Construction Of M~5C-related Prognostic Risk Model And Exploration Of Tumor Microenvironment Characteristics In Hepatocellular Carcinoma

Objective:5-Methylcytosine(m~5C)plays a crucial role in hepatocellular carcinoma(HCC),but the association between m~5C regulation and the tumor microenvironment(TME)characteristics in HCC has not been fully clarified.Here,we aim to construct a m~5C-related prognostic risk model to predict prognosis of HCC patients and investigate its relevance with the TME heterogeneity and potential compounds susceptibility,in order to achieve precision medicine for HCC.Methods:(1)We collected 371 HCC patients from The Cancer Genome Atlas(TCGA)database as the training cohort,and the landscapes of 18 m~5C regulators was investigated.(2)Through an unsupervised clustering algorithm,the training cohort was divided into different m~5C clusters,survival analysis was performed between clusters,and possible differences in pathways and biological processes between clusters were explored.(3)A prognostic risk model was established based on the selected differential genes related to prognosis by the Lasso algorithm,and the training cohort was divided into high-and low-risk groups by calculating the risk score.And we validated the prognostic risk model on an external dataset.(4)We then analyzed immune infiltration,immune checkpoint expression,tumor microenvironment-related pathway enrichment,and genomic alterations between high-and low-risk groups,and explored the relationship between risk scores and potential compounds susceptibility.(5)Finally,we collected HCC surgical specimens and their prognostic information to construct a retrospective validation cohort,validated the prognostic risk model by real-time quantitative PCR,and the differential expression of specific immune checkpoints in high-and low-risk patients were validated by immunohistochemical staining.Results:(1)In the TCGA-LIHC cohort,18 m~5C regulated genes except TET2 were significantly differentially expressed in tumors and normal tissues,and most of the m~5C regulated genes were significantly associated with the prognosis of HCC patients.(2)The training cohort was divided into two m~5C clusters,and there were significant differences in overall survival and progression-free survival between the two m~5C clusters.(3)A six-gene prognostic risk model was established by the Lasso algorithm.The overall survival and progression-free survival of HCC patients with high-risk scores were significantly shorter than those of patients with low-risk score.(4)Risk scores are closely related to immune infiltration,immune checkpoint expression,immune escape mechanisms,multiple tumor microenvironment-related pathways,and genomic alterations.Drug sensitivity analysis identified four potential drugs targeting high risk scores,including axitinib,vinblastine,docetaxel and YM-155.(5)Results consistent with the training cohort were obtained in the retrospective validation cohort,HCC patients with high-risk scores have shorter progression-free survival and significantly higher expression levels of PD-L1 and CTLA-4 than those with low-risk scores patients.Conclusions:The m~5C-related prognostic risk model constructed in this study can effectively predict the prognosis of patients with hepatocellular carcinoma.Combined with its predictive value for tumor microenvironmental characteristics and drug sensitivity,the model can be used as a clinical treatment reference to guide precision medicine for patients with hepatocellular carcinoma.