A Comprehensive Analysis of the ITIH Family Across Multiple Cancer Types and an Initial Investigation of ITIH1 in Gastric Cancer

Introduction: The ITIH family, crucial for extracellular matrix stability and cancer progression, is underexplored in multi-omic profiles and immune microenvironments; this study analyzes their roles across cancers and ITIH1’s function in gastric cancer to reveal diagnostic, prognostic, and therapeutic potential. Methods: We analyzed RNA-seq, protein expr ession, and clinical data from 33 cancer types and 24 non-cancerous conditions using TCGA, GTEx, GEO, CPTAC, and IMvigor210 datasets. Methods included differential expression analysis, ROC curve assessment for diagnostic potential, Cox regression and Kaplan-Meier survival analyses for prognostic value, GSEA for pathway enrichment, and molecular docking for ITIH1-targeted small molecule screening. Immune microenvironment interactions, tumor mutational burden (TMB), microsatellite instability (MSI), and immunotherapy response were evaluated. in vitro experiments validated ITIH1’s role in gastric cancer using qRT-PCR, Western blotting, siRNA knockdown, and functional assays. Results: ITIH family genes exhibited differential expression across cancers and non-- cancerous conditions, with ITIH1, ITIH4, and ITIH5 showing high diagnostic potential (AUC > 0.90 in multiple cancers). ITIH1 was a risk factor for poor survival in gastric cancer (p < 0.05). Lower ITIH scores correlated with improved survival in patients receiving immune checkpoint inhibitors (p < 0.05). ITIH genes showed strong correlations with immune checkpoints (PD-1, CTLA-4), TMB, and MSI. Molecular docking identified six small molecules, including Entinostat, with high binding affinity for ITIH1 (-8.4 kcal/mol). ITIH1 knockdown in gastric cancer cell lines (HGC-27, AGS) significantly reduced proliferation, migration, and invasion (p < 0.01). Discussion: This study underscores the ITIH family's critical role as diagnostic and prognostic biomarkers across various cancers and non-cancerous conditions, with ITIH1's therapeutic potential in gastric cancer highlighted through its impact on tumor progression, though limitations include discrepancies in some ITIH gene expressions between in vitro and in vivo settings, necessitating further validation. Conclusion: Our findings highlight the ITIH family's potential as diagnostic biomarkers, prognostic indicators, and therapeutic targets, particularly in gastric cancer. The identification of ITIH1 inhibitors and their association with immune checkpoints, TMB, and MSI paves the way for improved diagnostics, targeted therapies, and immunotherapy predictions, enhancing patient outcomes across diseases.