Effects of Hepatic Drug-metabolizing Enzyme Induction on Clinical Pathology Parameters in Animals and Man

Abstract

Hepatic drug-metabolizing enzyme (DME) induction is an adaptive response associated with changes in preclinical species; this response can include increases in liver weight, hepatocellular hyperplasia and hypertrophy, and upregulated tissue expression of DMEs. Effects of DME induction on clinical pathology markers of hepatobiliary injury and function in animals as well as humans are not well established. This component of a multipart review of the comparative pathology of xenobiotically mediated induction of hepatic metabolizing enzymes reviews pertinent data from retrospective and prospective preclinical and clinical studies. Particular attention is given to studies with confirmation of DME induction and concurrent evaluation of liver and/or serum hepatobiliary marker enzyme activities and histopathology. These results collectively indicate that in the rat, when histologic findings are limited to hepatocellular hypertrophy, DME induction is not expected to be associated with consistent or substantive changes in serum or plasma activity of hepatobiliary marker enzymes such as alanine aminotransferase, alkaline phosphatase, and gamma glutamyltransferase. In the dog and the monkey, published studies also do not demonstrate a consistent relationship across DME-inducing agents and changes in these clinical pathology parameters. However, increased liver alkaline phosphatase or gamma glutamyltransferase activity in dogs treated with phenobarbital or corticosteroids suggests that direct or indirect induction of select hepatobiliary injury markers can occur both in the absence of liver injury and independently of induction of DME activity. Although correlations between tissue and serum levels of these hepatobiliary markers are limited and inconsistent, increases in serum/plasma activities that are substantial or involve changes in other markers generally reflect hepatobiliary insult rather than DME induction. Extrahepatic effects, including disruption of the hypothalamic-pituitary-thyroid axis, can also occur as a direct outcome of hepatic DME induction in humans and animals. Importantly, hepatic DME induction and associated changes in preclinical species are not necessarily predictive of the occurrence, magnitude, or enzyme induction profile in humans.

Keywords

hepatic drug-metabolizing enzyme induction clinical chemistry alanine aminotransferase gamma glutamyltransferase alkaline phosphatase

Introduction

The biotransformation and elimination of lipophilic substances of endogenous or foreign origin is a critical activity performed by living organisms to guard against injury caused by intracellular accumulation of potential toxicants. In animals and man, biotransformation or xenobiotic metabolizing enzyme systems are present in most if not all tissues, with the highest concentration found in liver. Expression of these enzymes is influenced by a range of factors including diet, sex, age, environmental exposures, and most importantly, endobiotics and xenobiotics, including drugs and chemicals. Although drug-metabolizing enzyme (DME) systems are generally present at low levels in the liver, they can be rapidly and reversibly upregulated in response to endogenous or exogenous stimuli, a process known as enzyme induction (Dickins 2004; Xu et al. 2005). An extensive review of DME systems is beyond the scope of this article, however, additional information is available in the introduction to this series (Botts et al. 2010) and in several excellent reviews in the literature (Dickins 2004; Graham and Lake 2008; Handschin and Meyer 2003).

Although changes in liver morphology and hepatic DME, including CYP450 activities, in response to microsomal enzyme induction are well characterized in preclinical species, the effects of microsomal enzyme induction on clinical pathology markers of hepatic injury and function are less commonly described. Increased alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), and/or gamma glutamyltransferase (GGT) activities in liver parenchyma have been described in association with drug-induced CYP450 induction in the rat, the dog, and the human. However, reports of increased ALT, AST, ALP, or GGT activity in serum or plasma and their association with DME induction are conflicting in these species. Therefore, the objective of this review is to summarize information available in the literature on the effects of hepatic DME induction on clinical pathology parameters in these preclinical species and in humans. Additionally, select endocrine changes associated with DME induction will also be reviewed.

Hepatic Effects of DME Induction in Animals

Following administration of xenobiotics, enzyme induction is first observed in the liver because of its anatomical location, function, and high concentration of DMEs. Centrilobular hepatocellular hypertrophy, the most common histological change associated with enzyme induction in animals, is an effect considered adaptive and not injurious (Greaves 2007; Schulte-Hermann 1974). However, this adaptive response can be overcome following intense or prolonged induction stimuli, leading to hepatocellular degeneration, necrosis, or proliferation (Williams and Iatropoulos 2002). Hepatobiliary injury can also be an indirect consequence of enzyme induction, resulting from increased formation of highly reactive intermediates that cause oxidative injury or formation of adducts that covalently bind cellular proteins, nucleotides, or DNA/RNA (Klaunig et al. 1998). A more comprehensive review of the morphologic consequences of hepatic microsomal enzyme induction is provided in an accompanying article in this series (Maronpot et al. in press).

Commonly Used Biomarkers for Monitoring Hepatic Effects

Noninvasive monitoring for liver injury is routinely performed by measurement of hepatobiliary markers in plasma or serum. This paper will focus on the activities of ALT, ALP, and GGT, as these enzymes are generally liver associated and often reported to be altered in association with microsomal enzyme induction. Furthermore, these enzymes are recommended as indicators of hepatocellular (ALT activity) and hepatobiliary (ALP and GGT activities) injury in preclinical studies (Boone et al. 2005; Weingand et al. 1996). Glutamate dehydrogenase (GLDH) and α-glutathione S-transferase (αGST) have also been shown to be sensitive indicators of hepatocellular injury, however, use of these markers is less common, and data from prospective studies with prototypic inducer compounds are more limited (Giffen et al. 2002; O’Brien et al. 2002). Aspartate aminotransferase is also used as a marker of liver injury, particularly for sequential monitoring of hepatic injury, where declining values in serial measurements reflect recovery from hepatic injury owing to its shorter circulating half-life than ALT (Meyer and Harvey 2004). Aspartate aminotransferase is generally considered to be less sensitive and less specific for liver (Center 2007; Loeb 1999) and will not be discussed further for preclinical species.

Alanine aminotransferase is present in the highest concentrations in periportal hepatocytes and catalyzes a crucial reaction in gluconeogenesis and the urea cycle. Because of its predominantly cytosolic location, ALT can be rapidly released into the serum following irreversible hepatocellular injury or necrosis. Alanine aminotransferase release also occurs following reversible hepatocellular injury, generally thought to occur via cytoplasmic blebbing (Lemasters et al. 1990). Distinguishing among these conditions is generally not possible based on circulating ALT activity alone. However, reversible or less extensive injury is generally associated with changes of smaller magnitude than irreversible or widespread cellular injury (Lassen 2004; Solter 2005). Increases in serum ALT activity are not liver specific, as increased serum ALT has been reported following strenuous exercise or severe muscle necrosis in dogs and restraint in cynomolgus monkeys (Center 2007; Landi et al. 1990; Solter 2005; Valentine et al. 1990). In the future, separate analysis of the two isoforms of ALT in serum may provide a novel method for better discrimination between tissue origin, severity of hepatic injury, and mechanism of increase, as ALT1 and 2 isoenzymes with unique subcellular and tissue localizations and patterns of response to metabolic or xenobiotic stimuli are described in rodents, dogs, and humans (Glinghammar et al. 2009; Goldstein et al. 2007; Jadhao et al. 2004; Lindblom et al. 2007; Liu et al. 2009; Rajamohan et al. 2006; Thulin et al. 2008; Yang et al. 2009).

