title: Plasma autoantibodies against heat shock protein 70, enolase 1 and ribonuclease/angiogenin inhibitor 1 as potential biomarkers for cholangiocarcinoma.
abstract: The diagnosis of cholangiocarcinoma (CCA) is often challenging, leading to poor prognosis. CCA arises via chronic inflammation which may be associated with autoantibodies production. This study aims to identify IgG antibodies directed at self-proteins and tumor-associated antigens. Proteins derived from immortalized cholangiocyte cell line (MMNK1) and CCA cell lines (M055, M214 and M139) were separated using 2-dimensional electrophoresis and incubated with pooled plasma of patients with CCA and non-neoplastic controls by immunoblotting. Twenty five immunoreactive spots against all cell lines-derived proteins were observed on stained gels and studied by LC-MS/MS. Among these, heat shock protein 70 (HSP70), enolase 1 (ENO1) and ribonuclease/angiogenin inhibitor 1 (RNH1) obtained the highest matching scores and were thus selected for further validation. Western blot revealed immunoreactivity against HSP70 and RNH1 in the majority of CCA cases and weakly in healthy individuals. Further, ELISA showed that plasma HSP70 autoantibody level in CCA was significantly capable to discriminate CCA from healthy individuals with an area under the receiver operating characteristic curve of 0.9158 (cut-off 0.2630, 93.55% sensitivity and 73.91% specificity). Plasma levels of IgG autoantibodies against HSP70 were correlated with progression from healthy individuals to cholangitis to CCA (r = 0.679, P<0.001). In addition, circulating ENO1 and RNH1 autoantibodies levels were also significantly higher in cholangitis and CCA compared to healthy controls (P<0.05). Moreover, the combinations of HSP70, ENO1 or RNH1 autoantibodies positivity rates improved specificity to over 78%. In conclusion, plasma IgG autoantibodies against HSP70, ENO1 and RNH1 may represent new diagnostic markers for CCA.
authors: Department of Parasitology, Faculty of Medicine, Khon Kaen University, Khon Kaen, Thailand; Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen, Thailand.
authors: Rucksak
authors: R
authors: Rucksaken
authors: Department of Pathology, Faculty of Medicine, Khon Kaen University, Khon Kaen, Thailand; Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen, Thailand.
authors: Chawalit
authors: C
authors: Pairojkul
authors: Centre for Research and Development in Medical Diagnostic Laboratory, Faculty of Associated Medical Sciences, Khon Kaen University, Khon Kaen, Thailand; Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen, Thailand.
authors: Porntip
authors: P
authors: Pinlaor
authors: Department of Surgery, Faculty of Medicine, Khon Kaen University, Khon Kaen, Thailand; Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen, Thailand.
authors: Narong
authors: N
authors: Khuntikeo
authors: Proteomics Research Laboratory, Genome Institute, National Center for Genetic Engineering and Biotechnology, Pathumthani, Thailand.
authors: Sittiruk
authors: S
authors: Roytrakul
authors: Division of Rheumatology and Clinical Immunology, Humanitas Clinical and Research Center, Rozzano, Milan, Italy; BIOMETRA Department, University of Milan, Italy.
authors: Carlo
authors: C
authors: Selmi
authors: Department of Parasitology, Faculty of Medicine, Khon Kaen University, Khon Kaen, Thailand; Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen, Thailand.
authors: Somchai
authors: S
authors: Pinlaor
mesh_terms: D000368:Aged; D000818:Animals; D001323:Autoantibodies / Q000097:blood*; D001650:Bile Duct Neoplasms / Q000097:blood* / Q000175:diagnosis / Q000473:pathology; D014408:Biomarkers, Tumor / Q000097:blood* / Q000276:immunology; D002352:Carrier Proteins / Q000276:immunology*; D016022:Case-Control Studies; D002478:Cells, Cultured; D018281:Cholangiocarcinoma / Q000097:blood* / Q000175:diagnosis / Q000473:pathology; D004268:DNA-Binding Proteins / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D018840:HSP70 Heat-Shock Proteins / Q000276:immunology*; D006801:Humans; D008297:Male; D008875:Middle Aged; D010751:Phosphopyruvate Hydratase / Q000276:immunology*; D014200:Trematoda / Q000276:immunology / Q000502:physiology; D014201:Trematode Infections / Q000097:blood / Q000150:complications / Q000276:immunology; D025521:Tumor Suppressor Proteins / Q000276:immunology*
chemical_list: D001323:Autoantibodies; D014408:Biomarkers, Tumor; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D018840:HSP70 Heat-Shock Proteins; C501467:RNH1 protein, human; D025521:Tumor Suppressor Proteins; C095157:ENO1 protein, human; D010751:Phosphopyruvate Hydratase