[Combination therapy of experimental murine hepatoma with mIL-12 gene and MHC I gene mediated by liposome].
OBJECTIVE
Interleukin-12 gene and MHC I gene have been used in gene therapy for cancer individually. To explore the synergistic antitumor effects of these two genes, the therapeutic effects of mIL-12 gene combined with MHC I (mouse H-2K) gene mediated by cationic liposome for experimental murine hepatoma were investigated.


METHODS
Balb/C mouse liver cancer MM45T. Li cells were transfected with pcDNA3/mIL-12, pcDNA3/H-2Kb(H-2K of C57BL/6 mouse), and pcDNA3/GFP (reporter gene) mediated by lipofectAMINE 2000(LF 2000). The expressions of foreign genes in transfected cells were detected. Balb/C mice were inoculated subcutaneously with pcDNA3/mIL-12 and pcDNA3/H-2Kb transfected MM45T. Li. The tumorigenesis of the inoculated cells was detected. After intratumoral injection with LF2000-plasmid DNA complexes, the growth of murine tumor and the survival time of the tumor bearing mice were observated.


RESULTS
The optimal ratio of LF2000: DNA is 3:1(microgram: microgram). The transfection efficiency reached to 30%. RT-PCR result showed the specific amplified fragments of the mIL-12 cDNA and H-2Kb cDNA in the transfected cells. Western blot analysis showed the expression of H-2Kb protein at 57 kDa. ELISA assay showed that the secretory mIL-12 was 48 ng/ml/10(6) cells. The tumorigenesis was decreased for transfected MM45T. Li cells with pcDNA3/mIL-12 and pcDNA3/H-2Kb. FACS assay showed that the numbers of CD3+, CD4+, and CD8+ cells from murine spleen were increased more in therapeutic group than in control group. The tumors grow slowly. The mIL-12 gene combined with H-2Kb gene has stronger antitumor effect for mouse liver cancer than single gene.


CONCLUSIONS
The combination therapy with mIL-12 gene and MHC I gene mediated by LF-2000 have the positive synergistic antitumor effect for experimental murine hepatoma.
Wang
Ke-min
KM
Department of Biochemistry & Research Center for Human Gene Therapy, Shanghai Second Medical University, Shanghai 200025, P. R. China. kmwang0105@yahoo.com.cn
Xia
Ai-di
AD
Chen
Shi-shu
SS
D000818:Animals; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000473:pathology / Q000628:therapy*; D018076:DNA, Complementary / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D005805:Genes, MHC Class I / Q000235:genetics*; D015316:Genetic Therapy / Q000379:methods*; D049452:Green Fluorescent Proteins; D018664:Interleukin-12 / Q000235:genetics* / Q000378:metabolism; D008081:Liposomes / Q000008:administration & dosage; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008856:Microscopy, Fluorescence; D009374:Neoplasms, Experimental / Q000235:genetics / Q000473:pathology / Q000628:therapy*; D010957:Plasmids / Q000008:administration & dosage / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D014407:Tumor Cells, Cultured
D018076:DNA, Complementary; D008081:Liposomes; D008164:Luminescent Proteins; D012333:RNA, Messenger; D049452:Green Fluorescent Proteins; D018664:Interleukin-12