A Prognostic Lysine Crotonylation Signature Shapes the Immune Microenvironment in Hepatocellular Carcinoma.

Introduction: Hepatocellular carcinoma (HCC) has a poor prognosis due to late diagnosis and rapid progression, highlighting the need for a deeper understanding of its pathogenesis. Lysine crotonylation (Kcr), a unique post-translational modification, plays a crucial role in epigenetic regulation. However, the role of crotonylation-related genes (CRGs) in HCC remains poorly understood, necessitating an investigation of their prognostic and therapeutic relevance.

Methods: Transcriptomic and clinical data were obtained from TCGA and GEO databases. A CRG-based risk score was developed using Cox and LASSO regression analyses. To enhance survival prediction, a nomogram incorporating the risk score was constructed. Immune cell infiltration and drug sensitivity were assessed using CIBERSORT and 'OncoPredict.' Single-cell sequencing was employed to examine CRG expression within the HCC tumor microenvironment.

Results: An 8-gene risk score model (HDAC2, ACADS, HDAC1, ENO1, PPARG, ACADL, ACSL6, and AGPAT5) was established, effectively stratifying patients into high- and low-risk groups in the training set. Cox regression and Kaplan-Meier analyses validated its prognostic value in the test set. The nomogram demonstrated enhanced prognostic accuracy for survival prediction. Differences in immune cell infiltration and immune checkpoint expression between risk groups highlighted the association between CRGs and the tumor immune microenvironment. Single-cell sequencing revealed that CRGs were highly expressed in key immune cells within the HCC microenvironment. Additionally, drug sensitivity analysis suggested that specific targeted therapies may be more effective in HCC patients.

Discussion: Crotonylation-related gene signature demonstrates strong prognostic value in hepatocellular carcinoma (HCC), effectively stratifying patients into high- and low-risk groups and recapitulating known oncogenic roles of HDAC1/2, ENO1, PPARG, AGPAT5 and the protective functions of ACADS, ACADL, and ACSL6. It was found that crotonylation not only influences tumor cell metabolism and epigenetic regulation but also shapes the immune microenvironment, highlighted by distinct checkpoint expression, differential immune cell infiltration, and drug sensitivity profiles, which positions our model as a promising tool for personalized therapeutic decision-making. However, clinical translation will require standardized, reproducible assays for crotonylation measurement and rigorous validation across diverse HCC etiologies (e.g., viral vs. non-viral), along with mechanistic and longitudinal studies to dissect causality versus correlation, assess off-- target effects of crotonylation modulators, and confirm functional impacts on immune modulation before routine diagnostic or therapeutic use.

Conclusion: This study identifies a prognostic CRG signature for HCC and provides novel insights into personalized treatment strategies.