In animals, increases in serum ALP activity are most commonly associated with hepatobiliary or bone changes. Unlike ALT, ALP is a membranous enzyme located primarily on the canalicular surface attached by a glycosyl phosphatidyl inositol (GPI) anchor. In cholestatic disease, ALP release is mediated by endogenous phospholipases in association with increased serum concentrations of bile acids. Upregulated ALP expression in the absence of biochemical or morphologic evidence of cholestasis has also been observed on canalicular and basolateral surfaces of hepatocytes and biliary epithelia. Alkaline phosphatase activity is also not liver specific. In the rat, the intestinal isoform is the predominant ALP isoform in serum, with increases in serum ALP activity following food consumption and decreased activity associated with fasting (Center 2007; Hoffmann et al. 1999; Solter 2005). In the dog, increased serum ALP activity can also result from induction of a dog-specific, glucocorticoid-induced ALP isoenzyme in response to increases in endogenous or exogenous corticosteroid hormones (Solter et al. 1994).

The liver is the primary source of serum GGT activity, where the enzyme is a membrane-bound protein found on hepatocytes and biliary epithelia that plays a critical role in cyclic regeneration of amino acids from extracellular glutathione for synthesis of intracellular glutathione. As such, GGT plays a pivotal role in cellular detoxification. Similar to ALP, increased serum GGT activity during cholestasis is almost certainly a result of the release of membrane-bound GGT in part or wholly by bile acids. Increased GGT activity therefore can occur in animals as a result of both cholestasis and biliary epithelial injury. Gamma glutamyltransferase is considered less sensitive than ALP for detection of cholestasis in both the dog and the rat (Bain 2003; Center 2007; Leonard et al. 1984; Loeb 1999).

Overview of Hepatobiliary Biomarker Changes Associated with Prototypic Hepatic DME Induction in Preclinical Species

Reports associating induction of microsomal DMEs with increased liver or blood ALT, ALP, or GGT activities are available in the preclinical literature. However, this review revealed extensive differences in both study design and analytic methodologies across studies included in these reports. One of the goals of this article is to provide a retrospective evaluation of data from rat and dog studies of prototypic inducers where concurrent assessment of hepatic DME induction and liver and serum or plasma ALT, ALP, and/or GGT activities were reported. Retrospective studies of preclinical toxicology data from rats, dogs, and monkeys will also be reviewed. Additionally, a summary of findings for rats given prototypic inducer compounds as part of a recent prospective hepatotoxicogenomics project—including confirmation of DME induction, liver histopathology, and changes in ALT, ALP, and GGT activity—will be presented. Effects of DME, especially phase II enzyme induction on the thyroid-hypothalamic-pituitary axis, will also be discussed.

Corticosteroid-mediated Induction of Hepatobiliary Biomarker Enzyme Activities in the Rat and the Dog

The finding of marked increases in liver ALT activity in rats given cortisol acetate has long been considered as evidence that ALT is an inducible enzyme (Haining 1970; Rosen et al. 1959). Dexamethasone, a more potent glucocorticoid than cortisol and a potent inducer of CYP3A in the rat (Lake et al. 1998; Martignoni et al. 2004), is also associated with increases in hepatic and serum ALT activities in rats (Jackson et al. 2008; O’Brien et al. 2002). In these studies, increases in serum ALT activities consistently exceeded increases in hepatic ALT activities. Importantly, histopathologic findings were limited to hepatic glycogen deposition with minimal to mild multifocal areas of hepatocellular necrosis (Jackson et al. 2008). Because liver histologic findings were of mild severity and disproportionate relative to the large magnitude of concurrent serum ALT increases, pharmacologically mediated induction of hepatic gluconeogenesis was considered as a potential mechanism for these findings (Jackson et al. 2008). This mechanism was considered particularly relevant, as glucocorticoids are known to play a central role in the metabolic response to stress via induction of hepatic gluconeogenesis, primarily via upregulation of phosphoenolpyruvate carboxykinase (PEPCK) (Jin et al. 2004; Sapolsky et al. 2000).

In another study, in which male Sprague Dawley rats were given increasing doses of dexamethasone (0.01 to 300 mg/kg/day) for four days (Ennulat et al. 2010), hepatic expression of CYP3A3 and ALT1 mRNA (but not hepatic ALT activity) were evaluated in parallel with clinical chemistry and liver histopathology. In this study, progressive, dose-responsive increases in serum ALT activity were observed and greatly exceeded increases in liver ALT1 and CYP3A3 mRNA expression, both of which peaked at approximately three-fold of control means at doses ≥30 mg/kg/day (Figure 1). Hepatocellular single-cell necrosis of minimal severity was observed in rats given ≥3 mg/kg/day, with no increase in severity or incidence at doses ≥30 mg/kg/day. These findings are consistent with previous studies of glucocorticoids, where the magnitude of increases in serum ALT activities were larger than expected when based on the minimal severity of liver histopathologic changes (Jackson et al. 2008). Furthermore, comparable increases in serum ALT activity occurred in rats both with and without evidence of hepatocellular necrosis. Therefore, the progressive, dose-responsive increase in serum ALT activity in the absence of a dose-responsive increase in either hepatocellular necrosis or ALT1 or CYP3A3 expression suggest a post-transcriptional mechanism for the increase in serum ALT activities. Although dexamethasone is a known inducer of CYP3A in the rat, the data from these studies (Ennulat et al. 2010; Jackson et al. 2008) support that glucocorticoid-mediated induction of liver and/or serum ALT activities in these rats is more consistent with direct pharmacological or hepatotoxic events rather than changes associated with CYP450 induction.

Figure 1.

Serum alanine aminotransferase (sALT) and hepatic CYP3a3 and ALT1 mRNA changes in male Sprague Dawley rats given ascending doses of dexamethasone for four days. Mean hepatic CYP3a3 and ALT1 mRNA expression and individual rat serum ALT activities are normalized to concurrent control mean. For serum ALT values, open circles represent rats without microscopic evidence of hepatocellular necrosis, and solid circles represent rats with minimal hepatocellular single-cell necrosis.

The hepatic response of the dog to corticosteroid administration differs from that of other animals, particularly the rat and the monkey, in the following ways: (1) dexamethasone has no effect or reduces CYP activity in dog hepatocytes in vitro (Jayyosi et al. 1996; Lu and Li 2001); (2) the dog is also unique in expression of a corticosteroid-inducible isoenzyme of ALP (CIALP) in addition to the liver- and bone-specific ALP isoenzymes found in the circulation of the dog and other species (Hoffmann et al. 1999). In dogs given prednisone, increases in hepatic ALP and GGT, but not ALT, activities are seen, often along with increases in serum ALP, GGT, and ALT activities (Rutgers et al. 1995; Solter et al. 1994). Increased serum activity of the liver ALP isoenzyme (LALP) can occur following a single dose of prednisone in the dog (Wiedmeyer et al. 2002a); however, significant increases in serum CIALP activity do not occur before seven to ten days of prednisone administration (Solter et al. 1994; Wiedmeyer et al. 2002b). These differences are related in part to different regulatory mechanisms, as the onset of hepatic CIALP mRNA expression is delayed in comparison to LALP mRNA (Wiedmeyer et al. 2002b). Corticosteroid-mediated induction and release of both ALP and GGT into serum in the dog occurs independently of cholestasis, as both hepatic and serum bile acid concentrations are unchanged in dogs given prednisone (Solter et al. 1994). Although these increases in hepatic and serum ALP and GGT activities are attributed to induction, the increased serum ALT activities in these studies may be related to hepatotoxicity, because increases of hepatic ALT were not seen. Although histopathology was not performed in these studies, other studies of dogs given comparable doses of prednisone consistently demonstrate hepatocellular glycogenosis and hypertrophy with focal hepatocellular necrosis (Fittschen and Bellamy 1984; Rutgers et al. 1995). Therefore, corticosteroid-mediated induction of ALP and GGT in dogs is a unique, pharmacologically mediated event that is not related to cholestasis or induction of hepatic DME activity.

