【作者】 唐军；

【导师】 刘宁；

【作者基本信息】 四川大学 ， 无机化学， 2003， 硕士

【摘要】 加速器制备的α核素²¹¹At是最适宜于靶向内放射治疗的理想核素之一。用²¹¹At标记偶联胰岛素，将胰岛素的靶向载体作用和²¹¹At的极强细胞毒性相结合，对肝细胞癌的受体靶向治疗研究具有重要的科学意义和应用前景。 由于采用直接标记法制得的²¹¹At-多肽（蛋白质、抗体等）在体内不稳定，因此选择合成合适的偶联剂实现胰岛素的²¹¹At间接标记，是提高²¹¹At-胰岛素的体内外稳定性，从而进行²¹¹At-胰岛素受体靶向治疗研究的关键问题。At和I同属卤族元素，化学性质相似。但放射性碘(如¹³¹I或¹²⁵I)易得，且半衰期较长，因此在研究²¹¹At的标记方法时，往往以放射性碘代替²¹¹At进行探索。 为此，本论文以5-溴烟酸为起始物，合成了两种适用于蛋白质放射性砹（碘）标记的偶联剂—5-（三正丁基锡）-3-吡啶甲酸N-琥珀酰亚胺酯（SBPC）和5-（三甲基锡）-3-吡啶甲酸N-琥珀酰亚胺酯（SMPC），并用这两种双功能偶联剂，首次进行了¹³¹I标记胰岛素的研究。 实验以5-溴烟酸（5-bromonicotinic acid）为起始物，先与正丁基锂、三烷基锡反应，制得5-（三烷基锡）-3三烷基锡代吡啶甲酸酯。经分离纯化后再与N-羟基琥珀酰亚胺（N-Hydroxysuccinimide，NHS）、二环己基碳二亚胺（Dicyclohexyl-carbodiimide，DCC）反应，分别得到5-（三正丁基锡）-3-吡啶甲酸N-琥珀酰亚胺酯（N-Succinimidyl 5-（tributylstannyl）-3-pyridinecarboxylate，SBPC）和5-（三甲基锡）-3-吡啶甲酸N-琥珀酰亚胺酯（N-Succinimidyl 5-（trimethylstannyl）-3-pyridinecarboxylate，SMPC）。对SBPC和SMPC的合成及分离的条件进行了系统研究，并用核磁共振（NMR）、质谱（MS）、高效液相（HPLC）等对产物进行了表征。 以¹²⁵I代替²¹¹At，IgG为模拟蛋白进行了初步标记实验。结果表明，以5-（三正丁基锡）-3-吡啶甲酸N-琥珀酰亚胺酯为双功能偶联剂进行IgG的¹²⁵I标记偶联，其标记率可达30%以上。用SePhadexGS。柱分离得到的1251一IgG，放射化学纯度>99%。’z5l一IgG室温下放置3天，其放化纯度仍在97%以上，表明其在体外具有较高的稳定性。 在”，I标一记IgG的基础上，以sBPc和sMPc为双功能偶联剂，在国内外首次进行了’3’I标记胰岛素的研究。其标记率分别为16.8%和10.6%;经sePhade、G15柱分离后，13’I一胰岛素的放化纯度>98%。室温放置4天，其放射化学纯度均在94%以上，表明’3’I一insulin具有较高的体外稳定性。 通过本研究，建立了用间接标记方法进行放射性成/碘偶联多肤（蛋白质、抗体）、特别是小分子多肤的方法，为下一步川At标记胰岛素，以及进行肝细胞癌的受体靶向治疗研究奠定了基础。更多还原

【Abstract】 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.更多还原