Objective Osteoarthritis (OA) is a chronic and slowly progressive disease for which biomarkers may be able to provide a more rapid indication of therapeutic responses to therapy than is currently available; this could accelerate and facilitate OA drug discovery and development programs. The goal of this document is to provide a summary and guide to the application of in vitro (biochemical and other soluble) biomarkers in the development of drugs for OA and to outline and stimulate a research agenda that will further this goal. Methods The Biomarkers Working Group representing experts in the field of OA biomarker research from both academia and industry developed this consensus document between 2007–2009 at the behest of the Osteoarthritis Research Society International (OARSI FDA initiative). Results This document summarizes definitions and classification systems for biomarkers, the current outcome measures used in OA clinical trials, applications and potential utility of biomarkers for development of OA therapeutics, the current state of qualification of OA-related biomarkers, pathways for biomarker qualification, critical needs to advance the use of biomarkers for drug development, recommendations regarding practices and clinical trials, and a research agenda to advance the science of OA-related biomarkers. Conclusions Although many OA-related biomarkers are currently available they exist in various states of qualification and validation. The biomarkers that are likely to have the earliest beneficial impact on clinical trials fall into two general categories, those that will allow targeting of subjects most likely to either respond and/or progress (prognostic value) within a reasonable and manageable time frame for a clinical study (for instance within one to two years for an OA trial), and those that provide early feedback for preclinical decision-making and for trial organizers that a drug is having the desired biochemical effect. As in vitro biomarkers are increasingly investigated in the context of specific drug treatments, advances in the field can be expected that will lead to rapid expansion of the list of available biomarkers with increasing understanding of the molecular processes that they represent.

Osteoarthritis (OA) is a progressive disorder characterized by destruction of articular cartilage and subchondral bone, and by synovial changes. The diagnosis of OA is generally based on clinical and radiographic changes, which occur fairly late during disease progression and have poor sensitivity for monitoring disease progression. Progression of joint damage is likely to result primarily from an imbalance between cartilage degradation and repair, so measuring markers of these processes would seem a promising approach to improve the prediction of disease progression at the individual level. Moreover, genetic markers might be useful predictors of prognosis. The lack of fully effective, chondroprotective medications has limited the use of such potential markers to monitor the effect of treatment for OA. Nevertheless, owing to their dynamic changes in response to treatment, biological markers might provide relevant information more rapidly than imaging techniques (such as radiography and MRI) can, and should contribute to our understanding of mechanisms that underlie the clinical efficacy of OA treatments. Most of the identified genes involved in OA encode signal-transduction proteins, which provide the potential for novel therapeutic approaches. In this Review, we will use the recently proposed BIPED (i.e. burden of disease, investigative, prognostic, efficacy of intervention and diagnostic) classification of OA markers to describe the potential usage of a given marker.

Objective. The hallmark of osteoarthritis (OA) is the loss of articular cartilage. This loss arises from an imbalance between cartilage synthesis and cartilage degradation over a variable period of time. The aims of this study were to investigate the rates of these processes in patients with knee OA using two new molecular markers and to investigate whether the combined use of these markers could predict the progression of joint damage evaluated by both radiography and arthroscopy of the joints during a period of 1 year.Methods. Seventy-five patients with medial knee OA (51 women, 24 men; mean ؎ SD age 63 ؎ 8 years, mean ؎ SD disease duration 4.8 ؎ 5.2 years) were studied prospectively. At baseline, we measured serum levels of N-propeptide of type IIA procollagen (PIIANP) and urinary excretion of C-terminal crosslinking telopeptide of type II collagen (CTX-II) as markers of type II collagen synthesis and degradation, respectively. Joint space width (JSW) on radiography and medial chondropathy at arthroscopy (assessed using a 100-mm visual analog scale [VAS]) were measured in all patients at baseline and in 52 patients at 1 year. Progression of joint destruction was defined as a decrease of >0.5 mm in JSW on radiography and as increased chondropathy (an increase in the VAS score of >8.0 units) between the baseline and 1-year evaluations.Results. At baseline, compared with 58 healthy age-and sex-matched controls, patients with knee OA had decreased serum levels of PIIANP (20 ng/ml versus 29 ng/ml; P < 0.001) and increased urinary excretion of CTX-II (618 ng/mmole creatinine [Cr] versus 367 ng/ mmole Cr; P < 0.001). The highest discrimination between OA patients and controls was obtained by combining PIIANP and CTX-II in an uncoupling index (Z score CTX-II ؊ Z score PIIANP), which yielded a mean Z score of 2.9 (P < 0.0001). Increased baseline values in the uncoupling index were associated with greater progression of joint damage evaluated either by changes in JSW (r ‫؍‬ ؊0.46, P ‫؍‬ 0.0016) or by VAS score (r ‫؍‬ 0.36, P ‫؍‬ 0.014). Patients with both low levels of PIIANP (less than or equal to the mean ؊ 1 SD in controls) and high levels of CTX-II (greater than or equal to the mean ؉ 1 SD in controls) had an 8-fold more rapid progression of joint damage than other patients (P ‫؍‬ 0.012 and P < 0.0001 as assessed by radiography and arthroscopy, respectively) and had relative risks of progression of 2.9 (95% confidence interval [95% CI] 0.80-11.1) and 9.3 (95% CI 2.2-39) by radiography and arthroscopy, respectively.Conclusion. Patients with knee OA are characterized by an uncoupling of type II collagen synthesis and degradation which can be detected by assays for serum PIIANP and urinary CTX-II. The combination of these two new markers could be useful for identifying knee OA patients at high risk for rapid progression of joint damage.

