Abstract
Human carboxylesterase 1 (hCE1), one of the most important serine hydrolases distributed in liver and adipocytes, plays key roles in endobiotic homeostasis and xenobiotic metabolism. This study aimed to find potent and selective inhibitors against hCE1 from phytochemicals and their derivatives. To this end, a series of natural triterpenoids were collected and their inhibitory effects against human carboxylesterases (hCEs) were assayed using D-Luciferin methyl ester (DME) and 6,8-dichloro-9,9-dimethyl-7-oxo-7,9-dihydroacridin-2-yl benzoate (DDAB) as specific optical substrate for hCE1, and hCE2, respectively. Following screening of a series of natural triterpenoids, oleanolic acid (OA), and ursolic acid (UA) were found with strong inhibitory effects on hCE1 and relative high selectivity over hCE2. In order to get the highly selective and potent inhibitors of hCE1, a series of OA and UA derivatives were synthesized from OA and UA by chemical modifications including oxidation, reduction, esterification, and amidation. The inhibitory effects of these derivatives on hCEs were assayed and the structure-activity relationships of tested triterpenoids as hCE1 inhibitors were carefully investigated. The results demonstrated that the carbonyl group at the C-28 site is essential for hCE1 inhibition, the modifications of OA or UA at this site including esters, amides and alcohols are unbeneficial for hCE1 inhibition. In contrast, the structural modifications on OA and UA at other sites, such as converting the C-3 hydroxy group to 3-O-β-carboxypropionyl (compounds 20 and 22), led to a dramatically increase of the inhibitory effects against hCE1 and very high selectivity over hCE2. 3D-QSAR analysis of all tested triterpenoids including OA and UA derivatives provide new insights into the fine relationships linking between the inhibitory effects on hCE1 and the steric-electrostatic properties of triterpenoids. Furthermore, both inhibition kinetic analyses and docking simulations demonstrated that compound 22 was a potent competitive inhibitor against hCE1-mediated DME hydrolysis. All these findings are very helpful for medicinal chemists to design and develop highly selective and more potent hCE1 inhibitors for biomedical applications.
Highlights
Mammalian carboxylesterases (CEs) are important members of the serine hydrolase superfamily (E.C. 3.1.1.1), which catalyze the hydrolysis of a wide variety of endogenous and xenobiotics ester compounds (Satoh and Hosokawa, 1998; Redinbo and Potter, 2005)
A series of natural triterpenoids were collected and their inhibitory effects against human carboxylesterases were assayed by using D-Luciferin methyl ester (DME) and dihydroacridin-2-yl benzoate (DDAB) as specific optical substrate for Human carboxylesterase 1 (hCE1) and human carboxylesterase 2 (hCE2), respectively (Figure 1)
Replacement of the C-3 ethyl ester group with 3-O-β-carboxypropionyl in compound 20 led to a dramatically increase in the inhibitory effects against hCE1 (IC50, 17 nM) and the high selectivity over hCE2 (3296-fold against hCE2). These results suggested that the structural modifications on the C-3 hydroxyl group of oleanolic acid (OA) were more feasible for the development of potent and highly selective inhibitors against hCE1
Summary
Mammalian carboxylesterases (CEs) are important members of the serine hydrolase superfamily (E.C. 3.1.1.1), which catalyze the hydrolysis of a wide variety of endogenous and xenobiotics ester compounds (Satoh and Hosokawa, 1998; Redinbo and Potter, 2005). Two primary carboxylesterases including human carboxylesterase 1 (hCE1) and human carboxylesterase 2 (hCE2), have been found and extensively studied in the past decade (Imai, 2006). These two isoforms share 47% amino acid sequence identity, but exhibit differential tissue distribution and distinct substrate and inhibitor specificities (Hosokawa, 2008). HCE1 is primarily expressed in the liver and adipocytes, and demonstrates substrate specificity for a large, bulky acyl group and a small alcohol group (Satoh et al, 2002; Imai et al, 2006). The distribution and the catalytic property of hCE1 make this enzyme a key determinant for the bioactivation of numerous ester-containing drugs including oseltamivir (Shi et al, 2006), dabigatran etexilate (Hu et al, 2013), mycophenolate mofetil (Fujiyama et al, 2010), and trandolapril (Zhu et al, 2009), as well as for the metabolic inactivation and clearance of some esterified drugs, such as clopidogrel (Tang et al, 2006), methylphenidate (Sun et al, 2004), rufinamide (Williams et al, 2010), and oxybutynin (Sato et al, 2012)
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