Novel serum biomarkers for renal cell carcinoma NIH AACR

Background: Renal cell carcinoma (RCC) is often undetected until it is in an advanced stage. Currently there are no reliable blood biomarkers to monitor the presence of the disease and/or response to therapies. Mass spectrometry (MS)-based proteomics provides tools for sensitive and high-throughput screening of proteinaceous biological samples and greatly facilitates biomarker iscovery.

Materials and methods: Sera from patients diagnosed with RCC were obtained at the baseline (n=14), then from the same individuals at 1 week (n=10), 2 weeks (n=5), and 3 weeks (n=7) after radical nephrectomy. In addition, unrelated RCC (n=4) and healthy control sera (n=30) were collected. Levels of inflammatory cytokines and growth factors (interleukins 1β, IL 2-7, 10, 12, and 13, GM-CSF, interferon-γ, TNF-α) were quantified in multiplex immunoassays. SELDI TOF MS protein profiling of the whole sera and specimens immunoprecipitated with anti-kallikrein antibodies was done in IMAC-Cu arrays. Further purification and identification of differentially expressed proteins was achieved by combination of two-dimensional gel electrophoresis (2DPAGE), in-gel trypsin digestion and LC-ESIMS for peptide fingerprinting. LC-ESIMS/MS was then used for de novo peptide sequence analysis. Mass spectral data were reduced using the Mascot database search engine. Initial matches were refined and confirmed against raw data and searched against decoy databases to reinforce identifications. Data were further refined using Progenesis Stats.

Results: Serum levels of all growth factors and cytokines (except IL-6) increased above the baseline until 3 weeks post-nephrectomy, with significant (p<0.05)>

Conclusions: SAA, an inflammatory acute phase protein, has been viewed as a biomarker of host response in many human cancers. The recently reported presence of SAA in tumor tissue suggests alternative function of the protein. Sudden disappearance of SAA versus persistent upregulation of multiple inflammatory proteins in post-nephrectomy RCC sera supports this notion. The additional matches derived from the 2D gels of the immunoprecipitates are all either reasonably expected or supported by recent literature identifications in other pathologies. The identifications of the other targets are being confirmed by independent methods. Selective proteome isolation by targeted MS is a promising analytical tool with potential clinical applications.