Hepatobiliary Biomarker Findings in Association with Prototypic Hepatic Enzyme-inducing Agents in the Rat and the Dog

Our current understanding of the relationship between liver and serum hepatobiliary biomarker enzyme activities and DME induction in rats and dogs is based on numerous, yet widely varied published studies of prototypical DME inducers in these species (Tables 1–4). In rodents, the most common inducers evaluated were phenobarbital (PB), ethanol, and PPARα agonists (Tables 1 and 3). In dog, hepatic metabolizing enzyme induction was most commonly reported for anticonvulsant drugs such as PB, phenytoin (PHE), or primidone (PRI, Table 2).

Table 1.

Changes in liver parameters in rats treated with various drugs associated with hepatic drug-metabolizing enzyme induction

Compound, dose and duration	Liver Enzyme Activity/gram^a,b	Serum Enzyme Activity^a	Liver histopathology	Reference
Cortisol acetate, 1 mg, 7 days	↑ALT 5X	NM	Not evaluated	Rosen, 1959
Dexamethasone, 50 mg/kg, 4 days	↑ALT 3.2X	↑ALT 10.3X	Hepatocellular glycogenosis and single cell necrosis	Jackson, 2008
Dexamethasone, 100 mg/kg, 3 days	↑ALT 3X	↑ALT 5X	Not evaluated	O'Brien, 2002
Phenobarbital, 100 mg/kg, 1 day	↑ALT 2.2X	NM	Not evaluated	Boll, 1998
Phenobarbital, 100 m/kg, 4 days	↑GGT 3.4X^c	NM	Not evaluated	Ratanasavanh, 1978
Phenobarbital, 100 mg/kg, 5 days	↑GGT <1.7X	No change	Not evaluated	Goldberg, 1981
Phenobarbital, 100 mg/kg, 5 days	↑GGT ~;<1.8X^d	No change	Not evaluated	Roomi, 1981
Aminopyrine, 600 mg/kg, 20 days	↑GGT 13.1X	↑GGT 3.2X	Hepatocellular hypertrophy	Satoh, 1982
Wyeth-14643, 50 mg/kg, 3 days	No ↑ ALT, ↑ALP 3X	↑ALT 1.3X , ↑ ALP 2.3X	Not evaluated	O'Brien, 2002
WAY-144112, 130 mg/kg, 28 days	NM	↑ALT < 1.5X, ↑ALP < 1.7X	Hepatocellular hypertrophy	Peterson, 2004
Clofibrate, 800 mg/kg, 5 days	NM	↑ALT 1.5X, ALP NM	Hepatocellular hypertrophy	Kramer, 2003

Abbreviation: ALP, alkaline phosphatase; ALT, alanine aminotransferase; AST, aspartate aminotransferase; GGT, gamma glutamyltransferase; NM, not measured.

^a Fold change compared to concurrent controls.

^b Homogenate, per g liver weight unless otherwise specified.

^c Plasma membrane fraction.

^d Microsomal fraction.

Table 2.

Changes in liver parameters in dogs treated with various drugs associated with hepatic drug-metabolizing enzyme induction.

Compound, dose^a and duration	Liver CYP activity^b	Liver ALP or ALT activity/g^b	Serum enzyme activity^b	Liver histopathology	Reference
Phenobarbital, median dose of 5 mg/kg for 4 months to 6.5 years	11× ↑ BROD activity	No difference in ALP but ↓ ALT activity	Median sALP ↑ 4.7×, median sALT ↑ 2.1X	Hepatocellular hypertrophy, SER proliferation and exascerbation of background liver findings	Gaskill 2005
Phenobarbital, ascending dose from 5 to 40 mg/kg for 13 days, followed by 8-day recovery	15.3× ↑aminopyrene demethylase activity	≤12.3× ↑ALP on Day 14, 1.7× ↑ on recovery Day 8	≤2.8× ↑ALP on Day 14, ↑ 1.4X on recovery Day 8	No remarkable changes	Litchfield 1972
Phenobarbital, 40 mg/kg for 7 weeks	3.3× ↑ CYP protein content	Not measured	<2.7× ↑ in ALP	No remarkable changes	Balazs 1978
Phenytoin, 40 mg/kg for 7 weeks	1.75× ↑ CYP protein content	Not measured	No remarkable changes	No remarkable changes
Primidone, 40 mg/kg to week 3, 80 mg/kg from weeks 3 to7	2.3× ↑ CYP protein content	Not measured	No remarkable changes	No remarkable changes
Primidone, 55 or 165 mg/kg for 6 months	Not measured	No remarkable changes	No remarkable changes	Hepatocellular hypertrophy, SER proliferation and/or lipidosis	Bunch 1985
Phenytoin, 66 or 198 mg/kg for 6 months	Not measured	1.7 and 3.1× ↑ALP	Peak ALP ↑ by 9 weeks, gradual ↓ to plateau at 16 weeks for high dose	Hepatocellular hypertrophy, SER proliferation and/or lipidosis
Primidone (55 mg/kg) and phenytoin (66 or 198 mg/kg) for 6 months	Not measured	5.5× and 7.7× ↑ALP	Peak ALP ↑ during week 9-12, sustained in high dose and gradual ↓ at low dose	Hepatocellular hypertrophy, SER proliferation and/or lipidosis, with intrahepatic cholestasis and single cell hepatocellular necrosis in decedent dogs

^a Oral daily dose in mg/kg/day.

^b Fold change compared to concurrent controls.

Table 3.

Changes in liver parameters in rat models of chronic ethanol consumption.

Ethanol model, sex, strain, and duration	Liver enzyme activity/g^a	Serum enzyme activity^a	Liver histopathology	Reference
LDC diet, pair fed, 36% kcal^c, female SD, 6 weeks	↑GGT 1.7×, ↑ALT 1.7×, no ↑ALP	↑GGT 1.9×, ↑ALT 2.4×, ↑ALP 1.8×	Not evaluated	Nishimura 1982
LDC diet, pair fed, 36% kcal^c, female SD, 6 weeks	↑GGT 1.5×	↑GGT 2×	Not evaluated	Nishimura 1977
LDC diet, pair fed, 36% kcal^c, female SD, 4–5 weeks	↑GGT 1.6×, ↑ALT 1.5×	↑GGT 3.5×, no ↑ALT	Not evaluated	Teschke 1977
LDC diet, pair fed, 36% kcal^c, female SD, 6 weeks	↑GGT 1.5×, ↑ALT 1.4×, no ↑ALP	↑GGT 1.5×, ↑ALT 2.4×, ↑ALP 1.8×	Not evaluated	Teschke 1983
LDC diet, pair fed, 35% kcal^c, male SD, 6 weeks	Not measured	↑ALT 1.6×, ↑ALP 1.5×	Diffuse hepatocellular lipidosis	Oh 1998
LDC-based ethanol diet with low and high fat groups, pair fed, 35% kcal^c, male SD, 12 weeks	Slight ↑GGT relative to high fat diet control	Not measured	Centrilobular lipidosis and ↑GGT immunoreactivity	Millsbeck 1986
Modified LDC diet, 36% kcal^c, male SD, 4 weeks	↑GGT<3×, inversely related to CHO content	↑GGT <3.5×, inversely related to CHO content	Not evaluated	Yamada 1985
Ethanol, 10% v/v in drinking water with ad lib pelleted food and water, male and female SD, 4 weeks	↑GGT 2.5× in males, 1.8× in females	↑GGT <2.3× in males, 1.6× in females	Not evaluated	Lahrichi 1982
Ethanol in Alcock powdered alcohol diet and in drinking water, pair fed, female Wistar, 12 weeks	↑GGT 2.9× by 9 weeks, return to contol level by 12 weeks	↑GGT 3.4× by 9 weeks, return to contol level by 12 weeks	Not evaluated	Rambabu 1986

Abbreviation: LDC; Lieber DeCarli diet, CHO; carbohydrate.

