Abstract
Background
Anti-mitochondrial autoantibodies (AMA) detected by indirect immunofluorescence (IIF) on rodent tissues are the diagnostic marker of primary biliary cholangitis (PBC). However, up to 15% of patients with PBC are AMA-negative by IIF. In the effort to close the serological gap and improve the diagnostic sensitivity of PBC testing, recently, novel autoantibodies specific for PBC, such as kelch-like 12 (KLHL12, KLp epitope) and hexokinase 1 (HK1) have been described. In this study, we evaluated the autoantibody profile in a large cohort of PBC patients and in patients with other liver disease, including anti-HK1 and anti-KLp autoantibodies.
Methods
Sera of 194 PBC patients (126 AMA-IIF-positive and 68 AMA-IIF-negative) and 138 disease controls were tested for a panel of PBC-specific antibodies (MIT3, sp100, gp210, HK1, KLp) using a new automated particle-based multi-analyte technology (PMAT) assay on the Aptiva instrument (Inova).
Results
Selecting a cutoff yielding a specificity of >95% for all the markers, the sensitivity for anti-MIT3, anti-sp100, anti-gp210, anti-HK1 and anti-KLp in the PBC AMA-IIF-negative cohort was 20.6%, 16.2%, 23.5%, 22.0%, 17.6 and 13.2%, respectively. Six out of the 68 (8.8%) AMA-IIF negative sera were positive for anti-HK1 or anti-KLp alone. Using these new markers in addition to anti-MIT3, anti-sp100 and anti-gp210, the overall sensitivity in this cohort of AMA-IIF-negative patients increased from 53% to 61.8%, reducing the serological gap in AMA-negative PBC patients.
Conclusions
PBC antibody profiling, made possible by the new Aptiva-PMAT technology, allows recognition of a higher number of AMA-negative PBC patients than conventional immunoassays and may represent a useful tool to evaluate the prognostic significance of autoantibody association in PBC patients.
Acknowledgments
The authors thank Sandro Sulfaro from the Anatomic Pathology Department of the S. Maria degli Angeli Hospital, Pordenone, Italy, for providing the microphotographs of liver histology.
Author contributions: All the authors have accepted responsibility for the entire content of this submitted manuscript and approved submission.
Research funding: None declared.
Employment or leadership: Andrea Seaman, Marychel Tiongson, Charles Warren, Chelsea Bentow, Gary L. Norman and Michael Mahler are employees of Inova Diagnostics Inc.
Honorarium: None declared.
Competing interests: Inova Diagnostics Inc. provided financial sponsorship for commercial assay costs. Employees of these companies participated as coauthors of the manuscript by contributing to the study design, the acquisition, analysis, and interpretation of data, and the manuscript preparation. The authors independently collected the data, interpreted the results, and had the final decision to submit the manuscript for publication. However, publication of this article was not contingent upon approval by the company.
References
1. Kaplan MM, Gershwin ME. Primary biliary cirrhosis. N Engl J Med 2005;353:1261–73.10.1056/NEJMra043898Search in Google Scholar
2. Walker JG, Doniach D, Roitt IM, Sherlock S. Serological tests in diagnosis of primary biliary cirrhosis. Lancet 1965;1:827–31.10.1016/S0140-6736(65)91372-3Search in Google Scholar
3. Achenza MI, Meda F, Brunetta E, Selmi C. Serum autoantibodies for the diagnosis and management of autoimmune liver diseases. Exp Rev Gastroenterol Hepatol 2012;6:717–29.10.1586/egh.12.58Search in Google Scholar PubMed
4. Lindor KD, Gershwin ME, Poupon R, Kaplan M, Bergasa NV, Heathcote EJ. Primary biliary cirrhosis. Hepatology 2009;50:201–308.10.1002/hep.22906Search in Google Scholar PubMed
5. EASL. Clinical practice guidelines: management of cholestatic liver diseases. J Hepatol 2009;13:441–4.Search in Google Scholar
