Transcriptomic and proteo-metabolic determinants of the grading system in clear cell renal cell carcinoma

Background: Pathological grade is a morphological parameter of clear cell–renal cell carcinoma (ccRCC) and an independent predictor of cancer-specific survival. The aim of this study was to identify grade-dependent metabolic signatures and corresponding gene and protein expression changes that connect variations in cancer metabolism with nuclear grade, especially in high-grade tumors. Methods: Forty ccRCC samples were collected and stratified according to nuclear grade: 23 low-grade (LG = G1–G2) and 17 high-grade (HG = G3–G4) samples. In addition, 122 patients with sarcomatoid ccRCC (sRCC) were classified according to the abundance of sarcomatoid features as low sarcomatoid (LS; sarcomatoid component<20%; n = 67) or high sarcomatoid (HS; sarcomatoid component≥20%; n = 55). Untargeted metabolomic analysis was performed. To study the relative changes in gene and protein expression in HG vs. LG ccRCC, data from 4 different datasets were downloaded and stratified according to nuclear grade. Immunohistochemistry and immunofluorescence were used to evaluate protein expression. Cancer-specific survival (CSS) and progression-free survival (PFS) were calculated using Kaplan–Meier analysis. Multivariate analysis was performed using a Cox regression model. Results: The Warburg effect, in association with changes in Krebs cycle intermediates and related metabolites, was more prominent in HG ccRCC than in LG ccRCC. Additional alterations included metabolic reprogramming in the urea cycle and modulation of glutathione metabolism with the accumulation of reduced glutathione and carnitine derivatives in HG tumors, while the concentrations of long- and medium-chain fatty acids were greater in LG ccRCC. CSS and PFS were significantly decreased in patients with HS tumors. According to the multivariate analysis, the abundance of the sarcomatoid component was an adverse prognostic factor. Conclusions: ccRCC is characterized by a particular grade-dependent metabolic reprogramming. Metabolic pathways and associated molecular alterations are grade-specific and could represent potential therapeutic targets, especially in HG tumors. sRCC subclassification based on the abundance of sarcomatoid components into HS vs. LS tumors have prognostic value.