206. Frequency and Analysis of Liver Tumors in AAV-Treated B6C3F1 Hybrid Mice

Abstract

Top of pageAbstract We report on the frequency of liver tumors observed in a gene therapy study with AAV vectors in male mice of the B6C3F1 hybrid background, which are known to have a high frequency of spontaneous tumor formation in the liver. Because of their increased susceptibility to develop tumors, these animals should serve as sensitive models to reveal any potential tumorigenic properties of AAV vectors. Male mice with mutations in their OTC gene (spf and spf-ash) and their wild-type siblings had received AAV vectors expressing either the murine OTC (mOTC) or LacZ gene. Untreated control animals were included in the study. All experimental groups including wild-type and mOTC-deficient animals not treated with vector developed liver nodules. In the majority of the animals, H&E-stained liver samples were available that allowed to score the observed nodules according to their likelihood to represent hepatocellular carcinomas. In the spf group no statistically significant risk differences were found between vector-treated and untreated animals. Likewise, in the spf-ash group the administration of vectors expressing mOTC did not increase the risk of tumor development. However, these mice showed increased risks of tumor formation when treated with LacZ-expressing vectors compared to untreated animals or animals that had received vectors with mOTC as transgene, suggesting that the transgene but not the vector might play a role in the formation of tumors. In general, both the overall risk of developing liver nodules and the histological appearance as hepatocellular carcinoma was more frequent in mice that had received AAV-LacZ compared to untreated controls or animals treated with AAVmOTC. Tumor tissue contained consistently lower amounts of vector DNA than healthy liver tissue, and only one out of 16 tumors was found to express the transgene as determined by histochemistry to detect OTC activity and immunostaining against beta-galactosidase. We therefore conclude that AAV vectors alone do not contribute to the formation of tumors, even in mouse strains with a heightened susceptibility to the development of those lesions. However, the expression of LacZ alone or in combination with vector may be problematic.