Transduction of hepatocellular carcinoma (HCC) using recombinant adeno-associated virus (rAAV): in vitro and in vivo effects of genotoxic agents.
OBJECTIVE
Adeno-associated virus (AAV) is an attractive tool for gene therapy. Here we investigated the in vitro and in vivo transduction of hepatocellular carcinoma (HCC) cells by an AAV vector and the efficacy of different strategies to enhance the transduction of the tumor.


METHODS
Transduction efficiency was determined by analyzing AAV-mediated beta-galactosidase gene (rAAV/lacZ) expression.


RESULTS
Adenovirus help or pretreatment of HCC cells with y-irradiation or with the topoisomerase inhibitor etoposide resulted in marked enhancement of cell transduction in vitro. In vivo studies in nude mice with subcutaneous HCC tumors showed that HCC cells were not transduced by AAV vector alone. However, co-infection of the tumor with adenovirus allowed an efficient expression of the reporter gene but only at the sites of vector injection. Previous gamma-irradiation of subcutaneous tumors with 1800 rad was able to improve transduction of HCC cells (up to 30%) using recombinant AAV. Continuous i.p. infusion of etoposide in buffalo rats harboring HCC tumors in the liver resulted in transduction of normal liver tissue and also of very small neoplastic lesions (<2 mm) but no transduction was observed in tumors bigger than 2 mm. To analyze this phenomenon we determined etoposide concentration in hepatic tissue. Our results revealed high concentrations of the drug in non-tumoral tissue but almost undetectable levels in big tumor nodules.


CONCLUSIONS
Our results indicate that while both radiotherapy and etoposide enhance transduction of tumor cells by rAAV in vitro, only radiotherapy increases tumor transduction in vivo. Our data suggest the existence of a barrier which limits in vivo the diffusion of chemotherapeutic agents to well-established HCC nodules.
Peng
D
D
Department of Internal Medicine, Clinica Universitaria and Medical School, University of Navarra, Pamplona, Spain.
Qian
C
C
Sun
Y
Y
Barajas
M A
MA
Prieto
J
J
D000818:Animals; D000972:Antineoplastic Agents, Phytogenic / Q000493:pharmacokinetics / Q000494:pharmacology; D006528:Carcinoma, Hepatocellular / Q000235:genetics*; D002460:Cell Line; D000229:Dependovirus / Q000235:genetics*; D005047:Etoposide / Q000493:pharmacokinetics / Q000494:pharmacology; D005720:Gamma Rays; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006801:Humans; D008099:Liver / Q000378:metabolism; D008113:Liver Neoplasms / Q000235:genetics*; D051379:Mice; D008819:Mice, Nude; D009153:Mutagens / Q000494:pharmacology*; D009368:Neoplasm Transplantation; D051381:Rats; D011915:Rats, Inbred BUF; D011995:Recombination, Genetic; D014161:Transduction, Genetic; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000528:radiation effects
D000972:Antineoplastic Agents, Phytogenic; D009153:Mutagens; D005047:Etoposide