in pathologic conditions while preserving a wide range of VEGF-mediated physiological processes associated with the smaller isoforms [47–51].

Published Trials

The landmark phase III VEGF Inhibition Study In Ocular Neovascularization (VISION) trials were multicenter, dose-ranging studies that enrolled subjects with a wide range of neovascular lesions, including all angiographic subtypes and lesions up to 12 disc areas in size (including blood, scar, or atrophy, and neovascularization). Patients in the study had a best corrected visual acuity (VA) of 20/40–20/320 in their study eye [52, 53]. Subjects received sham injections or injections of intravitreal pegaptanib sodium (0.3, 1, or 3 mg) every 6 weeks for 54 weeks for the first year of the study.

In the combined trials, 1,186 subjects were enrolled and the results showed that intravitreal pegaptanib sodium decreased the loss vision, with 70% of treated patients losing fewer than 15 letters of visual acuity compared with 55% of controls [53, 54]. Moreover, 6% of pegaptanib sodium-treated patients gained at least 15 letters compared with 2% of the patients in the control group [53]. Fluorescein angiography at 30 and 54 weeks showed that the pegaptanib-treated group had a significant reduction (P < 0.01) in the rate of growth in the total area of their CNV and in the severity of leakage compared with the control group [53]. The 0.3 mg dose appeared more effective than the 1 mg or 3 mg doses, so the 0.3 mg became the FDA-approved dose.

Bevacizumab

Drug Overview

Research from the 1980s and 1990s has shown that VEGF inhibition using a murine and humanized monoclonal antibody against VEGF markedly suppressed tumor growth in vivo, thereby setting the stage for the development of bevacizumab. Bevacizumab is a humanized monoclonal antibody (IgG1) against human VEGF-A that selectively inhibits all isoforms and bioactive proteolytic breakdown products of VEGF-A. Bevacizumab is

an immunoglobulin G molecule that is comprised of amino acid sequences, which are about 93% human and 7% murine. FDA-approved in 2004 as treatment for metastatic colorectal cancer, bevacizumab was given intravenously at a dose of 5 mg/ kg and infused every two weeks in combination with 5-fluorouracil. Additional phase III clinical trials with bevacizumab have since resulted in FDA approval for the treatment of breast, lung, kidney, and brain cancers [55]. No evidence of an antibody immunogenic response to bevacizumab has been found in any clinical trials, confirming the success of the humanization technique.

Bevacizumab is commercially available as 100 and 400 mg preservative-free, single-use vials in a volume of 4 or 16 mL (25 mg/mL). The 100 mg product is formulated in 240 mg a(alpha),a(alpha)-trehalose dihydrate, 23.2 mg sodium phosphate (monobasic, monohydrate), 4.8 mg sodium phosphate (dibasic, anhydrous), and 1.6 mg polysorbate 20, and should be diluted in water prior to intravenous infusion. For off-la- bel ophthalmic use, bevacizumab is not diluted, but rather dispensed into individual syringes for intravitreal injection and the volume (dose) of injection ranges from 0.05 mL (1.25 mg) to 0.1 mL (2.5 mg).

The intravitreal pharmacokinetics of monoclonal antibodies was initially studied in monkeys and rabbits and found to be about 5.6 days [56, 57]. Bevacizumab has since been studied experimentally in rabbits and is shown to have a half-life of 4.32 days [58]. When 1.25 mg was injected in rabbits, concentrations of over 10 m(mu)g/mL bevacizumab were maintained in the vitreous for at least 30 days. Additional animal studies in rabbits and monkeys suggest a half-life of bevacizumab in the range of four– six days [59, 60]. A human study reported that the half-life of intravitreal bevacizumab was approximately three days, and also showed that a single dose of intravitreal bevacizumab was likely to provide complete intravitreal VEGF blockade for a minimum of four weeks [61]. Another human study has suggested a half-life of 6.7 days while yet another study has reported a half-life of as long as 9.8 days [62, 63]. The exact half-life of bevacizumab in the eye is uncertain at this time

and may vary depending on the extent of vitreous liquefication and the phakic status of the eye.

