Generation and characterization of ABBV642, a dual variable domain immunoglobulin molecule (DVD-Ig) that potently neutralizes VEGF and PDGF-BB and is designed for the treatment of exudative age-related macular degeneration.

Exudative age-related macular degeneration (AMD) is the most common cause of moderate and severe vision loss in developed countries. Intraocular injections of vascular endothelial growth factor (VEGF or VEGF-A)-neutralizing proteins provide substantial benefit, but frequent, long-term injections are needed. In addition, many patients experience initial visual gains that are ultimately lost due to subretinal fibrosis. Preclinical studies and early phase clinical trials suggest that combined suppression of VEGF and platelet-derived growth factor-BB (PDGF-BB) provides better outcomes than suppression of VEGF alone, due to more frequent regression of neovascularization (NV) and suppression of subretinal fibrosis. We generated a dual variable domain immunoglobulin molecule, ABBV642 that specifically and potently binds and neutralizes VEGF and PDGF-BB. ABBV642 has been optimized for treatment of exudative AMD based on the following design characteristics: 1) high affinity binding to all VEGF-A isoforms and both soluble and extracellular matrix (ECM)-associated PDGF-BB; 2) potential for extended residence time in the vitreous cavity to decrease the frequency of intraocular injections; 3) rapid clearance from systemic circulation compared with molecules with wild type Fc region for normal FcRn binding, which may reduce the risk of systemic complications; and 4) low risk of potential effector function. The bispecificity of ABBV642 allows for a single injection of a single therapeutic agent, and thus a more streamlined development and regulatory path compared with combination products. In a mouse model of exudative AMD, ABBV642 was observed to be more effective than aflibercept. ABBV642 has potential to improve efficacy with reduced injection frequency in patients with exudative AMD, thereby reducing the enormous disease burden for patients and society.

Creation of a S1P Lyase bacterial surrogate for structure-based drug design.

S1P Lyase (SPL) has been described as a drug target in the treatment of autoimmune diseases. It plays an important role in maintaining intracellular levels of S1P thereby affecting T cell egress from lymphoid tissues. Several groups have already published approaches to inhibit S1P Lyase with small molecules, which in turn increase endogenous S1P concentrations resulting in immunosuppression. The use of structural biology has previously aided SPL inhibitor design. Novel construct design is at times necessary to provide a reagent for protein crystallography. Here we present a chimeric bacterial protein scaffold used for protein X-ray structures in the presence of early small molecule inhibitors. Mutations were introduced to the bacterial SPL from Symbiobacterium thermophilum which mimic the human enzyme. As a result, two mutant StSPL crystal structures resolved to 2.8Å and 2.2Å resolutions were solved and provide initial structural hypotheses for an isoxazole chemical series, whose optimization is discussed in the accompanying paper.

Functional comparison of recombinant acidic mammalian chitinase with enzyme from murine bronchoalveolar lavage.

Acidic mammalian chitinase (AMCase) is an enzyme that selectively degrades the biopolymer chitin. Several chitinase enzymes are utilized by mammals to hydrolyze chitin encountered by inhalation and ingestion. AMCase is distinct from other mammalian chitinases as its activity is retained in strongly acidic conditions (pH <2.0). AMCase expression is induced by antigen-induced mouse models of allergic lung inflammation. This protein has also been implicated in the pathogenesis of asthma although its precise role is poorly defined. We describe a novel way to express and purify active murine AMCase. This material retains properties observed in mouse bronchoalveolar lavage (BAL) fluid with regard to pH preference of activity and its inhibition by cyclic peptide inhibitors argifin and argadin. We found that chitinase in BAL from both antigen-challenged and control animals have similar properties in this regard. This strongly supports the notion the same enzyme (AMCase) gives rise to chitinase activity in both challenged and unchallenged animals. We also describe expression of active human AMCase. The methods described in this paper provide a reliable source of recombinant AMCase that can be utilized to expand understanding of AMCase's role in regulating allergic inflammation.