Analysis of Autoantibody Profiles in Osteoarthritis Using Comprehensive Protein Array Concepts

Osteoarthritis (OA) is the most common rheumatic disease and one of the most disabling pathologies worldwide. To date, the diagnostic methods of OA are very limited, and there are no available medications capable of halting its characteristic cartilage degeneration. Therefore, there is a significant interest in new biomarkers useful for the early diagnosis, prognosis, and therapeutic monitoring. In the recent years, protein microarrays have emerged as a powerful proteomic tool to search for new biomarkers. In this study, we have used two concepts for generating protein arrays, antigen microarrays, and NAPPA (nucleic acid programmable protein arrays), to characterize differential autoantibody profiles in a set of 62 samples from OA, rheumatoid arthritis (RA), and healthy controls. An untargeted screen was performed on 3840 protein fragments spotted on planar antigen arrays, and 373 antigens were selected for validation on bead-based arrays. In the NAPPA approach, a targeted screening was performed on 80 preselected proteins. The autoantibody targeting CHST14 was validated by ELISA in the same set of patients. Altogether, nine and seven disease related autoantibody target candidates were identified, and this work demonstrates a combination of these two array concepts for biomarker discovery and their usefulness for characterizing disease-specific autoantibody profiles

Circulating Carnosine Dipeptidase 1 Associates with Weight Loss and Poor Prognosis in Gastrointestinal Cancer

<div><p>Background</p><p>Cancer cachexia (CC) is linked to poor prognosis. Although the mechanisms promoting this condition are not known, several circulating proteins have been proposed to contribute. We analyzed the plasma proteome in cancer subjects in order to identify factors associated with cachexia.</p><p>Design/Subjects</p><p>Plasma was obtained from a screening cohort of 59 patients, newly diagnosed with suspected gastrointestinal cancer, with (n = 32) or without (n = 27) cachexia. Samples were subjected to proteomic profiling using 760 antibodies (targeting 698 individual proteins) from the Human Protein Atlas project. The main findings were validated in a cohort of 93 patients with verified and advanced pancreas cancer.</p><p>Results</p><p>Only six proteins displayed differential plasma levels in the screening cohort. Among these, Carnosine Dipeptidase 1 (CNDP1) was confirmed by sandwich immunoassay to be lower in CC (p = 0.008). In both cohorts, low CNDP1 levels were associated with markers of poor prognosis including weight loss, malnutrition, lipid breakdown, low circulating albumin/IGF1 levels and poor quality of life. Eleven of the subjects in the discovery cohort were finally diagnosed with non-malignant disease but omitting these subjects from the analyses did not have any major influence on the results.</p><p>Conclusions</p><p>In gastrointestinal cancer, reduced plasma levels of CNDP1 associate with signs of catabolism and poor outcome. These results, together with recently published data demonstrating lower circulating CNDP1 in subjects with glioblastoma and metastatic prostate cancer, suggest that CNDP1 may constitute a marker of aggressive cancer and CC.</p></div

Correlations between CNDP1 and clinical parameters in cohort 2.

<p>Correlations were performed using simple linear regression, r- and p-values are shown.</p><p>Correlations between CNDP1 and clinical parameters in cohort 2.</p

Candidates found in the unbiased proteomic screen.

<p>Candidates found in the unbiased proteomic screen.</p

Clinical characteristics of cohort 1 and 2.

<p>Values are given as mean±S.D. except for gender where the actual number of males (M) and females (F) are given. Group values were compared using unpaired t-test or χ² test.</p><p>*Measured by bioimpedance in cohort 1 and by DXA in cohort 2.</p><p>Clinical characteristics of cohort 1 and 2.</p

Identification and validation of CNPD1 reduction in CC.

<p>Detected protein levels are shown for <b>A.</b> the screen using Sandwich Bead Arrays (SBA) and <b>B.</b> validation by Sandwich Immunoassays (SIA) and expressed as mean intensity fluorescence (MIF). <b>C.</b> There was an excellent correlation between results obtained by SBA and SIA. CNPD1 levels measured by SIA were not influenced by <b>D.</b> age or <b>E.</b> gender. <b>F.</b> Distribution of circulating CNDP1 concentrations determined by SIA in Cohort 2. WS = Weight stable, CC = Cancer cachexia. P- and rho-values are given.</p