Fan et al. Arthritis Research & Therapy (2019) 21:123 https://doi.org/10.1186/s13075-019-1906-y RESEARCHARTICLE Open Access Cysteine-rich 61 (Cyr61): a biomarker reflecting disease activity in rheumatoid arthritis Yong Fan†, Xinlei Yang†, Juan Zhao, Xiaoying Sun, Wenhui Xie, Yanrong Huang, Guangtao Li, Yanjie Hao and Zhuoli Zhang* Abstract Background: Numerous preclinical studies have revealed a critical role of cysteine-rich 61 (Cyr61) in the pathogenesis of rheumatoid arthritis (RA). But there is little literature discussing the clinical value of circulation Cyr61 in RA patients. The aim of our study is to investigate the serum Cyr61 level and its association with disease activity in RA patients. Methods: A training cohort was derived from consecutive RA patients who visited our clinic from Jun 2014 to Nov 2018. Serum samples were obtained at the enrollment time. To further confirm discovery, an independent validation cohort was set up based on a registered clinical trial. Paired serum samples of active RA patients were respectively collected at baseline and 12 weeks after uniformed treatment. Serum Cyr61 concentration was detected by enzyme-linked immunosorbent assay. The comparison of Cyr61 between RA patients and controls, the correlation between Cyr61 levels with disease activity, and the change of Cyr61 after treatment were analyzed by appropriate statistical analyses. Results: A total of 177 definite RA patients and 50 age- and gender-matched healthy controls were enrolled in the training cohort. Significantly elevated serum Cyr61 concentration was found in RA patients, demonstrating excellent diagnostic ability to discriminate RA from healthy controls (area under the curve (AUC) = 0.98, P < 0.001). In addition, the Cyr61 level in active RA patients was significantly lower than that in patients in remission/low disease activity, and it was inversely correlated with composite disease activity scores and almost all of the components in statistic. Further study in the validation cohort (n = 77) showed a significant increase of the Cyr61 level at 12 weeks in ACR responders (ACR20/50/70), while no significant change of the Cyr61 level from baseline was observed in non-responders. Conclusions: Serum Cyr61 levels were remarkably increased in RA patients compared with those in healthy controls. The Cyr61 concentration was inversely correlated with RA disease activity and upregulated in those therapeutic responders. Trial registration: Combination Therapy Prevents the Relapse of RA, NCT02320630. Registered 19 December 2014, Keywords: Rheumatoid arthritis, Cysteine-rich protein 61, Biomarker, Disease activity, Treatment response * Correspondence: [email protected] †Yong Fan and Xinlei Yang contributed equally to this work. Department of Rheumatology and Clinical Immunology, Peking University First Hospital, No.8, Xishiku Street, West District, Beijing 100034, China © The Author(s). 2019 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Fan et al. Arthritis Research & Therapy (2019) 21:123 Page 2 of 9 Introduction controls and to investigate the correlation between Rheumatoid arthritis (RA) is a chronic inflammatory serum Cyr61 level with clinical disease activity in our autoimmune disease that causes progressive articular training cohort and validation cohort. damage, functional loss, and comorbidity [1]. Although the development of effective biologics and Methods small-molecule kinase inhibitors in the past two decades Study population has substantially improved clinical outcomes of RA, a In this study, a training cohort was used for discovery considerable number of patients respond insufficiently and to identify some relationships, while an independent to the therapy [2]. Until now, the exact pathological pro- validation cohort was set up to further confirm. The cesses involved in RA remain incompletely understood. training cohort was derived from consecutive RA pa- Thus, we still need to investigate the pathological mech- tients who visited the Rheumatology clinic, Peking Uni- anisms and look for the appropriate markers to reflect versity First Hospital, China, from Jun 2014 to Nov disease activity as well as predict treatment response. 