^a Fold change compared with concurrent controls.

^b Homogenate.

^c Ethanol.

Table 4.

Pathology findings and CYP gene expression changes in rats given prototypic drug-metabolizing enzyme inducer compounds

Compound	TCDD^a	3-MC	BNP	CBZ	PB	PHE	CYP	PCN	RFP	CLF	DEHP	FEN	WY-14_643	TBZ	CZL	PBO	ORPH
Dose (mg/kg)	15	60	100	100	30	300	300	100	100	300	1500	30	100	1000	150	1000	300
1a1^b	20 (2)	1604 (81)	806 (157)	2.6 (4.3)	1.4 (0.3)	0.7 (0.1)	1.9 (1)	0.8 (0.6)	1.1 (0.3)	0.5 (0.1)	0.6 (0.3)	0.6 (0.1)	0.6 (0.1)	1399 (222)	1.6 (1)	1.3 (1.5)	215 (20)
1a2	1.7 (0.2)	5.2 (0.2)	3.2 (0.3)	0.7 (0.2)	0.5 (0.1)	0.5 (0.2)	0.2 (0.1)	0.4 (0)	0.3 (0.1)	0.4 (0.1)	0.2 (0)	0.2 (0)	0.1 (0)	2.9 (0.3)	0.1 (0.1)	0.3 (0.1)	0.2 (0.1)
2b2	1 (0.3)	0.5 (0.3)	1.2 (0.2)	5.7 (1.5)	11 (0.9)	9.5 (1)	2.2 (0.4)	ORPH
Dose (mg/kg)	15	60	100	100	30	300	300	100	100	300	1500	30	100	1000	150	1000	300
1a1^b	20 (2)	1604 (81)	806 (157)	2.6 (4.3)	1.4 (0.3)	0.7 (0.1)	1.9 (1)	0.8 (0.6)	1.1 (0.3)	0.5 (0.1)	0.6 (0.3)	0.6 (0.1)	0.6 (0.1)	1399 (222)	1.6 (1)	1.3 (1.5)	215 (20)
1a2	1.7 (0.2)	5.2 (0.2)	3.2 (0.3)	0.7 (0.2)	0.5 (0.1)	0.5 (0.2)	0.2 (0.1)	0.4 (0)	0.3 (0.1)	0.4 (0.1)	0.2 (0)	0.2 (0)	0.1 (0)	2.9 (0.3)	0.1 (0.1)	0.3 (0.1)	0.2 (0.1)
2b2	1 (0.3)	0.5 (0.3)	1.2 (0.2)	5.7 (1.5)	11 (0.9)	9.5 (1)	2.2 (0.4)	2.7 (0.2)	0.8 (0.3)	2.08 (0.4)	2.1 (0.3)	2.7 (0.1)	1.8(0.1)	5.7 (1.6)	4.1 (1.1)	5.8 (3)	5.7 (0.6)
3a2	0.9 (0.2)	1.2 (0.3)	1.1 (0)	1.3 (0)	1.2 (0.1)	1.5 (0.2)	0.9 (0.1)	1.2 (0.1)	0.7 (0.2)	1 (0.1)	1(0.1)	1.2(0.2)	1 (0.1)	0.4 (0.2)	0.8 (0.4)	1.7 (1.7)	1 (0.2)
3a3	1.2 (0.1)	1.2 (0.2)	1.1 (0.1)	1.8 (0.2)	2.2 (0.4)	2.7 (0.2)	2.3 (0.2)	2.4 (0.2)	1.7 (0.3)	1.1 (0.2)	1.5 (0.1)	1 (0.1)	0.8 (0.1)	1.6 (0.2)	2.4 (1.6)	3.1 (2.8)	1.7 (0.3)
4a1	0.8 (0.1)	0.7 (0.2)	0.7 (0.1)	0.7 (0.2)	1.1 (0.1)	0.3 (0.1)	0.2 (0)	0.4 (0.1)	0.6 (0.1)	1.9 (0.2)	1.8 (0.2)	2.7 (0.4)	1.9 (0.1)	0.4 (0.1)	0.3 (0.1)	0.7 (0.1)	0.5 (0.2)
ALT	0.8 (0.1)	0.7 (0.2)	0.9 (0.2)	1.2 (0.2)	0.9 (0.1)	1.4 (0.3)	1.4 (0.2)	0.8 (0.1)	2.1 (0.7)	1.1 (0.1)	1.5 (0.4)	1.1 (0.2)	1 (0.4)	1.4 (0.2)	9 (6)	1.6 (0.5)	4 (0)
ALP	0.8 (0)	0.9 (0.1)	0.8 (0.1)	0.8 (0.2)	1 (0.2)	1.1 (0.3)	0.9 (0.1)	0.9 (0.1)	1.5 (0.4)	1.3 (0.3)	1.4 (0.3)	1.2 (0.1)	1.7 (0.3)	1.1 (0.2)	1.7 (0.5)	1.3 (0.3)	1 (0.3)
GGT	0 (0)	0 (0)	0 (0)	0 (0)	0.4 (0.5)	0 (0)	0.4 (0.5)	0 (0)	0.2 (0.4)	0 (0)	0.6 (0.5)	0 (0)	0 (0)	0 (0)	8.3 (6.4)	23 (17)	0 (0)
AST	1.5 (0.3)	1.1 (0.1)	1.1 (0.2)	1.5 (0.7)	1 (0.2)	1.3 (0.3)	0.8 (0.1)	0.9 (0.2)	1.2 (0.6)	1.2 (0.6)	1.5 (0.8)	1.2 (0.1)	1.1 (0.3)	1.3 (0.4)	25 (24)	1.6 (0.5)	3.6 (1.5)
Tbili	0.8 (0)	1 (0)	1.2 (0.4)	1.6 (1.3)	1.2 (0.4)	1.4 (0.5)	0.7 (0.2)	1 (0)	13.5 (0.5)	1.2 (0.4)	2 (0.7)	1 (0)	1.4 (0.5)	5 (1.2)	7.8 (5.1)	6.9 (11.6)	3 (1)
SBA	1.4 (0.4)	0.6 (0.2)	0.5 (0.2)	0.9 (0.2)	0.7 (0.1)	1 (0.4)	2.1 (1.3)	0.5 (0.1)	3.1 (0.7)	0.9 (0.4)	0.8 (0.1)	2.2 (1.4)	1 (0.3)	0.8 (0.3)	2 (1)	1.5 (0.5)	2.2 (0.9)
Chol	1.6 (0.3)	1.8 (0.3)	1.3 (0.3)	1.2 (0.3)	1.3 (0.4)	1.4 (0.2)	1.5 (0.3)	1.2 (0.3)	3.2 (1.8)	0.7 (0.2)	1.7 (0.2)	0.6 (0.2)	0.7 (0)	2.6 (0.3)	0.5 (0.2)	2.7 (0.7)	1.1 (0.3)
Trig	2 (0.7)	0.5 (0.3)	0.5 (0.2)	0.8 (0.2)	0.7 (0.3)	0.7 (0.2)	NM	0.9 (0.4)	1.8 (0.7)	0.8 (0.3)	1.1 (0.2)	0.7 (0.2)	0.7 (0.2)	2.4 (1.1)	0.3 (0.1)	5.3 (9.5)	0.5 (0.2)
Body weight change (%)	−7.5 (1)	−7.6 (1.7)	−4.4 (1.5)	−5.1 (2.5)	−2.4 (0.5)	−8 (1.5)	NM	−3.8 (2.6)	−8.3 (1.3)	−1.6 (4)	−7.2 (2.6)	−3.9 (1.4)	−3.6 (1.7)	−15 (3.3)	−15 (1.3)	−10 (4.8)	−24 (1.7)
Relative liver weight change (%)	39 (14)	43 (14)	23 (2.6)	13.6 (6)	20 (4)	25 (5)	NM	37 (9.3)	26 (7)	49 (5)	64 (7.4)	48 (9)	71 (2)	73 (9.4)	118 (7)	121 (27)	71 (19)
Hypertrophy, centrilobular^c				4	5	5
Hypertrophy, panlobular	4									5	5	5	5	5	4		2
SER proliferation	2
Cytoplasmic rarefaction
Peroxisome proliferation										5	5	5	5
Hepatocellular mitoses					4	1				5	5	5	4		4
Kupffer cell activation	3														4
Lipidosis	4	5	5											3	4		3
Hepatocellular necrosis															4	3	3