Bevacizumab has a molecular weight of approximately 149 kDa and there was a question of whether such a large molecule could penetrate the retina. Han et al. were the first to show that a full-length immunoglobulin was capable of penetrating the rabbit retina after an intravitreal injection [64]. Subsequently, Sharar et al. used qualitative immunofluorescence to show that intravitreal bevacizumab was able to completely penetrate the retina by 24 h and was essentially absent at four weeks after an injection [65] Moreover, Dib et al. demonstrated subretinal detection of bevacizumab after an intravitreal injection in rabbit eyes. They detected bevacizumab molecules in the subretinal space of all six eyes studied 2 h after an intravitreal bevacizumab injection of 0.05 mL (1.25 mg), suggesting that the molecule could rapidly diffuse through the retina [66].

Published Studies

Although systemic bevacizumab (5 mg/kg) was shown to reduce leakage from CNV, decrease central retinal thickness (CRT) using optical coherence tomography (OCT), and significantly improve vision in neovascular AMD [67–70], the intravenous use of bevacizumab for neovascular AMD was never widely adopted because the intravitreal approach uses up to 500-fold less drug, is much less expensive, and is perceived to be safer due to the smaller dose of drug. In the first-reported case [71] of intravitreal bevacizumab, a patient with recurrent CNV secondary to AMD, who had previously been treated with verteporfin photodynamic therapy (PDT) in combination with triamcinolone acetonide and then treated with pegaptanib injections, was shown to experience a reduction in retinal thickness with resolution of subretinal fluid using OCT imaging and the visual distortion resolved within one week following a single injection of 1.0 mg bevacizumab. Subsequently, several retrospective [72–86] and prospective [73, 87–97] studies of intravitreal bevacizumab (dose range 1.0–2.5 mg) in neovascular AMD patients have been published, all demonstrating clinically significant

improvement in mean visual acuity, reduction in fluorescein angiographic leakage, resolution of OCT-visualized edema in up to 90% of bevaci- zumab-treated patients, and apparent overall clinical safety (see example of treatment effect in Fig. 6.1). Most studies have been small (up to 100 patients), uncontrolled studies with different retreatment criteria and outcome measures.

A randomized, prospective clinical trial compared verteporfin PDT with bevacizumab (2.5 mg) for the treatment of predominantly classic CNV secondary to AMD and found that at month 6, all 32 eyes (100%) receiving bevacizumab lost fewer than15 letters of visual acuity compared with 73.3% of the PDT-receiving eyes (P = 0.002) [98]. The OCT outcomes were significantly better at 3 and 6 months in patients treated with bevacizumab versus the PDT group (P = 0.04 and P = 0.002, respectively). The study showed overall benefit of treatment with bevacizumab compared with PDT. Another study showed the effect of previous PDT treatment on the response to bevacizumab injections. The authors compared treatment-naïve eyes (80) with eyes previously treated with PDT (29) and showed that both groups had equal anatomic and functional improvements. However, the eyes previously treated with PDT required fewer injections (4.22) when compared with treatment-naïve eyes (6.13) [99]. The rationale of using combined therapy is to either reduce the number of antiVEGF injections in wet AMD [98–100], or to improve the efficacy of anti-VEGF treatment in cases of exudative maculopathy such as polypoidal choroidal vasculopathy [100]. Combined therapy is now being evaluated by several prospective randomized trials and Chapter 7 discusses this topic in greater detail.

Systemic and ocular adverse events (AEs) attributable to intravitreal bevacizumab have been rare with the most common ocular side effects being endophthalmitis, uveitis, submacular hemorrhage, and RPE tears. In a recent retrospective safety assessment of intravitreal bevacizumab involving 1,173 patients, there were 18 (1.5%) reported systemic AEs, including five deaths (0.4%) and the ocular AEs included subconjunctival hemorrhage [838 cases (19% of 4,303

Fig. 6.1 A 72-year-old woman with age-related macular degeneration diagnosed with an occult lesion and a vascularized retinal pigment epithelium detachment (PED) in the right eye. She received four-monthly bevacizumab injections and then was treated every three months. Color fundus images with late-phase images of the fluorescein

and indocyanine angiograms at baseline are shown. Optical coherence tomography (OCT) responses from baseline to one year after the last bevacizumab injection are shown. Horizontal (left) and vertical (right) OCT B-scans through the central macula and visual acuity are shown. Resolution of the PED was observed at the last follow-up visit

injections)], increased intraocular pressure (IOP), endophthalmitis, and tractional retinal detachment [seven cases (0.16%)] each [85]. The low rates of systemic complications in these studies

were consistent with the rates of these lifethreatening adverse events in the general, untreated population and those reported in an earlier survey of 5,228 patients [101].