Minireview Biotransformation of Drugs in Human Skin

Although it is the largest organ of the human body, skin is often not considered in discussions of drug metabolism. However, there is growing evidence that most common drug-metabolizing enzymes are expressed in the skin. Evidence for expression of cytochromes P450, flavin monooxygenases, glutathione-S-transferases, N-acetyltransferases, and sulfotransferases in human skin and skin cells are presented. Additional discussion is focused on the evidence of actual metabolism of drugs. Finally, the potential clinical implications of metabolism within the skin are discussed briefly. Representing the largest organ in the human body, skin provides an important barrier role in protecting the body from external chemicals and pathogens that would otherwise impair critical functions needed for survival. Comprised of a complex cellular network, skin is capable of many metabolic functions common to visceral organs, including the biotransformation of drugs that penetrate through its most external layer. In addition to being an important target for pharmacotherapy, delivery through the skin is increasingly used as an effective means of delivering drugs to the systemic circulation. It is also evident that biotransformation within the skin may be an essential step to manifesting cutaneous toxicity to certain drugs and chemicals. Hence, an understanding of the capacity and impact of drug biotransformation within skin is an important component in assessing pharmacotherapy directed to or through the cutaneous environment. In the present article, discussion will be limited to evidence for biotransformation of drugs generated specifically in human skin or skin cells. Although many investigations have probed the ability of skin from numerous species to metabolize a variety of environmental chemicals, the focus of the present review is on evidence for metabolism of therapeutic agents. The reader is referred to the recent review of Oesch et al. (2007) for a comparison of xenobiotic-metabolizing enzymes in the skin of various species. Overview of the Structure and Function of Skin Existing as a dynamic and flexible barrier, human skin serves a multifaceted role as the organ with the greatest exposure to the external environment. In addition to providing the primary means for tactile evaluation of the immediate surroundings, skin serves a critical role in thermoregulation of the body. The plethora of infectious diseases with cutaneous manifestations attests to the important sentinel role of skin in protecting the body from bacterial, fungal, and viral pathogens. Although this diversity of function is recognized, it is not surprising that skin is populated by numerous types of specialized cells (see Table 1) that enable dynamic interaction with the internal and external environment. Because studies assessing the role of skin in drug metabolism often use model cells, it is important to begin this review with a brief discussion of skin structure and function. Three identifiable layers comprise human skin: epidermis, dermis, and hypodermis. Constituting the outermost region of the skin, the epidermis is primarily composed of keratinocytes (KCs), which make up 90 to 95% of cells in this layer of skin (Haake et al., 2000). Programmed differentiation of KCs as they progress through the epidermal layer produces a stratification of cell phenotype, ultimately giving rise to a highly keratinized dead cell at the surface of the skin. Whereas this terminally differentiated stage produces the initial skin barrier and is important for reducing water loss, cells at earlier stages of differentiation provide important regulatory signals for immune responses in the cutaneous environment. Due to their abundance and ready access from human tissue samples, KCs represent the most common skin cell used to assess skin metabolism of drugs and chemicals. As seen in other “ports of entry” to the body, the epidermis is punctuated by the presence of resident dendritic cells, which are known as Langerhans cells (LCs). Found in close association with KCs through E-cadherin receptors, these cells engulf antigens and, in the presence of appropriate stress signals, mobilize for migration to draining lymph nodes (Romani et al., 2003). Once present in the lymph node, these LCs present antigen to T cells, causing a clonal expansion of antigen-specific T cells that migrate to the cutaneous environment via the vascular system. Hence, LCs have the capacity to carry drug-protein conjugates formed in situ from the cutaneous environment to lymph organs that are essential in mediating a systemic response. Skin pigmentation is attributed to another important cell type Article, publication date, and citation information can be found at http://dmd.aspetjournals.org. doi:10.1124/dmd.108.024794. ABBREVIATIONS: KCs, keratinocytes; LCs, Langerhans cells; P450, cytochrome P450; FMO, flavin monooxygenase; GST, glutathione S-transferase; SULT, sulfotransferase; NAT, N-acetyltransferase; CE, carboxylesterase. 0090-9556/09/3702-247–253$20.00 DRUG METABOLISM AND DISPOSITION Vol. 37, No. 2 Copyright © 2009 by The American Society for Pharmacology and Experimental Therapeutics 24794/3428707 DMD 37:247–253, 2009 Printed in U.S.A. 247 at A PE T Jornals on A ril 2, 2017 dm d.aspurnals.org D ow nladed from present in the epidermis—the melanocyte. The active production of melanin by these cells provides a component of the protection from UV light afforded by the skin. There is evidence for intercellular communication between KCs and melanocytes in the skin, although the full nature of this is unclear at present (Nordlund and Boissy, 2000). A final cell type of importance in the epidermis is Merkel cells, which serve as the mechanoreceptors for the skin. As expected, the density of these cells is highest in regions of the skin that provide the greatest tactile response (e.g., fingertips). These cells have also been found in association with KCs in specific regions of the skin (e.g., hair bulb), suggesting that intercellular communication may occur within this pairing. Possessing an extensive vascular network, as well as a dense network of fibrils, the dermis endows the skin with tensile strength that provides mechanical resistance. The primary cells of the dermis are fibroblasts, which secrete the extracellular matrix that creates the aforementioned fibril network (Haake et al., 