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Objective: To construct an eukaryotic expression vector containing Smac gene and study the expression efficiency and speci- ficity of prostate specific antigen(PSA) enhancer/promoter in a possible targeted gene therapy scheme for prostate cancer. Meth- ods: PSA enhancer (PSAE) and promoter (PSAP) sequences were amplified using PCR method. CMV and T7 promoters were deleted from pcDNA3.1-Smac and replaced by the two specific fragments to generate pPSAE-PSAP-Smac. After transfection into different cell lines, the status of cells was observed. And then, we determined the relative concentration of Smac mRNA in RT-PCR. Results: The recombinant plasmid of pPSAE-PSAP-Smac was successfully constructed. And only the prostate cancer cell line PC-3 was suppressed after transfection with pPSAE-PSAP-Smac. However, other nonprostate lines were not. Moreover, the concentration of Smac mRNA regulated by PSA promoter and enhancer was higher in comparison to the CMV promoter-driven control vectors. Conclusion: An expression vector containing the Smac gene (based on elements of the PSA gene regulatory se- quences) has been developed and shown to function in prostate cancer cell lines which provides a solid platform for launching clin- ical studies.
Objective: To construct an eukaryotic expression vector containing Smac gene and study the expression efficiency and speci- ficity of prostate specific antigen (PSA) enhancer / promoter in a possible targeted gene therapy scheme for prostate cancer. Meth ods: PSA enhancer (PSAE) and the promoter (PSAP) sequences were amplified using PCR method. CMV and T7 promoters were deleted from pcDNA3.1-Smac and replaced by the two specific fragments to generate pPSAE-PSAP-Smac. After transfection into different cell lines, the status of cells was observed. And then, we determined the relative concentration of Smac mRNA in RT-PCR. Results: The recombinant plasmid of pPSAE-PSAP-Smac was successfully constructed. And only the prostate cancer cell line PC-3 was suppressed after transfection with pPSAE -PSAP-Smac. However, other nonprostate lines were not. Furthermore, the concentration of Smac mRNA by PSA promoter and enhancer was higher than that of CMV promoter-driven control vectors. Concl usion: An expression vector containing the Smac gene (based on elements of the PSA gene regulatory se- quences) has been developed and shown in function in prostate cancer cell lines which provide a solid platform for launching clinical studies.