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We have developed the first specific immunoassay for serum PIIANP which exhibits adequate technical performances. This assay detects specifically two immunoreactive forms both in healthy adults and patients with arthritis and does not cross react with other proteins with sequence homology with PIIANP. Levels of PIIANP were significantly decreased in patients with knee OA and RA suggesting that type IIA collagen synthesis may be altered in these arthritic diseases. The measurement of type IIA collagen synthesis with this new molecular marker may be useful for the clinical investigation of patients with joint diseases.

The stimulation of bovine articular cartilage explants with OSM/TNFalpha released aggrecan fragments both in an MMP and non-MMP-mediated route. These immunoassays carry a potential as diagnostic tools for the quantitative assessment of the cartilage turnover in RA patients in addition to their utility in ex vivo explant cultures.

The processing of human collagen type-V chains was studied using anti-peptide polyclonal antibodies raised against peptide sequences at the N-terminal non-triple-helical region of pro-al(V) and pro-a2(V) chains. The anti-peptide polyclonal antibody raised against positions 48 -57 of the N-terminal a2(V) sequence recognized the mature form of the human a2(V) chain extracted without any proteolytic treatment from several tissues in the presence of a mixture of protease inhibitors. It also recognized the pro-a2(V) and pN-a2(V) collagen chains secreted in the cell-culture media of the rhabdomyosarcoma A204 cell line. The pN-a2(V) collagen chain from this cell line migrated during electrophoresis with the a2(V) chain obtained from tissues. This demonstrates that the a2(V) chain in tissues is incompletely processed and is present as the pN-a2(V) collagen chain which lacks the C-propeptide. In comparison, an anti-peptide polyclonal antibody raised against residues at positions 284-299 of the N-terminal al(V) human sequence failed to recognize the mature form of the al(V) chain while it reacted with the pN-al(V) collagen chain form. These results suggest that the al(V) chain undergoes a processing event in the N-terminal region that involves the removal of at least the first 284 residues.Amino acid sequence analysis was performed on cyanogen-bromide-generated or trypsin-generated peptides of the two electrophoretic bands obtained for the tissue form of collagen V. The slower-migrating band corresponding to the intact al(V) chain gave, as expected, only sequences corresponding to the al(V) chain. However, the band previously considered to be the intact a2(V) chain also gave sequences for the al(V) chain in addition to the a2(V) chain. This result indicates the presence in tissue extracts of a further processed form of al(V) chain which migrates with the intact a2(V) chain. On further analysis, we observed that the two bands of the tissue form of collagen V occurred in a 1 : 1 ratio whereas, after the pepsin digestion to remove non-collagenous regions, two bands were observed with an al(V)la2(V) chain ratio of 3 : 1. These results indicate that the a1 (V) chain exists in an additional stoichiometry, different from [al(V)],a2(V). We suggest the existence of two different populations of type-V collagen molecules consisting of an [a1 (V)],a2(V) heterotrimer bearing considerable N-terminal non-triple-helical extensions of both al(V) and a2(V) chains and an [al(V)], homotrimer composed of fully processed al(V) chains.Collagen V is a quantitatively minor constituent of collagen fibrils in the extracellular matrix, It is mainly found in tissues that contain type-I collagen as the major collagen component. Collagen V is heterogeneous and a1 (V)-a3(V)