An anti-TNF-glucocorticoid receptor modulator antibody-drug conjugate is efficacious against immune-mediated inflammatory diseases.

Glucocorticoids (GCs) are efficacious drugs used for treating many inflammatory diseases, but the dose and duration of administration are limited because of severe side effects. We therefore sought to identify an approach to selectively target GCs to inflamed tissue. Previous work identified that anti-tumor necrosis factor (TNF) antibodies that bind to transmembrane TNF undergo internalization; therefore, an anti-TNF antibody-drug conjugate (ADC) would be mechanistically similar, where lysosomal catabolism could release a GC receptor modulator (GRM) payload to dampen immune cell activity.

Design and Development of Glucocorticoid Receptor Modulators as Immunology Antibody-Drug Conjugate Payloads.

Glucocorticoid receptor modulators (GRM) are the first-line treatment for many immune diseases, but unwanted side effects restrict chronic dosing. However, targeted delivery of a GRM payload via an immunology antibody-drug conjugate (iADC) may deliver significant efficacy at doses that do not lead to unwanted side effects. We initiated our α-TNF-GRM ADC project focusing on identifying the optimal payload and a linker that afforded stable attachment to both the payload and antibody, resulting in the identification of the synthetically accessible maleimide-Gly-Ala-Ala linker.

Dual Blockade of TNF and IL-17A Inhibits Inflammation and Structural Damage in a Rat Model of Spondyloarthritis.

The tumor necrosis factor (TNF) and IL-23/IL-17 axes are the main therapeutic targets in spondyloarthritis. Despite the clinical efficacy of blocking either pathway, monotherapy does not induce remission in all patients and its effect on new bone formation remains unclear. We aimed to study the effect of TNF and IL-17A dual inhibition on clinical disease and structural damage using the HLA-B27/human β2-microglobulin transgenic rat model of SpA.

Continuous IL-23 stimulation drives ILC3 depletion in the upper GI tract and, in combination with TNFα, induces robust activation and a phenotypic switch of ILC3.

Mutations in the Interleukin (IL)-23/IL-23 receptor loci are associated with increased inflammatory bowel disease (IBD) susceptibility, and IL-23 neutralization has shown efficacy in early clinical trials. To better understand how an excess of IL-23 affects the gastrointestinal tract, we investigated chronic systemic IL-23 exposure in healthy wildtype mice. As expected, IL-23 exposure resulted in early activation of intestinal type 3 innate lymphoid cells (ILC3), followed by infiltration of activated RORγt+ T helper cells.

A-770041, a novel and selective small-molecule inhibitor of Lck, prevents heart allograft rejection.

Lck, one of eight members of the Src family of tyrosine kinases, is activated after T cell stimulation and is required for T-cell proliferation and interleukin (IL)-2 production. Inhibition of Lck has been a target to prevent lymphocyte activation and acute rejection. Here, we report the pharmacologic characterization of 1-methyl-1H-indole-2-carboxylic acid (4-{1-[4-(4-acetyl-piperazin-l-yl)-cyclohexyl]-4-amino-1H-pyrazolo[3,4-d]pyrimidin-3-yl}-2-methoxy-phenyl)-amide (A-770041), an orally bioavailable pyrazolo[3,4-d]pyrimidine with increased selectivity for Lck compared with previously reported compounds.

A-420983: a potent, orally active inhibitor of lck with efficacy in a model of transplant rejection.

We have identified the pyrazolo[3,4-d]pyrimidine A-420983 (compound 7) as a potent inhibitor of lck. A-420983 exhibits oral efficacy in animal models of delayed-type hypersensitivity and organ transplant rejection.

Immunolocalization analysis of transforming growth factor-beta1 in the growth plates of broiler chickens with high and low incidences of tibial dyschondroplasia.

Immunolocalization of transforming growth factor-beta1 (TGF-beta1) was determined in growth plates of two lines of broiler chickens with low and high incidences of tibial dyschondroplasia (TD). Ultrathin sections of growth plates from each line were treated with a polyclonal antibody specific for TGF-beta1, followed by colloidal gold-labeled protein A. Immunolocalization for TGF-beta1 was observed in chondrocytes of all zones of growth plates of low and high TD incidence lines.

Experimental granulomatous colitis in mice is abrogated by induction of TGF-beta-mediated oral tolerance.

In previous studies we showed that a chronic colitis associated with a Th1 T cell response can be induced by the rectal administration of the haptenizing reagent 2,4,6-trinitrobenzene sulfonic acid (TNBS). We report here that oral administration of haptenized colonic proteins (HCP) before rectal administration of TNBS effectively suppresses the ability of the latter to induce colitis. This suppression (oral tolerance) appears to be due to the generation of mucosal T cells producing TGF-beta and Th2-type cytokines after oral HCP administration.

Transforming growth factor-beta inhibits interleukin-12-induced production of interferon-gamma by natural killer cells: a role for transforming growth factor-beta in the regulation of T cell-independent resistance to Toxoplasma gondii.

