Circulating PR3-Specific B Cells in Patients with Active ANCA-Associated Vasculitis

Alvise Berti¹, Sophie Hillion ², Amber Hummel ¹, Eva Carmona ¹, Tobias Peikert ¹, Carol Langford ³, Peter Merkel ⁴, Paul Monach ⁵, Philip Seo ⁶, Robert Spiera ⁷, E William St. Clair ⁸, Fernando Fervenza ¹, Kristina Harris ⁹, John Stone ¹⁰, Jacques-Olivier Pers ¹¹, Peter Heeringa ¹², Wayel Abdulahad ¹³, Ulrich Specks ¹⁴ and Divi Cornec ¹¹, ¹Mayo Clinic, Rochester, MN, ²Université de Bretagne Occidentale, Brest, Bretagne, France, ³Cleveland Clinic, Cleveland, OH, ⁴University of Pennsylvania, Philadelphia, PA, ⁵Brigham and Women's Hospital, Boston, MA, ⁶Johns Hopkins Medicine, Baltimore, MD, ⁷Hospital for Special Surgery, New York, NY, ⁸Duke, Raleigh, ⁹Immune Tolerance Network, Bethesda, ¹⁰Massachusetts General Hospital Rheumatology Unit, Harvard Medical School, Boston, MA, ¹¹Universite de Bretagne Occidentale, Brest, Bretagne, France, ¹²Department of Pathology and Medical Biology, University of Groningen, University Medical Center Groningen, Groningen, ¹³Department of Rheumatology and Clinical Immunology, University of Groningen, University Medical Center Groningen, Groningen, ¹⁴Mayo Clinic College of Medicine, Rochester, MN

Meeting: 2019 ACR/ARP Annual Meeting

Keywords: B cell, systemic vasculitides and auto-immunity

Session Information

Date: Monday, November 11, 2019

Title: B Cell Biology & Targets In Autoimmune & Inflammatory Disease Poster

Session Type: Poster Session (Monday)

Session Time: 9:00AM-11:00AM

Background/Purpose: We previously developed a method to recognize proteinase 3 (PR3)-specific B cells by flow cytometry. We reported that, in a small number of subjects, the proportion of PR3-specific B cells was higher in patients with PR3-ANCA-associated vasculitis (AAV) compared to healthy controls (HCs) (Cornec et al, J Autoimmunity 2017). The objectives of this study were to replicate our previous findings on a larger number of subjects, including anti-myeloperoxidase (MPO)-AAV patients as disease controls, and to study the association of PR3-specific B cell proportions with clinical and biological features in patients with PR3-AAV.

Methods: We analyzed samples from 161 patients who participated in the RAVE trial, including 110 patients with PR3-AAV and 51patients with MPO-AAV, as well as 27 HCs. We measured the proportion of PR3-specific B cells and subsets among cryopreserved PBMCs using a multi-color flow cytometry panel including CD19, IgD, CD27, CD38, CD24, and biotinylated PR3. SPADE (Spanning-tree Progression Analysis of Density-normalized Events) algorithm was used for the unsupervised clustering of flow cytometry data, based on the level of expression of each of the 6 markers on each cell.

Results: The proportion of PR3-specific B cells was higher in patients with PR3-AAV and MPO-AAV (median [25-75% interquartile range]: 2.82% [2.35-3.74] and 2.87 [2.15-3.84], respectively) compared to HCs (1.51 [1.25-1.73], both p< 0.001), while they did not differ substantially between PR3-AAV and MPO-AAV (p=0.517). We observed a significant shift towards a more mature phenotype of these PR3-specific B cells in patients with PR3-AAV compared to MPO-AAV and HCs, as represented by a higher proportion of PR3-specific B cells among the unswitched memory B-cell subsets (IgD+CD27+; 6.14% [4.27-8.74] versus 4.47% [3.11-6.87] versus 2.89% [2.19-4.23], p< 0.01 for all comparisons), switched memory B-cell subsets (IgD-CD27+; 2.98% [1.67-5.00] versus 2.1% [1.38-3.09] versus 1.19% [0.96-1.60], p< 0.001 for all comparisons), and double negative B-cell subsets (IgD-CD27-; 2.23% [1.54-3.76] versus 1.44% [0.83-2.53] versus 0.80% [0.00-1.34], p< 0.01 for all comparisons). In patients with PR3-AAV, no associations between PR3-specific B cell proportion and age, sex, new diagnosis vs relapsing disease, or disease activity (BVAS/WG) was observed. The SPADE clustering resulted in the definition of 200 distinct subsets detectable in each sample with variable proportions, and showed a strong segregation of patients and HCs, and PR3-AAV and MPO-AAV patients. A cluster belonging to the switched memory subset was PR3-reactive only in PR3-AAV (figure).

