Integrating single-cell and bulk RNA sequencing reveals CK19 + cancer stem cells and their specific SPP1 + tumor-associated macrophage niche in HBV-related hepatocellular carcinoma

• Published in: Hepatology international 2024-02, Vol.18 (1), p.73-90
• Main Authors: Yang, Cheng-Lei, Song, Rui, Hu, Jun-Wen, Huang, Jun-Tao, Li, Nan-Nan, Ni, Hang-Hang, Li, Yuan-Kuan, Zhang, Jie, Lu, Zhan, Zhou, Min, Wang, Jun-Duo, Li, Min-Jun, Zhan, Guo-Hua, Peng, Tao, Yu, Hong-Ping, Qi, Lu-Nan, Wang, Qiu-Yan, Xiang, Bang-De
• Format: Article
• Language: English
• Subjects: Angiogenesis; Cancer; Colorectal Surgery; Cytokeratin; Design of experiments; Experimental design; Gelatinase B; Gene sequencing; Hepatitis B; Hepatocellular carcinoma; Hepatology; Heterogeneity; Immunotherapy; Liver cancer; Macrophages; Malignancy; Matrix metalloproteinase; Matrix metalloproteinases; Medicine; Medicine & Public Health; Metalloproteinase; Metastases; Metastasis; Microenvironments; Original Article; Patients; Prognosis; Ribonucleic acid; RNA; Stem cells; Surgery; Transcriptomics; Tumors
• ISSN: 1936-0533; 1936-0541
• DOI: 10.1007/s12072-023-10615-9

Purpose Cytokeratin 19-positive cancer stem cells (CK19 + CSCs) and their tumor-associated macrophages (TAMs) have not been fully explored yet in the hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC). Experimental design Single-cell RNA sequencing was performed on the viable cells obtained from 11 treatment-naïve HBV-associated HCC patients, including 8 CK19 + patients, to elucidate their transcriptomic landscape, CK19 + CSC heterogeneity, and immune microenvironment. Two in-house primary HCC cohorts (96 cases-related HBV and 89 cases with recurrence), TCGA external cohort, and in vitro and in vivo experiments were used to validate the results. Results A total of 64,581 single cells derived from the human HCC and adjacent normal tissues were sequenced, and 11 cell types were identified. The result showed that CK19 + CSCs were phenotypically and transcriptionally heterogeneous, co-expressed multiple hepatics CSC markers, and were positively correlated with worse prognosis. Moreover, the SPP1  + TAMs (TAM_SPP1) with strong M2-like features and worse prognosis were specifically enriched in the CK19 + HCC and promoted tumor invasion and metastasis by activating angiogenesis. Importantly, matrix metalloproteinase 9 (MMP9) derived from TAM_SPP1, as the hub gene of CK19 + HCC, was activated by the VEGFA signal. Conclusions This study revealed the heterogeneity and stemness characteristics of CK19 + CSCs and specific immunosuppressive TAM_SPP1 in CK19 + HCC. The VEGFA signal can activate TAM_SPP1-derived MMP9 to promote the invasion and metastasis of CK19 + HCC tumors. This might provide novel insights into the clinical treatment of HCC patients. Graphical abstract Lay summary: Cytokeratin 19-positive hepatocellular carcinoma is a highly aggressive malignancy with poor therapeutic progress. Understanding the characteristics of CK19 + CSCs and their immune microenvironment better might facilitate the development of effective targeted and immunotherapy. This study identified the heterogeneity and characteristics of CK19 + CSCs and explored their unique immunosuppressive SPP1+TAM niche. The results suggested that the VEGFA signal activates TAM_SPP1-derived MMP9 to promote the invasion and metastasis of CK19 + HCC tumors. The study might have an important therapeutic value.