ARTICLE ABSTRACTMetabolomics is increasingly being used in cancer biology for biomarker discovery and identification of potential novel therapeutic targets. However, a systematic metabolomics study of multiple biofluids to determine their interrelationships and to describe their use as tumor proxies is lacking. Using a mouse xenograft model of kidney cancer, characterized by subcapsular implantation of Caki-1 clear cell human kidney cancer cells, we examined tissue, serum, and urine all obtained simultaneously at baseline (urine) and at, or close to, animal sacrifice (urine, tissue, and plasma). Uniform metabolomics analysis of all three “matrices” was accomplished using gas chromatography– and liquid chromatography–mass spectrometry. Of all the metabolites identified (267 in tissue, 246 in serum, and 267 in urine), 89 were detected in all 3 matrices, and the majority was altered in the same direction. Heat maps of individual metabolites showed that alterations in serum were more closely related to tissue than was urine. Two metabolites, cinnamoylglycine and nicotinamide, were concordantly and significantly (when corrected for multiple testing) altered in tissue and serum, and cysteine–glutathione disulfide showed the highest change (232.4-fold in tissue) of any metabolite. On the basis of these and other considerations, three pathways were chosen for biologic validation of the metabolomic data, resulting in potential therapeutic target identification. These data show that serum metabolomics analysis is a more accurate proxy for tissue changes than urine and that tryptophan degradation (yielding anti-inflammatory metabolites) is highly represented in renal cell carcinoma, and support the concept that PPAR-α antagonism may be a potential therapeutic approach for this disease. Cancer Res; 72(14); 3471–9. ©2012 AACR.