Photodynamic Therapy of Rabbit Corneal Neoascularization with Photosensitizer Conjugated to Antibody against the Extradomain–B of Fibronectin

Abstract

Abstract: : Purpose:The extradomain B (ED–B) containing fibronectin isoform (B–FN) is strongly expressed in choroidal neovascular membranes and is a candidate for targeted drug delivery to the neovasculature. Our objective was to evaluate the efficacy of an immunoconjugate composed of the photosensitizer (PS) Sn(IV)chlorin e6 and the high affinity human antibody against B–FN named SIPL19, in the photodynamic therapy of experimental corneal neovascularization in the rabbit. Methods: Immunoconjugate was prepared by incubating PS in the monoester form with SIPL19 and purified from non conjugated photosensitizer by size exclusion chromatography. The conjugate was characterized by its antibody/photosensitizer stechiometric ratio, migration profile in acrylamide gel and fast protein liquid chromatography (FPLC), immunoreactivity and capacity to produce singlet oxygen. Ocular biodistribution of radioisotope–labeled SIPL19 was studied in the rabbit corneal micropocket assay. Photodynamic therapy (PDT) of the corneal neovasculature was also performed with escalating light doses. PDT was followed–up by color photography, fluorescein angiography and histology. The formation of anti–immunoconjugate antibodies was evaluated by surface plasmon resonance. Results: The immunoconjugate migrated as a single band of 80kD, had an immunoreactivity of >75% and an antibody/PS ratio of 1/3.1 and selectively accumulated around newly developed corneal vessels as early as 4 hours post–administration. Occlusion of corneal neovascularization was obtained after 635nm diode laser irradiation with energies >=150J/cm2 at a fluence rate of 600mW/cm2. No antibodies were detected against immunoconjugates composed of photosensitizer and rabbit IgG. Conclusions:PDT of newly developed vessels was effective with the Sn(IV)chlorin e6/SIPL19 immunoconjugate in the rabbit corneal micropocket assay. We conclude that SIPL19–mediated photosensitizer delivery may be beneficial in the treatment of ocular neovascular diseases.

Keywords: photodynamic therapy • neovascularization • extracellular matrix