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Over the past several years, viral and non-viral gene delivery systems have been intensively developed to establish an ideal delivery vector for cancer gene therapy. Among the large variety of virus vectors currently being developed, the vector based on recombinant AAV (rAAV) is one of those that are closest to the ideal vector due to the following features; the lack of pathogenicity and toxicity, ability to infect dividing and non-dividing cells of various tissue origins, a very limited host immune response and long-term persistent expression. However, there is concern that two major obstacles of AAV serotype 2 (AAV2), the relative paucity of AAV receptors on certain cell types and the presence of pre-existing immunity to AAV2 in humans might limit its clinical applications in humans. To overcome these limitations, we first investigated in vitro transduction efficiencies for 7 different AAV serotypes (AAV1 – AAV6, AAV8) in 12 human tumor cell lines (MKN74, AGS, NCI-H460, HT1080, HepG2, SK-Hep1, HCT-116, 5637, UMUC3, HeLa, DU145 and MCF7). We employed “pseudotyped” AAV vector systems, in which rAAV2 vector genomes containing the CMV promoter/ enhancer, the green fluorescent protein (GFP) cDNA and SV40 poly (A) signal, flanked by AAV2 ITR sequences are cross-packaged into the capsid proteins of the other AAV serotypes (several plasmids are kindly provided by Dr. Katherine A. High, University of Pennsylvania Medical Center). As we reported previously, the efficiency of rAAV2 infection into most of the tested cancer cell lines was significantly high (>90%), whereas the transduction efficiencies for other 5 different rAAV serotypes were varied and greatly low. On the other hand the efficiency of rAAV5 transduction in various tumor cell lines were comparably high, mostly similar to that of rAAV2. Using the xenograft nude mice implanted with either NCI-H460 or HCT-116 cancer cells, we also studied in vivo transduction efficiency for several pseudotyped rAAV vectors, which were shown to have a host range different from rAAV2. Taken altogether our results suggest that the pseudotyped rAAV5 vector can be developed as one of the most efficient gene delivery systems for cancer gene therapy to overcome the anti-AAV2 immune response. (This study was supported by the grant from the Ministry of Science and Technology (M10534040005-05N3404-00511), Seoul, and the Chung-Buk Pioneering Bioindustry R&D Grant, Chung-Buk, Korea.)Molecular Therapy (2006) 13, S368–S368; doi: 10.1016/j.ymthe.2006.08.1047 [ABSTRACT FROM AUTHOR]

Naringin, an active flavonoid found in citrus fruit extracts, has pharmacological utility. The present study identified a novel mechanism of the anticancer effects of naringin in urinary bladder cancer cells. Naringin treatment resulted in significant dose-dependent growth inhibition together with G1-phase cell-cycle arrest at a dose of 100 μM (the half maximal inhibitory concentration) in 5637 cells. In addition, naringin treatment strongly induced p21WAF1 expression, independent of the p53 pathway, and downregulated expression of cyclins and cyclin dependent kinases (CDKs). Moreover, treatment with naringin induced phosphorylation of extracellular signal-regulated kinase (ERK), p38 mitogen-activated protein kinase and c-Jun N-terminal kinase. Among the pathways examined, only PD98059, an ERK-specific inhibitor, blocked naringin-dependent p21WAF1 expression. Consistently, blockade of ERK function reversed naringin-mediated inhibition of cell proliferation and decreased cell-cycle proteins. Furthermore, naringin treatment increased both Ras and Raf activation. Transfection of cells with dominant-negative Ras (RasN17) and Raf (RafS621A) mutant genes suppressed naringin-induced ERK activity and p21WAF1 expression. Finally, the naringin-induced reduction in cell proliferation and cell-cycle proteins also was abolished in the presence of RasN17 and RafS621A mutant genes. These data demonstrate that the Ras/Raf/ERK pathway participates in p21WAF1 induction, subsequently leading to a decrease in the levels of cyclin D1/CDK4 and cyclin E–CDK2 complexes and naringin-dependent inhibition of cell growth. Overall, these unexpected findings concerning the molecular mechanisms of naringin in 5637 cancer cells provide a theoretical basis for the therapeutic use of flavonoids to treat malignancies. [ABSTRACT FROM AUTHOR]