Purpose: To develop an injectable and long-acting poly(lactic-co-glycolic acid) (PLGA) implant formulation of anti-vascular endothelial growth factor (VEGF) monoclonal antibody (MAb, bevacizumab, Avastin®) to reduce administration frequency for treatment of wet age-related macular degeneration (wet AMD).
Methods: Preparation of implants: MAb powder (trehalose:MAb = 1.5:1 w/w) was suspended in PLGA 50/50 (Mw = 54 kD) solution in acetone with 3% MgCO3 as an antacid to control microclimate pH. The suspension was then extruded in silicone tubing (I.D. = 0.8 mm) and dried to obtain core cylindrical implants. For coated implants, a pure PLGA solution at various concentrations in acetone was extruded over the dried core implants in the tubing to coat the surface and dried.
In vitro release study: Release kinetics of MAb from the PLGA implants in PBST (PBS, 0.02 % Tween 80) at 37 °C without agitation was measured by size-exclusion chromatography (SEC). Monomer content, secondary structure and immunoreactivity of the encapsulated and released MAb were evaluated by SEC, circular dichroism (CD) and enzyme-linked immunosorbent assay (ELISA), respectively.
Anti-VEGF efficacy evaluation in a rabbit retinal vascular leakage model: Anti-VEGF efficacy of the PLGA-bevacizumab implants was examined in the rabbit VEGF-induced retinal vascular leakage model (male Dutch Belted rabbits, 2 animals, 4 eyes, 8 blood vessels/group). The 30 % PLGA coated implants (400 µg of bevacizumab) were injected into the rabbit vitreous. To compare with free MAb, the same dose of Avastin® solution was injected into the free MAb control group. The no-treatment control group received no drug. On day 42, recombinant human VEGF (1000 ng) was injected into each eye to induce retinal leakage. On day 45, fluorescein angiography (FA) was performed and the degrees of fluorescein leakage and tortuosity of retinal blood vessels were scored on 4-point scale. On days 56 and 70, the same assay was performed for the implant group. For these time points, two untreated rabbits were added as a new control group at each time point and the previous no-treatment control and free MAb groups were not tested further since their retinal blood vessels were damaged during the 42-day VEGF challenge.
Results: The optimized implant formulations with 30-50 % PLGA coating concentration achieved high MAb loading (7.6 to 8.2 % w/w) and demonstrated continuous in vitro release kinetics over six weeks with a total cumulative release of 82 ± 8 to 89 ± 4 %. Little changes in monomer content, immunoreactivity, and secondary structure of the released MAbs during the 6-week in vitro release period were observed. Anti-VEGF efficacy of the optimized MAb implant was compared to the same dose of free MAb in the rabbit VEGF-induced retinal leakage model. Six weeks after intravitreal injection of both formulations, only the MAb implant protected retinal blood vessels while significant leakage was observed in the no-treatment control and free MAb groups. The retinal blood vessels of the implant group were still protective over 8 weeks, but not at 10 weeks.
Conclusion: This approach may be useful for development of long-acting anti-VEGF therapy. Owing to the structural similarity of therapeutic monoclonal antibodies, the coated PLGA implants with addition of sugars and antacids as stabilizers have the potential to serve as delivery platforms for sustained release of other therapeutic monoclonal antibodies as well.
Jennifer Walker– University of Michigan, Ann Arbor, Michigan
David Antonetti– Professor, Department of Ophthalmology and Visual Sciences, University of Michigan, Michigan
Jeffrey Jamison– CEO, Ophthy-DS, Inc., Michigan
Steven Schwendeman– Professor, Department of Pharmaceutical Sciences and the Biointerfaces Institute, University of Michigan, Michigan