2000). Microscopically, they are readily differentiated from KCs by their elongated nature. In addition to fibroblasts, the dermis possesses a mononuclear phagocytic system comprised of monocytes, macrophages, and dermal dendrocytes. As immunocompetent cells, dermal dendrocytes play an important role in fibrotic and inflammatory conditions of the skin. An additional cell type, mast cells, is present throughout the dermal layer and serves as the primary effector cells for allergic reactions. The hypodermis plays several important roles, including insulation, cushioning, and energy supply. Composed primarily of adipocytes, this layer of skin also possesses the apocrine and eccrine sweat glands. Evidence for the Expression of Drug-Metabolizing Enzymes in Skin and Skin Cells The first line of evidence suggesting that drug metabolism may occur in human skin arises from studies that demonstrate the expression of drug-metabolizing enzymes in cutaneous tissue. Studies have varied in terms of the sources of tissue, ranging from skin biopsies to cultures of isolated cells from skin. Tables 2 and 3 summarize the evidence for specific enzyme expression from these various sources. As will be discussed, whereas message expression has been detected for many genes of drug-metabolizing enzymes, evidence for expression of the actual enzyme is more limited. Interpretation of studies that present evidence for expression of enzymes in particular subpopulations of skin cells must be undertaken cautiously. Isolation of an uncontaminated sample of a specific skin cell type is experimentally challenging. In our experience, commercially available sources of specific cell types may be contaminated with a low frequency of other cell types. Most studies noted herein have not incorporated (or at least described) rigorous assessment of cell populations to assure the homogeneity of the cell samples. Hence, low-level contamination of samples by other skin cell types may result in an erroneous conclusion of expression of enzyme in the cell type of interest. Thus, whereas the studies referenced in Tables 2 and 3 provide ample evidence for expression of these enzymes in skin cells, the certainty of expression in specific cell types is less than uniform. Cytochromes P450. Because cytochromes P450 (P450) are the most abundant of the drug-metabolizing enzymes found in liver, numerous investigations have naturally focused on probing for the presence of members of this enzyme family in human skin and skin cells. Early work focused on demonstrating P450-like activity and responsiveness to some inducers of these enzymes. As molecular biology techniques have advanced, probes for specific enzymes have been used and demonstrated the presence of numerous members of this important enzyme family. Table 2 provides a summary of evidence to date for the presence of these enzymes in human skin biopsy samples or cultures of specific skin cells, together with representative references supporting these findings. Evidence for the presence of the CYP1A enzymes has been sought by several investigators. Early studies demonstrating the ability of animal skin to metabolize benzo[a]pyrene suggested expression of CYP1A1 in skin. Indeed, CYP1A1 mRNA has been found in all human skin sources probed to date, although data supporting expression of CYP1A1 protein is limited to KCs (Baron et al., 2001; Saeki et al., 2002; Yengi et al., 2003). In contrast, studies probing for CYP1A2 mRNA have found that message for expression of this gene in skin or skin cells is undetectable (Baron et al., 2001; Saeki et al., 2002). CYP1B1 mRNA has been readily detected in skin biopsy samples and all skin cells examined to date (Baron et al., 2001; Saeki et al., 2002; Smith et al., 2003; Yengi et al., 2003). Studies probing for the presence of CYP2A6 have been limited to probes for message and in cultured skin cells, with mixed results. Although CYP2A6 mRNA was not detected in KCs, these same investigators observed message in fibroblasts and melanocytes (Saeki et al., 2002). Studies to date provide evidence for the expression of CYP2B6 mRNA in skin biopsy samples and KCs, with the latter also demonstrating the presence of CYP2B6 protein (Baron et al., 2001; Yengi et al., 2003). Probes for the expression of the CYP2C families of enzymes have yielded varied results. Although CYP2C9 mRNA was demonstrated in skin biopsy samples, message was not detectable in KCs or HaCaT cells, an immortalized keratinocyte cell line (Yengi et al., 2003; Vyas et al., 2006a). Assessment of the expression of CYP2C9 protein in KCs and HaCaT cells by immunoblot were inconclusive (Vyas et al., 2006a). CYP2C18 and CYP2C19 mRNA have both been detected in skin biopsy samples, although assessment of protein expression has yet to be reported. Members of the important CYP3A family of enzymes have also been detected in human skin or skin cells. CYP3A5 message and protein have been identified in all skin cell types and biopsy samples probed to date (Baron et al., 2001; Saeki et al., 2002; Smith et al., 2003; Yengi et al., 2003; Vyas et al., 2006a). It is interesting to note that whereas CYP3A4 message is readily detected in KCs, it has not been observed in HaCaT cells (Vyas et al., 2006a). Because experimentalists commonly use this immortalized cell line, it is important to recognize that the pattern of metabolism in this cell line may not match that seen in primary cells and in skin itself. In general, studies that have probed both skin biopsy samples and primary skin cell cultures have been consistent with regard to message expression for specific CYP450s—expression is either present or absent in both sample types. Two exceptions to this observation are CYP2C9 and CYP2D6. Whereas message for these two enzymes was observed in skin biopsy samples from the same laboratory, other investigators have not detected message for these enzymes in cultured KCs. It is unclear at this time whether this difference is due to a loss of gene expression upon culture or methodological differences for probing message between research groups. Hence, data to date suggests that cultured cell models are appropriate for investigations of TABLE 1 Primary cells in human skin as a function of layer Epidermis Dermis Hypodermis KCs Fibroblasts Adipocytes LCs Dendritic cells Melanocytes Monocytes Merkel cells Dendrocytes Mast cells 248 SVENSSON at A PE T Jornals on A ril 2, 2017 dm d.aspurnals.org D ow nladed from