Abbreviations: BNP, beta-napthoflavone; CBZ, carbamazepine; CLF, clofibrate; CYP, cyproterone; CZL, clotrimazole; DEHP, di(2-ethylhexyl) phthalate; FEN, fenofibrate; 3-MC, 3-methylcholanthrene; NM, not measured; ORPH, orphenadrine; PB, phenobarbital; PBO, piperonyl butoxide; PCN, pregnenolone-16α-carbonitrile; PHE, phenytoin; RFP, rifampin; TBZ, thiabendazole; TCDD, 2_3_7_8-tetrachlorodibenzo-p-dioxin.

^a μg/kg/day

^b Mean (SD)

^c For histopathology findings, number of rats with finding (five per group). Bold font for CYP isoform data indicates CYP induction based on magnitude of gene expression change at level of group mean or individual rat.

There are rare reports in which PB was shown to minimally increase hepatic ALT activity in the rat (Boll et al. 1998). However, effects on serum activity are unknown, as concurrent serum ALT activity was not evaluated, and to our knowledge, hepatic ALT activities have not been reported in repeat-dose studies in rats given PB. In contrast, rats given comparable doses of PB for up to five days exhibit increased liver GGT activity in association with hepatic DME induction without concomitant increases in serum GGT activity (Goldberg et al. 1981; Ratanasavanh et al. 1979; Roomi and Goldberg 1981). Notably, the magnitude of hepatic GGT increase was different between these three studies and was related to whether the whole liver or the hepatic plasma membrane fraction, the predominant location of liver GGT, was evaluated. The lack of concordance between liver and serum GGT activities is not surprising, given the low baseline activity (Lahrichi et al. 1982) and exceptionally short circulating half-life of GGT in the rat (Boyd 1983; Huseby 1992; Loeb 1999).

In the dog, induction of hepatic DME activity with increasing doses of PB was associated with increases in both hepatic and serum ALP activities (Litchfield and Conning 1972). Similar increases in hepatic and serum ALP activities were noted in dogs given comparable doses of PB for up to nine weeks (Balazs et al. 1978; Unakami et al. 1987). In contrast, in a prospective study of dogs given therapeutic doses of PB for up to six years, no effect on hepatic ALP or ALT activities were observed, despite increases in serum ALP and ALT activities and confirmation of hepatic DME induction by benzyloxyreso-rufin-O-dealkylase activity as a measure of CYP2B11, and CYP2B immunoblotting in liver homogenate (Gaskill et al. 2005). The cause of this discrepancy is unclear, but it may be related to methodologic differences in tissue extraction techniques, as whole homogenates rather than membrane fractions were evaluated in the latter study (Gaskill et al. 2005). In dogs given PHE alone or in combination with PRI, induction of hepatic DME activity was associated with dose-responsive increases in both hepatic and serum ALP activities (Bunch et al. 1985); however, serum ALP activities declined to baseline levels over time. For all of these studies, the predominant histologic findings at study termination were hepatocellular hypertrophy and SER proliferation, with exacerbation of spontaneous or background hepatic pathology in dogs given the lower pharmacologic doses of PB, and hepatocellular necrosis and intrahepatic cholestasis in the decedent dogs given PRI and PHE in combination. Based on the lack of prominent hepatobiliary pathology in all but dogs given the combination PHE/PRI, a relationship between hepatic metabolizing enzyme induction and increases in serum and/or liver ALP activities in the dogs in these studies cannot be excluded. Furthermore, although hepatic ALP activity in dogs given PB or PHE alone or in combination appears to not correlate with serum ALP activities, the disproportionate changes in liver and serum hepatobiliary marker enzyme activities in these studies do not exclude the potential for serum and hepatic levels to both reflect induction. Rather, differences in serum and hepatic enzyme activities may be related to methodologic differences in tissue extraction or to differences between serum and tissue half-life, as rate of clearance is a major determinant of serum activity of hepatobiliary marker enzymes (Boyd 1983).

Other microsomal enzyme inducers have also been studied for effects on liver and/or serum hepatobiliary marker enzyme activity. Fibrate hypolidemics and other PPARα agonists are potent inducers of CYP4A in the rat (Peterson et al. 2004; Sabzevari et al. 1996). Minimal increases in serum ALT and/or ALP activity were seen in rats given WY-14643, WAY-144112, and clofibrate and were associated with increased hepatic ALP, but not ALT, activities for WY-14643 and WAY-14112 (Kramer et al. 2003; O’Brien et al. 2002; Peterson et al. 2004). Importantly, increases in enzyme activities for these PPARα agonists were accompanied only by centrilobular hypertrophy. Chronic administration of aminopyrine, a nonspecific CYP substrate in rats, also causes CYP450 induction, based on increases in both microsomal enzyme activities and protein content, and was associated with an up to eight-fold increase of hepatic GGT activity, and an approximately three-fold increase in serum GGT activity (Satoh et al. 1982). As above, histologic findings in this study were limited to hepatocellular hypertrophy. These examples are difficult to further interpret in terms of relationship between DME induction and serum hepatobiliary marker enzyme activity because of differences in CYP enzyme induction profile and/or mechanism of induction, and lack of corroborative information from other studies.

Chronic ethanol consumption in rodents and man is commonly associated with induction of CYP2E1 and increases in serum and liver GGT activities (Roberts et al. 1995; Teschke et al. 1983). Despite extensive methodological differences between rat chronic ethanol consumption models, particularly with regard to ethanol dosage and dietary carbohydrate content, increased hepatic GGT activities consistently paralleled increases in serum GGT activities (Table 3). In contrast, increases in hepatic and/or serum ALT activities were less commonly observed, and no effects on liver ALP activities were reported despite sporadic mild increases in serum ALP activities. Although induction of hepatic DME activity was not assessed in these rat chronic ethanol consumption studies, in comparable rat dietary models of chronic ethanol consumption, ethanol was shown to cause up to a 2.5-fold increase in CYP2E1 protein (Roberts et al. 1995). When histopathology was assessed, microscopic findings were limited to lipid accumulation and increased membrane GGT activity in centrilobular hepatocytes (Misslbeck et al. 1986), or panlobular hepatocellular lipidosis in association with increased hepatic CYP2E1 protein (Oh et al. 1998). These data suggest that hepatic GGT induction can be associated with CYP induction and increases in serum GGT activity in the rat.

In summary, these studies of prototypic inducer compounds in the rat and the dog demonstrate that the relationship between induction of hepatic DME activity and increases in liver or serum hepatobiliary marker enzyme activities is both compound and species-specific.

Retrospective Analyses of Rat, Dog, and Monkey Toxicology Study Data

Retrospective analyses of toxicology studies on new chemical entities provide an additional resource for evaluation of the relationship between hepatic microsomal enzyme induction and clinical pathology parameters in rats, dogs, and monkeys. For each toxicology study, end points evaluated included liver weight and histopathology, clinical chemistry, and hepatic CYP450 content and activity. Importantly, hepatic hepatobiliary marker enzyme activity was not assessed, as this is not a standard end point in preclinical toxicology studies used for human risk assessment (Boone et al. 2005; Weingand et al. 1996).

