The study described here had two major goals: to characterize the pathophysiological role of TNF-α versus VEGF in the development of laser-induced CNV in cynomolgus monkeys and to explore the feasibility of preventing such CNV formation by topical administration of a TNF-inhibitory scFv antibody fragment.
Inhibition of VEGF by intravitreal ranibizumab injection is highly effective in preventing CNV formation in this model.
19 As opposed to bevacizumab, a full-size antibody, ranibizumab is a Fab antibody fragment of approximately 48 kDa molecular weight.
34,35 This may translate to a reduced intravitreal half-life of the molecule compared with conventional full-length monoclonal antibodies that have a molecular weight of approximately 150 kDa.
31 Such pharmacokinetic differences between antibody formats may possibly influence duration of effect after intravitreal administration.
36 When the experiment was conducted, all marketed anti–TNF-α antibodies were of the full-length format, (infliximab, adalimumab). Consequently, we decided to use the full-length anti-VEGF antibody bevacizumab (which is highly effective in the treatment of AMD as well) instead of ranibizumab as a positive VEGF inhibitor control in our experiment. Because infliximab has an extremely narrow species specificity, neutralizing only human and chimpanzee TNF-α,
28 we used adalimumab to characterize the role of TNF-α in this cynomolgus model. This choice allowed exclusion of potential antibody format-specific effects when assessing the pathophysiological role of TNF-α because two potent molecules with comparable intravitreal pharmacokinetics (bevacizumab, adalimumab) but different target specificity were used. Hence, monkeys were treated with intravitreal injection of 1 mg bevacizumab or adalimumab, respectively, 1 week before (day 1) and 1 week (day 15) and 3 weeks (day 29) after argon green laser photocoagulation to induce CNV lesions. Development of grade 4 lesions was assessed by fluorescence angiography 2, 3, and 4 weeks (days 22, 28, and 36) after laser treatment, as described previously.
19 Intravitreal bevacizumab completely blocked the formation of grade 4 lesions at all times. Intravitreal adalimumab was clearly less effective than bevacizumab but reduced the number of laser-induced grade 4 lesions by >50%, demonstrating an important contributing pathophysiological role of TNF-α in the model (
Fig. 1).
We also injected ESBA105,
31–33 a potent anti–TNF-α single-chain (scFv) antibody, intravitreally into the eyes of monkeys. ESBA105, based on its low molecular weight of 27 kDa, carries entirely distinct pharmacokinetic properties and represents the first topically applied antibody in clinical trials in ophthalmology.
37,38 Intravitreal ESBA105 was injected at the same time points as bevacizumab and adalimumab. Given that the intravitreal half-life of ESBA105 in rabbits was determined to be in the range of 1 day only,
31 whereas the intravitreal half-life of full-length antibodies (such as bevacizumab and adalimumab) was determined to be in the range of 5.5 to 8.5 days.
34,39,40 ESBA105 (2 mg) was injected at each of the dosing events to partially compensate for the anticipated lower exposure. Similar to adalimumab, intravitreal ESBA105 reduced the formation of grade 4 CNV lesions at all times by approximately half (
Fig. 1), confirming previous data demonstrating that ESBA105 shows in vivo potency similar to that of marketed TNF-α inhibitors in suitable animal models.
33
Most important, these data on intravitreal adalimumab and ESBA105 define the anticipated maximal pharmacologic effect size for the prevention of grade 4 lesion formation that a TNF-α inhibitor can possibly achieve in this model because the expected minimal concentration (C
min) in the vitreous throughout this study is at least equal to that of adalimumab during steady state in rheumatoid arthritis therapy.
41 This definition was crucial because the second objective of this study was to assess the feasibility of topical administration of a suitable scFv antibody fragment in this model.
Thus, another group of animals was treated with 10 daily eyedrops of ESBA105 for the entire study duration. We found that topical ESBA105 reduced the number of developing grade 4 lesions by approximately 30% in this experiment compared with saline-treated control animals (
Fig. 1).