Are Serum Peptidome Patterns Reproducible Enough to Monitor Cancer?

抄録

Proteomics has generated great anticipation in the cancer field as it may hold promise to establish simple blood tests through mass spectrometric (MS)-based profiling of patient proteomes/peptidomes. However, several concerns have been voiced regarding biological (patient demographics, environmental, dietary and psychological factors), technological (sample collection, processing and storage, analytical chemistry) and data mining artifacts that may introduce bias. We have established an automated platform to measure serum peptides using magnetic beads for selective capture and release, followed by a MALDI-TOF MS read-out. It is more sensitive than SELDI chips as spherical particles have larger combined surface areas than small-diameter spots. Hundreds of peptides can be detected in a droplet of serum; use of robotics facilitates throughput and ensures reproducibility. Unfortunately, serum preparation, handling and storage remain problematic, especially among archived samples. Thus, we have made great efforts to identify and remedy potential sources of bias, such as blood collection tubes, clotting times and temperature, number of freeze-thaw cycles, surface chemistries of magnetic beads, MALDI sample crystallization, laser irradiation, and various automation steps. In addition, adequate signal processing of the spectra (e.g. peak alignment, binning, etc.) proved more difficult and rate limiting than the actual statistical analysis. We therefore developed a minimal entropy-based algorithm that simplifies and improves alignment of spectra. Overall, our current SOP and informatics pipeline has greatly reduced handler variability and induced error typically associated with peptide measurements of clinical samples. A different problem of oncopeptidomics is that most of the serum peptidome as measured using high-throughput, low-resolution MS is the result of exopeptidase activities targeted at coagulation and complement activation byproducts. Formation of these peptides can be deregulated in disease and is easily affected by sample collection and handling. We have therefore designed and optimized a reliable in vitro activity test, uniquely suited to probe the altered balance of exopeptidases and/or their modulators in the blood of cancer patients.