Serum peptidome for cancer detection: spinning biologic trash into diagnostic gold

Abstract

The low molecular weight region of the serum peptidome contains protein fragments derived from 2 sources: (a) high-abundance endogenous circulating proteins and (b) cell and tissue proteins. While some researchers have dismissed the serum peptidome as biological trash, recent work using mass spectrometry-based (MS-based) profiling has indicated that the peptidome may reflect biological events and contain diagnostic biomarkers. In this issue of the JCI, Villanueva et al. report on MS-based peptide profiling of serum samples from patients with advanced prostate, bladder, or breast cancer as well as from healthy controls. Surprisingly, the peptides identified as cancer-type-specific markers proved to be products of enzymatic breakdown generated after patient blood collection. The impact of these results on cancer biomarker discovery efforts is significant because it is widely believed that proteolysis occurring ex vivo should be suppressed because it destroys endogenous biomarkers. Villanueva et al. now suggest that this suppression may in fact be preventing biomarker generation.

Figure 1

Proteinases generate biomarker fragments. Circulating protein fragments generated in the diseased tissue microenviroment may serve as diagnostic protein markers. Proteolytic cascades within the tissue (a product of the interacting cellular ecology such as stromal-epithelial interactions), immune cell MHC presentation, or apoptosis generate protein fragments that passively diffuse into the circulation. Shed LMW peptides are protected from kidney-mediated clearance by sequestration on abundant resident blood proteins such as albumin. According to the results presented by Villanueva et al. (18) in this issue of the JCI, diagnostic protein fragments can also be generated ex vivo by circulating enzymes derived from the diseased tissue microenvironment acting on exogenously derived peptides produced by serum collection methodology (see Figure 10 in ref. 18).

Figure 2

Immuno-MS provides a means for rapidly determining the specific size and identity of each member of a panel of peptide marker fragments present within patient sera. As part of a high-throughput assay performed in clinical diagnostic laboratories, patient serum would be applied to a multiplexed plate of microcolumns filled with antibody-immobilized beads. Each microcolumn captures both the parental and fragment isoforms of the candidate marker since they both contain the antibody recognition site. The captured population of analytes, including the fragment(s) with potential for disease detection and/or discrimination, are eluted and analyzed directly by MALDI-TOF MS. The presence of the specific peptide biomarker at its precise mass/charge ratio (m/z) would be used as a diagnostic test result. This analysis could be performed rapidly by simple software that determines if the ion peak is present or absent at the predefined m/z location.