Using synovial fluid biomarkers to define a phenotype of osteoarthritis in the hip

Osteoarthritis (OA) is a chronic, multi-factorial, progressive joint disease. The understanding of OA has evolved away from a wear and tear phenomenon to a complex disease of the joint taken as a whole. A method of stratifying disease, and monitoring progression, is necessary for clinical decision making and for use as end points in future research studies to develop disease modifying OA drugs (DMOADs). The creation of such a tool requires detailed understanding of the disease biopathogenesis. Synovial fluid (SF) is the most likely target to generate the insight required due to its proximity to the intra-articular structures, whose changes are fundamental to the symptomatology of the disease. No work to date has identified any useful single biomarkers of OA and for progress to be made, a consideration of a number of proteins as a panel of markers, whose interrelations describe a signature of different disease states, is required. The emergence of the slow off-rate modified aptamer scan (SOMAscan), a broad spectrum proteomic platform, presents the opportunity to analyse 1310 proteins on a single sample of fluid that measures less than 1 ml. This thesis aims to examine the utility of the SOMAscan in OA biomarker discovery by validation against traditional bench assays, use multivariate statistical analysis methods to identify candidate markers of OA disease states, and contextualise their best clinical timing in the patient pathway by examining the treatment pathways of patients from initial presentation to final treatment.
Synovial fluid samples from hip and knee joints were collected from patients with different stages of clinically classified disease including knee injury, early and late OA cohorts, and hip early and late OA cohorts with the early hip OA group requiring the development of an entirely new collection method. The performance of the SOMAscan was validated using ELISA and MSD assays that were performed on a curated list of proteins and compared to the same markers in the SOMAscan showing robust comparison. Multivariate regression analysis showed separation of the knee injury and late OA cohorts using PLS-DA, but the early group were not separable from the injury group. Markers identified by LASSO that were strongly associated with late disease included elevated TSG-6, TIMP-1 and R-spondin 4. These markers were also identified in the PLS-DA model generating potential targets for use in future OA studies.
Hip SF analysis showed separation between the early and late OA groups on PLS-DA. LASSO analysis revealed 6 markers: TSG-6, DDR-2, Ephrin-A5, S100A7, MICB, IDS all of which were present in the top 50 marker model produced using PLS-DA. This combined analysis approach has shown differences between the hip groups, differences between hip and knee OA biology and suggested novel targets for future OA studies.
An epidemiological study investigating the patient pathways, from presentation in primary care through to eventual arthroplasty, was conducted to identify where a diagnostic test would be best placed in the patient journey. Results showed marked variation in the rates of treatments offered to patients during their disease, with wide variation in analgesia, physiotherapy and intermediate surgical interventions. Most notably it showed that there was a continued increase in the use of opioid analgesia overall, particularly in the most deprived patients, indicating a need to investigate ways to harmonise this patient experience such that patients can be identified reliably at earlier stages of disease.
In summary, the works in this thesis have validated the SOMAscan. A new system of biomarker discovery in OA, for use in SF analysis. The SOMAscan analysis of SF has shown a variety of differently expressed markers and pathways including well described OA processes in both knee and hip. It has suggested that early knee OA appears biologically closer to joint injury than to late knee OA, and that hip and knee OA appear to be biologically distinct processes with common thematic pathways as well as differences. This work has collected a new and unique SF cohort of human early hip OA SF to define a biological phenotype of late OA that discriminates between the groups studied. This work has further shown that the patient journey from presentation to definitive treatment is hugely varied, both in duration and treatments delivered, demonstrating that ongoing work is needed not only to identify biomarkers of early OA but to identify means of accessing patients earlier in their clinical disease states. Together these critical elements of disease description and patient identification require optimisation such that future OA biomarkers may be utilised as disease diagnosis, stratification and outcome tools in DMOAD studies.

Keywords: Knee
Hip
Biomarkers
Osteoarthritis
Synovial Fluid
Advisor: Hulley, P
Glyn-Jones, S
Judge, A