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1.
J Pharmacol Exp Ther ; 361(3): 429-440, 2017 06.
Artigo em Inglês | MEDLINE | ID: mdl-28442583

RESUMO

Phosphoinositide 3-kinases (PI3K) are key signaling enzymes regulating cellular survival, development, and function. Expression of the PI3Kδ isoform is largely restricted to leukocytes and it plays a key role in immune cell development and function. Seletalisib is a novel small-molecule inhibitor of PI3Kδ that was evaluated in biochemical assays, cellular assays of adaptive and innate immunity, and an in vivo rat model of inflammation. Our findings show that seletalisib is a potent, ATP-competitive, and selective PI3Kδ inhibitor able to block protein kinase B (AKT) phosphorylation following activation of the B-cell receptor in a B-cell line. Moreover, seletalisib inhibited N-formyl peptide-stimulated but not phorbol myristate acetate-stimulated superoxide release from human neutrophils, consistent with a PI3Kδ-specific activity. No indications of cytotoxicity were observed in peripheral blood mononuclear cells (PBMCs) or other cell types treated with seletalisib. Findings from cellular assays of adaptive immunity demonstrated that seletalisib blocks human T-cell production of several cytokines from activated T-cells. Additionally, seletalisib inhibited B-cell proliferation and cytokine release. In human whole blood assays, seletalisib inhibited CD69 expression upon B-cell activation and anti-IgE-mediated basophil degranulation. Seletalisib showed dose-dependent inhibition in an in vivo rat model of anti-CD3-antibody-induced interleukin 2 release. Collectively, these data characterize seletalisib as a selective PI3Kδ inhibitor and potential therapeutic candidate for the treatment of B-cell malignancies and autoimmune diseases driven by dysregulated proinflammatory cytokine secretion.


Assuntos
Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Inibidores de Fosfoinositídeo-3 Quinase , Piridinas/química , Piridinas/farmacologia , Quinolinas/química , Quinolinas/farmacologia , Animais , Células Cultivadas , Classe I de Fosfatidilinositol 3-Quinases , Relação Dose-Resposta a Droga , Humanos , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/enzimologia , Masculino , Fosfatidilinositol 3-Quinases/metabolismo , Ratos , Ratos Endogâmicos Lew , Ratos Wistar
2.
MAbs ; 15(1): 2289681, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-38084840

RESUMO

Gremlin-1, a high-affinity antagonist of bone morphogenetic proteins (BMP)-2, -4, and -7, is implicated in tumor initiation and progression. Increased gremlin-1 expression, and therefore suppressed BMP signaling, correlates with poor prognosis in a range of cancer types. A lack of published work using therapeutic modalities has precluded the testing of the hypothesis that blocking the gremlin-1/BMP interaction will provide benefits to patients. To address this shortfall, we developed ginisortamab (UCB6114), a first-in-class clinical anti-human gremlin-1 antibody, currently in clinical development for the treatment of cancer, along with its murine analog antibody Ab7326 mouse immunoglobulin G1 (mIgG1). Surface plasmon resonance assays revealed that ginisortamab and Ab7326 mIgG1 had similar affinities for human and mouse gremlin-1, with mean equilibrium dissociation constants of 87 pM and 61 pM, respectively. The gremlin-1/Ab7326 antigen-binding fragment (Fab) crystal structure revealed a gremlin-1 dimer with a Fab molecule bound to each monomer that blocked BMP binding. In cell culture experiments, ginisortamab fully blocked the activity of recombinant human gremlin-1, and restored BMP signaling pathways in human colorectal cancer (CRC) cell lines. Furthermore, in a human CRC - fibroblast co-culture system where gremlin-1 is produced by the fibroblasts, ginisortamab restored BMP signaling in both the CRC cells and fibroblasts, demonstrating its activity in a relevant human tumor microenvironment model. The safety and efficacy of ginisortamab are currently being evaluated in a Phase 1/2 clinical trial in patients with advanced solid tumors (NCT04393298).


