Collagen Type II C-telopeptide Fragments as an Index of Cartilage Degradation S. CHRISTGAU, 1 P. GARNERO, 2 C. FLEDELIUS, 1 C. MONIZ, 3 M. ENSIG, 4 E. GINEYTS, 2 C. ROSENQUIST, 1 and P. QVIST 1 1 Osteometer BioTech A/S, Herlev, Denmark 2 Department of Biochemistry, INSERM Unit 403, Lyon, France 3 Kings College Hospital, London, UK 4 Centre for Clinical and Basic Research, Ballerup, Denmark We report the development of an assay for measurement of the urinary concentration of collagen type II C-telopeptide frag- ments. This assay was developed for providing a specific marker of joint metabolism. A monoclonal antibody, recogniz- ing a linear six amino acid epitope from the middle region of the collagen type II C-telopeptide was used in a competitive en- zyme-linked immunoassay (ELISA) format for measurement of urine samples. The technical performance and specificity of the assay was evaluated and a panel of samples from patients with rheumatoid arthritis (RA) (n  27), osteoarthritis (OA) (n  29), Paget’s disease (n  9), and healthy controls (n  428) was measured in the assay. The ELISA was specific for the peptide EKGPDP derived from collagen type II C-telopeptide: it did not recognize peptides from the N-telopeptide of the molecule or from other collagen types. Collagen type II C-telopeptide frag- ments measured in the assay resisted seven freeze-thaw cycles and >20 h of storage at room temperature. RA and OA patients showed significant 2.33-fold (95% confidence interval [CI] 1.50 –3.16) and 1.53-fold (CI 1.24 –1.82) elevations in CartiLaps concentration, respectively. Paget’s disease patients did not have elevated CartiLaps levels. RA patients with radiological evidence of cartilage damage had significantly higher (1.79-fold, CI 1.04 –2.54) CartiLaps levels than RA patients without radio- logical evidence of cartilage destruction. The Cartilaps assay showed high technical precision and an ability to differentiate populations with an elevated joint metabolism from normal controls. This suggests that the assay may have clinical value in assisting in the diagnosis of joint diseases and in monitoring progression and therapy in RA and OA. (Bone 29:209 –215; 2001) © 2001 by Elsevier Science Inc. All rights reserved. Key Words: Collagen type II; Cartilage degradation; Biochem- ical marker; Immunoassay. Introduction One of the major clinical manifestations in rheumatoid arthritis (RA) and osteoarthritis (OA) is abnormal and degraded cartilage in affected joints, but this aspect of the disease can be difficult to quantify. Clinical assessment of patients with RA and OA is performed by assessing pain and mobility problems caused by joint destruction and in RA patients, by evaluating inflammation. A number of standardized rating methods have been introduced for evaluation of pain and symptoms associated with joint dis- eases (i.e., HAQ, DAS, VAS, and ACR scores), but it is difficult to quantify these parameters. 13,22 Markers used for assessment of inflammation in RA patients, such as erythrocyte sedimentation rate (ESR) or C-reactive protein (CRP) and rheumatoid factors, are not specific for the inflammatory process involved in the disease, and they are probably not directly related to the level of cartilage destruction. 24 At present, radiological examination is one of the best ways to obtain information about the status of the joints and especially the progression of articular cartilage de- struction in diseased joints of RA and OA patients. 20 To improve diagnosis and monitoring of arthritis, significant efforts have focused on development of biochemical markers of cartilage destruction. Measurement of cartilage-derived metabolites, such as hyaluronates and aggrecan fragments, has been report- ed. 13,24 The clinical usefulness of these markers, however, remains to be proven. Recently, new commercially available assays, mea- suring other circulating components of the cartilage matrix, have been described. One assay, the Chrondrex enzyme-linked immuno- assay (ELISA), 6,9,10 measures a small glycoprotein (YKL-40 or GP-39) released by chondrocytes and synovial lining cells. Hence, this marker may not be a true marker of cartilage degradation, but rather of metabolic activity in the joint tissue. Another assay specific for cartilage oligomeric matrix protein (COMP) has been shown to provide a measure of elevated cartilage degradation. 14,18 This pro- tein is a constituent of articular cartilage, but is also synthesized by tendon fibroblasts and synoviocytes. 7 Other cartilage-derived pro- teins have been described in the literature, but at present their potential as biomarkers for human joint diseases has not been thoroughly evaluated. 13 Herein we describe the development and preliminary clinical evaluation of a specific immunoassay for measurement of the urinary concentration of collagen type II C-telopeptide degradation products. Type II collagen is localized almost exclusively in carti- lage, where it is a major structural component of the tissue. 16 Hence, measurements of fragments derived from this protein may poten- tially represent a specific index for cartilage degradation. A monoclonal antibody specific for an epitope derived from the collagen type II C-telopeptide was used in a competitive ELISA format for measurement of urinary samples. The collagen type II fragments measured in the assay were significantly Address for correspondence and reprints: Dr. Stephan Christgau, Oste- ometer BioTech A/S, Osteopark, Herlev Hovedgade 207, 2730 Herlev, Denmark. E-mail: sc@osteobio.com Bone Vol. 29, No. 3 September 2001:209 –215 209 © 2001 by Elsevier Science Inc. 8756-3282/01/$20.00 All rights reserved. PII S8756-3282(01)00504-X