Fetal gene therapy: Efficacy, toxicity, and immunologic effects of early gestation recombinant adenovirus

Background/Purpose: Advancements in gene transfer technology and prenatal d iagnosis have allowed investigators to consider an in utero gene therapy ap proach for fatal genetic diseases. The authors sought to develop fetoscopic techniques for gene delivery and investigate the efficacy and safety of re combinant adenoviral vectors in the fetus. Methods: Fetal sheep between 60 and 130 days' gestation (dGA) underwent eit her fetoscopic intratracheal injection or umbilical Vein (UV) injection of recombinant adenovirus, AdCMV-lacZ At death, fetal organs were examined for P-galactosidase expression, histopathology, and CD45 immunostaining. Fetal serum was compared with preimmune serum for transaminase levels and the pr esence of antiadenoviral neutralizing antibodies. Results: Fetoscopic intratracheal delivery of AdCMVlacZ in late-gestation s heep fetuses resulted in efficient alveolar gene transfer, but, antiadenovi ral immunologic reactions limited the longevity of transgene expression to 14 days. This prompted an examination of whether early gestational exposure could induce tolerance in the fetus to adenoviral and transgene antigens. AdCMVlacZ (1 x 10(11) particles) was injected via UV into fetuses at 60 dGA . Within 3 days, beta-galactosidase expression was localized to the fetal l iver, adrenal glands, kidneys, and endocardium. Although adrenal expression was nearly constant over 28 days, expression in fetal liver disappeared wi thin 14 to 28 days. Loss of hepatic expression did not appear to be immune mediated because there was no evidence of hepatic inflammation or appearanc e of antiadenoviral neutralizing antibodies. Fetuses injected with AdCMVlac Z at 60 dGA were reinjected with 1 X 10(13) particles at 125 dGA and antiad enoviral humoral immune responses were recorded. Despite early-gestation ad enovirus injection, fetuses still responded to the late-gestation adenovira l exposure, developing antiadenoviral neutralizing antibodies similar to co ntrol fetuses. Conclusions: The authors developed fetoscopic access for pulmonary adenovir us delivery in late-gestation sheep. Although initial alveolar transduction was highly efficient, antiadenoviral immune responses limited the duration of transgene expression. In contrast, early-gestation adenoviral delivery did not elicit antiadenoviral immune responses despite achieving efficient transduction of many fetal tissues. Furthermore, early-gestation adenovirus delivery did not affect late-gestation antiadenoviral immune responses. Th ese findings suggest that the early-gestation sheep fetus is not amenable t o adenoviral tolerance induction by UV injection and that it is incompetent of immunologic response to adenovirus. For the purposes of in utero gene t herapy, recombinant adenovirus may be applied optimally to genetic diseases requiring transient in utero expression.