Using intron splicing trick for preferential gene expression in transduced cells: an approach for suicide gene therapy.

Suicide gene therapy is one of the most innovative approaches in which a potential toxic gene is delivered to the targeted cancer cell by different target delivery methods. We constructed a transfer vector to express green fluorescent protein (GFP) in transduced cells but not in packaging cells. We placed gfp under the control of the cytomegalovirus (CMV) promoter, which is positioned between the two long-terminal repeats in reverse direction. The intron-2 sequence of the human beta globin gene with two poly-A signals and several stop codons on the antisense strand was placed on the leading strand between the CMV promoter and gfp. For lentiviral production, the HEK293T and line were co-transfected with the PMD2G, psPAX2 and pLentiGFP-Ins2 plasmids. The HEK293T and line were transduced with this virus. PCR was performed for evaluation of intron splicing in transduced cells. The GFP expression was seen in 65% of the cells transduced. The PCR amplification of the genomic DNA of transduced cells confirmed the splicing of intron 2. The strategy is significant to accomplish our goal for preserving the packaging cells from the toxic gene expression during viral assembly and the resultant reduction in viral titration. Also it serves to address several other issues in the gene therapy.

Possible association of the CD4 gene polymorphism with vitiligo in an Iranian population.

BACKGROUND: Vitiligo is an acquired idiopathic and polygenic disorder with progressive depigmentation of circumscribed patches. Its exact pathogenesis is unknown. The CD4 gene plays an important role in the cell-mediated immune response and its association with type 1 diabetes mellitus, which is an autoimmune disease, has been previously reported. METHODS: Based on the assumption that autoimmunity is also involved in vitiligo, the CD4 gene was selected for study using a candidate gene approach. The pyrimidine-rich pentanucleotide repeat length polymorphism located in the promoter of the gene was studied. We screened 144 unrelated Iranian patients with vitiligo and 144 healthy matched controls by PCR. RESULTS: The CD4*A4 allele has a susceptibility association with the development of vitiligo in the Iranian population (OR = 1.68, 95% CI 1.18-2.42; P < 0.01, P(c) = 0.02). When we compared CD4*A4-containing genotypes in the case and control groups, even more significant positive association was identified (OR = 2.02, 95% CI 1.26-3.22; P < 0.01 and P(c) < 0.01). The CD4 gene polymorphism has a modest association with the development of vitiligo in Iranian patients.