Clonorchis sinensis infection amplifies hepatocellular carcinoma stemness, predicting unfavorable prognosis
Authors: Lin Qiumei , Tang Zeli , Qin Yuling , Deng Xueling , Wei Caibiao , Liu Fengfei , Pan Xiaolan , Liu Dengyu , Zhan Tingzheng , Fang Min 
Published: January 2024

Background
Extensive evidence links Clonorchis sinensis (C. sinensis) to cholangiocarcinoma; however, its association with hepatocellular carcinoma (HCC) is less acknowledged, and the underlying mechanism remains unclear. This study was designed to investigate the association between C. sinensis infection and HCC and reveal the relationship between C. sinensis infection and cancer stemness.

Methods
A comprehensive analysis of 839 HCC patients categorized into C. sinensis (-) HCC and C. sinensis (+) HCC groups was conducted. Chi-square and Mann–Whitney U tests were used to assess the association between C. sinensis infection and clinical factors. Kaplan–Meier and Cox regression analyses were used to evaluate survival outcomes. Immunohistochemistry was used to determine CK19 and EpCAM expression in HCC specimens.

Results
Compared to C. sinensis (-) HCC patients, C. sinensis (+) HCC patients exhibited advanced Barcelona Clinic Liver Cancer (BCLC) stage, higher male prevalence and more liver cirrhosis as well as elevated alpha-fetoprotein (AFP), carbohydrate antigen 19–9 (CA19-9), eosinophil, complement 3 (C3), and complement 4 (C4) values. C. sinensis infection correlated with shorter overall survival (OS) (p < 0.05) and recurrence-free survival (RFS) (p < 0.05). Furthermore, Cox multivariate analysis revealed that C. sinensis infection was an independent prognostic factor for OS in HCC patients. Importantly, C. sinensis infection upregulated the expression of HCC cancer stem cell markers CK19 and EpCAM.

Conclusion
HCC patients with C. sinensis infection exhibit a poor prognosis following hepatectomy. Moreover, C. sinensis infection promotes the acquisition of cancer stem cell-like characteristics, consequently accelerating the malignant progression of HCC.

Author summary
Clonorchis sinensis (C. sinensis) is a prominent food-borne parasite prevalent in regions such as China, particularly in Guangxi. C. sinensis has been associated with various hepatobiliary system injuries, encompassing inflammation, periductal fibrosis, cholangiocarcinoma and even hepatocellular carcinoma (HCC). A substantial body of evidence links C. sinensis to cholangiocarcinoma, However, the connection between C. sinensis and HCC and the intricate mechanisms underlying its contribution to HCC development remain incompletely elucidated. Our study demonstrates clear clinicopathological associations between C. sinensis and HCC, such as gender, BCLC stage, liver cirrhosis, MVI, AFP, CA19-9, circulating eosinophils and complements. Furthermore, we found that the co-occurrence of C. sinensis exhibited a significant association with shorter OS and RFS in patients diagnosed with HCC. A major finding was that C. sinensis infection promotes the acquisition of cancer stem cell-like characteristics, consequently accelerating the malignant progression of HCC. Our results provide a more comprehensive comprehension of the interplay between C. sinensis and HCC, shedding fresh light on the carcinogenic potential of C. sinensis.