A dual-targeting PDGFRβ/VEGF-A molecule assembled from stable antibody fragments demonstrates anti-angiogenic activity in vitro and in vivo

Abstract

Targeting angiogenesis is a promising approach to the treatment of solid tumors and age-related macular degeneration (AMD). Inhibition of vascularization has been validated by the successful marketing of monoclonal antibodies (mAbs) that target specific growth factors or their receptors, but there is considerable room for improvement in existing therapies. Combination of mAbs targeting both the VEGF and PDGF pathways has the potential to increase the efficacy of anti-angiogenic therapy without the accompanying toxicities of tyrosine kinase inhibitors and the inability to combine efficiently with traditional chemotherapeutics. However, development costs and regulatory issues have limited the use of combinatorial approaches for the generation of more efficacious treatments. The concept of mediating disease pathology by targeting two antigens with one therapeutic was proposed over two decades ago. While mAbs are particularly suitable candidates for a dual-targeting approach, engineering bispecificity into one molecule can be difficult due to issues with expression and stability, which play a significant role in manufacturability. Here, we address these issues upstream in the process of developing a bispecific antibody (bsAb). Single-chain antibody fragments (scFvs) targeting PDGFRbeta and VEGF-A were selected for superior stability. The scFvs were fused to both termini of human Fc to generate a bispecific, tetravalent molecule. The resulting molecule displays potent activity, binds both targets simultaneously, and is stable in serum. The assembly of a bsAb using stable monomeric units allowed development of an anti-PDGFRB/VEGF-A antibody capable of attenuating angiogenesis through two distinct pathways and represents an efficient method for rapid engineering of dual-targeting molecules.

Hypoxia promotes adipose-derived stem cell proliferation via VEGF

“…Thus, we evaluated PDGFRβ expression in hypoxic ADSCs and showed that ADSCs highly expressed this receptor. In previous reports, PDGFRβ has been demonstrated as a facultative receptor for VEGF (Ball et al, 2007;Mabry et al, 2010;Pfister et al, 2012). Ball et al (2007) have shown that VEGF-A could bind to both PDGFRα and PDGFRβ, inducing tyrosine phosphorylation.…”

Nanocell targeting using engineered bispecific antibodies

“…21 To produce more complex BsAbs compared to the tandem scFv BsAb, manipulation of the standard IgG mAb format has produced a plethora of Fc-containing BsAbs, whereby designs generally rely on the principle that Fc-containing BsAbs will dimerize through the Fc, similar to native, whole mAbs. 11,46,47,63,64 Features of these antibodies include engineering Fc components to improve or remove various related functions, forced heterodimerisation, removal of redundant domains and addition of other binding motifs to the IgG molecule. 47,63,[65][66][67] We used an IgG1 Fc-domain to link and separate the 2 scFvs spatially (ABX-EGF scFv at the N-terminus of human IgG1 Fc domain, and an 1H10 scFv at the C-terminus) to create the ABX-EGF-Fc-1H10 homodimeric Fc-containing BsAb, similar to the scFv-Fc-scFv BsAb dimer described in Jendreyko et al and the Emergent Biosolutions TM product termed ADAPTIR TM Multi-Specific.…”

Dual targeting strategies with bispecific antibodies

“…98,99 In another study, a bispecific antibody was applied to simultaneously inhibit PDGFRβ and VEGF-A. 100 The antibody was generated by fusing different scFvs to the N-terminus and C-terminus of an Fc fragment (scFv-Fc-scFv), resulting in a tetravalent, bispecific molecule with the two chains covalently linked by disulfide bonds of the hinge region. This bispecific antibody inhibited For example, dual targeted toxins were generated fusing IL-13 and EGF with Diphtheria toxin.…”