Integrative Analysis of Sialylation and Amino Acid Metabolism Signatures Reveals Prognostic Drivers and Immunotherapy Responses in Colorectal Cancer

Background CRC, or colorectal cancer, exhibits substantial biological diversity, with varied metabolic programs and inconsistent benefit from immunotherapy. Pathways involving sialylation together with metabolism of amino acids shape tumor development and regulation of anti-tumor immunity. However, the prognostic significance of this combined metabolic signature for CRC remains clinically undefined in practice. Methods We assembled genes related to sialylation plus those linked to metabolism of amino acids from public databases to construct a comprehensive candidate list. Multi-omics profiling integrated bulk RNA-seq, single-cell transcriptome profiling, and spatially resolved transcriptomic data to examine their functions in CRC. Differential expression testing, survival modeling, and consensus-based clustering were used to define metabolic–epigenetic tumor categories and compare clinical outcomes. From these markers, we derived a multigene risk score centered on six regulators connected with sialylation and amino-acid metabolic pathways. Its prognostic performance was rigorously evaluated within several external colorectal carcinoma cohorts using multiple survival endpoints. We further examined the key regulator PLOD3 in patient specimens and cultured cells using qRT-PCR, immunoblotting, flow cytometry, colony formation, migration, and invasion assays. Results We identified six prognosis-related genes (AOC3, CDO1, PHGDH, MAT2B, ASRGL1, and PLOD3) that function as principal metabolic determinants in CRC. For unsupervised clustering, non-negative matrix factorization separated tumors into six molecular subgroups, among which Cluster 3 showed the most unfavorable survival. Single-cell transcriptomics and spatially resolved transcriptomic analyses demonstrated marked metabolic and immune diversity across the tumor microenvironment. These datasets revealed distinct architectures of immune-cell infiltration and region-specific metabolic activity within tumor tissue. The six-gene risk model displayed high prognostic accuracy, outperforming conventional clinicopathologic variables and previously proposed CRC signatures. Patients assigned to the high-risk group exhibited increased tumor mutational burden, pronounced activation of immune pathways, and perturbed immune responses, indicating exploitable therapeutic opportunities. In experimental validation, PLOD3 expression was markedly elevated in peripheral blood and neoplastic tissue from individuals with CRC. Stable depletion of PLOD3 in colorectal cancer cell lines reduced cell proliferation, migratory capacity, invasive potential, and epithelial–to–mesenchymal plasticity. It simultaneously enhanced apoptotic cell death in colorectal cancer cells under in vitro experimental conditions. Conclusions Our investigation emphasizes the central contribution of sialylation and amino-acid–related metabolic programs to colorectal cancer progression and immune modulation. The integrative prognostic framework, together with mechanistic dissection of PLOD3, supports refined risk stratification and informs individualized therapeutic planning for patients with CRC. Trial registration Not applicable. This study was not a clinical trial because no participants were prospectively assigned to any healthrelated intervention.