Gene-viral vectors: a promising way to target tumor cells and express anticancer genes simultaneously / 中华医学杂志(英文版)

<p><b>OBJECTIVE</b>To develop a new kind of vector system called gene-viral vector, which combines the advantages of gene and virus therapies.</p><p><b>METHODS</b>Using recombinant technology, an anti-tumor gene was inserted into the genome of replicative virus specific for tumor cells. The cell killing effect, reporter gene expression of the green fluorescence protein, anti-tumor gene expression of mouse interleukin-12 (mIL-12) and replication of virus were observed by the methods of cell pathology, fluorescence microscopy, ELISA and electron microscopy, respectively.</p><p><b>RESULTS</b>A new kind of gene-viral vector system of adenovirus, in which the E1b-55 kD gene was deleted but the E1a gene was preserved, was constructed. The vector system, like the replicative virus ONYX-015, replicated and proliferated in tumor cells but not in normal ones. Our vector had an advantage over ONYX-015 in that it carried different kinds of anti-tumor genes to enhance its therapeutic effect. The reporter gene expression of the green fluorescence protein in tumor cells was much better than the adenovirus vector employed in conventional gene the rapy, and the expression in our vector system was as low as or even less than that in the conventional adenovirus gene therapy system. Similar results were observed in experiments with this vector system carrying the anti-tumor gene mIL-12. Replication and proliferation of the virus carrying the mIL-12 gene in tumor cells were confirmed by electron microscopy.</p><p><b>CONCLUSIONS</b>Gene-viral vectors are new vectors with an anti-tumor gene inserted into the genome of replicative virus specific for tumor cells. Because of the specific replication and proliferation of the virus in tumor cells, expression of the anti-tumor gene is increased hundreds to thousands of times. This approach takes full advantages of gene therapy and virus therapy to enhance the effect on the tumor. It overcomes the disadvantages of conventional gene therapy, such as low transfer rate, low gene expression, lack of target tropism, and low anti-tumor activity. We believe that this is a promising means for future tumor treatment.</p>

Injection of NKG5SV gene to inhibit growth and metastasis of hepatocellular carcinoma / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To study the injection of NKG5SV gene to inhibit growth and metastasis of hepatocellular carcinoma (HCC).</p><p><b>METHODS</b>NKG5SV gene was inserted into retroviral vector pLXSN by normal methods. LacZ gene was used as control. LCI-D20 tumor together with saline, pLXSN-LacZ DNA or pLXSN-NKG5SV was subcutaneously inoculated to the nude mice. Tumor formation rate and tumor size were noted 35 days after inoculation. LCI-D20 tumor was inoculated subcutaneously. Saline, pLXSN-LacZ DNA or pLXSN-NKG5SV was intratumorally injected respectively 10 days after inoculation. Tumor growth was observed 35 days after inoculation. Liver cancer was resected 22 days after intrahepatic inoculation. Saline, pLXSN-LacZ DNA or pLXSN-NKG5SV was respectively injected at incisal margin or intraspleen. Mice were killed 35 days after inoculation to observe tumor recurrence at incisal margin, intrahepatic metastasis and extrahepatic metastasis.</p><p><b>RESULTS</b>Tumor formation rate and tumor diameter(cm) were 1.76 +/- 0.11, 1.51 +/- 0.34, 0.33 +/- 0.04 in the control group, LacZ group, NKG5SV group respectively when tumor and different cDNA were inoculated together. Tumor diameter(cm) and weight(g) were 0.87 +/- 0.08, 0.83 +/- 0.05, 0.26 +/- 0.04; 0.43 +/- 0.06, 0.38 +/- 0.04, 0.08 +/- 0.06 in the control group, LacZ group, NKG5SV group respectively when different cDNA were injected into the LCI-D20 tumor. Sites with extrahepatic metastasis nidi, incisal margin recurrence tumor size(cm), intrahepatic metastasis nidi, metastasis involved hepatic lobes in the control group, LacZ group, NKG5SV group were 4.25 +/- 1.48, 4.25 +/- 1.04, 0.63 +/- 0.51; 1.51 +/- 0.27, 1.35 +/- 0.17, 0.81 +/- 0.17; 2.50 +/- 1.41, 2.38 +/- 1.06, 1.25 +/- 0.71; 2.13 +/- 0.99, 2.00 +/- 0.75, 1.38 +/- 0.74 respectively when NK cells were injected at incise margin. They were 4.38 +/- 1.85, 4.25 +/- 1.48, 1.00 +/- 0.75; 1.13 +/- 0.23, 0.97 +/- 0.29, 0.76 +/- 0.16; 2.50 +/- 1.41, 2.05 +/- 1.12, 0; 2.13 +/- 0.83, 1.75 +/- 0.88, 0 respectively when NK cell were injected intrasplenicly.</p><p><b>CONCLUSIONS</b>NKG5SV gene can inhibit HCC growth and postoperative metastasis and recurrence.</p>

Construction of double-copy and x gene deleted hepatitis B virus DNA expression plasmid and cells transfection study / 第二军医大学学报

Objective:To construct double copy and x gene deleted HBV expression plasmid and study its expression in Hep3B cell line. Methods:The double copy HBV DNA ( adr Ⅰ) was used to inactivate HBV x gene by inserting mutation and gene recombination. The inserted 55 bp DNA sequence was synthesized artificially; the insertion point was ApaL Ⅰ of x gene area. After recombination, an x gene defected HBV plasmid containing single P, S and C gene was constructed,which can express in mammalian cell line. Another plasmid carrying double copy HBV DNA with normal x gene was constructed as contrast. Both were used to transfect Hep3B cells. Then the cells were screened by G418 and HBV virus in culture medium were isolated and detected by fluorescence quantitative PCR. Results: Plasmids pcDNA3 KN F1F2 and pcDNA3 ES HBV2 were constructed successfully. After cell transfection, the HBV DNA was highly expressed with both plasmids on the 3 rd, 6 th,14th day. Conclusion: The plasmids constructed can express in Hep3B cell line and cause HBV replication; x gene defected HBV gene has no effect on HBV replication in Hep3B cell line.