Human Carboxylesterase Isozymes: Catalytic Properties and Rational Drug Design

Abstract

Human carboxylesterase 1 (hCE-1, CES1A1, HU1) and carboxylesterase 2 (hCE-2, hiCE, HU3) are a serine esterase involved in both drug metabolism and activation. Although both hCE-1 and hCE-2 are present in several organs, the hydrolase activity of liver and small intestine is predominantly attributed to hCE-1 and hCE-2, respectively. The substrate specificity of hCE-1 and hCE-2 is significantly different. hCE-1 mainly hydrolyzes a substrate with a small alcohol group and large acyl group, but its wide active pocket sometimes allows it to act on structurally distinct compounds of either large or small alcohol moiety. In contrast, hCE-2 recognizes a substrate with a large alcohol group and small acyl group, and its substrate specificity may be restricted by a capability of acyl-hCE-2 conjugate formation due to the presence of conformational interference in the active pocket. Furthermore, hCE-1 shows high transesterification activity, especially with hydrophobic alcohol, but negligible for hCE-2. Transesterification may be a reason for the substrate specificity of hCE-1 that hardly hydrolyzes a substrate with hydrophobic alcohol group, because transesterification can progress at the same time when a compound is hydrolyzed by hCE-1. From the standpoint of drug absorption, the intestinal hydrolysis by CES during drug absorption is evaluated in rat intestine and Caco2-cell line. The rat in situ single-pass perfusion shows markedly extensive hydrolysis in the intestinal mucosa. Since the hydrolyzed products are present at higher concentration in the epithelial cells rather than blood vessels and intestinal lumen, hydrolysates are transported by a specific efflux transporter and passive diffusion according to pH-partition. The expression pattern of CES in Caco-2 cell monolayer, a useful in vitro model for rapid screening of human intestinal drug absorption, is completely different from that in human small intestine but very similar to human liver that expresses a much higher level of hCE-1 and lower level of hCE-2. Therefore, the prediction of human intestinal absorption using Caco-2 cell monolayers should be carefully monitored in the case of ester and amide-containing drugs such as prodrugs. Further experimentation for an understanding of detailed substrate specificity for CES and development of in vitro evaluation systems for absorption of prodrug and its hydrolysates will help us to design the ideal prodrug.

Hyperpolarized [1,3‐13C2]ethyl acetoacetate is a novel diagnostic metabolic marker of liver cancer

“…Unregulated cellular uptake as well as cosubstrate independent enzymatic conversion of EAA, made this substrate highly useful as a hyperpolarized 13 C-MR marker. This could be appreciated by the signal-tonoise (SNR) obtained from EAA, which was comparable to the SNR reported in a literature liver cancer study with state-of-the-art hyperpolarized substrate, [1][2][3][4][5][6][7][8][9][10][11][12][13] C]pyruvate. Also, the contrast-to-noise (CNR) in the EAA based metabolic ratio images was significantly improved compared with the CNR in equivalent images reported using [1- Hepatocellular carcinoma (HCC) is the most common form of primary liver cancer.…”

“…With this hyperpolarized substrate it was shown how a liver cancer implanted in rat could be detected due to a higher substrate-to-product ratio in the cancer tissue compared with the surrounding healthy tissue. The signal-tonoise ratio (SNR) in the images, was comparable to the SNR obtained in a reported liver cancer study using hyperpolarized [1][2][3][4][5][6][7][8][9][10][11][12][13] C]pyruvate, 16 the most widely used d-DNP-MR substrate. 17 is a popular animal model.…”

“…Finland radical, free acid (1.8 mg, 1.7 lmol) was dissolved in [1,[3][4][5][6][7][8][9][10][11][12][13] C 2 ]ethyl acetoacetate (100 ll, 104 mg, 0.79 mmol). To the solution was added the gadolinium complex Gadoteridol (2.3 mg of a 100 lmol/g solution in DMSO).…”

“…This work led to the identification of a class of substrates, low molecular weight ethyl-esters, which showed high specificity for carboxyl esterases and proved in many cases to possess good properties for signal enhancement. In particular, hyperpolarized [1,[3][4][5][6][7][8][9][10][11][12][13] C 2 ]ethyl acetoacetate (EAA) was shown to provide a metabolic fingerprint of HCC. Using this substrate a liver cancer implanted in rats was diagnosed as a consequence of an~4 times higher metabolic substrate-to-product ratio than in the surrounding healthy tissue, (p 5 0.009).…”

“…15 In the reported study, we evaluate the potential for using the d-DNP-MR method to diagnose liver cancer with carboxyl esterase activity. [1,[3][4][5][6][7][8][9][10][11][12][13] C 2 ]ethyl acetoacetate was developed into a hyperpolarized substrate for the liver isoform of carboxyl esterase (CE-1). With this hyperpolarized substrate it was shown how a liver cancer implanted in rat could be detected due to a higher substrate-to-product ratio in the cancer tissue compared with the surrounding healthy tissue.…”