Bile and urine peptide marker profiles: access keys to molecular pathways and biological processes in cholangiocarcinoma

Abstract

Background: Detection of cholangiocarcinoma (CCA) remains a diagnostic challenge. We established diagnostic peptide biomarkers in bile and urine based on capillary electrophoresis coupled to mass spectrometry (CE-MS) to detect both local and systemic changes during CCA progression. In a prospective cohort study we recently demonstrated that combined bile and urine proteome analysis could further improve diagnostic accuracy of CCA diagnosis in patients with unknown biliary strictures. As a continuation of these investigations, the aim of the present study was to investigate the pathophysiological mechanisms behind the molecular determinants reflected by bile and urine peptide biomarkers.Methods: Protease mapping and gene ontology cluster analysis were performed for the previously defined CE-MS based biomarkers in bile and urine. For that purpose, bile and urine peptide profiles (from samples both collected at the date of endoscopy) were investigated from a representative cohort of patients with benign (n = 76) or CCA-associated (n = 52) biliary strictures (verified during clinical follow-up). This was supplemented with a literature search for the association of the individual biomarkers included in the proteomic patterns with CCA or cancer progression.Results: For most of the peptide markers, association to CCA has been described in literature. Protease mapping revealed ADAMTS4 activity in cleavage of both bile and urine CCA peptide biomarkers. Furthermore, increased chymase activity in bile points to mast cell activation at the tumor site. Gene ontology cluster analysis indicates cellular response to chemical stimuli and stress response as local and extracellular matrix reorganization by tissue destruction and repair as systemic events. The analysis further supports that the mapped proteases are drivers of local and systemic events. Conclusions: The study supports connection of the CCA-associated peptide biomarkers to the molecular pathophysiology and indicates an involvement in epithelial-to-mesenchymal transition, generation of cancer-associated fibroblasts and activation of residual immune cells. Proteases, extracellular matrix components, inflammatory cytokines, proangiogenic, growth and vasoactive factors released from the tumor microenvironment are drivers of systemic early events during CCA progression.

Pilot Multi-Omic Analysis of Human Bile from Benign and Malignant Biliary Strictures: A Machine-Learning Approach

“…If validated in subsequent analyses, the evaluation of a well-selected panel of these biomarkers may increase the diagnostic accuracy of biliary stenosis. On the other hand, bile proteomics can also contribute to a better understanding of the mechanisms of the tumorigenic process [45]. Taken together, these findings reveal the complexity of the bile proteome and attest to the interest of its characterization from both physiological, pathological and diagnostic points of view.…”

Discovery, validation and sequencing of urinary peptides for diagnosis of liver fibrosis—A multicentre study

“…Finally, when the activity of the predicted proteases was assessed based on the associated peptide marker expressions in the patient's CE-MS peptide profiles [25] , significance could be reached for the MMP's 2, 3, 13 and 14, as well as CTSB. Since for all of these proteases a decrease in activity was predicted in the case of LC, this leads us to the hypothesis based on literature findings [ 43 , 44 ] that mainly TIMP-2 might be the inducer and/or hepatorenal connector leading to the observed changes associated with liver fibrosis in the urinary peptidome.…”

“…Protease activity was assessed in the patient's CE-MS peptide profiles to gain fold changes between LC cases versus normal controls based on the average of associated peptide intensities. This method is described in detail by Voigtl€ ander et al [25].…”

Role of Endoscopy in Primary Sclerosing Cholangitis

“…In addition, developments in ancillary investigations are improving outcomes, and ERCP-based tissue sampling gives endoscopy a unique opportunity to play a substantial role in translational medicine. Improved analyses of brush material and better intraductal biopsies will help to improve clinical decision making, while identification of tumor markers in bile may support our diagnostic matrix in the future [ 59 , 60 ]. ERCP in these patients will remain challenging and should be increasingly centralized to transplant or other high-competence centers with high volumes of PSC patients.…”