Construction of MAGE-3 Tumor Vaccine and Its Effect on CTL Activity in HLA-A * 0201 Mice

Objective 　To obtain the DNA and protein vaccine of human MA GE-3 gene and to observe the effect on CTL activity induction in HLA-A * 0201 mice. Methods 　The MAGE-3 gene fragment was obtained by RT-PCR f rom human melanoma A375, and then cloned into the pcDNA3. 1/ V5-His and p ET32a vectors, respectively. The expression vectors were confirmed by rest riction enzyme digestion analysis.The mRNA expression of MAGE-3 gene was detected af ter transfection of the recombinant karyotic expression vector into B16 tumor cells and the expression of p ET-32a-MAGE-3 fusion protein was analyzed in E. coli by SDS-PAGE. The fusion protein was purified by Ni ²⁺ affinity chromatography. According to the DNA-priming and protein-boosting immune protocol, HLA-A * 0201 transgenic mice were vaccinated with DNA and followed by the protein of MAGE-3. Then, the activity of CTL against specific peptide pulsed by SW480 tumor cells was tested by the method of LDH. Results 　Rest riction enzyme digestion analysis showed that the recombinant expression vectors pcDNA3. 1-MAGE-3 and p ET32a-MAGE-3 were successfully const ructed and expressed in B16 or E. coli as a fusion protein. The DNA and purified protein vaccine were obtained respectively and the CTL activity in HLA-A *0201 transgenic mice was obviously observed. Conclusion 　MAGE-3 tumor vaccine was obtained successfully, which is helpful for the further study of the MAGE-3 vaccine for tumor therapy.