Biomarkers in Early-stage Spondyloarthritis
Biomarkers in Early-stage Spondyloarthritis
Axial spondyloarthritis (axSpA) is a group of inflammatory rheumatic conditions with
 inflammatory back pain caused by inflammation in the sacroiliac joints (SIJ) and back as
 hallmark.

 Currently, no laboratory test or biomarker is cable of differentiating between patients with
 early-stage axSpA and persisting low back pain of other causes.

 The objective of this study is to investigate the predictive value of baseline levels of the
 biomarkers Complement C3d and high-sensitive C-reactive protein (HsCRP) identifying bone
 marrow oedema (BME) at MRI of the SIJ at baseline.
Currently, no laboratory test or biomarker is cable of differentiating between patients with
 early-stage axSpA and persisting low back pain of other causes. Human leukocyte antigen (HLA)
 B27 is not an acute biomarker, but a prognostic marker for the development of axSpA.

 C-reactive protein (CRP) is commonly used as an acute biomarker in different inflammatory
 conditions. Previous studies have shown that only 40-60% of ankylosing spondylitis (AS)
 patients with clinical signs of active disease have elevated levels of CRP.

 Earlier studies regarding the complement C3 split product C3d have shown a correlation
 between inflammatory diseases like AS and elevation of C3d. Those studies were based on a
 small number of cases, but a newer study from 2016 has shown a correlation between different
 biomarkers, including C3 and new fatty lesions at MRI in Golimumab treated AS patients.

 Objective To investigate the predictive value of baseline levels of C3d and hsCRP identifying
 bone marrow oedema (BME) at MRI of the sacroiliac joints (SIJ) at baseline.

 Design:

 The study is a prospective cohort study including patients, aged 18-45 years, having
 unspecific low back pain registered in the Spines of Southern Denmark (SSD) cohort including
 patients referred to the Spine Centre of Southern Denmark.

 At baseline, all patients filled in a standardised questionnaire and were interviewed
 identifying inflammatory symptoms followed by a clinical examination focusing on symptoms
 related to the SIJ. Blood samples were taken and analysed for HLA-B27 (prognostic biomarker).
 Furthermore, all patients underwent an MRI of the spine including the SIJ.

 In total, 1037 patients were included, 696 had blood tests taken and gave consent for further
 use for research purposes.

 For research purposes, the blood samples were stored in the Molecular Biology of Infectious
 Agents in the Early Diagnosis of Spondyloarthritis biobank (MICSA). The biobank is hosted by
 the research group at Graasten Rheumatological Hospital. Of the 696 patients who gave
 consent,188 patients were examined by a Rheumatologist at baseline. Of those 96 patients
 fulfilled the criteria of having axSpA according to The Assessment of SpondyloArthritis
 international Society (ASAS) criteria, 38 patients had inflammatory changes at MRI or had one
 feature consisting with spondyloarthritis, but did not fulfil the ASAS criteria. 82 randomly
 selected patients with low back pain were included as a control group.

 In total, 216 patients were defined as the cohort included for further analysis in this
 study.

 Data:

 The Department of Clinical Biochemistry at Vejle Hospital has previously assisted in the
 collection and storage of MICSA samples. It is technically possible to use the stored frozen
 MICSA samples for analysing C3d.

 In total, 184 (of 216) baseline samples were accessible for further analysis. The baseline
 samples will be analysed for C3d with the use of EDTA (ethylenediaminetetraacetic acid)
 plasma and treated according to the local protocol for such analyses.

 MRI

 All patients in the MICSA cohort have had an MRI of the spine including the SIJ at baseline.
 Two experienced radiologists have systematically described the MRI regarding signs of
 inflammatory changes including BME.

 Statistics:

 STATA (version 15.0) will be used for statistical analysis. Parametric data will be reported
 as the mean and standard derivation (SD) or One-way analysis of variance (ANOVA). In
 between-group comparisons for continuous and categorical demographic variables will be
 performed with the independent sample t-test and Pearson Chi-square test or Fisher´s exact
 test. The Kruskal-Wallis test and interquartile range will be used to describe nonparametric
 data. Logistic regression analysis will be used to test independent variables as predictive
 factors for BME detected by MRI, resulting in odds-ratios, sensitivity, specificity, and area
 under the curve (AUC). This will be followed by multivariate logistic regression to evaluate
 the potential effect of other variables. A p-value < 0.05 is considered statistically
 significant.

 Sample size and power calculation:

 A previous study with patients suspected of having spondyloarthritis has found a significant
 difference in mean levels of C3 between groups with 23 and 22 participants, respectively.

 By the use of means and SD from the above study and alfa= 0.05, beta= 0.2, power 0.8 and
 enrollment ratio 1:2 the sample size in our study is calculated to n:10 and n:20
 respectively.

 It is therefore expected, that a significant difference in C3d levels between the three
 groups in the present study can be detected based on the size of each group in the MISCA
 cohort.
Inclusion criteria:

 -  patients from the Spines of Southern Denmark cohort who underwent MRI, blood samples,
 and clinical examination by a rheumatologist.

 -  aged 18-45

 -  three months of low back pain prior to inclusion

 Exclusion criteria:

 -  < 3 months of low back pain

 -  not wish to participate/ did not sign consent form