To obtain human sodium/iodide symporter gene cDNA for studying its potential ability as a radioiodide treatment for melanoma, the hNIS gene cDNA was amplified with total RNA from human thyroid tissue by RT-PCR. The hN...To obtain human sodium/iodide symporter gene cDNA for studying its potential ability as a radioiodide treatment for melanoma, the hNIS gene cDNA was amplified with total RNA from human thyroid tissue by RT-PCR. The hNIS cDNA was inserted into cloning vector pUCm-T and subcloned into eukaryotic expression vector pc-DNA3. The pc-DNA3-hNIS and pc-DNA3 were transduced into melanoma cells (B16) by electroporation, and two cell lines termed B16-A and B16-B respectively were established. The uptake and efflux of iodide was examined in vitro. The three cell lines (B16-A, B16-B, B16) were injected subcutaneously into the right flank of C57 mice. Biodistribution study and tumor imaging were performed when the tumor reached approximately 10mm in diameter. The cloned hNIS cDNA sequence was identical with the published sequence. Two novel cell lines named 16-A containing pc-DNA3-hNIS and B16-B containing pc-DNA3 only were established. The resultant cell line B16-A accumulated 17 and 19 times more radioiodide in vitro than B16 and B16-B respectively. The iodide uptake reached the half-maximal level within 10 min, and reached a plateau at 30 min. The efflux of iodide was also rapid (T1/2eff=10min). The imaging shows in vivo uptake in expected sites including the salivary glands, thyroid, stomach, and hNIS-transduced tumor, whereas the nontransduced tumor was not visualized. The %ID/g of B16-A tumors at 1, 2, 4, 12, and 24h after injec- tion of 125I were 12.22±0.71, 10.91±0.72, 8.73±0.99, 1.24±0.29, and 0.19±0.03, respectively, which were signifi- cantly higher percentages than those for controlling tumors, p<0.01. However, biologic T1/2 was about 6 h. Our pre- liminary data indicate that the transduction of the hNIS gene per se is sufficient to induce iodide transport in mela- noma cells both in vitro and in vivo, but T1/2eff is short.展开更多
文摘To obtain human sodium/iodide symporter gene cDNA for studying its potential ability as a radioiodide treatment for melanoma, the hNIS gene cDNA was amplified with total RNA from human thyroid tissue by RT-PCR. The hNIS cDNA was inserted into cloning vector pUCm-T and subcloned into eukaryotic expression vector pc-DNA3. The pc-DNA3-hNIS and pc-DNA3 were transduced into melanoma cells (B16) by electroporation, and two cell lines termed B16-A and B16-B respectively were established. The uptake and efflux of iodide was examined in vitro. The three cell lines (B16-A, B16-B, B16) were injected subcutaneously into the right flank of C57 mice. Biodistribution study and tumor imaging were performed when the tumor reached approximately 10mm in diameter. The cloned hNIS cDNA sequence was identical with the published sequence. Two novel cell lines named 16-A containing pc-DNA3-hNIS and B16-B containing pc-DNA3 only were established. The resultant cell line B16-A accumulated 17 and 19 times more radioiodide in vitro than B16 and B16-B respectively. The iodide uptake reached the half-maximal level within 10 min, and reached a plateau at 30 min. The efflux of iodide was also rapid (T1/2eff=10min). The imaging shows in vivo uptake in expected sites including the salivary glands, thyroid, stomach, and hNIS-transduced tumor, whereas the nontransduced tumor was not visualized. The %ID/g of B16-A tumors at 1, 2, 4, 12, and 24h after injec- tion of 125I were 12.22±0.71, 10.91±0.72, 8.73±0.99, 1.24±0.29, and 0.19±0.03, respectively, which were signifi- cantly higher percentages than those for controlling tumors, p<0.01. However, biologic T1/2 was about 6 h. Our pre- liminary data indicate that the transduction of the hNIS gene per se is sufficient to induce iodide transport in mela- noma cells both in vitro and in vivo, but T1/2eff is short.
