摘要
OBJECTIVE： Little effort has been made to study the protein-encoding genes isolated from traditional Chinese medicine（TCM） drugs, and the delivery of these genes into malignant cells through recombinant adeno-associated virus（r AAV） vectors has not been attempted. METHODS： We synthesized the c DNAs of five known cytotoxic proteins isolated from TCM drugs and the FLAG epitope-tagged c DNAs were subcloned into a r AAV plasmid vector. The protein expression was confi rmed by Western blot assay. Various cancer cell lines were transfected with the above plasmids and cell growth was monitored both in vitro and in vivo. The best cytotoxic gene was further packaged into r AAV vectors, under the control of a liver cancer-specifi c promoter. The liver tumor growth was then monitored following intratumor administration of the r AAV vectors.RESULTS： The expression plasmids, encoding individual potential cytotoxic genes tagged with FLAG epitope, were successfully generated and sequenced. Among these genes, trichosanthin（TCS） gene yielded the most promising results for the inhibition of cancer cell growth in vitro. The over-expressed TCS functioned as a type I ribosome-inactivating protein, followed by inducing apoptosis that is associated with the Bcl-PARP signaling pathway. Furthermore, intratumor injection of r AAV vectors containing the TCS gene signifi cantly inhibited the growth of human hepatocellular carcinoma tumors in a murine xenograft model.CONCLUSION： Our studies suggest that the use of TCM cytotoxic genes is a useful therapeutic strategy for treating human cancers in general, and liver tumors in particular.
OBJECTIVE： Little effort has been made to study the protein-encoding genes isolated from traditional Chinese medicine（TCM） drugs, and the delivery of these genes into malignant cells through recombinant adeno-associated virus（r AAV） vectors has not been attempted. METHODS： We synthesized the c DNAs of five known cytotoxic proteins isolated from TCM drugs and the FLAG epitope-tagged c DNAs were subcloned into a r AAV plasmid vector. The protein expression was confi rmed by Western blot assay. Various cancer cell lines were transfected with the above plasmids and cell growth was monitored both in vitro and in vivo. The best cytotoxic gene was further packaged into r AAV vectors, under the control of a liver cancer-specifi c promoter. The liver tumor growth was then monitored following intratumor administration of the r AAV vectors.RESULTS： The expression plasmids, encoding individual potential cytotoxic genes tagged with FLAG epitope, were successfully generated and sequenced. Among these genes, trichosanthin（TCS） gene yielded the most promising results for the inhibition of cancer cell growth in vitro. The over-expressed TCS functioned as a type I ribosome-inactivating protein, followed by inducing apoptosis that is associated with the Bcl-PARP signaling pathway. Furthermore, intratumor injection of r AAV vectors containing the TCS gene signifi cantly inhibited the growth of human hepatocellular carcinoma tumors in a murine xenograft model.CONCLUSION： Our studies suggest that the use of TCM cytotoxic genes is a useful therapeutic strategy for treating human cancers in general, and liver tumors in particular.
基金
supported in part by the Alex’s Lemonade Foundation and the Bankhead-Coley Cancer Research Program, 3BN04, Florida Department of Health
Public Health Service Grants R01 HL-097088 and R21 EB-015684 from the National Institutes of Health
an institutional grant from the Children’s Miracle Network
the National Natural Science Foundation of China grant No. 81273881
