Michael Steward

A dual-targeting antibody against EGFR-VEGF for lung and head and neck cancer treatment†

An antibody simultaneously targeting epidermal growth factor receptor (EGFR) and vascular endothelial growth factor (VEGF), two major tumor growth‐driving machineries, may provide a novel effective strategy for optimizing tumor targeting and maximizing potential clinical benefits. Human domain antibodies selected against VEGF and EGFR were formatted into a fully human dual‐targeting IgG (DT‐IgG) to directly target both antigens in a single molecule. We evaluated the efficacy of DT‐IgG in comparison with bevacizumab and cetuximab alone and in combination in the lung cancer cell line A549 (low EGFR expression and KRAS mutant) and the head and neck squamous cell carcinoma (HNSCC) cell line Tu212 (high EGFR expression and KRAS wild type) in vitro and in vivo. DT‐IgG suppressed Tu212 and A549 cell growth, inhibited EGFR activation and induced apoptosis as effectively as cetuximab, and neutralized VEGF as effectively as bevacizumab. DT‐IgG induced EGFR‐dependent VEGF internalization, constituting a novel antiangiogenesis mechanism. In xenograft models with lung and head and neck cancer cell lines, DT‐IgG displayed efficacy equivalent to bevacizumab in diminishing tumor growth despite its short serum half‐life (36 hr in rats) and both agents may constitute preferable alternatives to cetuximab in KRAS‐mutant tumors. Immunofluorescence staining revealed that localization of DT‐IgG was similar to that of cetuximab, largely associated with EGFR+tumor cells. Our proof of principle study suggests a DT‐IgG against EGFR and VEGF as an alternative therapeutic strategy with potentially enhanced clinical benefit.

Novel Bispecific Domain Antibody to LRP6 Inhibits Wnt and R-spondin Ligand-Induced Wnt Signaling and Tumor Growth.

Aberrant WNT signaling is associated with the formation and growth of numerous human cancer types. The low-density lipoprotein receptor-related protein 6 (LRP6) is the least redundant component of the WNT receptor complex with two independent WNT ligand-binding sites. Using domain antibody (dAb) technology, a bispecific antibody (GSK3178022) to LRP6 was identified that is capable of blocking stimulation in the presence of a range of WNT and R-spondin (RSPO) ligands in vitro. GSK3178022 was also efficacious in reducing WNT target gene expression in vivo, in both cancer cell line and patient-derived xenograft models, and delays tumor growth in a patient-derived RSPO fusion model of colorectal cancer. Implications: This article demonstrates the inhibition of a key oncogenic receptor, intractable to mAb inhibition due to multiple independent ligand interaction sites, using an innovative dAb approach. Mol Cancer Res; 14(9); 859–68. ©2016 AACR.

Pharmacodynamics of DT-IgG, a dual-targeting antibody against VEGF-EGFR, in tumor xenografted mice

PurposeDT-IgG is a fully humanized dual-target therapeutic antibody being developed to simultaneously target epidermal growth factor receptor (EGFR) and vascular endothelial growth factor (VEGF), important signaling molecules for tumor growth. The antitumor pharmacodynamics (PD) of DT-IgG was studied in nude mice bearing human tumor xenografts with different EGFR and VEGF expressions and K-ras oncogene status and compared with bevacizumab, cetuximab and bevacizumabxa0+xa0cetuximab.MethodsMice bearing human oral squamous cell carcinoma (Tu212), lung adenocarcinoma (A549), or colon cancer (GEO) subcutaneous xenografts were administered with the antibodies intraperitoneally (i.p.), and tumor volumes were measured versus time. Nonlinear mixed effects modeling (NONMEM) was used to study drug potencies (IC50) and variations in tumor growth.ResultsThe PD models adequately described tumor responses for the antibody dose regimens. In vivo IC50 values varied with EGFR and K-ras status. DT-IgG had a similar serum t1/2 as cetuximab (~1.7 vs. 1.5xa0day), was more rapid than bevacizumab (~6xa0day), and had the largest apparent distribution volume (DT-IgGxa0>xa0cetuximabxa0>xa0bevacizumab). The efficacy of DT-IgG was comparable to bevacizumab despite lower serum concentrations, but was less than bevacizumabxa0+xa0cetuximab.ConclusionsA lower IC50 of DT-IgG partially compensated for lower serum concentrations than bevacizumab and cetuximab, but may require higher doses for comparable efficacy as the combination. The model adequately predicted variations of tumor response at the DT-IgG doses tested and could be used for targeting specific tumor efficacies for future testing.

'In-Format' screening of a novel bispecific antibody format reveals significant potency improvements relative to unformatted molecules.

ABSTRACT Bispecific antibodies (BsAbs) are emerging as an important class of biopharmaceutical. The majority of BsAbs are created from conventional antibodies or fragments engineered into more complex configurations. A recurring challenge in their development, however, is the identification of components that are optimised for inclusion in the final format in order to deliver both efficacy and robust biophysical properties. Using a modular BsAb format, the mAb-dAb, we assessed whether an ‘in-format’ screening approach, designed to select format-compatible domain antibodies, could expedite lead discovery. Human nerve growth factor (NGF) was selected as an antigen to validate the approach; domain antibody (dAb) libraries were screened, panels of binders identified, and binding affinities and potencies compared for selected dAbs and corresponding mAb-dAbs. A number of dAbs that exhibited high potency (IC50) when assessed in-format were identified. In contrast, the corresponding dAb monomers had ∼1000-fold lower potency than the formatted dAbs; such dAb monomers would therefore have been omitted from further characterization. Subsequent stoichiometric analyses of mAb-dAbs bound to NGF, or an additional target antigen (vascular endothelial growth factor), suggested different target binding modes; this indicates that the observed potency improvements cannot be attributed simply to an avidity effect offered by the mAb-dAb format. We conclude that, for certain antigens, screening naïve selection outputs directly in-format enables the identification of a subset of format-compatible dAbs, and that this offers substantial benefits in terms of molecular properties and development time.