For the rat, data were reviewed from toxicology studies in male and/or female Fischer or Sprague Dawley rats given ten new chemical entities from five different therapeutic classes for approximately two weeks to three months (Amacher et al. 1998). Centrilobular or panlobular hepatocellular hypertrophy was associated with increases in absolute liver weights of >20%. However, there was no relationship between the magnitude of liver weight increase or hepatocellular hypertrophy and the degree of CYP450 induction. Importantly, CYP450 induction, hepatocellular hypertrophy, and increased absolute liver weights, in the absence of other histologic findings, were not associated with changes in serum ALT activity or other measured serum hepatic enzymes. For the compounds used in these studies, the results indicated that CYP450 induction and secondary hepatic adaptive changes were not associated with changes in serum indicators of hepatic injury in the rat.

In the retrospective analyses of preclinical toxicology study data for candidate or investigative drugs in the dog (Amacher et al. 2001) and monkey (Amacher et al. 2006), increased microsomal enzyme activity, consistent with hepatic DME induction, was seen in one or both sexes for eight of nine compounds from five different pharmacological classes in the dog, and for nine of ten compounds from four different pharmacological classes in the monkey. Microsomal enzyme induction in both species was variably associated with hepatocellular hypertrophy and relative liver weight increases, but no changes in clinical chemistry parameters or histopathology findings indicative of hepatotoxicity were seen in either species.

Prospective Evaluation of the Association between CYP Induction and Hepatobiliary Marker Changes in the Rat

Data collected from rats given prototypic inducer compounds as part of a recent hepatotoxicogenomics project were also evaluated to assess the relationship between DME induction and changes in serum ALT, ALP, and GGT activity (Ennulat et al. 2010). In this study, male Sprague Dawley rats (N = 254) were given seventeen prototypic inducer compounds for four days by oral gavage (Table 4). Study end points collected on Day 5 included histopathology, clinical pathology, body and relative liver weight, and CYP450 gene expression. Data were normalized across studies to concurrent control mean, such that individual values for treated animals were expressed as a fold change of the control mean for liver weight, CYP450 mRNA expression, and serum ALT and ALP activity. Absolute rather than normalized values were assessed for GGT because of the low level of GGT activity in the rat (Huseby et al. 1992; Loeb 1999). CYP450 gene thresholds used to identify rats with DME induction were ≥ five-fold upregulation of CYP450 1A1 or 2B2 and ≥ two-fold upregulation of 1A2, 3A2, or 4A1 mRNA. Based on these criteria, enzyme induction was present in all seventeen compounds. Microscopic findings consisted primarily of hepatocellular hypertrophy and lipid or glycogen accumulation. Peroxisome proliferation and increased hepatocellular mitoses were observed for several of the PPARα agonists, Kupffer cell activation was noted in rats given 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin and clotrimazole, and a dose-responsive increase in hepatocellular single-cell necrosis was identified in rats given clotrimazole, piperonyl butoxide, and orphenadrine. With the exception of rats given treatments which caused necrosis, minimal (<1.5-fold) increases in ALT, ALP, and GGT activities were the most common serum clinical chemistry findings. Rats given clotrimazole, piperonyl butoxide, and orphenadrine had concurrent dose-responsive increases in total bilirubin and bile acids of up to eight- and 2.2-fold, respectively.

Because of the strong association between ALT changes and hepatocellular necrosis, changes in serum ALT, ALP, and GGT activity were evaluated for all seventeen inducer compounds, and then separately for the three compounds that caused necrosis and the fourteen inducer compounds that did not elicit necrosis (Figure 2). Mean serum ALT, ALP, and GGT activities were only minimally increased in treated rats relative to controls, however, the greatest increases and variability of these enzymes was seen in rats given the inducer compounds that also caused hepatocellular necrosis. When the compounds that caused necrosis were excluded, ALT and GGT and, to a lesser extent, ALP activities decreased to levels comparable to those seen in control rats. These findings provide further support that hepatic DME induction in the absence of hepatocellular degeneration or necrosis is not associated with noteworthy increases in serum ALT, ALP, or GGT activities in the rat.

Figure 2.

Serum alanine aminotransferase, alkaline phosphatase, and gamma glutamyltransferase changes for prototypic DME inducer compounds in the presence and absence of hepatocellular necrosis. Boxplots of individual animal fold change data for alanine aminotransferase, alkaline phosphatase, and gamma glutamyltransferase from left to right for all animals (N = 17 compounds, <244 rats), animals given inducer compounds that caused necrosis (n<42 rats given clotrimazole, piperonyl butoxide or orphenadrine), and animals given prototypic inducer compounds that did not cause necrosis (n = 15, <202 rats). The top, bottom, and line through the middle of the box correspond to the 75th percentile (top quartile), 25th percentile (bottom quartile), and 50th percentile (median), respectively. The whiskers extend to the 10th percentile and 90th percentiles, and outlier values are represented by red circles.

Effects of Enzyme Induction on Endogenous Thyroid Hormones in Preclinical Species

In rodents and other species, alteration of the hypothalamus-pituitary-thyroid axis can be a secondary consequence of enzyme induction. Most notably, PB administration in rats has been associated with decreased thyroxine (T₄) and tri-iodothyronine (T₃) owing to induction of liver UDP-glucuronyltransferase (UGT) and accelerated biliary excretion (O’Connor et al. 1999). Decreases in functional T₄ and T₃ levels were further associated with compensatory increases in circulating thyroid-stimulating hormone (TSH) concentration and increased thyroid gland weights following four weeks of PB administration (O’Connor et al. 1999). These findings have been replicated in numerous comparable studies in Sprague Dawley rats (Barter and Klaassen 1994; Capen 1997; Hood et al. 2003, Hood, Hashimi et al. 1999; Hood, Liu et al. 1999; McClain et al. 1989). Development of thyroid follicular hypertrophy, hyperplasia, and progression to neoplasia occured with chronic PB administration in rats (Capen 1997; McClain et al. 1989; Meek et al. 2003); however, similar patterns of thyroid hormone changes without thyroid tumorigenesis were seen in rats given other microsomal enzyme inducing compounds including 3-methylcholanthrene and pregnenolone-16-α-carbonitrile (Barter and Klaassen 1994; De Sandro et al. 1992; Saito et al. 1991).

The Sprague Dawley rat appears to be unique in the development of thyroid tumors with chronic PB administration. Mice and other laboratory rat strains chronically given PB do not develop thyroid neoplasia (Kato et al. 2005; Kato et al. 2008; Lecureux et al. 2009; Meek et al. 2003), presumably owing to negligible effects on TSH and thyroid follicular cell proliferation following chronic PB administration (Hood et al. 2003; Viollon-Abadie et al. 1999). Thyroid tumorigenesis is also not reported in dogs, which develop similar thyroid hormone changes with chronic PB administration (Gaskill et al. 1999). Further, thyroid carcinogenesis has not been seen clinically in patients on chronic PB regimens (Capen 1997; Meek et al. 2003).

The marked differences in thyroid tumor susceptibility between different animal species and rat strains treated with PB is probably related to differences in thyroid hormone homeostatic mechanisms, as well as the specific DME changes that influence thyroid function. Notably, although induction of T₄- and T₃-UGTs has been the most well-studied mechanism proposed to explain thyroid hormonal effects of PB in rats, additional factors affecting thyroid hormone metabolism may be linked to hepatic DME induction, including induction of sulfotransferases and hepatic T₄ and T₃ transporters, inhibition of deiodinases, and differences in circulating half-life of thyroid hormones or binding proteins (Kato et al. 2005; Kato et al. 2008; Lecureux et al. 2009; McClain et al. 1988; McClain 1995; Qatanani et al. 2005).