6. EASL. Clinical practice guidelines: The diagnosis and management of patients with primary biliary cholangitis. J Hepatol 2017;67:145–72.10.1016/j.jhep.2017.03.022Search in Google Scholar PubMed
7. Muratori P, Muratori L, Gershwin ME, Czaja AJ, Pappas G, MacCariello S, et al. True antimitochondrial antibody-negative primary biliary cirrhosis, low sensitivity of the routine assay, or both? Clin Exp Immunol 2004;135:154–8.10.1111/j.1365-2249.2004.02332.xSearch in Google Scholar PubMed PubMed Central
8. Invernizzi P, Selmi C, Ranftler C, Podda M, Wesierska-Gadek J. Antinuclear antibodies in primary biliary cirrhosis. Semin Liver Dis 2005;25:298–310.10.1055/s-2005-916321Search in Google Scholar PubMed
9. Muratori P, Muratori. Autoantibodies in antimitochondrial antibody negative primary biliary cholangitis. Clin Res Hepatol Gastroenterol 2017;41:e47.10.1016/j.clinre.2017.04.016Search in Google Scholar PubMed
10. Gershwin ME, Mackay IR, Sturgess A, Coppel RL. Identification and specificity of a cDNA encoding the 70kd mitochondrial antigen recognized in primay biliary cirrhosis. J Immunol 1987;138:3525–31.10.4049/jimmunol.138.10.3525Search in Google Scholar
11. Leung PS, Choi J, Yang G, Woo E, Kenny TP, Gershwin ME. A contemporary perspective on the molecular characteristics of mitochondrial autoantigens and diagnosis in primary biliary cholangitis. Expert Rev Mol Diagn 2016;16:697–705.10.1586/14737159.2016.1164038Search in Google Scholar PubMed PubMed Central
12. Bassendine MF, Jones DE, Yeaman SJ. Biochemistry and autoimmune response to the 2-oxoacid dehydrogenase complexes in primary biliary cirrhosis. Semin Liver Dis 1997;17:49–60.10.1055/s-2007-1007182Search in Google Scholar PubMed
13. Miyakawa H, Tanaka A, Kikuchi K, Matsushita M, Kitazawa E, Kawaguchi N, et al. Detection of antimitochondrial autoantibodies in immunofluorescent AMA-negative patients with primary biliary cirrhosis using recombinant autoantigens. Hepatology 2001;34:243–8.10.1053/jhep.2001.26514Search in Google Scholar PubMed
14. Gabeta S, Norman GL, Liaskos C, Papamichalis PA, Zografos T, Garagounis A, et al. Diagnostic relevance and clinical significance of the new enhanced performance M3 (MIT3) ELISA for the detection of IgA and IgG antimitochondrial antibodies in primary biliary cirrhosis. J Clin Immunol 2007;27:378–87.10.1007/s10875-007-9092-0Search in Google Scholar PubMed
15. Dähnrich C, Pares A, Caballeria I, Rosemann A, Schlumberger W, Probst C, et al. New ELISA for detecting primary biliary cirrhosis-specific antimitochondrial antibodies. Clin Chem 2009;55:978–85.10.1373/clinchem.2008.118299Search in Google Scholar PubMed
16. Leung PS, Coppel RI, Ansari A, Munoz S, Gershwin ME. Antimitochondrial antibodies in primary biliary cirrhosis. Semin Liver Dis 1997;17:61–9.10.1055/s-2007-1007183Search in Google Scholar PubMed
17. Bizzaro N, Covini G, Rosina F, Muratori P, Tonutti E, Villalta D, et al. Overcoming a “probable” diagnosis in antimitochondrial antibody negative primary biliary cirrhosis: a study of 100 sera and review of the literature. Clin Rev Allergy Immunol 2012;42:288–97.10.1007/s12016-010-8234-ySearch in Google Scholar PubMed
18. Villalta D, Sorrentino MC, Girolami E, Tampoia M, Alessio MG, Brusca I, et al. Autoantibody profiling of patients with primary biliary cirrhosis using a multiplexed line-blot assay. Clin Chim Acta 2015;438:135–8.10.1016/j.cca.2014.08.024Search in Google Scholar PubMed
19. Zhang Q, Liu Z, Wu S, Duan W, Chen S, Ou X, et al. Meta-analysis of antinuclear antibodies in the diagnosis of antimitochondrial antibody-negative primary biliary cholangitis. Gastroenterol Res Pract 2019;2019:8959103. Doi: 10.1155/2019/8959103. eCollection 2019.Search in Google Scholar PubMed PubMed Central
