IMMUNOPATHOLOGY The significance of ANCA positivity in patients with inflammatory bowel disease WEI-I. LEE 1,2 ,KAVITHA SUBRAMANIAM 3,4 ,CAROLYN A. HAWKINS 1,2,4 , KATRINA L. RANDALL 1,2,4 1 Department of Immunology, Canberra Hospital, ACT, Australia; 2 Department of Immunopathology and Infectious Immunoassay, ACT Pathology, ACT, Australia; 3 Gastroenterology and Hepatology Unit, Canberra Hospital, ACT, Australia; 4 Australian National University Medical School, Australian National University, ACT, Australia Summary Traditionally anti-neutrophil cytoplasmic antibodies (ANCA) are used to subtype patients with inflammatory bowel disease (IBD) and to predict primary sclerosing cholangitis (PSC). The clinical utility of this testing in the Australian context is not known. Our retrospective, cross- sectional study looked at the results of ANCA testing performed during routine clinical review and aimed to retrospectively review (1) the distribution of different ANCA subtypes for IBD patients, (2) the temporal change of ANCA status, and (3) the predictive value of ANCA for PSC. Sixty-four IBD patients attending our hospital gastroenterology clinic between 2012 and 2016 had at least one ANCA test requested. Surprisingly, >80% of the IBD patients in our cohort who underwent ANCA testing had a positive ANCA result and a significant proportion had positive PR3 antibodies. However, no specific ANCA pattern predicted a specific IBD subtype or clinical course. Pairing ANCA and anti-Saccharomyces cerevi- siae (ASCA) did not add value in subtyping IBD for these patients. Our study suggests that there is little value in ordering an ANCA for patients with IBD. Key words: ANCA; MPO; PR3; inflammatory bowel disease. Received 21 November 2018, revised 11 July, accepted 18 July 2019 Available online: xxx INTRODUCTION Anti-neutrophil cytoplasmic antibodies (ANCA) were initially discovered as a useful marker for systemic vascu- litides such as granulomatous with polyangiitis (GPA) or microscopic polyangiitis (MPA) in the 1980s. 1,2 In these diseases, the primary antigens that are recognised are pro- teinase 3 (PR3) and myeloperoxidase (MPO). 3 The Interna- tional Consensus Statement on Testing and Reporting of ANCA published in 1999 recommended that indirect immunofluorescence (IIF) be performed on all ANCA re- quests, with further testing for those with typical or atypical fluorescent patterns with enzyme-linked immunosorbent assays (ELISAs) that detect PR3 or MPO antibodies. 3 In the 1990s it was noted that a significant proportion of patients with inflammatory bowel disease (IBD) had positive ANCA on IIF. This was more prevalent in patients with ul- cerative colitis (UC) than patients with Crohn’s disease (CD) as reviewed by Bossuyt. 4 Furthermore, anti-Saccharomyces cerevisiae (ASCA) was found to be highly specific for CD. 5,6 This led to the conventional teaching that paired ANCA and ASCA might be helpful in subtyping UC from CD. 4,7 As a significant proportion of patients with both inflammatory bowel disease and primary sclerosing cholangitis (PSC) are ANCA positive, ANCA status had been said to be of pre- dictive value for PSC. 6,8 As can be seen from Table 1, the prevalence of ANCA and MPO/PR3 in primary studies in inflammatory bowel disease varies significantly across different studies (also reviewed in Bossuyt). 4 This may be secondary to differences in study population age, ethnicity, inter-assay and inter-laboratory differences. Furthermore, the distinction between atypical and typical perinuclear ANCA (p-ANCA) was not routinely made until the 2000s. 9 As a result, studies in the 1990s reported p-ANCA as the dominant ANCA pattern in IBD patients with ulcerative colitis, whereas studies in the 2000s identified atypical p-ANCA as the dominant ANCA pattern in these patients. 4,8,10 – 12 While it is generally acknowledged that patients with IBD may have a positive ANCA by immunofluorescence, a sig- nificant number have also been found to be MPO and PR3 positive (Table 1); however, little is known about the sig- nificance of these observations. Do these patients have a concurrent vasculitis or do they have a different disease course for their IBD from those who are MPO and PR3 negative? As the association between IBD and MPO and PR3 positivity has not been reviewed from the Australian context, the aim of our study was to look at the characteristics of patients with both IBD and a positive ANCA, in particular the rate of MPO/PR3 positivity amongst the ANCA positive patients, to see whether there appeared to be any clinical correlations in this subgroup. MATERIAL AND METHODS Patients who attended the IBD clinic at the Canberra Hospital between 2012 and 2016 and had ANCA ordered during the same time frame were included. Demographic information, and clinical characteristics [including type of IBD, all ANCA testing (not just in the 4 year interval), ASCA testing, extra- intestinal manifestations, medication regime, response to treatment, and surgical history] were retrieved retrospectively from the medical records for these patients. This retrospective analysis was approved by the ACT Health Low Risk Ethics Committee (ETHLR.17.124). The patient’s previous ANCA testing was retrieved from the laboratory in- formation system. In the testing laboratory, IIF is performed using the Print ISSN 0031-3025/Online ISSN 1465-3931 © 2019 Published by Elsevier B.V. on behalf of Royal College of Pathologists of Australasia. DOI: https://doi.org/10.1016/j.pathol.2019.07.002 Pathology (- xxxx) xxx(xxx), xxx Please cite this article as: Lee W-I et al., The significance of ANCA positivity in patients with inflammatory bowel disease, Pathology, https://doi.org/10.1016/ j.pathol.2019.07.002