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L.L. Wei, M. Hamilton, D. McVey, C.R. King, D. Brough; Repeat Dosing of Adenovector in the Eye . Invest. Ophthalmol. Vis. Sci. 2004;45(13):5057.
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Purpose: To determine the feasibility of repeat dosing with an adenovector administered intraocularly to the eye and to test whether transgenes can be expressed from an adenovector expression system in the presence of pre–existing neutralizing antibodies to adenovectors. Methods: To assess whether repeat administration of an adenovector (Ad) is possible using intravitreous (IVT) injection, adult C57BL/6 mice were injected with AdNull (empty cassette) on Day 1 and then 2 wks later with AdLuciferase (AdL). Animals were sacrificed 24 hrs post–injection, eyes enucleated, and stored until assayed. As a positive control, some mice were injected intravitreously with AdL only on Day 1, sacrificed at 24 hrs, and eyes collected. As a negative control, naïve eyes were used. Serum samples were also taken to determine if intravitreal adenovector injections lead to induction of neutralizing Ad–antibody. Neutralizing Ad–antibody titres were determined by interference of green fluorescent protein expression. To determine if pre–existing neutralizing Ad–antibody would block transgene expression, animals were pre–immunized by intramuscular (IM) injection with AdNull (1 e 10 pu/animal). Fourteen days later, when circulating neutralizing antibody titres are expected to exist, mice were given a single IVT injection of AdL, animals sacrificed, and eyes collected for further analyses. Moreover, to rigorously test the effect of pre–immunization of animals, mice were pre–immunized with 2 or 3 IM exposures to AdNull. In both cases, AdL was administered 2 wks post–IM injection, animals sacrificed at 24 hrs, and eyes collected. Results: Repeat dose administration experiments showed that IVT injection with adenovectors was possible at 2 wk intervals. Moreover, measurement of neutralizing antibody titres revealed that single and multiple adenovector injections did not result in major increases of neutralizing antibody titres in all animals. Interestingly, studies also indicate that transgene expression from non–immunized animals versus animals that had been pre–immunized up to 3 boosts were the same. Conclusions: These studies provide evidence that repeat adenovector administration in to the eye, an immunoprivileged organ, is possible and that IVT injection does not cause a major induction of circulating neutralizing antibody titres as measured in serum. Moreover, pre–immunization of animals by IM delivery of adenovector, likewise, does not impair transgene expression in the eye. These studies indicate that repeat administration of adenovectors into the eye is likely to be possible in humans and that the pre–existence of neutralizing Ad–antibody will likely not affect transgene expression.
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