Conclusions on DME Induction and Clinical Pathology Biomarkers in Preclinical Species

Induction of DME activity can be associated with upregulation of liver activities of the hepatobiliary biomarker enzymes ALT, ALP, and GGT in rats, but the pattern and magnitude of increase of these enzymes in both the liver and the serum vary greatly between different compound classes in the rat. Additionally, the discrimination between whether increases in these hepatobiliary biomarkers in the serum is owing to upregulation within the liver or release from hepatocytes following liver injury requires both concurrent assessment of liver histopathology with clinical chemistry and consideration of marker biology and compound pharmacology. Based on data from published rat studies and in particular, results of the prospective study with prototypic inducer compounds, the weight of evidence suggests that in the absence of liver histopathologic findings other than hypertrophy, CYP induction alone is not associated with changes in serum or plasma activity of hepatobiliary marker enzymes in the rat.

In nonrodents, based on retrospective evaluation of data from dog and monkey preclinical toxicology studies in which hepatic DME induction was confirmed, there is also no consistent relationship between hepatic microsomal induction and hepatotoxicity, as reflected in changes in clinical chemistry parameters. However, induction of liver ALP activity in both phenobarbital- or corticosteroid-treated dogs suggests that release of serum liver enzymes can occur independently of liver injury or induction of DME activity.

Hepatic Metabolic Enzyme Induction and Associated Clinical Pathology Changes in Humans

In contrast with published preclinical information, clinical reports on DME induction have focused predominantly on effects on pharmacokinetics properties, drug–drug interactions, and bioactivation specific to a xenobiotic rather than relationship to liver pathology. Among clinical investigations of enzyme induction that have included effects on routine laboratory findings, reports with comparative liver histologic or functional end points are particularly limited. Thus, interpretation of the significance of the clinical pathology findings in humans exposed to prototypic DME-inducing agents has been largely based on empirical and indirect observations. These reports on associations between DME induction and human clinical pathology findings have predominantly focused on CYP families 1–3, the phase I enzymes responsible for metabolism of a majority of marketed drugs (Dresser et al. 2000; Tanaka 1998). The phase II UGT enzymes, which are responsible for the majority of drug conjugation reactions, have also received attention for influencing human clinical pathology (Evans and Relling 1999). Administration of prototypical agents that induce these DMEs and associated alterations in laboratory biomarkers thus forms the basis for the current understanding of the relationship between induction and clinical pathology results in humans. As with preclinical studies, collective results of these published clinical reports have generally not supported an effect on routine laboratory end points that is directly or consistently linked with phase I and/or II DME induction.

Hepatobiliary Biomarker Findings in Association with Prototypic Hepatic-inducing Agents in Humans

The drugs that have been most commonly studied clinically for the relationship between DME and laboratory findings are the early generation antiepileptics (Table 5 ), which include PB, PHE, PRI, and carbamazepine (CBZ) (Perucca et al. 1984). Clinical consequences of DME induction with the anti-mycobacterial rifampicin are also described in the literature. Although these agents induce a variety of human CYPs, they share the potential of profound induction of CYP3A4, rendering CYP3A4 a particularly common focus of studies on clinical effects of DME induction (Mohutsky et al. 2010). Extrapolation of findings related to the CYP3A subfamily from humans to animals is problematic. For example, CYP3A4 is highly inducible through PXR ligand binding (Hewitt et al. 2007; Jones et al. 2000) and only weakly induced through CAR in humans (Faucette et al. 2006), in contrast with induction of CYP3A by both PXR and CAR in rodents (Graham and Lake 2008). Even within human populations, CYP3A4 inductive responses vary widely (Hewitt et al. 2007; Wilkinson 1996). Beyond inter-individual differences in exposure to inducers, age, obesity, and even exercise level may affect CYP3A4 activity (Anderson 2008; Kotlyar and Carson 1999; Meijer et al. 2001). Acute-phase inflammatory reactants can profoundly inhibit the extent of CYP3A4 induction (Cooper et al. 2008; Jover et al. 2002). These variables likely contribute to a greater extent to the conflicting information on clinical pathology findings with DME induction in man than in traditional preclinical studies in comparatively homogeneous laboratory animal species with comparable CYP-inducing agents.

Table 5.

Serum or plasma hepatobiliary marker enzyme changes in adults and children given anticonvulsants.

Drug ^a (number of patients)	Dosing duration	Age^b, number, and sex ^c	Testing interval and comparator	GGT	ALP	AST	ALT	Liver biopsy, other	Reference
PB (4), DPH (19), MD (35)	>12 months	Children, 1.5–19, n=34M, 29F	Approximately every 6 weeks for patients with persistent serum enzyme elevations, relative to laboratory reference values	2-2.4× ↑ GGT in all 56 patients with therapeutic serum levels of PB or DPH	Not reported	Slight persistent ↑ AST in 6/63, transient ↑ mean AST in 5/63	Slight persistent ↑ ALT in 6/63, transient ↑ mean ALT in 5/63	Uniform hepatocelluar swelling by light microscopy; proliferation of smooth ER by TEM.	Aiges 1980
DPH (54), CBZ (56)	2 months to 8 years	Children, 5-12, n=110	Single interval, relative to laboratory reference values	<10× ↑ mean GGT in 91% of patients on DPH and 64% on CBZ	<1.7× ↑ mean ALP in 39% of patients on DPH, <1.3× ↑in 14% on CBZ	<1.7× ↑ in 11% of patients on DPH, <1.4× ↑ <1% on CBZ	<4.5× ↑ in 28% of patients on DPH, <1.2× ↑ in 9% on CBZ		Aldenhovel 1988
PMD (103), DPH (63), VPA (32)	1 month to 7.7 years	Children, 1-18,n=198	Single interval, relative to 36 untreated epileptic control children	<5.1× ↑ mean GGT for DPH group and 1.8× ↑ for PMD	Not reported	No treatment-related effect	<1.5× ↑ mean ALT for DPH, <1.2× ↑ for PMD group		Deutsch 1986
CBZ (22)	12 months	Children,5-12,13M, 9F	3, 6, 12 months, relative to pre-treatment values	↑ mean GGT at 6 and 12 months	↑ mean ALP (total, liver, bone and/or intestinal isozymes) at 3, 6, and 12 months	No treatment-related effect	No treatment-related effect		Voudrris 2005
CBZ (9)	5 weeks	Children,6-14,n=9	Single interval, relative to pre-treatment values	No treatment- related effect	Not reported	Not reported	Not reported	3.5× ↑ 6-β-hydroxycortisol	Moreland 1982
VPA (42), CBZ (36), VPA and CBZ (36)	5 months to 7.7 years	Children,5-15,n=114	Single interval, relative to 20 age-matched, healthy controls	<2.1× ↑ mean GGT with CBZ or CBZ+VPA, <1.5× ↑ for VPA alone	Not reported	<1.5× ↑ mean AST with CBZ alone or CBZ+VPA: 1.5× <2 ↑ for VPA alone	<1.9× ↑ mean ALT with CBZ alone, <1.4× ↑ for CBZ +VPA, <2× ↑ for VPA alone		Cepelak 1998
DPH (44), CBZ (13), PB (3), VPA (11), MD (53)	1 month to 10 years	Adults,16-82(mean 56),n=56M, 67F	Single interval, relative to sex- and age-matched controls	2.4× ↑ mean GGT (n = 30 for patient and control groups)	No treatment-related effect	No treatment-related effect	1.2× ↑ mean ALT (n = 123 for patient and control groups)		Mendis 1993
DPH (32), CBZ (31), VPA (24), MD (29)	1-28 years (mean 7 years)	Children and Adults, 7-78 (mean 37), n=116	Single interval, relative to control adults	↑ mean GGT in 26% of patients on CBZ, and 78% of patients on DPH	↑ mean ALP in 16% of patients on CBZ; 25% on DPH; and 4% on VPA	No treatment-related effect	Not reported	↑ Serum F protein in 6% of patients on CBZ; 13% on DPH, and 22% on VPA	Callaghan 1994
PB, DPH and VPA (45), PB and VPA (41), VPA (40)	6-86 months (mean 32.2 months)	Children and Adults,0.5-28 (mean 12.5), n=70M, 56F	Variable sampling among patients, relative to laboratory reference values	Not reported	Not reported	<2× ↑AST for 28.3% of patients on PB, DPH and VPA combination; 19.5% on PB + VPA; and 20.0% on VPA alone	<3× ↑ ALT for 26.1% of patients on PB, DPH, and VPA combination; 7.3% on PB + VPA; and 0.0% on VPA alone		Haidukewych 1986
DPH (70), PB (60), CBZ (33), VPA (8)	> 10 years	Adults, mean 38 years, n=56M, 34F	Single interval, relative to 49 age-/sex-matched controls	Not reported	Not reported	1.1× ↑ mean AST	1.3× ↑ mean ALT	Mean 1.8× ↑ α−GST and ADH	Tutor 2004

