The code used in the analyses.

<b>Background:</b> Focal segmental glomerulosclerosis (FSGS) is a clinicopathological syndrome manifesting glucocorticoid-resistant nephrosis, which requires new therapies targeting specific pathways. The Columbia classification stratifies FSGS into 5 variants: Collapsing, Tip, Cellular, Perihilar and NOS. Differences in their renal prognosis imply distinct underlying molecular mechanisms, which remain unclear.<b>Objective:</b> To investigate factors associated with the pathogenesis of FSGS and explore potential therapeutic targets.<b>Methods:</b> A total of 73 kidney biopsy tissues were analyzed by LC-MS/MS: FSGS (n=23), renal donors (n=14) as a healthy control group, and minimal change disease (n=15) and IgA nephropathy (n=21) as disease control groups. We validated the proteome data with immunohistochemistry and the Nephroseq microarray data.<b>Results:</b> Our proteomics platform identified a total of 4,168 proteins. A comparison between the FSGS with renal donor groups revealed 175 proteins increased in the FSGS group, which associated with the complement activation and immune responses. Together with comparisons against the disease control groups, complement factor D (CFD) emerged as a distinctive factor. Immunohistochemistry and the Nephroseq microarray analyses uncovered CFD production elevated primarily in glomeruli. Furthermore, the protein profiles demonstrated molecular differences underlying the Columbia variants. In the Cellular and Tip variants, factors involved in the complement pathway were enriched, while proteins associated with the ribosome, thermogenesis, and ferroptosis were predominant in the Collapsing variant.<b>Conclusion:</b> The elevated CFD and complement activation in glomeruli potentially contribute to the pathogenesis of FSGS. The molecular heterogeneity among the Columbia variants may enable to stratify treatment of FSGS as a new precision medicine approach.