Severe-combined immune deficient (SCID) mice have been found to resist infection with the intracellular protozoan parasite Toxoplasma gondii via interleukin (IL)-12 stimulation of interferon (IFN)-gamma production by natural killer (NK) cells. Previously, we demonstrated the presence of increased levels of transcripts for transforming growth factor-beta (TGF-beta) in the brains and lungs of SCID mice infected with T. gondii, leading us to investigate the role of TGF-beta in the mechanism of resistance to T.

Spontaneous elaboration of transforming growth factor beta suppresses host defense against bacterial infection in autoimmune MRL/lpr mice.

Infection with gram-negative and gram-positive bacteria remains a leading cause of death in patients with systemic lupus erythematosis (SLE), even in the absence of immunosuppressive therapy. To elucidate the mechanisms that underly the increased risk of infection observed in patients with systemic autoimmunity, we have investigated host defense against bacterial infection in a murine model of autoimmunity, the MRL/Mp-lpr/lpr (MRL/lpr) mouse. Our previous study implicated transforming growth factor beta (TGF-beta) in a novel acquired defect in neutrophil function in MRL/lpr but not congenic MRL/Mp-+/+ (MRL/n) mice (Gresham, H.

Growth acceleration and stem cell expansion in Dexter-type cultures by neutralization of TGF-beta.

Hematopoietic lineage-restricted stem cell growth has been shown to be significantly inhibited by the addition of exogenous transforming growth factor-beta (TGF-beta) to Dexter-type long-term murine bone-marrow cultures. In order to examine whether TGF-beta produced by these cells has a role in hematopoietic growth regulation, Dexter cultures have been treated with either 1D11.16, a monoclonal antibody that neutralizes the biological activity of TGF-beta types 1, 2, and 3, or with a control antibody.

Immunocytochemical localization and serologic detection of transforming growth factor beta 1. Association with type I procollagen and inflammatory cell markers in diffuse and limited systemic sclerosis, morphea, and Raynaud's phenomenon.

Objective: To determine the presence of transforming growth factor beta 1 (TGF beta 1) and inflammatory cell markers (HLA-DR and Factor XIIIa) and to compare these with the presence of type I procollagen, in clinically uninvolved and involved skin from patients with different subsets of systemic sclerosis (SSc), and to analyze circulating levels of TGF beta 1 in SSc patients.

Methods: TGF beta 1, HLA-DR, Factor XIIIa, and type I procollagen were detected in skin biopsy sections using a biotin-streptavidin-peroxidase system. Levels of circulating TGF beta 1 were measured using a capture enzyme-linked immunosorbent assay technique.

Transforming growth factor-beta 1 is required for secretion of IgG of all subclasses by LPS-activated murine B cells in vitro.

Addition of transforming growth factor-beta 1 (TGF-beta 1) to in vitro cultures of murine B cells activated with bacterial LPS selectively stimulates IgG2b and IgA class switching and decreases cellular proliferation. To assess a possible role for endogenous TGF-beta in modulating the Ig isotypes produced by LPS-activated cells, we utilized a neutralizing anti-TGF-beta mAb to abrogate endogenous TGF-beta activity. Anti-TGF-beta antibody, over a range of relatively low cell densities, strikingly inhibited both IgG3 and IgG2b production in response to LPS, with little or no change in the concentrations of secreted IgM.

Transforming growth factor beta 1 (TGF-beta 1) receptor expression on resting and mitogen-activated T cells.

Transforming growth factor beta 1 (TGF-beta 1) is a potent autocrine growth inhibitor of lymphocytes. In this study, the expression of TGF-beta 1 binding proteins was characterized on murine splenic T cells. With an affinity cross-linking method and by neutralizing antibodies to TGF-beta 1, [125I] TGF-beta 1 was found to bind to three cell surface-binding proteins (280-200 kD, 95-85 kD, 65 kD) that were differentially expressed on resting and mitogen-stimulated T cells.

Monoclonal antibodies recognizing transforming growth factor-beta. Bioactivity neutralization and transforming growth factor beta 2 affinity purification.

Four mAb able to recognize transforming growth factor-beta 2 (TGF-beta)2 were obtained. One of these mAb, 1D11.16, was able to neutralize the biological activity of both TGF-beta 1 and beta 2 in vitro.

Transforming growth factor-beta s are equipotent growth inhibitors of interleukin-1-induced thymocyte proliferation.

The effects of two forms of transforming growth factor-beta, TGF-beta 1 and TGF-beta 2, upon the proliferative response of murine thymocytes were investigated in this study. TGF-beta 1 and TGF-beta 2 were found to be equipotent growth inhibitors of interleukin-1 (IL-1)- and phytohemagglutinin (PHA)-stimulated thymocytes when added at the initiation of the cultures. These factors suppressed the proliferative response in a dose-dependent fashion between 0.

The use of congenital immunologic mutants to probe autoimmune disease in New Zealand mice.

To further our understanding of autoimmunity, many laboratories have concentrated on the study and manipulation of murine lupus in several strains. Although direct extrapolation of data from animal to man must proceed with caution, the use of such animal systems, both in vitro and in vivo, has been an enormous help in the development of immunologic concepts. Our laboratory has been studying murine lupus by selective breeding of specific genetic immune defects onto New Zealand mice.