Combined analytical assays for the characterization of drugs binding to human IgE: Applicability to omalizumab-bearing biosimilar candidates assessment.

Analytical and functional comparison is key for substantiating the level of convergence (essential sameness) or divergence between versions or variants of a given biological medicine. Accordingly, an overlapping biological activity between products meant to be equal probably reflects a highly similar structure and anticipates a comparable pharmacodynamic behavior. We developed an orthogonal approach to compare the human IgE binding features of different lots and versions of Xolair® (omalizumab), an anti-human IgE monoclonal antibody. The IgE binding affinity and kinetics were measured by surface plasmon resonance. Ability to prevent mast cell activity was assessed in vitro and in vivo in mast cell-based models. The variability of monoclonal antibodies with identical amino acid sequences produced either in Chinese hamster ovarian cells or in human HEK293 cells, was compared. Monoclonal antibodies from the two sources exhibited slightly different human IgE binding and neutralizing features. A known variant exhibiting a three amino acid replacement in the Fab region had lower IgE binding affinity than the original omalizumab. The lower binding affinity translated into reduced IgE neutralizing capacity and, in turn, a difference in the ability to prevent mast cell activation in vitro and in vivo. The proposed set of analytical and functional assays was sensitive enough to detect Fab-linked differences between anti-IgE antibody versions exhibiting an identical aminoacid sequence. In addition to add value to the comparative assessment of biosimilar candidates bearing omalizumab, these methods can aid pre-assessments of new anti-IgE agents that aim to improve therapeutic performance.

In Vitro and In Vivo Validation of EP2-Receptor Agonism to Selectively Achieve Inhibition of Mast Cell Activity.

PURPOSE: Agonism of the prostaglandin E2 receptor, E-prostanoid receptor 2 (EP2), may represent an alternative protective mechanism in mast cell (MC)-mediated diseases. Previous studies have suggested that activation of the MC EP2 receptor prevents pathological changes in the murine models of allergic asthma. This work aimed to analytically validate the EP2 receptor on MCs as a therapeutic target. METHODS: Murine MC lines and primary cultures, and MCs bearing the human immunoglobulin E (IgE) receptor were subjected to IgE-mediated activation subsequent to incubation with selective EP2 agonists. Two molecularly unrelated agonists, butaprost and CP-533536, were tested either in vitro or in 2 in vivo models of allergy. RESULTS: The diverse range of MC populations was consistently inhibited through selective EP2 agonism in spite of exhibiting a heterogeneous phenotype. Such inhibition occurred in both mouse and human IgE (hIgE)-mediated activation. The use of molecularly unrelated selective EP2 agonists allowed for the confirmation of the specificity of this protective mechanism. This effect was further demonstrated in 2 in vivo murine models of allergy where MCs are a key to pathological changes: cutaneous anaphylaxis in a transgenic mouse model expressing the hIgE receptor and aeroallergen-induced murine model of asthma. CONCLUSIONS: Selective EP2 agonism is a powerful pharmacological strategy to prevent MCs from being activated through IgE-mediated mechanisms and from causing deleterious effects. The MC EP2 receptor may be an effective pharmacological target in allergic and other MC-mediated conditions.

8-Mercaptoquinoline as a Ligand for Enhancing the Photocatalytic Activity of Pt(II) Coordination Complexes: Reactions and Mechanistic Insights.

A family of quinoline-platinum(II) complexes as efficient photocatalysts is presented. Their key characteristic is their easy preparation by coordination of the readily available 8-hydroxy- or 8-thio-quinoline ligands, which are well known for their strong chelating ability to different metal ions. In the different photochemical transformations investigated, such as cross-dehydrogenative coupling, oxidation of arylboronic acids, and asymmetric alkylation of aldehydes, 8-mercaptoquinoline-Pt(II) complex proved to be the most general catalyst. Moreover, quenching experiments showed that, contrary to related methods reported in the literature, these complexes followed an oxidative quenching mechanism in all transformations studied. Besides, simulations performed with high-level ab initio methods of the complexes have helped to understand their photocatalytic activity.

Locally administered prostaglandin E2 prevents aeroallergen-induced airway sensitization in mice through immunomodulatory mechanisms.

Prostaglandin E2 attenuates airway pathology in asthmatic patients and exerts a protective effect in antigen-sensitized mice when administered systemically. We aimed to establish the consequences of intranasal PGE2 administration on airway reactivity to aeroallergens in mice and reveal the underlying immunoinflammatory mechanisms. PGE2 was administered either daily during a 10-day exposure to house dust mite (HDM) extracts or for limited intervals. Airway hyperreactivity was measured by whole-body and invasive plethysmography. The phenotypes of lung immune cells and cytokine production were analysed by flow cytometry and ELISA, respectively. Airway hyperreactivity was sustainably reduced only when PGE2 administration was restricted to the initial 5 days of exposure to HDM. Lung inflammation, IL-4 production, and airway mast cell activity were also prevented under this early short-term treatment with PGE2. Interestingly, a Th2 response was already committed on day 5 of exposure to HDM. This was paralleled by GM-CSF and osteopontin upregulation and a decreased number of plasmacytoid dendritic and T regulatory cells, as well as a trend towards reduced IL-10 expression. Local PGE2 administration prevented the increase of airway IL-13 and osteopontin and kept lung plasmacytoid dendritic cell counts close to baseline. GM-CSF and Tregs were unaffected by the treatment. These findings suggest that the protection provided by PGE2 is a result of the modulation of early lung immunomodulatory mechanisms, and possibly a shift in the balance of dendritic cells towards a tolerogenic profile.