Assuntos
Neoplasias , Transdução de Sinais , Humanos , Animais , Camundongos , Linhagem Celular , Neoplasias/tratamento farmacológico , Microambiente Tumoral
3.
MAbs ; 10(7): 1111-1130, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-30130439

RESUMO

Rozanolixizumab (UCB7665), a humanized high-affinity anti-human neonatal Fc receptor (FcRn) monoclonal antibody (IgG4P), has been developed to reduce pathogenic IgG in autoimmune and alloimmune diseases. We document the antibody isolation and compare rozanolixizumab with the same variable region expressed in various mono-, bi- and trivalent formats. We report activity data for rozanolixizumab and the different molecular formats in human cells, FcRn-transgenic mice, and cynomolgus monkeys. Rozanolixizumab, considered the most effective molecular format, dose-dependently and selectively reduced plasma IgG concentrations in an FcRn-transgenic mouse model (no effect on albumin). Intravenous (IV) rozanolixizumab dosing in cynomolgus monkeys demonstrated non-linear pharmacokinetics indicative of target-mediated drug disposition; single IV rozanolixizumab doses (30 mg/kg) in cynomolgus monkeys reduced plasma IgG concentration by 69% by Day 7 post-administration. Daily IV administration of rozanolixizumab (initial 30 mg/kg loading dose; 5 mg/kg daily thereafter) reduced plasma IgG concentrations in all cynomolgus monkeys, with low concentrations maintained throughout the treatment period (42 days). In a 13-week toxicology study in cynomolgus monkeys, supra-pharmacological subcutaneous and IV doses of rozanolixizumab (≤ 150 mg/kg every 3 days) were well tolerated, inducing sustained (but reversible) reductions in IgG concentrations by up to 85%, with no adverse events observed. We have demonstrated accelerated natural catabolism of IgG through inhibition of IgG:FcRn interactions in mice and cynomolgus monkeys. Inhibition of FcRn with rozanolixizumab may provide a novel therapeutic approach to reduce pathogenic IgG in human autoimmune disease. Rozanolixizumab is being investigated in patients with immune thrombocytopenia (NCT02718716) and myasthenia gravis (NCT03052751).


Assuntos
Anticorpos Monoclonais Humanizados/química , Antígenos de Histocompatibilidade Classe I/imunologia , Imunossupressores/química , Miastenia Gravis/tratamento farmacológico , Púrpura Trombocitopênica Idiopática/tratamento farmacológico , Receptores Fc/imunologia , Animais , Anticorpos Monoclonais Humanizados/genética , Anticorpos Monoclonais Humanizados/metabolismo , Ensaios Clínicos como Assunto , Antígenos de Histocompatibilidade Classe I/genética , Humanos , Imunoglobulina G/sangue , Imunossupressores/metabolismo , Macaca fascicularis , Camundongos , Camundongos Transgênicos , Ligação Proteica , Receptores Fc/genética , Transgenes/genética
4.
Sci Rep ; 7(1): 9963, 2017 08 30.
Artigo em Inglês | MEDLINE | ID: mdl-28855577

RESUMO

Fibrosis is a common driver of end-stage organ failure in most organs. It is characterised by excessive accumulation of extracellular matrix (ECM) proteins. Therapeutic options are limited and novel treatments are urgently required, however current cell-based high-throughput screening (HTS) models to identify molecules affecting ECM accumulation are limited in their relevance or throughput. We report a novel sensitive approach which combines in situ fluorescent staining of accumulated decellularised ECM proteins with automated high-content microscopy. Using this method to measure ECM accumulation in a kidney cell model, we demonstrated good agreement with established radiolabelled amino acid incorporation assays: TGFß1 delivered a potent pro-fibrotic stimulus, which was reduced by TGFß antibody or the anti-fibrotic nintedanib. Importantly, our method also provides information about matrix organisation: the extent of ECM accumulation was unaffected by the BMP antagonist Gremlin-1 but a pronounced effect on matrix fibrillar organisation was revealed. This rapid, straightforward endpoint provides quantitative data on ECM accumulation and offers a convenient cross-species readout that does not require antibodies. Our method facilitates discovery of novel pro- and anti-fibrotic agents in 384-well plate format and may be widely applied to in vitro cell-based models in which matrix protein deposition reflects the underlying biology or pathology.


Assuntos
Matriz Extracelular/química , Fibrose/patologia , Nefropatias/patologia , Microscopia de Fluorescência/métodos , Proteínas/análise , Automação Laboratorial/métodos , Células Cultivadas , Células Epiteliais/patologia , Humanos , Modelos Biológicos , Coloração e Rotulagem/métodos
5.
Sci Rep ; 7(1): 18111, 2017 12 21.
Artigo em Inglês | MEDLINE | ID: mdl-29269854

RESUMO

A correction to this article has been published and is linked from the HTML version of this paper. The error has been fixed in the paper.

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