Adenovirus-Mediated Gene Transfer to the Respiratory Tract of Fetal Sheep In Utero

Abstract

Many human genetic diseases, such as congenital surfactant protein B deficiency, manifest in the perinatal period. Prenatal gene therapy may be necessary to minimize morbidity in these diseases. We hypothesized that bacterial β-galactosidase (β-Gal) gene could be transferred to and expressed in the pulmonary epithelium of fetal sheep in utero using a replication-deficient adenovirus (Av1LacZ4). We instilled Av1LacZ4 (1.5 × 10¹¹ plaque-forming units, n = 10) or saline (n = 2) intratracheally to chronically instrumented fetal sheep at 112–134 days gestation (term = 145 days). Lung fluid was collected before and after Av1LacZ4 administration for cytological analysis. Lung tissue was examined for transgenic β-Gal activity and evidence of toxicity. Transgenic β-Gal activity was visualized as blue nuclear staining of tissue treated with X-Gal and was detected in the lungs of 5 animals for up to 14 days after administration. Transgenic β-Gal activity was not detected in the lungs of animals analyzed beyond 14 days after treatment. Pulmonary histopathology was detected in most Av1LacZ4-treated animals and manifested as a mixed cellular infiltrate consisting of neutrophils, macrophages, and lymphocytes. Fetal lung fluid analysis revealed a predominantly lymphocytic response in most Av1LacZ4-treated animals within 3 days (2.88 × 10⁶ vs. 4 × 10³ total cells/ml in control animals). We have demonstrated that adenovirus vectors can direct gene transfer to the lungs of fetal sheep in utero. The transferred gene expression was transient and possibly limited by the induced inflammatory response.

Overview summary

Certain diseases, such as cystic fibrosis and congenital surfactant protein B deficiency, have significant morbidity and mortality soon after birth. It is desirable to initiate treatment early in development before chronic lung damage develops. The purpose of this study was to determine the potential utility of adenoviral vectors to mediate gene transfer in the developing fetus. A replication-deficient adenoviral vector, Av1LacZ4, transferred expression of bacterial β-Gal throughout the lungs of fetal sheep. Gene expression was detectable between 3 and 14 days after vector administration. These studies provide the experience and insight for future strategies to treat lung diseases initiated during the perinatal period. The general approach used to transfer gene expression to the fetal lung may be applied to future strategies for gene transfer to other fetal organs.