No evidence for germ-line transmission following prenatal and early postnatal AAV-mediated gene delivery.
BACKGROUND
Recombinant adeno-associated viruses have been used successfully in a number of pre-clinical and clinical gene therapy studies. Since there is a broad consensus that gene therapy must not lead to germ-line transmission, the potential of such vectors for inadvertent gene transfer into germ cells deserves special attention. This applies in particular to pre- or perinatal vector application which has been considered for diseases presenting with morbidity already at birth.


METHODS
AAV serotype 2 derived vectors carrying a beta-galactosidase reporter gene or human clotting factor IX cDNA were injected intraperitoneally or via a yolk sac vein into mouse fetuses or administered intravascularly to newborn mice. Tissue samples of the treated animals including the gonads as well as sperm DNA, obtained by differential lysis of one testis of each male animal, and the offspring of all treated mice were investigated for the presence of vector DNA by nested PCR. In positive samples, the copy number of the vector was determined by quantitative real-time PCR.


RESULTS
AAV vectors administered intraperitoneally or intravascularly to fetal or newborn mice reached the gonads of these animals and persisted there for time periods greater than one year. Intravascular injection of the vector resulted more frequently in gene transfer to the gonads than intraperitoneal injection. Vector copy numbers in the gonads ranged from 0.3 to 74 per 10(4) cell equivalents. However, neither in isolated sperm DNA from the treated animals nor in their offspring were vector sequences detectable.


CONCLUSIONS
These data suggest the risk of inadvertent germ-line transmission following prenatal or early postnatal AAV type 2 mediated gene delivery to be very low.
Jakob
Marcus
M
Department of Experimental Medicine I, Nikolaus Fiebiger Centre of Molecular Medicine, University of Erlangen-Nuernberg, Germany.
Mühle
Christiane
C
Park
Jung
J
Weiss
Susi
S
Waddington
Simon
S
Schneider
Holm
H
D000818:Animals; D000831:Animals, Newborn; D004274:DNA, Recombinant / Q000008:administration & dosage / Q000235:genetics* / Q000633:toxicity; D004279:DNA, Viral / Q000008:administration & dosage / Q000235:genetics* / Q000633:toxicity; D000229:Dependovirus / Q000235:genetics*; D004622:Embryo, Mammalian / Q000821:virology; D005164:Factor IX / Q000235:genetics / Q000378:metabolism; D005260:Female; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods; D005822:Genetic Vectors; D005854:Germ Cells / Q000821:virology*; D006066:Gonads / Q000821:virology; D018445:Infectious Disease Transmission, Vertical; D008297:Male; D051379:Mice; D010322:Parvoviridae Infections / Q000635:transmission*; D016133:Polymerase Chain Reaction; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D004274:DNA, Recombinant; D004279:DNA, Viral; D005164:Factor IX; D001616:beta-Galactosidase