Evaluating the Potential of Germ Line Transmission after Intravenous Administration of Recombinant Adenovirus in the C3H Mouse

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Abstract
The goal of this study is to assess the likelihood that an adenoviral vector disseminated to gonads will be transmitted to offspring. This study is based on the observation that systemically administered vector can be detected in both ovaries and testes, using sensitive nested PCR techniques. Although the extent of vector dissemination to gonads is extremely small, as it is detectable only by nested PCR, it is unclear where it is located within these tissues and whether the DNA is capable of integration and transmission to offspring. A protocol was developed in C3H mice to address this question. Both male and female C3H mice were injected with a high dose of H5.001CBhOTC, an E1- and E4-deleted vector expressing human ornithine transcarbamylase. This dose of vector was sufficient to target 80% of hepatocytes (Gao et al., J. Virol. 1996; 70:8934–8943) and disseminate, at low levels, to both ovaries and testes in 94% of animals as determined by PCR. Vector-administered animals and controls were mated and 814 offspring were evaluated for germ line transmission of the adenoviral vector by DNA hybridization of total cellular DNA extracted from the fetus. Southern blot analysis showed no evidence of germ line transmission in 578 offspring of crosses in which either one or both parents received recombinant adenovirus. Direct administration of vector to the patient (i.e., in vivo gene therapy) will be required in the treatment of most diseases in which gene transfer is employed. An important safety concern is the distribution of vector beyond the target organ. This is particularly important if vector distributes to gonads, raising the possibility of inadvertent germ line transmission. We evaluated the potential of this scenario in which E1- and E4-deleted vector was administered at high doses in the venous blood of C3H mice. Vector did distribute to both ovaries and testes, although no vector sequence was detected in the offspring by Southern blot analysis. This study would predict that inadvertent germ line transmission of adenovirus is an unlikely event.