Author information
1Duke Clinical Research Institute, Durham, North Carolina; Liver Cell Biology, Centenary Institute, University of Sydney, Sydney, NSW, Australia. Electronic address: keyur.patel@duke.edu.
2GlaxoSmithKline, RTP, North Carolina.
3OpGen, Gaithersburg, MD.
4BioAgilytix Labs, RTP, NC.
5Institute for Genome Sciences and Policy, Duke University, Durham, North Carolina.
6Duke Clinical Research Institute, Durham, North Carolina; Gilead Sciences, Foster City, CA.
7NIHR Biomedical Research Unit in Gastrointestinal and Liver Diseases at Nottingham University Hospitals NHS Trust and the University of Nottingham, Nottingham, UK.
Abstract
BACKGROUNDS & AIMS:
Non-invasive tests cannot differentiate between adjacent stages of fibrosis, which limits assessment of disease progression and regression during therapy. We investigated whether levels of cytokines and extracellular matrix proteins in serum and biopsy samples can be used to determine actual stage of liver fibrosis in patients with chronic hepatitis C (CHC), and in prognosis.
METHODS:
We collected data from 383 treatment-naïve patients with CHC from the Duke Hepatology Clinical Research Database and Biorepository, from 2006 through 2009, for use in the training set. Serum samples were obtained from 100 individuals without CHC (controls). We selected 37 serum biomarkers for customized array analysis, using the SearchLight multiplex sandwich ELISA. Data from 434 treatment-naïve patients with CHC, obtained from the Trent HCV cohort, were used in the validation analysis. Multivariable modeling, marker selection, and validation included RandomForest and Obuchowski measures, with independent comparison to FibroSURE.
RESULTS:
Four serum markers (levels of hyaluronic acid, VCAM1, A2M, and RBP4) and age associated with fibrosis stage (F0-1, F2-3, or F4); these had Obuchowski measures of 0.85-0.89, with misclassification rates of 38% and 29% in training and validation sets, compared to 50% for the FibroSURE test. In the training set, area under the curve (AUC) values for the multiplex markers were similar to those from the FibroSURE test: stages F0 vs F1 (0.51 vs 0.53), F1 vs F2 (0.60 vs 0.59), F2 vs F3 (0.69 vs 0.72), and F3 vs F4 (0.51 vs 0.52). AUC values were similar in the validation cohort. In longitudinal analyses of 133 paired biopsies, 9 markers (level of alanine aminotransferase, γ-glutamyltranspeptidase, hyaluronic acid, ICAM1, interleukin (IL)4, CXCL10, CXCL9, and VCAM1) were associated with change in the histologic activity index (P values ranging from .000 to .049) and 4 (GMCSF, IL12, IL2, and MMP13) were associated with a change in fibrosis stage (P values ranging from .001 to .042).
CONCLUSIONS:
We identified serum biomarkers that can be measured by mutiplex ELISA to determine levels of fibrosis in patients with CHC, although misclassification is frequent and results are comparable with those from the FibroSURE test. Changes in protein levels in biopsy samples associated with progression of fibrosis in patients.