Determination of Circulating CPN Cleavage Peptides Enables Early Diagnosis of Breast Cancer

The concentration of circulating peptides produced by CPN (carboxypeptidase N) cleavage reflects the CPN activity in tumors, and analysis of these biomarkers demonstrates potential for the noninvasive and early diagnosis of breast cancer.

CPN is important in regulating vasoactive peptide hormones, growth factors, and cytokines by specifically cleaving their C-terminal basic residues. Investigators at Weill Cornell Medical College (New York, NY, USA) investigated whether circulating peptides specifically cleaved by CPN in the tumor microenvironment could be stage-specific indicators of breast cancer.

The investigators identified peptide fragments produced by CPN using an ex vivo peptide cleavage assay. They incubated a synthesized C3f peptide (His6-C3f_S1304-R1320-His6) in interstitial fluids taken from breast tumors and adjacent normal breast tissues in mice with orthotopic implantation of the human breast cancer cell line MDA-MB-231. The nature and extent of peptide cleavage by CPN was investigated by fragment profiling using nanopore fractionation and mass spectrometry.

Results revealed that generation of the C3f_R1310-L1319 cleavage peptide specifically correlated with the CPN expression level. In both the mouse and clinical patient samples, CPN was clearly increased in tumor tissues compared with normal breast tissue, whereas corresponding CPN abundance in blood remained constant. Concentrations of six CPN-catalyzed peptides predominantly increased in sera taken from the mice at two weeks after orthotopic implantation. Six homologous peptides displayed significantly higher expression in the patients' plasma as early as the first pathologic stage of breast cancer.

“Our results represent a first demonstration, to our knowledge, that clearly links the proteolytic activity of CPN, particularly at tumor sites, to the cleavage patterns of its catalytic substrates in the blood,” said senior author Dr. Ye Hu, assistant professor of cell and developmental biology at Weill Cornell Medical College. “These biomarkers show strong potential for the noninvasive and early diagnosis of breast cancer. We advocate their use … certainly to be detected and identified before metastasis, and perhaps even before the tumor presents with any observable characteristics commonly used in the clinic.”

The study was published in the January 2014 issue of the journal Clinical Chemistry.

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