC (DP) patients. Additionally, we integrated publicly available scRNA-seq and ST-seq datasets from HBV-associated (HP) patients. Immunofluorescence, immunohistochemistry and in vitro experiments were conducted to validate inter-tumor heterogeneity among the three HCC subtypes.

Results: C. sinensis Infection is significantly associated with poorer prognosis in HCC patients. Multi-omics analyses revealed distinct inter-tumor heterogeneity in epithelial, immune, and stromal compartments across different HCC subtypes. Tumor cells in the DP group exhibited more malignant marker expression, higher copy number variation scores, increased activation of p53 pathway, and worse survival outcomes. Compared with Other HCC subtypes, the TME in DP samples was enriched with SPP1+ macrophages, exhausted CD8+ T cells and COL1A1+ fibroblasts. In contrast, the CP and HP groups showed higher proportions of M2-like macrophages and ENPP2+ liver vascular endothelial cells, respectively.

Conclusion: These findings decipher the cellular signatures and their interactions within the TME, shedding light on the inter-tumoral heterogeneity driven by different infections, and the development of targeted therapies for infectious HCC.

Keywords

Clonorchis sinensis; Hepatitis B virus; Hepatocellular carcinoma; Inter-tumoral heterogeneity; Single cell RNA sequencing; Spatial transcriptomics.

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