Conclusion: Circulating PR3-specific B cells can be detected in the peripheral blood. The proportion of PR3-specific B cells was higher in patients than in HCs. Interestingly, PR3-specific B cell repartition is shifted towards a memory phenotype in PR3-AAV patients. Further studies are ongoing to characterize the dynamics of the PR3-specific B cells after therapy, and to understand the differences of these autoreactive B cells between patients with PR3-AAV and HCs.

SPADE analysis of B-cell subsets. PR3-reactive subsets are represented in red. One post-switch memory B-cell subset is PR3-reactive only in PR3-AAV patients.

Disclosure: A. Berti, None; S. Hillion, None; A. Hummel, None; E. Carmona, None; T. Peikert, None; C. Langford, Bristol-Myers Squibb, 2, GlaxoSmithKline,, 2, ChemoCentryx, 2, Genentech, 2, Bristol-Myers Squibb, 5, 9, Abbvie, 9, AstraZeneca, 9; P. Merkel, AbbVie, 5, AstraZeneca, 2, 5, Biogen, 5, Boeringher-Ingelheim, 2, 5, Bristol-Myers Squibb, 2, 5, Celgene, 2, 5, ChemoCentryx, 2, 5, CSL Behring, 5, Genentech/Roche, 2, 5, Genzyme/Sanofi, 5, GlaxoSmithKline, 2, 5, InflaRx, 5, Insmed, 5, Jannsen, 5, Kiniksa, 5, Kypha, 2, TerumoBCT, 2, UpToDate, 7; P. Monach, None; P. Seo, None; R. Spiera, BMS, 2, Boehringer Ingelheim, 2, Bristol-Myers Squibb, 2, ChemoCentryx, 2, 5, Chemocentryx, 2, Corbus, 2, CSL Behring, 5, Cytori, 2, Formation Biologics, 2, Genentech, Inc., 2, Genzyme/Sanofi, 2, 5, GlaxoSmithKline, 2, 5, GSK, 2, 5, Hoffman-La Roche Ltd, 2, Janssen, 5, Mitsubishi, 5, Roche, 2, Roche Genetech, 2, 5, Roche/Genetech, 2, 5, Roche-Genentech, 2, 5, Roche-Genetech, 2, 5, Sanofi, 5, Sanofi-Aventis, 5; E. St. Clair, None; F. Fervenza, None; K. Harris, None; J. Stone, Genentech, 2, 5, Roche, 2, 5, Xencor, 2, 5; J. Pers, None; P. Heeringa, None; W. Abdulahad, None; U. Specks, None; D. Cornec, None.

To cite this abstract in AMA style:

Berti A, Hillion S, Hummel A, Carmona E, Peikert T, Langford C, Merkel P, Monach P, Seo P, Spiera R, St. Clair E, Fervenza F, Harris K, Stone J, Pers J, Heeringa P, Abdulahad W, Specks U, Cornec D. Circulating PR3-Specific B Cells in Patients with Active ANCA-Associated Vasculitis [abstract]. Arthritis Rheumatol. 2019; 71 (suppl 10). https://acrabstracts.org/abstract/circulating-pr3-specific-b-cells-in-patients-with-active-anca-associated-vasculitis/. Accessed .

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