Human plasma arguably contains one of the most complex human proteomes and possesses the ability to reflect progress of human diseases, potentially including early detection of cancers. Colorectal cancer (CRC) is a particularly devastating condition, claiming ~700,000 lives/year. Although early detection significantly increases survival, it is rarely detected at early stage. This study aims to build technologies (SWATH-MS) that are potentially capable of identifying plasma prognostic biomarkers for early CRC detection.
To maximise CRC plasma protein identification from SWATH plasma library, 14 highest abundant proteins (HAPs) were depleted from pooled staged CRC plasmas. Depleted plasmas were tryptically digested and fractioned using 4 different independent methods (high pH reversed-phase, strong anion exchange, strong cation exchange and size exclusion chromatography). Quantitative SWATH-MS analyses were performed on 100 CRC patient plasma pools (n=20 patients in each of CRC AJCC stages I-IV and age- and sex-matched healthy controls). SWATH-MS was performed on both depleted (14 HAP) and non-depleted plasmas.
A total of 529 unique, high-quality, plasma proteins were identified from 4 different peptide fractionation methods with 1% FDR level. Out of these, 363 proteins were quantified from depleted and 315 from non-depleted CRC plasma. Statistical analysis (BH-adjusted p-value <0.05 and fold change >1.5) showed 8 protein candidates exhibited differential expression across all CRC stages compared to healthy controls from non-depleted and 9 candidates from depleted plasma samples (SAA2 was common in both plasma samples). We are currently validating these candidates.
Our analysis revealed some known prognostic candidates effectively validating the methodology employed. We also uncovered number of novel candidates. We are extending SWATH-MS analysis to the 100 individual CRC and control plasmas to identify biomarkers that associate with CRC patient early stage disease and with clinical outcomes (e.g., survival, recurrence and chemotherapy effectiveness).