Ferroptosis-mitochondria-related molecular subtypes and prognostic signature reveal immune and pathway heterogeneity in hepatocellular carcinoma

Abstract

BACKGROUND

Hepatocellular carcinoma (HCC) is highly heterogeneous, and conventional clinicopathological factors incompletely capture its biological diversity. Ferroptosis and mitochondrial dysfunction converge on oxidative stress, metabolism, and regulated cell death, but their integrated prognostic relevance remains unclear. We hypothesized that genes shared by ferroptosis and mitochondrial programs would define biologically distinct HCC subtypes and support prognostic stratification.

AIM

To characterize ferroptosis-mitochondria-related heterogeneity and develop a prognostic signature for HCC.

METHODS

Ferroptosis-related genes from FerrDb were intersected with mitochondrial genes from MitoCarta 3.0 and analyzed in The Cancer Genome Atlas liver HCC cohort. Differential expression, Cox regression, consensus clustering, immune scoring, pathway enrichment, least absolute shrinkage and selection operator Cox modeling, 7:3 internal validation, and external evaluation in GSE14520 and the International Cancer Genome Consortium Liver Cancer-RIKEN Japan (ICGC-LIRI-JP) cohort were performed. A nomogram was constructed and evaluated using calibration and decision curve analyses; exploratory analysis of public proteomic data was also performed.

RESULTS

Forty-seven overlapping genes were identified, significantly exceeding the overlap expected by chance (1.71-fold enrichment; P = 2.41 × 10^-4). Four molecular subtypes showed distinct survival and immune score patterns. A six-gene score comprising NT5DC2, TIMM9, CISD1, LONP1, PRDX6, and LIG3 was constructed. In the internal validation cohort, the model achieved 1-, 3-, and 5-year areas under the curve of 0.767, 0.740, and 0.575, respectively, with a C-index of 0.654. In the external ICGC-LIRI-JP cohort, the corresponding areas under the curve were 0.682, 0.648, and 0.581, respectively. In both GSE14520 and ICGC-LIRI-JP, the continuous risk score remained significantly associated with overall survival, whereas the dichotomized risk groups did not show statistically significant Kaplan-Meier separation. High-risk tumors showed proliferative and metabolic pathway enrichment.

CONCLUSION

Ferroptosis-mitochondria-related genes capture prognostic and immunological heterogeneity in HCC, although the six-gene score requires further prospective and experimental validation.

Keywords: Hepatocellular carcinoma; Ferroptosis; Mitochondria; Prognostic signature; Immune cell scores; Bioinformatics

Core Tip: This study integrated ferroptosis- and mitochondria-related genes to characterize hepatocellular carcinoma heterogeneity and develop a six-gene prognostic score. Four molecular subtypes showed distinct survival and immune score patterns. Internal validation supported short- and intermediate-term discrimination, whereas performance weakened at 5 years, and Kaplan-Meier separation after risk-group dichotomization was not statistically significant in the external cohorts. The principal contribution is a compact, biologically interpretable framework linking ferroptosis, mitochondrial dysfunction, immune remodeling, and metabolic pathway alterations in hepatocellular carcinoma.