Generation of a Porcine Antibody Fab Fragment Using Protein Engineering to Facilitate the Evaluation of Ocular Sustained Delivery Technology

Mol Pharm. 2022 May 2;19(5):1540-1547. doi: 10.1021/acs.molpharmaceut.2c00048. Epub 2022 Apr 8.

Abstract

Treatment of age-related macular degeneration (AMD) with anti-vascular endothelial growth factor (VEGF) biologic agents has been shown to restore and maintain visual acuity for many patients afflicted with wet AMD. These agents are usually administered via intravitreal injection at a dosing interval of 4-8 weeks. Employment of long-acting delivery (LAD) technologies could improve the therapeutic outcome, ensure timely treatment, and reduce burden on patients, caregivers, and the health care system. Development of LAD approaches requires thorough testing in pre-clinical species; however, therapeutic proteins of human origin may not be well tolerated during testing in non-human species due to immunogenicity. Here, we have engineered a surrogate porcine antibody Fab fragment (pigG6.31) from a human antibody for testing ocular LAD technologies in a porcine model. The engineered Fab retains the VEGF-A-binding and inhibition properties of the parental human Fab and has stability properties suitable for LAD evaluation. Upon intravitreal injection in minipigs, pigG6.31 showed first-order clearance from the ocular compartments with vitreal elimination rates consistent with other molecules of this size. Application of the surrogate molecule in an in vivo evaluation in minipigs of a prototype of the port delivery (PD) platform indicated continuous ocular delivery from the implant, with release kinetics consistent with both the results from in vitro release studies and the efficacy observed in human clinical studies of the PD system with ranibizumab (PDS). Anti-drug antibodies in the serum against pigG6.31 were not detected over exposure durations up to 16 weeks, suggesting that this molecule has low porcine immunogenicity.

Keywords: long-acting delivery; ocular clearance; pharmacokinetics.

MeSH terms

  • Angiogenesis Inhibitors*
  • Animals
  • Humans
  • Immunoglobulin Fab Fragments / therapeutic use
  • Intravitreal Injections
  • Protein Engineering
  • Ranibizumab / therapeutic use
  • Swine
  • Swine, Miniature / metabolism
  • Technology
  • Vascular Endothelial Growth Factor A / metabolism
  • Wet Macular Degeneration* / drug therapy

Substances

  • Angiogenesis Inhibitors
  • Immunoglobulin Fab Fragments
  • Vascular Endothelial Growth Factor A
  • Ranibizumab