Antibody-drug conjugates enhance the antitumor effects of antibodies and reduce adverse systemic effects of potent cytotoxic drugs. However, conventional drug conjugation strategies yield heterogenous conjugates with relatively narrow therapeutic index (maximum tolerated dose/curative dose). Using leads from our previously described phage display-based method to predict suitable conjugation sites, we engineered cysteine substitutions at positions on light and heavy chains that provide reactive thiol groups and do not perturb immunoglobulin folding and assembly, or alter antigen binding. When conjugated to monomethyl auristatin E, an antibody against the ovarian cancer antigen MUC16 is as efficacious as a conventional conjugate in mouse xenograft models. Moreover, it is tolerated at higher doses in rats and cynomolgus monkeys than the same conjugate prepared by conventional approaches. The favorable in vivo properties of the near-homogenous composition of this conjugate suggest that our strategy offers a general approach to retaining the antitumor efficacy of antibody-drug conjugates, while minimizing their systemic toxicity.
Bronchoalveolar lavage fluid from mice with experimentally induced allergic pulmonary inflammation contains a novel 9.4 kDa cysteine‐rich secreted protein, FIZZ1 (found in inflammatory zone). Murine (m) FIZZ1 is the founding member of a new gene family including two other murine genes expressed, respectively, in intestinal crypt epithelium and white adipose tissue, and two related human genes. In control mice, FIZZ1 mRNA and protein expression occur at low levels in a subset of bronchial epithelial cells and in non‐neuronal cells adjacent to neurovascular bundles in the peribronchial stroma, and in the wall of the large and small bowel. During allergic pulmonary inflammation, mFIZZ1 expression markedly increases in hypertrophic, hyperplastic bronchial epithelium and appears in type II alveolar pneumocytes. In vitro, recombinant mFIZZ1 inhibits the nerve growth factor (NGF)‐mediated survival of rat embryonic day 14 dorsal root ganglion (DRG) neurons and NGF‐induced CGRP gene expression in adult rat DRG neurons. In vivo, FIZZ1 may modulate the function of neurons innervating the bronchial tree, thereby altering the local tissue response to allergic pulmonary inflammation.
Ranibizumab represents a novel therapy that, for the first time, appears to have the potential to enable many AMD patients to obtain a meaningful and sustained gain of vision. On June 30 2006, ranibizumab was approved by the US Food and Drug Administration for the treatment of wet AMD.
Our findings suggest that the character of antigen-antibody interfaces is similar to that of other protein-protein interfaces, such as ligand-receptor interactions; in the case of VEGF, the principal difference is that the residues essential for binding to the Fab fragment are concentrated in one continuous segment of polypeptide chain, whereas those essential for binding to the receptor are distributed over four different segments and span across the dimer interface.
The pH-dependent binding of Igs to the neonatal FcR (FcRn) plays a critical role in the in vivo homeostasis of IgGs. Modulating the interaction between Fc and FcRn through protein engineering is one method for improving the pharmacokinetics of therapeutic Abs. Recent studies disputed the direct relationship between increasing FcRn affinity and improved pharmacokinetic properties. In this work, we studied the pharmacokinetics of two human IgG1 Fc variants in cynomolgus monkey to further clarify the affinity-pharmacokinetic relationship. First, we report a number of novel Fc point mutations and combination variants, including some with primate-specific FcRn-binding improvements. By studying these variants along with some previously described variants across a wide range of affinities, we discovered a direct correlation of pH 6 affinity improvements with neutral pH improvements, suggesting that all of the tested variants exhibit similar pH dependency in FcRn binding. We then evaluated the pharmacokinetics of variants N434A and N434W, which, respectively, gave ∼4- and 80-fold improvements in pH 6-binding affinity to both human and nonhuman primate FcRn. Surprisingly, clearance of N434W was similar to that of wild type. N434W is the first variant studied in primates that exhibits significant binding to FcRn at pH 7.4, and its clearance substantiates the principle that too much affinity improvement, i.e., beyond that of N434W, does not yield improved pharmacokinetics. In contrast, N434A exhibited a ∼2-fold decrease in clearance in cynomolgus monkey, supporting the notion that modest increases in pH 6 FcRn affinity can result in improved pharmacokinetics in primates.
The results offer the first details at an atomic level of the interaction between a chemokine and its receptor. Consideration of other biochemical data allow extrapolation to a model for the interaction of IL-8 with the full-length receptor. In this model, the heparin-binding residues of IL-8 are exposed, thereby allowing presentation of the chemokine from endothelial cell-surface glycosaminoglycans. This first glimpse of how IL-8 binds to its receptor provides a foundation for the structure-based design of chemokine antagonists.
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