Gerald H. Nabozny

Polymorphism of the MHC class II Eb gene determines the protection against collagen-induced arthritis

Collagen-induced arthritis (CIA) is an animal model of auto immune polyarthritis, sharing similarities with rheumatoid arthritis (RA). Paradoxally, susceptibility to mouse CIA is controlled by the H2A loci (DQ homologous) while RA is linked to HLA.DR genes (H2E homologous). We recently showed that the Eβd molecule prevents CIA development in susceptible H2q mice. We addressed the question of whether H2Eb polymorphism will influence CIA incidence as HLA.DRB1 polymorphism does in RA. In F1 mice, only H2Ebd and H2Ebs molecules showed protection. Using recombinant B10.RDD (Ebd/b) mice, we found that CIA protection was mediated by the first domain of the Eβd molecule. Using peptides covering the third hypervariable region of the Eβ chain, we found a perfect correlation between presentation of Eβ peptides by the H2Aq molecule and protection on CIA. Therefore, the mechanism by which H2Eb protects against CIA seems to rely on the affinity of Eβ peptides for the H2Aq molecule.

Nonsteroidal Dissociated Glucocorticoid Agonists Containing Azaindoles as Steroid A-Ring Mimetics

Syntheses and structure-activity relationships (SAR) of nonsteroidal glucocorticoid receptor (GR) agonists are described. These compounds contain azaindole moieties as A-ring mimetics and display various degrees of in vitro dissociation between gene transrepression and transactivation. Collagen induced arthritis studies in mouse have demonstrated that in vitro dissociated compounds (R)-16 and (R)-37 have steroid-like anti-inflammatory properties with improved metabolic side effect profiles, such as a reduced increase in body fat and serum insulin levels, compared to steroids.

The Immunogenetic Basis of Collagen Induced Arthritis in Mice: An Experimental Model for the Rational Design of Immunomodulatory Treatments of Rheumatoid Arthritis

Elucidating the genetic influences and immunological processes involved in the induction and pathogenesis of rheumatoid arthritis (RA) has been an area of intense investigation over the past two decades. During this period, various experimental animal models of RA have been studied which have contributed greatly to our basic understanding of this disease. One model of particular interest is the mouse model of collagen induced arthritis (CIA). Following the initial description of the model in rats,1 the successful induction of CIA in mice was achieved by Courtenay et al.2 CIA is usually induced by the intradermal injection of heterologous or homologous type II collagen (CII) in complete Freund’s adjuvant.3,4 Both T and B cell reactivity to CII is detected and, approximately 4–6 weeks post-immunization, mice develop a polyarthritis initially characterized by erythema and edema followed by joint distortion and, in some cases, culminating in joint ankylosis.5 Histologic examination of afflicted paws from mice with CIA reveals marked synovial proliferation, pannus formation and subsequent joint destruction with replacement by mononuclear cells.6 These clinical and histopathologic features of CIA strongly resemble those observed in human RA. Of particular importance has been the fact that the availability of numerous inbred mouse strains has enabled scientists to undertake a systematic and detailed analysis of the genetic factors contributing to this disease. Thus, understanding the immunogenetic characteristics of murine CIA strengthens the potential of this model for developing specific immune intervention strategies in human RA.

Influence of deletion of t cell receptor vβ genes on the theiler's virus model of multiple sclerosis

To determine the role of TCR V beta genes in a model of multiple sclerosis (MS), we studied Theilers virus infection in congenic mice with deletion of TCR V beta chromosome. Congenic mice expressing the V beta a [50% deletion of TCR V beta] or V beta c 70% deletion of TCR V beta] haplotype were generated in mice resistant [B10 (H-2b)], intermediate [B10.K (H-2k), B10.RIII (H-2r)] or susceptible [B10.S (H-2s), and B10.Q (H-2q)] to Theilers virus induced demyelination. Deletion of TCR V beta genes (V beta a or V beta c) did not convert B10 or B10.K congenic mice to susceptibility. In contrast, congenic B10.RIII-V beta c developed prominent demyelination and 10- to 100-fold increase in virus-antigen expression in spinal cord compared to B10.RIII mice. No effect on the extent of demyelination was observed in B10.S-V beta a, B10.S-V beta c or B10.Q-V beta c mice. These experiments illustrate the critical interactions between MHC, TCR, and background genes in susceptibility to immune-mediated disease.

