Dipartimento di Scienze Farmaceutiche Pietro Pratesi, Università degli Studi di Milano, Via Mangiagalli 25, 20133 Milan, Italy.
ChemMedChem. 2011 Jul 4;6(7):1269-82. doi: 10.1002/cmdc.201100042. Epub 2011 Jun 1.
β-Alanyl-D-histidine (D-CAR, the enantiomer of the natural dipeptide carnosine) is a selective and potent sequestering agent of reactive carbonyl species (RCS) that is stable against carnosinase, but is poorly absorbed in the gastrointestinal tract. Herein we report a drug discovery approach aimed at increasing the oral bioavailability of D-CAR. In our study we designed, synthesized, and evaluated a series of novel lipophilic D-CAR prodrugs. The considered prodrugs can be divided into two categories: 1) derivatives with both terminal groups modified, in which the carboxyl terminus is always esterified while the amino terminus is protected by an amidic (N-acetyl derivatives) or a carbamate (ethyloxy or benzyloxy derivatives) function; 2) derivatives with only one terminus modified, which can be alkyl esters as well as amidic or carbamate derivatives. The prodrugs were designed considering their expected lipophilicity and their hydrolysis predicted by docking simulations on the most important human carboxylesterase (hCES1). The stability and metabolic profile of the prodrugs were studied by incubating them with rat and human serum and liver fractions. The octyl ester of D-CAR (compound 13) was chosen as a candidate for further pharmacological studies due to its rapid hydrolysis to the bioactive metabolite in vitro. Pharmacokinetic studies in rats confirmed the in vitro data and demonstrated that the oral bioavailability of D-CAR is increased 2.6-fold if given as an octyl ester relative to D-CAR. Compound 13 was then found to dose-dependently (at daily doses of 3 and 30 mg kg(-1) equivalent of D-CAR) decrease the development of hypertension and dyslipidemia, to restore renal functions of Zucker fa/fa obese rats, and to inhibit the carbonylation process (AGEs and pentosidine) as well as oxidative stress (urinary 8-epi-prostaglandin F2α and nitrotyrosine). A plausible mechanism underlying the protective effects of 13 is RCS sequestration, as evidenced by the significant increase in the level of adduct between CAR and 4-hydroxy-trans-2-nonenal (HNE, the main RCS generated by lipid oxidation) in the urine of treated animals.
β-丙氨酰-D-组氨酸（D-CAR，天然二肽肌肽的对映体）是一种选择性和有效的活性羰基物种（RCS）的封闭剂，对肌肽酶稳定，但在胃肠道中吸收不良。本文报道了一种旨在提高 D-CAR 口服生物利用度的药物发现方法。在我们的研究中，我们设计、合成并评价了一系列新型亲脂性 D-CAR 前药。所考虑的前药可分为两类：1）两端基团均修饰的衍生物，其中羧基末端总是酯化，而氨基末端被酰胺（N-乙酰衍生物）或氨基甲酸酯（乙氧基或苄氧基衍生物）保护；2）仅一端修饰的衍生物，可以是烷基酯以及酰胺或氨基甲酸酯衍生物。根据预期的亲脂性以及对接模拟预测的水解情况，设计了前药，对接模拟针对最重要的人羧酸酯酶（hCES1）进行。通过与大鼠和人血清和肝部分孵育，研究了前药的稳定性和代谢特征。由于其在体外迅速水解为生物活性代谢物，因此选择 D-CAR 的辛基酯（化合物 13）作为进一步药理学研究的候选药物。在大鼠中的药代动力学研究证实了体外数据，并表明与 D-CAR 相比，作为辛基酯给药可使 D-CAR 的口服生物利用度提高 2.6 倍。然后发现化合物 13 以剂量依赖性方式（每天给予 3 和 30 mg kg(-1) 相当于 D-CAR 的剂量）降低高血压和血脂异常的发展，恢复 Zucker fa/fa 肥胖大鼠的肾功能，并抑制羰基化过程（AGEs 和戊糖素）以及氧化应激（尿 8-epi-前列腺素 F2α 和硝基酪氨酸）。13 具有保护作用的合理机制是 RCS 封闭，这从处理动物尿液中 CAR 与 4-羟基-trans-2-壬烯醛（HNE，脂质氧化产生的主要 RCS）之间加合物水平的显著增加得到证实。
