ALTERED METABOLIC PATHWAYS IDENTIFIED IN CLEAR CELL RENAL CELL CARCINOMA BY MASS SPECTROMETRY-BASED FOOTPRINTING

MARÍA ELENA KNOTT; MALENA MANZI; MARIO O. SALAZAR; LYDIA INÉS PURICELLI; MARÍA EUGENIA MONGE

Conferencia; Pittcon 2018; 2018

In-vitro cell models can be used to investigate altered central metabolic networks that contribute to cell proliferation, growth and survival in cancer. Clear cell renal cell carcinoma (ccRCC) is the most common (75%) histological subtype of kidney cancer. In this study, we have optimized a protocol for harvesting, and extraction of extracellular metabolites from human renal cell lines, and we have utilized a discovery-based metabolomics approach to profile the exometabolome by means of ultraperformance liquid chromatography coupled to quadrupole-time-of-flight mass spectrometry (UPLC-QTOF-MS). Conditioned media were obtained from a non-tumor human embryonic kidney cell line, HEK-293, which was used as control; and from a ccRCC cell line 786-O, which derives from primary ccRCC tumor. A total of 22 conditioned media samples were analyzed for each cell type, including 2 biological replicates. Metabolite extracts were lyophilized and reconstituted prior to reversed-phase UPLC-QTOF-MS analysis. Metabolic features (Rt, m/z pairs) were analyzed using a cross-validated orthogonal projection to latent structures-discriminant analysis model using a genetic algorithm variable selection method. A panel of 12 compounds with tentative chemical identification distinguished sample classes with 100% specificity, sensitivity and accuracy. In addition, 5 of these compounds were present in 10 human serum samples from ccRCC patients and healthy controls. Discriminant metabolic features suggest alterations of the glutathione, and phenylalanine metabolism, and alteration of the tryptophan degradation pathway and the pentose phosphate pathway. Our current work involves the validation of the tentative identification with chemical standards.