Brensocatib (an oral, reversible inhibitor of dipeptidyl peptidase-1) attenuates disease progression in two animal models of rheumatoid arthritis

Abstract

Rheumatoid arthritis (RA) is a painful and incurable disease characterized by chronic joint inflammation and a progressive destruction of cartilage and bone. Although current treatments have improved clinical outcomes for some patients, the high relapse rates and sizeable proportion of non-responders emphasize the need for further research. Arthritic joints are massively infiltrated by neutrophils, which influence inflammatory and immune processes by releasing cytokines, chemokines, eicosanoids, and neutrophil serine proteases (NSPs) - all of which are known to contribute to RA initiation and progression. Active NSPs are generated from zymogens at the promyelocytic stage of neutrophil differentiation under the action of dipeptidyl peptidase 1 (DPP-1) and DPP-1 knockout mice are resistant to the development of arthritis. Thus, DPP-1 inhibition represents a promising therapeutic approach in RA. In this study, we assessed the efficacy of a potent and highly selective DPP-1 inhibitor, brensocatib, in two well established RA models - rat collagen-induced arthritis (CIA) and mouse collagen antibody-induced arthritis (CAIA). In both models, brensocatib at 3 and 30 mg/kg/day significantly reduced bone marrow NSP levels, in keeping with prior pharmacodynamic studies in rodents. More importantly, brensocatib treatment significantly improved disease score at both dosages in both rodent models. In the mouse CAIA model, brensocatib even proved at least as potent as anti-TNF antibodies in diminishing both the histopathological score and neutrophil infiltration into arthritic joints. Together, these results show that brensocatib alters RA disease progression in rodents and supports the need for its further evaluation as a potential therapeutic option, or to complement existing RA treatments.

Keywords: DPP-1; cathepsin C; inflammation; migration; neutrophil serine proteases; neutrophils.

Figure 1

Effect of brensocatib on classical DPP-1 substrates in two animal models. (A) Rats were treated in the CIA model and samples were analyzed as described in Methods. Briefly, rats were immunized twice with a collagen emulsion to induce arthritis or with vehicle (non-disease group). In animals developing arthritis, brensocatib was given orally at 0.3, 3.0 and 30 mg/kg/day; as a control, a group was given dexamethasone (“Dex”) orally once daily at 0.3 mg/kg. Bone marrow was isolated on day 30 or 31 and NSP activities were assessed (NE, neutrophil elastase; PR3, proteinase 3; CatG, cathepsin G). Mean ± SEM of at least 10 animals per group. (B) Mice were treated in the CAIA model and samples were analyzed as described in Methods. Briefly, mice were immunized on day 0 with i.v. injection of a collagen antibody cocktail or with vehicle (non-disease group). On day 3, all mice that received the collagen antibodies also received an injection of 25 μg LPS i.p.; non-disease control animals received an injection of vehicle instead. Vehicle control or brensocatib at 3 or 30 mg/kg/day was given orally from day -10 to day 1 BID, and from day 2 to day 20 QD. Anti-TNFα (“aTNF”) was administered 3x/week at 5 mg/kg, starting on day 0 and continued through day 21. Bone marrow was isolated on day day 21 and NSP activities were assessed. Mean ± SEM of at least 8 animals per group, except for non-diseased mice (5 animals). Statistical differences in this figure were determined using Kruskal-Wallis test with Dunn’s multiple comparisons post-hoc test. *p< 0.05; **p< 0.01; ***p< 0.001, relative to diseased animals receiving vehicle only.

Figure 2

Effect of brensocatib on body weight and disease parameters in the rat CIA model. Rats were treated as described in Figure 1 for arthritis induction and inhibitor administration. The following parameters were monitored: (A) body weight; (B) paw thickness; (C) clinical score (sum of the two hind paws). Mean ± SEM of at least 10 animals per group. Statistical differences were determined using multiple t test comparisons and a false discovery rate of 1% as determined by the two-stage set-up method of Benjamini, Krieger, and Yekutieli. *p< 0.05; **p< 0.01 relative to diseased animals receiving vehicle only.

Figure 3

Effect of brensocatib on body weight and disease parameters in the mouse CAIA model. Mice were treated as described in Figure 1 for arthritis induction and inhibitor administration. The following parameters were monitored: (A) body weight; (B) paw thickness; (C) clinical score (sum of the two hind paws). Mean ± SEM of 12 animals per group, except for non-diseased mice (5 animals). Statistical differences were determined using multiple t test comparisons and a false discovery rate of 1% as determined by the two-stage set-up method of Benjamini, Krieger, and Yekutieli. *p< 0.05; **p< 0.01; ***p< 0.001 relative to diseased animals receiving vehicle only.

Figure 4

Effect of brensocatib on histopathology parameters in the mouse CAIA model. Mice were treated as described in Figure 1 for arthritis induction and inhibitor administration. (A-D) Various histopathological scores were assessed on day 21; a summed score was also compiled (E). Mean ± SEM of at least 10 animals per group, except for non-diseased mice (5 animals). Statistical differences were determined using Kruskal-Wallis test with Dunn’s multiple comparisons post-hoc test. *p< 0.05; **p< 0.01 relative to diseased animals receiving vehicle only.

Figure 5

Effect of brensocatib on neutrophil infiltration in the mouse CAIA model. Mice were treated as described in Figure 1 for arthritis induction and inhibitor administration and neutrophil infiltration was assessed by immunohistochemistry on day 21, using either (A) myeloperoxidase (MPO); or (B) Ly6G as readouts. Mean ± SEM of at least 10 animals per group, except for non-diseased mice (5 animals). Statistical differences were determined using Kruskal-Wallis test with Dunn’s multiple comparisons post-hoc test. *p< 0.05 relative to diseased animals receiving vehicle only.