Syntheses And Characterization Of SBPC And SMPC As Linkers For Labeling Proteins With Astatine Or Iodine Radioisotopes And Their Application In ～(125/131)I Labeled Insulin

The cyclotron-produced21'At, which decays by the emission of a-particles, is one of the most feasible radionuclides in targeted endoradiotherapy of malignant tumors. It will be very appealing to receptor-targeted radiotherapy for hepatocellular carcinomas if 211At was conjugated with insulin, due to the powerful cytotoxic effect of 211At and the excellent specific localization in hepatocellular carcinomas of insulin as a carrier.However, if prepared by direct labeling method, 211At-peptides (proteins, antibodies, etc) will deastatine obviously in vitro and in vivo. In order to improve the stability of 211At labeled insulin and further investigate receptor-targeted radiotherapy of the radiopharmaceutics, it necessitates suitable bi-functional linkers to label insulin with 211At by indirect method. Additionally, iodine radioisotopes are usually used to be the substitute in the labeling method study of 21lAt, due to the similar chemical properties to astatine as halogens, longer half life and the commercial availability.In this study, two linkers for labeling proteins with astatine or iodine radioisotopes, one is the N-Succinimidyl 5-(tributylstannyl) -3-pyridinecarboxylate (SBPC), the other is N-succinimidyl 5-(trimethylstannyl) -3- pyridinecarboxylate (SMPC), have been synthesized. With the bi-functional esters, the first attempt to label insulin with 131I by indirect method was completed.5-Bromonicotinic acid was used as the starting material for syntheses of SBPC and SMPC. Firstly, 5-Bromonicotinic acid was reacted with n-BuLi, followed by RsSnCl (R=n-Bu or Me) to produce intermediates as 5-(trialkylstannyl)-3-pyridinecarboxylate. After purified by silica gel column, theintermediates were reacted with Dicyclohexyl-carbodiimide(DCC) and N-Hydroxysuccinimide(NHS), to yield N-Succinimidyl 5-(tributylstannyl)-3-pyridinecarboxylate (SBPC) or N-succinimidyl 5-(trimethyIstannyl)-3-pyridinecarboxylate (SMPC). The experimental conditions, especially the purification procedure of compounds were investigated considerably. The products were characterized by NMR, MS and HPLC.In order to explore the indirect 211At labeling of insulin with these bi-functional esters, 125/131I or IgG is used to be the substitute of 211At or insulin, respectively. With SBPC as linker, 125I labeled human IgG is performed in a labeling yield of approximately 30%, with radiochemical purity of more than 99% after purification by Sephadex G50. Even stayed for 3 days at room temperature (RT), the 125I-IgG maintained constantly stable in vitro, with radiochemical purity of more than 97%.After then, 131I labeled cattle insulin was performed using SBPC and SMPC as linker in a yield of 16.8% and 10.6%, respectively. 131I-insulin was purified by Sephadex G15 with radiochemical purity more than 98%. The radiochemical purity of more than 94% at RT after 4 days indicated that 131I-insulin has considerable stability in vitro.In summary, this study establishes an indirect method for astatine or iodine radioisotopes to label peptides (proteins and antibodies), especially for small molecular peptides. The results will be very important to 211At labeling of insulin and its further receptor-targeted radiotherapy for hepatocellular carcinomas.