在大反应体积中进行高剂量标记时的最佳质量（131）I-单克隆抗体，并使用IODO-GEN对抗体进行临时包被。

Optimal quality (131)I-monoclonal antibodies on high-dose labeling in a large reaction volume and temporarily coating the antibody with IODO-GEN.

作者信息

Visser G W, Klok R P, Gebbinck J W, ter Linden T, van Dongen G A, Molthoff C F

机构信息

Radionuclide Center Vrije Universiteit and Departments of Obstetrics and Gynecology, Otolaryngology, and Nuclear Medicine/PET Center, University Hospital Vrije Universiteit, Amsterdam, The Netherlands.

出版信息

J Nucl Med. 2001 Mar;42(3):509-19.

PMID:11337531

Abstract

UNLABELLED

A novel, facile procedure for efficient coupling of high doses of (131)I to monoclonal antibodies (MAbs) was developed with minimal chemical and radiation damage.

METHODS

To diminish the radiation and chemical burden during labeling, iodination was performed in a large reaction volume and by temporarily coating the MAb with a minimal amount of IODO-GEN. The MAb was coated by injection of IODO-GEN (dissolved in acetonitrile [MeCN]) into the aqueous MAb solution, and the coating was subsequently removed by addition of ascorbic acid. For chemoprotection before, during, and after PD-10 purification of the (131)I-MAbs, ascorbic acid and human serum albumin were used. The effects of autoradiolysis in the starting (131)I solution were countered by treatment with NaOH and ascorbic acid. For this so-called IODO-GEN-coated MAb method, the sensitive chimeric MAb MOv18 (c-MOv18) and the more robust murine MAbs K928 and E48 were used. The high-dose (131)I-labeled MAbs were characterized for radiochemical purity and MAb integrity by thin-layer chromatography, high-performance liquid chromatography, and sodium dodecyl sulfate polyacrylamide gel electrophoresis followed by phosphor imager quantification. The high-dose (131)I-labeled MAbs were also characterized for immunoreactivity. The radiopharmacokinetics and biodistribution of (131)I-c-MOv18 were analyzed in human tumor-bearing nude mice. For comparison, (131)I-c-MOv18 batches were made using the conventional chloramine-T or IODO-GEN-coated vial method.

RESULTS

Conventional high-dose labeling of 5 mg c-MOv18 with 4.4 GBq (131)I resulted in a labeling yield of 60%, a radiochemical purity of 90%, an immunoreactive fraction of 25% (72% being the maximum in the assay used), and the presence of aggregation and degradation products. Using similar amounts of (131)I and MAb in the IODO-GEN-coated MAb method, 85%-89% overall radiochemical yield, at least 99.7% radiochemical purity, and full preservation of MAb integrity and immunoreactivity were achieved. For this labeling, 5 mg MAb were coated with 35 microg IODO-GEN during 3 min in a reaction volume of 6 mL. Also, biodistribution was optimal, and tumor accumulation was superior to that of coinjected (125)I-c-MOv18 labeled according to the conventional IODO-GEN-coated vial method.

CONCLUSION

A new, facile, high-dose (131)I-labeling method was developed for production of (131)I-labeled MAbs with optimal quality for use in clinical radioimmunotherapy.

摘要

未标记

开发了一种新颖、简便的方法，可将高剂量的（131）I高效偶联至单克隆抗体（MAb），同时使化学和辐射损伤降至最低。

方法

为减少标记过程中的辐射和化学负担，碘化反应在大反应体积中进行，并通过用最少量的IODO-GEN临时包被MAb来实现。通过将IODO-GEN（溶解于乙腈[MeCN]）注入MAb水溶液中来包被MAb，随后通过添加抗坏血酸去除包被。在（131）I-MAb的PD-10纯化之前、期间和之后，使用抗坏血酸和人血清白蛋白进行化学保护。通过用NaOH和抗坏血酸处理来对抗起始（131）I溶液中的自动辐射分解效应。对于这种所谓的IODO-GEN包被的MAb方法，使用了敏感的嵌合MAb MOv18（c-MOv18）以及更稳定的鼠源MAb K928和E48。通过薄层色谱、高效液相色谱以及十二烷基硫酸钠聚丙烯酰胺凝胶电泳随后进行磷成像仪定量，对高剂量（131）I标记的MAb进行放射化学纯度和MAb完整性表征。还对高剂量（131）I标记的MAb进行免疫反应性表征。在荷人肿瘤裸鼠中分析（131）I-c-MOv18的放射药代动力学和生物分布。为作比较，使用传统的氯胺-T或IODO-GEN包被小瓶法制备（131）I-c-MOv18批次。

结果

用4.4 GBq（131）I对5 mg c-MOv18进行传统的高剂量标记，标记产率为60%，放射化学纯度为90%，免疫反应分数为25%（在所使用的检测中最大值为72%），并且存在聚集和降解产物。在IODO-GEN包被的MAb方法中使用相似量的（131）I和MAb，实现了85%-89%的总放射化学产率、至少99.7%的放射化学纯度，以及MAb完整性和免疫反应性的完全保留。对于该标记，在6 mL反应体积中，5 mg MAb在3分钟内用35 μg IODO-GEN包被。此外，生物分布最佳，肿瘤蓄积优于按照传统IODO-GEN包被小瓶法标记的共注射（125）I-c-MOv18。