TNF-α antagonists differentially induce TGF-β1-dependent resuscitation of dormant-like Mycobacterium tuberculosis

TNF-α- as well as non-TNF-α-targeting biologics are prescribed to treat a variety of immune-mediated inflammatory disorders. The well-documented risk of tuberculosis progression associated with anti-TNF-α treatment highlighted the central role of TNF-α for the maintenance of protective immunity, although the rate of tuberculosis detected among patients varies with the nature of the drug. Using a human, in-vitro granuloma model, we reproduce the increased reactivation rate of tuberculosis following exposure to Adalimumab compared to Etanercept, two TNF-α-neutralizing biologics. We show that Adalimumab, because of its bivalence, specifically induces TGF-β1-dependent Mycobacterium tuberculosis (Mtb) resuscitation which can be prevented by concomitant TGF-β1 neutralization. Moreover, our data suggest an additional role of lymphotoxin-α-neutralized by Etanercept but not Adalimumab-in the control of latent tuberculosis infection. Furthermore, we show that, while Secukinumab, an anti-IL-17A antibody, does not revert Mtb dormancy, the anti-IL-12-p40 antibody Ustekinumab and the recombinant IL-1RA Anakinra promote Mtb resuscitation, in line with the importance of these pathways in tuberculosis immunity

Patients with naproxen-induced liver injury display T-cell memory responses toward an oxidative (S)-O-Desmethyl Naproxen metabolite but not the acyl glucuronide

Background Exposure to nonsteroidal anti-inflammatory drugs (NSAIDs) such as ibuprofen (IBU) and naproxen (NAP) is associated with idiosyncratic drug-induced liver injury (DILI). Carboxylate bioactivation into reactive metabolites (e.g., acyl glucuronides, AG) and resulting T-cell activation is hypothesized as causal for this adverse event. However, conclusive evidence supporting this is lacking. Methods In this work, we identify CD4+ and CD8+ T-cell hepatic infiltration in a biopsy from an IBU DILI patient. Lymphocyte transformation test and IFN-γ ELIspot, conducted on peripheral blood mononuclear cells (PBMCs) of patients with NAP-DILI, were used to explore drug-specific T-cell activation. T-cell clones (TCC) were generated and tested for drug specificity, phenotype/function, and pathways of T-cell activation. Cells were exposed to NAP, its oxidative metabolite 6-O-desmethyl NAP (DM-NAP), its AG or synthesized NAP-AG human-serum albumin adducts (NAP-AG adduct). Results CD4+ and CD8+ T-cells from patients expressing a range of different Vβ receptors were stimulated to proliferate and secrete IFN-γ and IL-22 when exposed to DM-NAP, but not NAP, NAP-AG or the NAP-AG adduct. Activation of the CD4+ TCC was HLA-DQ-restricted and dependent on antigen presenting cells (APC); most TCC were activated with DM-NAP-pulsed APC, while fixation of APC blocked the T-cell response. Cross-reactivity was not observed with structurally-related drugs. Conclusion Our results confirm hepatic T-cell infiltrations in NSAID-induced DILI, and show a T-cell memory response toward DM-NAP indicating an immune-mediated basis for the adverse event. Whilst bioactivation at the carboxylate group is widely hypothesized to be pathogenic for NSAID associated DILI, we found no evidence of this with NAP

Brucella abortus strain RB51 infection of bovine trophoblast epithelial cells: characterization of receptor signaling and bacterial outer membrane proteins

In 1995, a bovine brucellosis vaccine containing Brucella abortus strain RB51 was accepted for calfhood vaccination. However there is no objective serologic tests, such as ELISA, which detects antibodies specific for RB51-vaccinated cattle, and intravenous inoculation of pregnant animals with strain RB51 is followed by placentitis;To create an objective diagnostic test, antigenic outer membrane proteins from strain RB51 were isolated by Concanavalin A lectin affinity chromatography. One eluted protein of 35 kDa reacted in western blot analysis to every individual serum from strain RB51 and 2308 infected cattle. This protein may be a useful candidate antigen in developing a specific test for detecting antibody to RB51 in RB51-vaccinated cattle;Invasion and replication of bacteria in trophoblastic epithelial cells are key events in placentitis. To identify the receptor involved in adhesion of strain RB51 to bovine trophoblastic epithelial cells and the signal pathway that mediates bacterial phagocytosis, a bioassay and double immunofluorescence labeling were used. In bioassays, adhesion of B. abortus to trophoblastic cells was inhibited by fibronectin, RGDS peptide, and antibodies to [alpha] 5 or [beta] 1 integrin subunits. Uptake of bacteria was inhibited by genistein, an inhibitor of tyrosine kinases. Ultrastructurally, intracellular bacteria were located in the rough endoplasmic reticulum or in phagolysosomes of trophoblasts. These results indicate that strain RB51 binds to the trophoblastic epithelial cell surface by an [alpha] 5[beta] 1 integrin, that uptake is mediated by tyrosine kinases, and that bacteria replicate intracellularly within cisternea of the rough endoplasmic reticulum;The consequences of Brucella induced-placentitis on the binucleate trophoblastic cell population has never been studied. Numbers of Binucleate cells stained with Dolichos Biflorus agglutinin lectin were quantified by morphometric analysis in placentomes from non-infected and strain RB51-infected pregnant cattle. There was diffuse decreased numbers of cells in sections of placentomes with placentitis. Immunohistochemistry for proliferating cell nuclear antigen and terminal deoxy dUTP nick end-labeling showed reduced proliferation of progenitor mononuclear cytotrophoblasts in arcade zone, that could explain binucleate cell depletion, but not increased programmed cell death. Because binucleate cells produce placental lactogen, decreased numbers of cells could participate to mechanisms of abortion.</p

Safety biomarkers in preclinical development: translational potential

The identification, application, and qualification of safety biomarkers are becoming increasingly critical to successful drug discovery and development as companies are striving to develop drugs for difficult targets and for novel disease indications in a risk-adverse environment. Translational safety biomarkers that are minimally invasive and monitor drug-induced toxicity during human clinical trials are urgently needed to assess whether toxicities observed in preclinical toxicology studies are relevant to humans at therapeutic doses. The interpretation of data during the biomarker qualification phase should include careful consideration of the analytic method used, the biology, pharmacokinetic and pharmacodynamic properties of the biomarker, and the pathophysiology of the process studied. The purpose of this review is to summarize commonly employed technologies in the development of fluid- and tissue-based safety biomarkers in drug discovery and development and to highlight areas of ongoing novel assay development

Assessing immunocompetence in minipigs in translational safety sciences – a perspective

The porcine immune system has been studied especially with regard to infectious diseases of the domestic pig, highlighting the economic importance of the pig in agriculture. Recently, in particular minipigs have received attention as alternative species to dogs or non-human primates in drug safety evaluations. The increasing number of new drug targets investigated to modulate immunological pathways has triggered renewed interest to further explore the porcine immune system. Comparative immunological studies of minipigs with other species broaden the translational models investigated in drug safety evaluations. The porcine immune system overall seems functionally similar to other mammalian species, but there are some anatomical and immunophenotypical differences. Here, we briefly review current knowledge of the innate and adaptive immune system in (mini)pigs. In conclusion, more systematic and cross-species comparisons are needed to assess the significance of immunological findings in minipigs in the context of translational safety sciences