Stereoselective Characteristics and Mechanisms of Epidermal Carboxylesterase Metabolism Observed in HaCaT Keratinocytes

Abstract

The advantages of transdermal drug delivery over other routes of administration have been well documented, including avoidance of variable absorption rates and first-pass hepatic metabolism with oral delivery and improved therapeutic activity.1) In the past, skin penetration studies usually focused only on the physicochemical factors affecting the transport of drugs across the skin. However, recently, many researchers have also been considering the concurrent epidermal metabolism. Various aspects of the metabolism of xenobiotics in the skin have been reviewed, 2) and it has been demonstrated that skin contains various enzymes including esterase capable of hydrolyzing different esters.3) Ester prodrugs for carboxylic acid-or hydroxyl-containing drugs are widely used for the enhancement of percutaneous absorption and have been noted to show stereoselective hydrolysis in various tissue preparations 4) ; surprisingly, the molecular mechanism of stereoselective hydrolysis in skin has been overlooked.A great deal of the current knowledge on the metabolic clearance of drugs in the skin is based on studies with homogenates of excised tissues, including excised animal skin or human skin. Inherent drawbacks of these models are the doubtful comparability of animal skin versus human skin, the restricted access to human skin, and the potentially high variability of enzyme activity of human skin samples along with the often poorly defined cellular viability of excised tissues. 5)It has been proposed that using cultured HaCaT keratinocytes as a readily available and reproducible in vitro metabolism model may circumvent some of these disadvantages.6) The transformed human HaCaT keratinocyte is a well-established cell model of dermal toxicity and metabolism studies in vitro, because it is easily cultured and has the characteristics of basal epidermal keratinocytes. 7)Carboxylesterase is a group of serine esterases found in numerous animal species and a variety of mammalian tissues. These enzymes hydrolyze many different endogenous and xenobiotic compounds and play a role in the metabolism of numerous drugs. 8) Some researchers have reported the purification and partial characterization of two distinct human carboxylesterases designated human carboxylesterase-1 (hCE-1) and human carboxylesterase-2 (hCE-2).9) These enzymes are expressed in human liver and are both members of the 60-kDalton serine esterase superfamily, but they differ substantially. Sequence homology between the two enzymes is only 48%. hCE-1 is an 180 kDalton trimer with an isoelectric point of 5.8, whereas hCE-2 is a monomer with an isoelectric point of 4.9. Substantial differences in substrate specificity also exist between the two isoforms. 10)Ketoprofen is a nonsteroidal antiinflammatory drug (NSAID) and is used through oral or suppository routes for the treatment of pain and inflammation. Given orally, gastrointestinal side effects are most frequently seen. Therefore percutaneous administration of ketoprofen has been studied to minimize gastrointestinal and o...

Biotransformation Capacity of Carboxylesterase in Skin and Keratinocytes for the Penta-Ethyl Ester Prodrug of DTPA

“…Of the remaining keratinocyte cultures, our results demonstrated that whereas HEKa, HEKn, and HaCaT express CES1, CES2 was more abundant across the various keratinocyte cell lines. These findings are consistent with the work of Zhu et al (2007), who identified CES2 as the main CES in HaCaT cells. Enzyme expression and activity in keratinocytes change over time in culture; cytochrome P450 enzymes are particularly vulnerable, but esterases and conjugated enzymes are also affected (Williams, 2008).…”

“…In humans, CES1 and CES2 expression is ubiquitous; however, CES1 predominates in most organs (Satoh et al, 2002). Although CESs are known to be expressed in human skin, information on their role in the metabolism of topically applied drugs and prodrugs is limited (Zhu et al, 2007).…”

Biotransformation of Drugs in Human Skin

“…To date, the only study that examined the expression of CEs was conducted in an immortalized keratinocyte cell line (HaCaT). In this study, CE-2 mRNA was readily detected, whereas CE-1 message was undetectable (Zhu et al, 2007). …”

Xenobiotic-metabolizing enzymes in the skin of rat, mouse, pig, guinea pig, man, and in human skin models

“…Both are expressed in the liver, where CE1 greatly exceeds CE2 (Laizure et al 2013 ). However, the study by Zhu et al ( 2007 ) suggests that in human skin, only CE2 may be expressed.…”

“…Zhu et al ( 2007 ) report that in the human keratinocyte cell line HaCaT, only CE2 mRNA was detected.…”