2018. All patients were > 18 years of age and satisfied the Cysteine-rich 61 (Cyr61, also called CCN1), a novel se- 2010 American College of Rheumatology (ACR) classifi- creted matricellular protein, is encoded by a growth cation criteria for RA [15]. Serum samples were col- factor-inducible immediate-early gene. Cyr61 expression lected at the time of enrollment. is at low levels in most adult tissues under homeostatic To further confirm discovery, a validation cohort con- conditions, and remarkably elevated with the induction sisted of RA patients from the screening phase of our of cytokines and growth factors [3]. Numerous studies prospective randomized and controlled clinical trial have revealed that Cyr61 played critical roles in cardio- (ClinicalTrials.gov identifier: NCT02320630). Briefly, the vascular development, inflammation, injury repair, and aim of this trial was to compare the efficacy and phar- cancer. In RA, it was first reported by Haas et al. that macoecomonic between combined conventional syn- Cyr61 mRNA was strongly increased in lymphoblastoid thetic DMARDs and tumor necrosis factor (TNF) B cell lines derived from RA discordant monozygotic inhibitor in preventing relapse of RA. Active RA patients twins, being one of the three most overexpressed genes who met the inclusion and exclusion criteria in the [4]. An increasing number of studies found elevated ex- screening phase uniformly received therapy (methotrex- pression of Cyr61 protein in fibroblast-like synoviocytes ate and Yisaipu [recombinant human TNF receptor: Fc (FLS), synovial fluid, and peripheral blood mononuclear fusion protein, Guojian Pharmceutical company, China]) cells from RA patients [5, 6]. Preclinical studies discov- for 12 weeks. Those patients achieving clinical remission ered that Cyr61 played a pivotal role in IL-17-dependent or low disease activity (DAS28 < 3.2) at 12 weeks were proliferation of FLS in RA and blocking Cyr61 with the subsequently randomized into different treatment arms. specific neutralizing antibody ameliorated the inflamma- Paired serum samples were collected at the time before tion of the collagen-induced arthritis mice in vivo [7, 8]. (at baseline) and after 12 weeks’ therapy respectively, In addition, it was found that Cyr61 could not only while RA disease activity were also evaluated. After uni- stimulate IL-6 production by FLS via the Cyr61/αvβ5/ formed treatment for 12 weeks, each RA patient was cat- Akt/NF-κB signaling pathway [8], but also promote neu- egorized as ACR20 responders, ACR50 responders, trophil infiltration via upregulation of IL-8 production in ACR70 responders, and ACR non-responders. ACR re- RA-FLS [9]. A recent study demonstrated Cyr61 pro- sponse criteria are widely used to assess and establish moted vascular endothelial growth factor expression in the improvement in tender or swollen joint counts along osteoblasts through negative regulation of miR-126 via with improvement in three of the following five parame- the PKC-α signaling pathway and increased endothelial ters: Acute phase reactant, Patient assessment, Physician progenitor cell angiogenesis in RA [10]. But the role of assessment, Pain scale, and Disability/functional ques- Cyr61 is paradoxical. In inflammatory liver diseases, tionnaire. Achieving ACR20/50/70 means patients Cyr61 contributed to myeloid-derived suppressor cell ac- achieved at least 20/50/70% improvement in tender or tivation and expansion which served as negative regula- swollen joint counts, as well as 20/50/70% improvement tors of excessive T cell responses [11]. Moreover, Cyr61 in three of the other five parameters. And those RA pa- could strongly inhibit immune cell migration [12] and tients who achieve ACR 20/50/70 would have a signifi- process anti-osteoclastogenic [13] and anti-inflammatory cant decrease of disease activity. By using self-controlled properties [14]. It is noteworthy that emerging preclin- design, we are able to catch the dynamic changes of the ical studies have discussed the important roles of Cyr61 Cyr61 level along with RA disease activity. in RA pathogenesis, but there is little literature discuss- Subjects with conditions known to influence Cyr61 ing the clinical value of circulation Cyr61 in RA patients. levels were excluded, for instance, other autoimmune Therefore, in this study, we aim to look at the serum diseases, cancer, diabetes mellitus, infection, liver dis- Cyr61 levels in RA patients in comparison with healthy eases, and coronary heart diseases. Fan et al. Arthritis Research & Therapy (2019) 21:123 Page 3 of 9 This study was approved by the Institutional Medical Cyr61 was assessed by receiver operating characteristic Ethics Review Board of Peking University First Hospital (ROC) curves based on data from the training cohort, (2014-785), and all clinical investigations