Supplementary Material for: Prognostic relevance of MDK and TIMP1 with immune infiltration in lung adenocarcinoma

Background: LUAD is a common and fatal form of lung cancer, with varying expressions of MDK and TIMP1 across different cancers. However, their specific roles in LUAD progression and tumor immunity remain unclear. Methods: We analyzed RNA-seq data from TCGA using the ggpubr R package to compare MDK and TIMP1 expression in normal versus LUAD tissues. This was validated through qRT-PCR and western blot. LUAD patients were categorized into high and low expression groups for MDK and TIMP1, and their impacts on overall survival (OS), disease-free interval (DFI), progression-free interval (PFI), and disease-specific survival (DSS) were assessed using Kaplan-Meier curves and ROC curves. KEGG and GO enrichment analyses were performed for 50 genes similar to MDK and TIMP1, and a gene-gene interaction network was created with GeneMANIA, examining DEGs across both expression groups. We also evaluated the mutational landscape and immune cell infiltration correlations. Finally, the relationship between MDK, TIMP1 and immune cells was explored by immunohistochemical (IHC) experiments. Results: MDK and TIMP1 were significantly overexpressed in LUAD. Patients with low MDK and TIMP1 expressions had better OS, DFI, DSS, and PFI. The AUC values for MDK and TIMP1 were 0.943 and 0.875, respectively, with TIMP1 identified as a risk factor for OS. Genes similar to MDK were enriched in the Proteasome pathway, while those akin to TIMP1 were linked to endopeptidase activity. No survival impact was noted from mutations in these genes. Higher expression of MDK and TIMP1 correlated with reduced tumor purity and altered immune scores, suggesting increased immune dysfunction in the high TIMP1 group. IHC results showed that when MDK and TIMP1 expression levels were higher, B cell and TREG cell infiltration was stronger, but macrophage infiltration was weaker. Conclusion: MDK and TIMP1 significantly influence the prognosis and progression of LUAD and immune cell infiltration, highlighting their potential as targets for immunotherapy.