Nonclinical immunogenicity risk assessment for knobs-into-holes bispecific IgG antibodies.

Bispecific antibodies, including bispecific IgG, are emerging as an important new class of antibody therapeutics. As a result, we, as well as others, have developed engineering strategies designed to facilitate the efficient production of bispecific IgG for clinical development. For example, we have extensively used knobs-into-holes (KIH) mutations to facilitate the heterodimerization of antibody heavy chains and more recently Fab mutations to promote cognate heavy/light chain pairing for efficient assembly of bispecific IgG in single host cells.

Introducing dendritic cell antibody internalization as an immunogenicity risk assessment tool.

Immunogenicity risk assessment assays are powerful tools that assess the relative immunogenicity of potential biotherapeutics. We detail here the development of a novel assay that measures the degree of antibody internalization by antigen-presenting cells as a predictor of immunogenicity. The assay uses the fluorescence signal from the antibody bound to the outside of the cell as well as inside the cell to determine internalization.

Development of an ultra-sensitive human IL-33 biomarker assay for age-related macular degeneration and asthma drug development.

Background: Over the past decade, human Interleukin 33 (hIL-33) has emerged as a key contributor to the pathogenesis of numerous inflammatory diseases. Despite the existence of several commercial hIL-33 assays spanning multiple platform technologies, their ability to provide accurate hIL-33 concentration measurements and to differentiate between active (reduced) and inactive (oxidized) hIL-33 in various matrices remains uncertain. This is especially true for lower sample volumes, matrices with low hIL-33 concentrations, and matrices with elevated levels of soluble Interleukin 1 Receptor-Like 1 (sST2), an inactive form of ST2 that competes with membrane bound ST2 for hIL-33 binding.

TGFβ2 and TGFβ3 isoforms drive fibrotic disease pathogenesis.

Transforming growth factor-β (TGFβ) is a key driver of fibrogenesis. Three TGFβ isoforms (TGFβ1, TGFβ2, and TGFβ3) in mammals have distinct functions in embryonic development; however, the postnatal pathological roles and activation mechanisms of TGFβ2 and TGFβ3 have not been well characterized. Here, we show that the latent forms of TGFβ2 and TGFβ3 can be activated by integrin-independent mechanisms and have lower activation thresholds compared to TGFβ1.

An Allosteric Anti-tryptase Antibody for the Treatment of Mast Cell-Mediated Severe Asthma.

Severe asthma patients with low type 2 inflammation derive less clinical benefit from therapies targeting type 2 cytokines and represent an unmet need. We show that mast cell tryptase is elevated in severe asthma patients independent of type 2 biomarker status. Active β-tryptase allele count correlates with blood tryptase levels, and asthma patients carrying more active alleles benefit less from anti-IgE treatment.

Molecular characterization of human anti-hinge antibodies derived from single-cell cloning of normal human B cells.

Anti-hinge antibodies (AHAs) are an autoantibody subclass that, following proteolytic cleavage, recognize cryptic epitopes exposed in the hinge regions of immunoglobulins (Igs) and do not bind to the intact Ig counterpart. AHAs have been postulated to exacerbate chronic inflammatory disorders such as inflammatory bowel disease and rheumatoid arthritis. On the other hand, AHAs may protect against invasive microbial pathogens and cancer.

Fibroblast Activation Protein Cleaves and Inactivates Fibroblast Growth Factor 21.

FGF21 is a stress-induced hormone with potent anti-obesity, insulin-sensitizing, and hepatoprotective properties. Although proteolytic cleavage of recombinant human FGF21 in preclinical species has been observed previously, the regulation of endogenously produced FGF21 is not well understood. Here we identify fibroblast activation protein (FAP) as the enzyme that cleaves and inactivates human FGF21.