Experimental Study On HIL-12 And HB7-1 Gene Therapy For Melanoma

The lymphocyte cells from bone mellow of Chinese people were isolated and cultured. After that, the transcription of IL-12-p40mRNA,IL-12-p35mRNA and B7-lmRNA was induced. The cDNA of these three kinds of cytokines was cloned and sequenced with the method of RT-PCR. The results showed that the sequences were not indetical with those of GeneBank. This suggested the mighty of gene multility and Chinese person's own charicteristic.The cloned cDNA was inserted into Vector pCI and adinovirou shuttle vector pAd and pAd-track-CMV. These three kinds of vector system were constructed successfully. We name them pCII-p40-p35, pCII-B7-l, pAd-IL-12, pAd-B7-l, pAd-track-CMV-IL-12 and p Ad-track CMV-B7-1.The adenovirus shuttle vectors were capsuled and amplified . The titer of adinoviruswas 5X 1010. When hela cells and 3T3 cells were transfected with pCI vector the purposegene can trascript identified with the method of RT-PCR. The amount of IL-12 protein is Ing/ml . 106.24h analyzed by the ELASA method. In the experiment ex vivo, the effect of tomour cells killing amount to 70% when Hela cells were transfected with IL-12 and B7-1 and innoculated with lymphocytes (target cells: effector =1:10).In situ tumor gence therapy used the model of Scid mouse burdended human melanoma (HuPBML-Scid) and administrated IL-12,B7-1 and human blood lymphocyte subcutenously, the rate of tumor suppression was 93.4%. In the surrounding tissue of the tumor , a large amount of human lymphocytes can be observed.Because of the toxity of adenovirus vector, this kind of gene therapy is more difficult in clinical practice. In our experiment, we used a low toxical trans-gene vector in order to avoid those sever side effect The reason we use the Scid mouse moudle burdended human melanoma is it is more fesible for the research of gene therapy at present. In order to evaluate the fesibility and effect of the gene therapy we describe in this paper in clinical practice, further study on colon and gastric melignet tumor is being in progress in our lab.