Abstract
Adeno-associated virus (AAV) vectors can transduce hepatocytes efficiently in vivo in various animal species, including humans. Few reports, however, have examined the utility of pigs in gene therapy. Pigs are potentially useful in preclinical studies because of their anatomical and physiological similarity to humans. Here, we evaluated the utility of microminipigs for liver-targeted gene therapy. These pigs were intravenously inoculated with an AAV8 vector encoding the luciferase gene, and gene expression was assessed by an in vivo imaging system. Robust transgene expression was observed almost exclusively in the liver, even though the pig showed a low-titer of neutralizing antibody (NAb) against the AAV8 capsid. We assessed the action of NAbs against AAV, which interfere with AAV vector-mediated gene transfer by intravascular delivery. When a standard dose of vector was administered intravenously, transgene expression was observed in both NAb-negative and low-titer (14×)-positive subjects, whereas gene expression was not observed in animals with higher titers (56×). These results are compatible with our previous observations using nonhuman primates, indicating that pigs are useful in gene therapy experiments, and that the role of low-titer NAb in intravenous administration of the AAV vector shows similarities across species.
Highlights
Research on gene therapy using adeno-associated virus (AAV) vectors is currently being conducted worldwide
The AAV8 vector carrying a luciferase gene driven by a liver-specific promoter/enhancer (AAV8-HCRHAAT-luciferase) was used to observe transgene expression in microminipigs
The plate was washed with PBS and 100 μl of serum at a 1:2000 dilution was added and incubated at the AAV8-HCRHAAT-luciferase vector was injected intravenously and transgene expression was assessed at 1 week
Summary
Research on gene therapy using adeno-associated virus (AAV) vectors is currently being conducted worldwide. Successful cases of hemophilia A gene therapy were reported in 2017 [7] These therapeutic strategies targeted the liver, but they can be applied to other disease conditions; the best AAV serotype for human application has yet to be determined. We observed suppression of transgene expression in NAb-positive monkeys [24] These studies demonstrate the importance of NAb upon systemic AAV vector administration. The microminipig was created as the world’s smallest minipig for biomedical research [28] As their body weight at maturity is ~10 kg, large-scale breeding facilities are unnecessary, smaller amount of reagents are sufficient in experiments, and handling is easy. We evaluated the threshold titer of NAb related to gene expression
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