20. Hu CJ, Song G, Huang W, Liu GZ, Deng CW, Zeng HP, et al. Identification of new autoantigens for primary biliary cirrhosis using human proteome microarrays. Mol Cell Proteomics 2012;11:669–80.10.1074/mcp.M111.015529Search in Google Scholar PubMed PubMed Central
21. Norman GL, Yang CY, Ostendorff HP, Shums Z, Lim MJ, Wang J, et al. Anti-kelch-like and anti-hexokinase 1: novel autoantibodies in primary biliary cirrhosis. Liver Int 2015;35:642–51.10.1111/liv.12690Search in Google Scholar PubMed PubMed Central
22. Bombaci M, Pesce E, Torri A, Carpi D, Crosti M, Cordiglieri C, et al. Novel biomarkers for primary biliary cholangitis to improve diagnosis and understand underlying regulatory mechanisms. Liver Int 2019;39:2124–35.10.1111/liv.14128Search in Google Scholar PubMed
23. Norman GL, Shums Z, Encabo S, Milo J, Mahler M, Swain MG, et al. Examination of Canadian primary biliary cholangitis (PBC) patients by conventional and novel-PBC-specific biomarkers. Hepatol 2016;64 (suppl 1):383Search in Google Scholar
24. Hennes EM, Zeniya M, Czaja AJ, Parés A, Dalekos JN, Krawitt EL, et al. Simplified criteria for the diagnosis of autoimmune hepatitis. Hepatology 2008;48:169–76.10.1002/hep.22322Search in Google Scholar PubMed
25. Yiman KK, Bowlus CI. Diagnosis and classification of primary sclerosing cholangitis. Autoimmune Rev 2014;13:445–50.10.1016/j.autrev.2014.01.040Search in Google Scholar PubMed
26. Richards M, Garcia-De La Torre I, Gonzalez-Bello YC, Vazquez-Del MM, Andrade-Ortega L, Medrano-Ramirez G, et al. Autoantibodies to Mi-2 alpha and Mi-2 beta in patients with idiopathic inflammatory myopathy. Rheumatology 2019;76:867–8.10.1093/rheumatology/kez092Search in Google Scholar PubMed
27. Milkiewicz M, Caballeria I, Smyk DS, Milkiewicz P. Predicting and preventing autoimmunity: the case of anti-mitochondrial antibodies. Auto Immun Highlights 2012;3:105–12.10.1007/s13317-012-0038-zSearch in Google Scholar PubMed PubMed Central
28. Liu H, Norman GL, Shums Z, Worman HJ, Krawitt EL, Bizzaro N, et al. PBC screen: an IgG/IgA dual isotype ELISA detecting multiple mitochondrial and nuclear autoantibodies specific from primary biliary cirrhosis. J Autoimmun 2010;35:346–2.10.1016/j.jaut.2010.09.005Search in Google Scholar PubMed
29. Robey RB, Hay N. Mitochondrial hexokinases: guardians of the mitochondria. Cell Cycle 2005;4:654–8.10.4161/cc.4.5.1678Search in Google Scholar PubMed
30. Gupta VA, Beggs AH. Kelch proteins: emerging roles in skeletal muscle development and diseases. Skeletal Muscle 2014;4:11–2.10.1186/2044-5040-4-11Search in Google Scholar PubMed PubMed Central
31. Dhanoa BS, Cogliati T, Satish AG, Bruford EA, Friedman JS. Update on the Kelch-like (KLHL) gene family. Hum Genomics 2013;7:13.10.1186/1479-7364-7-13Search in Google Scholar PubMed PubMed Central
32. Hu SL, Zhao FR, Hu Q, Chen WX. Meta-analysis assessment of gp210 and Sp100 for the diagnosis of primary biliary cirrhosis. PLoS One 2014;9:e101916.10.1371/journal.pone.0101916Search in Google Scholar PubMed PubMed Central
33. Norman GL, Reig A, Viñas O, Mahler M, Wunsch E, Milkiewicz P, et al. The prevalence of anti-hexokinase-1 and anti-kelch-like 12 peptide antibodies in patients with primary biliary cholangitis is similar in Europe and North America: a large international, multi-center study. Front Immunol 2019;10:662.10.3389/fimmu.2019.00662Search in Google Scholar PubMed PubMed Central
34. Reig A, Garcia M, Shums Z, Milo J, Encabo S, Bentow C, et al. The novel hexokinase (HK-1) antibodies are useful for diagnosis and associated with bad prognosis in primary biliary cholangitis. J Hepatol 2017;(Suppl 1):S355–6.10.1016/S0168-8278(17)31049-8Search in Google Scholar
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