Greene D L, Hau V S, Abbruscato T J, Bartosz H, Misicka A, Lipkowski A W, Hom S, Gillespie T J, Hruby V J, Davis T P
Department of Pharmacology, University of Arizona, College of Medicine, Tucson, USA.
J Pharmacol Exp Ther. 1996 Jun;277(3):1366-75.
To improve the blood-brain barrier penetration of the delta-opioid receptor peptides [D-Pen2, D-Pen5]enkephalin (DPDPE) and [D-Pen2, L-Cys5]enkephalin (DPLCE), various prodrug forms were synthesized to increase lipophilicity and drug delivery to the brain. The aims of this study were 3-fold, 1) to assess the metabolic conversion of various DPDPE and DPLCE prodrugs in vitro using mouse brain homogenate and mouse serum, 2)to characterize the proteolytic enzymes responsible for cleaving prodrugs to the parent compounds using select peptidase inhibitors and 3)to assess the blood-brain barrier permeability of prodrugs, compared with their parent compounds, using the in vitro bovine brain microvessel endothelial cell culture model. The prodrugs with carboxyl-terminal phenylalanine residues (DPDPE-Phe and DPLCE-Phe) had significantly longer metabolic conversion times in both mouse serum and brain homogenates than did the prodrugs with amino-terminal phenylalanine residues. Inhibition of leucine aminopeptidase with bestatin in the serum increased the conversion time of Phe0-DPDPE from 6.8 min to 92.2 min. Inhibition of aminopeptidase M with amastatin in the brain homogenate increased the conversion time of Phe0-DPDPE from 3.9 min to > 450 min. The long half-life of DPLCE-Arg-Pro-Ala in serum (317 min) vs. brain (9.2 min) can be explained by the high levels of the degradative endopeptidase 24.15 (EC 3.4.24.15) in the central nervous system but not in plasma. The data also showed that, for specific prodrugs of DPDPE such as Phe0-DPDPE and DPDPE-Arg-Gly, the prodrug shows a significant improvement in permeability, compared with the parent compound. Therefore, these data provide evidence that prodrugs or prodrug-enzyme inhibitor combinations may optimize the delivery of peptide and/or protein drugs to the central nervous system.
为提高δ-阿片受体肽[D-青霉胺2，D-青霉胺5]脑啡肽（DPDPE）和[D-青霉胺2，L-半胱氨酸5]脑啡肽（DPLCE）的血脑屏障穿透性，合成了各种前药形式以增加亲脂性并促进药物向脑内递送。本研究的目的有三个：1）使用小鼠脑匀浆和小鼠血清在体外评估各种DPDPE和DPLCE前药的代谢转化；2）使用特定的肽酶抑制剂鉴定负责将前药裂解为母体化合物的蛋白水解酶；3）使用体外牛脑微血管内皮细胞培养模型，与母体化合物相比，评估前药的血脑屏障通透性。具有羧基末端苯丙氨酸残基的前药（DPDPE-Phe和DPLCE-Phe）在小鼠血清和脑匀浆中的代谢转化时间均明显长于具有氨基末端苯丙氨酸残基的前药。在血清中用抑氨肽酶抑制亮氨酸氨肽酶可使Phe0-DPDPE的转化时间从6.8分钟增加到92.2分钟。在脑匀浆中用阿马astatin抑制氨肽酶M可使Phe0-DPDPE的转化时间从3.9分钟增加到>450分钟。DPLCE-Arg-Pro-Ala在血清（317分钟）与脑（9.2分钟）中的长半衰期可以用中枢神经系统中高水平的降解性内肽酶24.15（EC 3.4.24.15）来解释，但血浆中没有。数据还表明，对于DPDPE的特定前药，如Phe0-DPDPE和DPDPE-Arg-Gly，与母体化合物相比，前药的通透性有显著改善。因此，这些数据提供了证据，表明前药或前药-酶抑制剂组合可能优化肽和/或蛋白质药物向中枢神经系统的递送。
