Use of quantitative shotgun proteomics to identify fibronectin 1 as a potential plasma biomarker for clear cell carcinoma of the kidney

Background: Early detection would be one of the most effective means to improve the outcome of renal cell carcinoma (RCC). We searched for a new plasma marker for RCC using a label-free quantitative shotgun proteomics method. Methods: Plasma proteins were digested by trypsin, and the resulting peptides were analyzed by 2-Dimensional Image Converted Analysis of Liquid chromatography mass spectrometry (2DICAL). An identified biomarker candidate was subjected to validation using the Amplified Luminescent Proximity Homogeneous Assay (AlphaLISA). Results: Among a total of 23,407 independent MS peaks, we found that the mean intensity of 59~peaks significantly differed between 20 clear cell RCC patients and 20~healthy controls. MS/MS spectra from 16 of the 59~peaks matched the amino acid sequences of the fibronectin 1 (FN1) gene product. {The increased plasma level of FN1 in RCC patients was validated in a cohort of in 77 patients and 130 healthy controls (p<0.0001).} Conclusions: The FN1 is considered to be a promising biomarker candidate for clear cell RCC. Furthermore, AlphaLISA is an alternate to the conventional enzyme-linked immunosorbent assay and should prove useful for the rapid validation of biomarker candidates.