Exploratory Profiling of Carnitine and Acylcarnitine Metabolism in HCT116 Colorectal Cancer and Normal Colon-Derived Cells

Insights into Mitochondrial Acyl-Group Homeostasis and Metabolic Adaptation in HCT116 Colorectal Cancer Cells

Abstract

ABSTRACT

Background: Metabolic reprogramming is a hallmark of colorectal cancer and involves alterations in mitochondrial energy metabolism, fatty acid oxidation, amino acid catabolism, and acyl-group homeostasis. This study compared the carnitine and acylcarnitine profiles of HCT116 colorectal cancer cells and CCD normal colon cells to identify metabolic alterations associated with malignant transformation.

Materials and Methods: Twenty-seven carnitine and acylcarnitine metabolites were quantified using targeted LC–MS/MS. Metabolomic data were analysed using MetaboAnalyst 6.0 following median normalization and Pareto scaling. Principal component analysis (PCA), partial least squares–discriminant analysis (PLS-DA), fold-change analysis, false discovery rate (FDR)-adjusted statistical testing, hierarchical clustering, and correlation network analyses were performed.

Results: Although several metabolites demonstrated nominal differences between groups, none remained statistically significant after false discovery rate (FDR) correction. Free carnitine (C0) was increased in HCT116 cells at the nominal significance level but did not retain significance after multiple-testing correction. Fold-change analysis suggested increased C0, C3, and C6 levels and decreased C8:1 and C18:2 levels in HCT116 cells. PCA explained 74.0% of total variance but did not clearly separate the groups, whereas PLS-DA demonstrated partial discrimination.  Variable importance in projection analysis identified C10, C6, and C3 as the primary contributors to group separation. Correlation network analysis revealed a modular acylcarnitine interaction pattern, with C4DC, C4, and C2 emerging as hub metabolites. A

Conclusions: Although no individual metabolite remained statistically significant after FDR correction, the findings suggest exploratory alterations in free carnitine and short-/medium-chain acylcarnitine metabolism in colorectal cancer cells. These results should be considered hypothesis-generating and require validation in larger experimental models and functional metabolic studies.

Keywords: Colorectal cancer; HCT116 cells; CCD cells; carnitine; acylcarnitine; metabolomic