Anti-citrullinated histone monoclonal antibody CIT-013, a dual action therapeutic for neutrophil extracellular trap-associated autoimmune diseases.

Neutrophil extracellular traps (NETs) contribute to the pathophysiology of multiple inflammatory and autoimmune diseases. Targeting the NETosis pathway has demonstrated significant therapeutic potency in various disease models. Here, we describe a first-in-class monoclonal antibody (CIT-013) with high affinity for citrullinated histones H2A and H4, which inhibits NETosis and reduces tissue NET burden in vivo with significant anti-inflammatory consequences. We provide a detailed understanding of the epitope selectivity of CIT-013. Detection of CIT-013 epitopes in rheumatoid arthritis (RA) synovium provides evidence that RA is an autoimmune disease with excessive citrullinated NETs that can be targeted by CIT-013. We show that CIT-013 acts upon the final stage of NETosis, binding to its chromatin epitopes when plasma membrane integrity is compromised to prevent NET release. Bivalency of CIT-013 is necessary for NETosis inhibition. In addition, we show that CIT-013 binding to NETs and netting neutrophils enhance their phagocytosis by macrophages in an Fc-dependent manner. This is confirmed using a murine neutrophilic airway inflammation model where a mouse variant of CIT-013 reduced tissue NET burden with significant anti-inflammatory consequences. CIT-013's therapeutic activity provides new insights for the development of NET antagonists and indicates the importance of a new emerging therapy for NET-driven diseases with unmet therapeutic needs.

Functional characterization of the selective pan-allele anti-SIRPα antibody ADU-1805 that blocks the SIRPα-CD47 innate immune checkpoint.

BACKGROUND: Accumulating preclinical data indicate that targeting the SIRPα/CD47 axis alone or in combination with existing targeted therapies or immune checkpoint inhibitors enhances tumor rejection. Although several CD47-targeting agents are currently in phase I clinical trials and demonstrate activity in combination therapy, high and frequent dosing was required and safety signals (acute anemia, thrombocytopenia) were recorded frequently as adverse events. Based on the restricted expression pattern of SIRPα we hypothesized that antibodies targeting SIRPα might avoid some of the concerns noted for CD47-targeting agents. METHODS: SIRPα-targeting antibodies were generated and characterized for binding to human SIRPα alleles and blockade of the interaction with CD47. Functional activity was established in vitro using human macrophages or neutrophils co-cultured with human Burkitt's lymphoma cell lines. The effect of SIRPα versus CD47 targeting on human T-cell activation was studied using an allogeneic mixed lymphocyte reaction and a Staphylococcus enterotoxin B-induced T-cell proliferation assay. Potential safety concerns of the selected SIRPα-targeting antibody were addressed in vitro using a hemagglutination assay and a whole blood cytokine release assay, and in vivo in a single-dose toxicity study in cynomolgus monkeys. RESULTS: The humanized monoclonal IgG2 antibody ADU-1805 binds to all known human SIRPα alleles, showing minimal binding to SIRPß1, while cross-reacting with SIRPγ, and potently blocking the interaction of SIRPα with CD47. Reduced FcγR binding proved critical to retaining its function towards phagocyte activation. In vitro characterization demonstrated that ADU-1805 promotes macrophage phagocytosis, with similar potency to anti-CD47 antibodies, and enhances neutrophil trogocytosis. Unlike CD47-targeting agents, ADU-1805 does not interfere with T-cell activation and is not expected to require frequent and extensive dosing due to the restricted expression of SIRPα to cells of the myeloid lineage. ADU-1805 is cross-reactive to cynomolgus monkey SIRPα and upon single-dose intravenous administration in these non-human primates (NHPs) did not show any signs of anemia, thrombocytopenia or other toxicities. CONCLUSIONS: Blocking the SIRPα-CD47 interaction via SIRPα, while similarly efficacious in vitro, differentiates ADU-1805 from CD47-targeting agents with respect to safety and absence of inhibition of T-cell activation. The data presented herein support further advancement of ADU-1805 towards clinical development.

KH176 Safeguards Mitochondrial Diseased Cells from Redox Stress-Induced Cell Death by Interacting with the Thioredoxin System/Peroxiredoxin Enzyme Machinery.

A deficient activity of one or more of the mitochondrial oxidative phosphorylation (OXPHOS) enzyme complexes leads to devastating diseases, with high unmet medical needs. Mitochondria, and more specifically the OXPHOS system, are the main cellular production sites of Reactive Oxygen Species (ROS). Increased ROS production, ultimately leading to irreversible oxidative damage of macromolecules or to more selective and reversible redox modulation of cell signalling, is a causative hallmark of mitochondrial diseases. Here we report on the development of a new clinical-stage drug KH176 acting as a ROS-Redox modulator. Patient-derived primary skin fibroblasts were used to assess the potency of a new library of chromanyl-based compounds to reduce ROS levels and protect cells against redox-stress. The lead compound KH176 was studied in cell-based and enzymatic assays and in silico. Additionally, the metabolism, pharmacokinetics and toxicokinetics of KH176 were assessed in vivo in different animal species. We demonstrate that KH176 can effectively reduce increased cellular ROS levels and protect OXPHOS deficient primary cells against redox perturbation by targeting the Thioredoxin/Peroxiredoxin system. Due to its dual activity as antioxidant and redox modulator, KH176 offers a novel approach to the treatment of mitochondrial (-related) diseases. KH176 efficacy and safety are currently being evaluated in a Phase 2 clinical trial.

FSH receptor genotype is associated with pregnancy but not with ovarian response in IVF.

Two very common single nucleotide polymorphisms at positions 307 and 680 in exon 10 of the FSH receptor gene have been associated with ovarian response in IVF. This observational study evaluated the role of the FSH receptor genotype in the prediction of poor response and clinical pregnancy in IVF in comparison with other markers, such as age, basal FSH, anti-Müllerian hormone and antral follicle count. In addition, the in-vitro cAMP response towards recombinant FSH in cultured granulosa cells of patients with different FSH receptor genotypes was determined. A total of 105 IVF patients undergoing ovarian stimulation in a long suppression protocol were included in the study. The ovarian response was comparable between patients with different FSH receptor genotypes. Patients with polymorphism Ser/Ser had implantation and pregnancy rates that were three times higher compared with patients with polymorphism Asn/Asn. FSH receptor genotype was not associated with a poor response in IVF, but showed a positive association with pregnancy, independent of age. There was no difference in cAMP production in cultured granulosa cells of patients with different FSH receptor genotypes (n=62). It is concluded that FSH receptor genotype is associated with pregnancy in IVF, but not with ovarian response.

Identification of substituted 6-amino-4-phenyltetrahydroquinoline derivatives: potent antagonists for the follicle-stimulating hormone receptor.

Substituted 6-amino-4-phenyl-tetrahydroquinoline derivatives are described that are antagonists for the G(s)-protein-coupled human follicle-stimulating hormone (FSH) receptor. These compounds show high antagonistic efficacy in vitro using a CHO cell line expressing the human FSH receptor. Antagonist 10 also showed a submicromolar IC(50) in a more physiologically relevant rat granulosa cell assay and was found to significantly inhibit follicle growth and ovulation in an ex vivo mouse model. This compound class may open the way toward a novel, nonsteroidal approach for contraception.