Investigations Of Cytotoxicity Of ～(125)I-[Tyr～3]octreotide In NCI-H446 Cell Lines

Purpose: To investigate the expressing of Somatostatin receptor(SSTR) on small cell lung cancer(SCLC) NCI-H446 cell lines, and capacity of NCI-H446 cell lines bound and internalized [Tyr3]octreotide (TOC), and the effect of cytotoxicity of 125I-TOC in NCI-H446 cell lines. To assess the therapeutic radiopharmaceutical potential of 125I-TOC for SSTR-positive tumors.Methods: (1)125I-TOC, radioligand, was labeled with chloramin T techniques and purified by Sephadex G-10. SSTR on NCI-H446 cell lines was detected using radioligand binding assay. The maximum binding sits (Bmax) and the dissociation constant (Kd) of SSTR with 125I-TOC were obtained by Scatchard analysis. (2) To detect the total binding of 125I-TOC on cells, NCI-H446 cell lines were incubated together with 125I-TOC at the different time points of 0.5h, Ih, 2h, 4h and 24h at 37 ℃.Then cell membrane bound 125I-TOC was washed off with acidic buffer and the amount of internalized 125I-TOC was detected with y counter. Cellular nucleus were obtained after cells broken to determined the bound of 125I-TOC on the cellular nucleus. (3)Effect of viability of cell was analyzed by a 3-(4,5-dimethy (thiazol-2-yl) -2, 5-diphenyltetrazolium bromide (MTT) assay at different time points with various doses of 125I-TOC, free 125I and TOC.Results: (1) The labeling efficiency of 125I-TOC was 76%-80% by using chloramin T. The radiochemical purity of 125I-TOC was more than 95% after purification with Sephadex G-10 column.(2) Bmax of SSTR in NCI-H446 cell lines was (1.17².0)xl05 sits per cell and Kd was (0.56².0)xl0^-11mol/L.(3)The binding speed of I25I-TOC with SSTR on NCI-H446 cell lines is rapidly. The saturant binding was appeared at 0.5h after incubate. No significant difference of 125I-TOC binding amount was found between 24h and 0.5h. 125I-TOC of internalized into nucleus andbound on the nucleus showed time dependent. 125I-TOC of bound on the nucleus increased to the highest value at 24h. (4) Cytotoxicity of 125I-TOC in SSTR positive NCi-H446 cell lines shown dose effect and time effect. The supreme effect of cytotoxity was found at 96h with 74kBq 12 I-TOC. The survival ratio of cell was reduced to 44.8% 7.2%.Conclusion: 125I-TOC can be internalized into SSTR positive cells mediated by SSTR, the Auger electron emitting from 123I can kill the cells. Effect of cytoxity shown dose effect and time effect. Our experiments provide an initial evidience for I-TOC as a radiopharmaceutical for therapy of SSTR positive tumors.