Combined Therapy of KLK10 and Iodine-131 in PC3 Cell Line

1358 Objectives The tumour suppressor role of KLK10 gene in PC3 cell line has been proven in our previous study, by repressing proliferation, enhancing apoptosis and decreasing glucose metabolism. Radiotherapy is also an important treatment method for prostate cancer. Based on this, our study is to investigate the effect of combined therapy of KLK10 gene and radioactive iodine 131 in PC3 cell line. Methods To transfer KLK10 and sodium/iodide symporter (hNIS) gene into PC-3 with the recombinant lentiviral vector (pLVX-KLK10-IRES-hNIS-puro) to up-regulate the KLK10 and hNIS expressions. The high-purity, high KLK10 and hNIS stable expressing cell line, named as PC-3-KLK10-hNIS, was obtained by puromycin selection, and confirmed by western blot. Via the same method, PC-3-0-hNIS cell line was obtained as radiation therapy control, PC-3-KLK10-0 cell line as single gene therapy control, as well as PC-3-CON cell line as negative control. Iodine 125 uptake assay, iodine 125 uptake-inhibition by NaClO4 assay, iodine 125 efflux assay were undergone in these four kind of cell lines to detect the function of hNIS protein. Then iodine 131 was utilized in these cell lines to mimic the iodine 131 radiation therapy. Experiments of CCK-8 cell proliferation and cell clone formation assays were done with these radiation and non-radiation cell lines to evaluate the treatment effect. TUNEL assay was done to detected cell apoptosis. Results The stable NES1 expression in PC-3-KLK10-0 and PC-3-KLK10-hNIS cell lines was validated by western blot. Interestingly, the hNIS expression could be detected in PC-3-KLK10-0 and PC-3-CON cell lines, although it was much lower than that in the hNIS over-expressed cell lines (PC-3-0-hNIS and PC-3-KLK10-hNIS). However, the function of hNIS protein in PC-3-KLK10-0 and PC-3-CON cell lines was almost invalid respectively, because the uptake level of iodine 125 was extremely low. The hNIS over-expressed cell lines had the obvious iodine 125 uptake ability. The peak value and peak time in PC-3-0-hNIS was 6.2 times than that in PC-3-CON and at 60 min, as well as those in PC-3-KLK10-hNIS was 4.6 times and at 120 min. Both these two cell lines still had the iodine 125 uptake abilities more than 24 h. And NaClO4 could inhibit the iodine 125 uptake abilities of these two cell lines. Iodine 125 efflux from cell cytoplasm to extracellular could be detected constantly lasting for 2 hours. After 16 h radiation of iodine 131(3.7 MBq per 4[asterisk]105 cells), the growth rate of PC-3-KLK10-hNIS and PC-3-KLK10-0 were lower than other cell lines (including PC-3-CON and PC-3-0-hNIS, as well as four cell lines without radiation of iodine 131) in CCK-8 cell proliferation assay. And the growth rate of PC-3-KLK10-hNIS after radiation was the lowest. The result of cell clone formation assay was consistent with CCK-8 cell proliferation assay. Cell apoptosis could be detected in PC-3-KLK10-0 and PC-3-KLK10-hNIS cell lines, as well as these two cell lines after radiation. The apoptotic rate of PC-3-KLK10-hNIS after radiation was highest. Conclusions The effect of combined therapy of KLK10 gene and radioactive iodine 131 in PC3 cell line was better than single method treatment.