Targeted In-Gel MRM: A Hypothesis Driven Approach for Colorectal Cancer Biomarker Discovery in Human Feces

Abstract

Colorectal cancer (CRC) is the second most common cause of cancer-related deaths in both men and women. The fecal occult blood test is currently the first line method for CRC screening but has an unacceptably low sensitivity and specificity. Improved screening tests are therefore urgently required for early stage CRC screening. We have described a hypothesis-driven approach for a rapid biomarker discovery process whereby selected proteins previously implicated as colorectal cancer-associated proteins (CCAP), which can potentially be shed into the feces from a colorectal tumor, are targeted for excision from 1D-SDS-PAGE based on their predicted molecular weight followed by directed identification and relative quantification using multiple reaction monitoring (MRM). This approach can significantly reduce the time for clinical assay development with the added advantage that many proteins will have been validated by previous in vitro and/or in vivo studies. Sixty potential CCAPs were selected from the literature and appropriate MRM conditions were established for measurement of proteotypic peptides. Nineteen of these proteins were detected in the feces from a patient with colorectal cancer. Relative quantitation of these 19 CCAP across 5 CRC patients and 5 healthy volunteers were carried out, revealing hemoglobin, myeloperoxidase, S100A9, filamin A and l-plastin to be present only in the feces of CRC patients.

Regulation of the Transcriptional Coactivator FHL2 Licenses Activation of the Androgen Receptor in Castrate-Resistant Prostate Cancer

“…Calpain expression and activity is increased in human colorectal cancer and in colorectal polyps, where it may be an early tumorigenesis event (47) and we predict this may increase filamin cleavage and promote nuclear FHL2. Filamin is a potential diagnostic biomarker for colorectal cancer as it is shed by tumors into patient feces (48). Although this latter study did not examine the presence of the 90 kDa filamin fragment generated by calpain cleavage, of note was the observation that filamin appeared at a lower molecular weight than predicted.…”

Plastin Family of Actin-Bundling Proteins: Its Functions in Leukocytes, Neurons, Intestines, and Cancer

“…Cancer screening and diagnosis methods assessing the expression of plastins as a biomarker have been described. These include choroids plexus tumors, urinary bladder cancer, ovarian cancer, and colorectal cancer (Table 5) [71–74]. Furthermore, gene therapy experiments targeting the L-plastin gene or L-plastin promoters in cancer cells have been started, as shown in Table 5 [75–78].…”

The application of selective reaction monitoring confirms dysregulation of glycolysis in a preclinical model of schizophrenia

“…Here, we have shown that SRM mass spectrometry is a useful platform for achieving this goal. This approach has already been used in clinical studies such as those screening for the levels of dihydroartemisinin (DHA) in plasma of malaria patients [28], for determining the levels of apolipoproteins in human plasma [29] and for cancer biomarker screening [30]. Finally, although in a small number of samples, the results of the analysis support a role for the PCP rat as a good model for some aspects of schizophrenia, including metabolic dysfunction, as this has shown dysregulation of the glycolysis-related enzymes as previously shown in brain tissue from schizophrenia patients.…”