Chae Su Young, Jin Cheng-Hao, Shin Han Jong, Youn Yu Seok, Lee Seulki, Lee Kang Choon
Drug Targeting Laboratory, College of Pharmacy, SungKyunKwan University, 300 Chonchon-dong, Jangan-ku, Suwon City 440-746, Korea.
Bioconjug Chem. 2008 Jan;19(1):334-41. doi: 10.1021/bc700292v. Epub 2007 Dec 14.
Glucagon-like peptide-1 (GLP-1) (7-36) is a type of incretin hormone with unique antidiabetic potential. The introduction of orally active GLP-1 offers substantial benefits in the treatment of type 2 diabetes over conventional injection-based therapies. Because the intestinal absorption of GLP-1 is restricted by its natural characteristics, we developed a series of GLP-1 analogues via the site-specific conjugation of biotin-NHS and/or of biotin-poly(ethylene glycol)-NHS at Lys 26 and Lys 34 of GLP-1 (7-36), respectively, in order to improve oral delivery. The resultant GLP-1 analogues, Lys 26,34-DiBiotin-GLP-1 (DB-GLP-1) and Lys 26-Biotin-Lys 34-(Biotin-PEG)-GLP-1 (DBP-GLP-1), were prepared and studied in terms of their chemical, structural, and biological properties. DBP-GLP-1 demonstrated superior proteolytic stability against trypsin, intestinal fluid, and the major GLP-1 inactivation enzyme (dipeptidyl peptidase-IV (DPP-IV)) to native GLP-1 or DB-GLP-1 ( p < 0.001). The in vitro insulinotropic effects of DB-GLP-1 and DBP-GLP-1 showed potent biological activity in a dose-dependent manner, which resembled that of native GLP-1 in terms of stimulating insulin secretion in isolated rat islets of Langerhans. Intraperitoneal glucose tolerance tests (IPGTT) after the oral administration of GLP-1 analogues in diabetic db/db mice demonstrated that DB-GLP-1 and DBP-GLP-1 significantly reduced the AUC 0-180 min of glucose for 3 h by 14.9% and 24.5% compared to that of native GLP-1, respectively ( p < 0.01). In particular, DBP-GLP-1 concentration in plasma rapidly increased 30 min after oral administration in rats, presumably due to improved intestinal absorption. These findings revealed that site-specific biotinylated and biotin-PEGylated GLP-1 is absorbed by intestine and that it has biological activity in vivo. Therefore, we propose that this orally active bioconjugated GLP-1 might be considered as a potential oral antidiabetic agent for type 2 diabetes mellitus.
胰高血糖素样肽-1（GLP-1）（7-36）是一种具有独特抗糖尿病潜力的肠促胰岛素激素。与传统的基于注射的疗法相比，口服活性GLP-1的引入在2型糖尿病治疗中具有显著优势。由于GLP-1的肠道吸收受其天然特性限制，我们分别在GLP-1（7-36）的第26位赖氨酸和第34位赖氨酸处通过生物素-NHS和/或生物素-聚乙二醇-NHS的位点特异性偶联开发了一系列GLP-1类似物，以改善口服给药效果。制备了所得的GLP-1类似物，即赖氨酸26,34-双生物素-GLP-1（DB-GLP-1）和赖氨酸26-生物素-赖氨酸34-（生物素-聚乙二醇）-GLP-1（DBP-GLP-1），并对其化学、结构和生物学性质进行了研究。与天然GLP-1或DB-GLP-1相比，DBP-GLP-1对胰蛋白酶、肠液和主要的GLP-1失活酶（二肽基肽酶-IV（DPP-IV））表现出更高的蛋白水解稳定性（p<0.001）。DB-GLP-1和DBP-GLP-1的体外促胰岛素作用呈剂量依赖性地显示出强大的生物活性，在刺激分离的大鼠胰岛朗格汉斯细胞分泌胰岛素方面与天然GLP-1相似。在糖尿病db/db小鼠口服GLP-1类似物后进行的腹腔内葡萄糖耐量试验（IPGTT）表明，与天然GLP-1相比，DB-GLP-1和DBP-GLP-1在3小时内分别使葡萄糖的AUC 0-180分钟显著降低了1４.9%和24.5%（p<0.01）。特别是，大鼠口服给药30分钟后血浆中DBP-GLP-1浓度迅速升高，推测是由于肠道吸收改善。这些发现表明，位点特异性生物素化和生物素-聚乙二醇化的GLP-1可被肠道吸收并在体内具有生物活性。因此，我们提出这种口服活性生物共轭GLP-1可被视为2型糖尿病的一种潜在口服抗糖尿病药物。
