Rhesus Monkey Model for Fetal Gene Transfer: Studies with Retroviral- Based Vector Systems
Authors: Alice F. Tarantal, John P. O'Rourke, Scott S. Case, Garret C. Newbound, Jun Li, Chang I. Lee, Carole R. Baskin, Donald B. Kohn, Bruce A. Bunnell
Journal: Molecular Therapy (2001)

Many life-threatening conditions that can be diagnosed early in gestation may be treatable in utero using gene therapy. In order to determine in utero gene transfer efficiency and safety, studies were conducted with fetal rhesus monkeys as a model for the human. Included in these studies were Moloney murine leukemia virus (MLV)-based amphotropic retrovirus, vesicular stomatitis virus-G (VSV-G) pseudotyped MLV, and a VSV-G pseudotyped HIV-1-based vector, all expressing the enhanced green fluorescent protein (EGFP) as a reporter gene and driven by a cytomegalovirus-immediate early promoter (N = 16). Rhesus monkey fetuses were administered viral vector supernatant preparations by the intraperitoneal (ip) (N = 14) or intrahepatic (ih) (N = 2) routes via ultrasound guidance at 55 ± 5 days gestation (late first trimester; term 165 ± 10 days). Fetuses were monitored sonographically, specimens were collected prenatally and postnatally, and tissue harvests were performed at birth or 3 or 6 months postnatal age (3–10 months post-gene transfer). PCR analyses demonstrated that transduced cells were present at ∼1.2% in peripheral blood mononuclear cells from fetuses administered amphotropic MLV, <0.5% in fetuses receiving MLV/VSV-G, and ∼4.2% for the lentiviral vector, which decreased to 2% at birth. Hematopoietic progenitors showed that overall (mean of all time points assessed), ∼25% of the collected colonies were positive for the EGFP transgene with the lentiviral vector, which was significantly greater than results achieved with the MLV-based vector systems (4–9%; P ≤ 0.001–0.016). At necropsy, 0.001–10% of the total genomic DNA was positive for EGFP in most tissues for all groups. EGFP-positive fluorescent cells were found in cell suspensions of thymus, liver, spleen, lymph nodes, cerebral cortex, and bone marrow (0.5–6%). Overall, the results of these studies have shown: (1Bennett M.J. Reliable prenatal diagnosis of Canavan disease (aspartoacylase deficiency): Comparison of enzymatic and metabolite analysis.J. Inherit. Metab. Dis. 1993; 16: 831-836Crossref PubMed Scopus (37) Google Scholar) healthy infants expressing vector sequences up to 10 months post-gene transfer, (2Cheung M.-C. Goldberg J.D. Kan Y.W. Prenatal diagnosis of sickle cell anaemia and thalassaemia by analysis of fetal cells in maternal blood.Nat. Genet. 1996; 14: 264-268Crossref PubMed Scopus (243) Google Scholar) fetal primate administration of retroviral vectors results in gene transfer to multiple organ systems, (3Desnick R.J. First-trimester biochemical and molecular diagnoses using chorionic villi: High accuracy in the U.S. collaborative study.Prenat. Diagn. 1992; 12: 357-372Crossref PubMed Scopus (16) Google Scholar) the highest level of gene transfer to hematopoietic progenitors was observed with the lentiviral vector system, and (4Evans M.I. Drugan A. Koppitch F.C. Zador I.E. Sachs A.J. Sokol R.J. Genetic diagnosis in the first trimester: The norm for the 1990s.Am. J. Obstet. Gynecol. 1989; 160: 1332-1339Abstract Full Text PDF PubMed Scopus (51) Google Scholar) there was no evidence of transplacental transfer of vector sequences into the dams. The rhesus monkey is an important preclinical primate model system for exploring gene transfer approaches for future applications in humans.