An RORγt Oral Inhibitor Modulates IL-17 Responses in Peripheral Blood and Intestinal Mucosa of Crohn's Disease Patients

Background and Aims: Despite the negative results of blocking IL-17 in Crohns disease (CD) patients, selective modulation of Th17-dependent responses warrants further study. Inhibition of retinoic acid-related orphan receptor gamma (RORγt), the master regulator of the Th17 signature, is currently being explored in inflammatory diseases. Our aim was to determine the effect of a novel oral RORγt antagonist (BI119) in human CD and on an experimental model of intestinal inflammation. Methods: 51 CD patients and 11 healthy subjects were included. The effects of BI119 were tested on microbial-stimulated peripheral blood mononuclear cells (PBMCs), intestinal crypts and biopsies from CD patients. The ability of BI119 to prevent colitis in vivo was assessed in the CD4+CD45RBhigh T cell transfer model. Results: In bacterial antigen-stimulated PBMCs from CD patients, BI119 inhibits Th17-related genes and proteins, while upregulating Treg and preserving Th1 and Th2 signatures. Intestinal crypts cultured with supernatants from BI119-treated commensal-specific CD4+ T cells showed decreased expression of CXCL1, CXCL8 and CCL20. BI119 significantly reduced IL17 and IL26 transcription in colonic and ileal CD biopsies and did not affect IL22. BI119 has a more profound effect in ileal CD with additional significant downregulation of IL23R, CSF2, CXCL1, CXCL8, and S100A8, and upregulation of DEFA5. BI119 significantly prevented development of clinical, macroscopic and molecular markers of colitis in the T-cell transfer model. Conclusions: BI119 modulated CD-relevant Th17 signatures, including downregulation of IL23R while preserving mucosa-associated IL-22 responses, and abrogated experimental colitis. Our results provide support to the use of RORγt antagonists as a novel therapy to CD treatment.

BTK inhibition ameliorates kidney disease in spontaneous lupus nephritis

Lupus nephritis is a common disease manifestation of SLE, in which immune complex deposition and macrophage activation are important contributors to disease pathogenesis. Brutons tyrosine kinase (BTK) plays an important role in both B cell and FcgammaR mediated myeloid cell activation. In the current study, we examined the efficacy of BI-BTK-1, a recently described irreversible BTK inhibitor, in the classical NZB × NZW F1 (NZB/W) and MRL/lpr spontaneous mouse models of SLE. NZB/W mice were randomly assigned to a treatment (0.3 mg/kg, 1 mg/kg, 3 mg/kg and 10 mg/kg) or control group and began treatment at 22 weeks of age. The experimental setup was similar in MRL/lpr mice, but with a single treated (10 mg/kg, beginning at 8-9 weeks of age) and control group. A separate experiment was performed in the MRL/lpr strain to assess the ability of BI-BTK-1 to reverse established kidney disease. Early treatment with BI-BTK-1 significantly protected NZB/W and MRL/lpr mice from the development of proteinuria, correlating with significant renal histological protection, decreased anti-DNA titers, and increased survival in both strains. BI-BTK-1 treated mice displayed a significant decrease in nephritis-associated inflammatory mediators (e.g. LCN2 and IL-6) in the kidney, combined with a significant inhibition of immune cell infiltration and accumulation. Importantly, BI-BTK-1 treatment resulted in the reversal of established kidney disease. BTK inhibition significantly reduced total B cell numbers and all B cell subsets (immature, transitional, follicular, marginal zone, and class switched) in the spleen of NZB/W mice. Overall, the significant efficacy of BI-BTK-1 in ameliorating multiple pathological endpoints associated with kidney disease in two distinct murine models of spontaneous lupus nephritis provides a strong rationale for BTK inhibition as a promising treatment approach for lupus nephritis.

Collagen Arthritis in T Cell Receptor Congenic Mice

Identifying the genes responsible for susceptibility to human rheumatoid arthritis (RA) has been an area of intense investigation. Evidence showing that genetic factors play a role in RA susceptibility is certainly irrefutable (1) and the association between particular HLA class II alleles of the major histocompatibility complex (MHC) and RA is widely accepted (2). However, genetic analysis of RA patients and multiplex families indicates that MHC genes alone account for approximately 50% of RA heritability (3). Thus, additional non-MHC genes appear to play an important role in RA. Given the intimate association between the T cell receptor (TCR), antigenic peptide and MHC, a logical candidate which may also influence RA are the V alpha (VA) and V beta (VB) genes of the TCR. Analysis of the TCR repertoire from lymphocytes infiltrating RA synovium suggest, in some cases, biased VB gene utilization (4). Also, population studies indicate that certain VB polymorphisms may be more common in RA patients. Most recently, analysis of multiplex RA families suggested that a gene within or near the TCRB locus may confer RA susceptibility (5). Unfortunately, these findings, albeit compelling, cannot address the functional consequences of possessing such genetic markers. Therefore, understanding the mechanisms underlying a particular genetic association in RA may lead to the development of a highly specific therapy to treat this disease.