^a PB, phenobarbital; DPH, diphenylhydantoin; CBZ, carbamazepine; VPA, valproic acid; PMD, primidone; MD, multidrug regimen;

^b years,

^c where specified.

A majority of clinical studies reporting clinical pathology alterations with CYP3A4-inducing antiepileptics have involved children. Children are considered particularly susceptible to DME induction, partly because of a greater liver-to-body weight ratio and differences with adults in metabolic and protein binding capacity, and renal and hepatic function (Ginsberg et al. 2004; Kennedy 2008; Moreland et al. 1982). Hormonal changes with puberty are thought to further influence CYP3A4 regulation (Kennedy, 2008). In an exemplary study of children receiving long-term PB and/or PHE (Aiges et al. 1980), increased activity of one or more routinely measured serum hepatobiliary marker enzymes was reported in all subjects. In particular, greater GGT activity was seen in all of the patients, whereas increases in ALP, ALT, and/or AST activity occurred in a far lower proportion. Other studies of children or infants of nursing mothers given these antiepileptics have similarly shown increases, most consistently in serum GGT, and more variably in other serum hepatobiliary marker enzymes (Bartels et al. 1975; de Wolff et al. 1982; Frey et al. 2002; Merlob et al. 1992). The incidence of the serum enzyme increases among immature subjects across these studies tended to be greater than that reported in adults on comparable regimens of these antiepileptics (Table 5). However, the predominance of GGT increases over other laboratory changes, and the magnitude of change in all of the serum enzymes evaluated (typically ≤ three-fold reference values in the absence of other findings supportive of hepatic injury) were generally similar between children and adults in these reports (Table 5).

The predominance of increases in GGT over changes in other serum enzymes with administration of potent CYP3A4 inducers has also been supported by findings in humans given rifampicin (Bartels et al. 1975; Ellis et al. 1979; Perucca et al. 1988). Further, correlation analyses between serum enzymes in a few of these clinical studies have demonstrated no association between GGT and ALP and only an occasional association between GGT and ALT (Acheampong-Mensah 1976; Deutsch et al. 1986; Haidukewych and John 1986; Sano et al. 1981; Tutor et al. 1988). However, DME induction as a direct cause of GGT increases is contradicted by the wide variability in serum GGT activity among human subjects receiving these inducing agents, and lack of correlation between patients' serum GGT activities and DME induction based on urinary excretion of D-glucaric acid or antipyrine clearance (Frezza et al. 1989; Heinemeyer et al. 1986; Hermida et al. 2002; Hildebrandt et al. 1975; Ohnhaus and Studer 1983). In addition, weak to no correlation between serum GGT activity and drug dose or plasma drug concentration has been reported in those clinical studies that comparatively evaluated these end points (Aldenhövel 1988; Braide and Davies 1987; Cepelak et al. 1986; Sano et al. 1981). Collectively, these findings suggest that these increases in serum activity of GGT, as well as other routine hepatobiliary marker enzymes, were not directly related to the DME induction potential of the drugs.

Hepatobiliary Biomarker Findings in Association with (CYP2E1-inducing) Ethanol Consumption in Humans

Human CYP2E1 is of particular clinical importance because of its ready induction by ethanol consumption and its role in the metabolism of well-known, potentially hepatotoxic xenobiotics (Meskar et al. 2001; Mohutsky et al. in press). In contrast with CYP3A4, CYP2E1 is generally comparable between man and laboratory animals in both ligand profile and induction potential (Martignoni et al. 2006). However, CYP2E1 is responsible for metabolism of only 2% to 4% of pharmaceutical agents (Kalra 2007) and shows prominent polymorphism. In humans, CYP2E1 induction has been the purported cause of altered serum hepatobiliary biomarkers, particularly GGT, in heavy alcohol consumers without detectable liver disease. Increased serum GGT has long been considered a sensitive measure of alcohol intake and is even proposed as a marker of alcohol abuse (Borg 1996; Gheno and Mazzei 1983). However, variation in serum GGT response to alcohol is prominent with age, sex, and ethnic influences that are distinct from those known to affect CYP2E1 activity (Braide and Davies 1987; Lucas et al. 1995; Whitfield 2001). In addition, several studies have shown that the magnitude of GGT activity in alcohol-consuming subjects correlated with other circulating hepatobiliary biomarkers and/or the severity of morphologic hepatic injury (Alatalo et al. 2009; Banciu et al. 1983; Conigrave et al. 2002; Daeppen et al. 1999; Salaspuro 1987; Whitfield et al. 1981). These latter findings suggest that prominent increases in GGT, especially when accompanied by other hepatobiliary marker changes, likely reflect true hepatic injury in alcoholics. Serum GGT in subjects with moderate to heavy alcohol consumption has shown particularly close correlation with AST, and serum AST has been significantly linked with hepatic pathology based on biopsy (de Lédinghen et al. 2004; Gomes et al. 2005; Kazemi-Shirazi et al. 2008; Teschke et al. 1983; Vădan et al. 2003) or computed tomography (Allaway et al. 1988). Notably, serum ALT has traditionally been considered less reliable than AST in patients with potential nutritional deficiencies such as chronic alcoholism because of the role of vitamin B6 as cofactor for ALT (Moh and Watson 1989). A comparable pattern of serum enzyme change and corresponding histologic findings has also been found in some rat chronic ethanol consumption studies, with more pronounced serum GGT increases in comparison with AST and ALT in the rodents (Devi et al. 2009).

Gamma Glutamyltransferase Increases in Association with Prototypic Hepatic-inducing Agents in Humans—The Role of Glutathione Consumption

The relatively common occurrence of increases in GGT activity in association with DME-inducing drugs and ethanol has led to further research on the causes of the serum enzyme change in man. Increased circulating GGT at the lower levels seen in these clinical studies is now generally considered to reflect increased tissue glutathione (GSH) consumption. In support of this hypothesis, alcohol-consuming patients have been shown to have significantly lower plasma GSH that is inversely correlated to circulating GGT activity (Whitfield 2001). Induction of CYP2E1 may also increase GSH tissue demand due to oxidative stress (Lu and Cedarbaum 2008). Enhanced potential for increased GSH consumption and ROS generation have also recently been reported in children given CBZ, DPH, or PB (Aycicek and Iscan 2007). Notably in human alcoholics, the increases in serum or plasma GGT activity have been consistently associated with much less of a change in hepatic tissue GGT activity (Ishii et al. 1986; Ivanov et al. 1980; Selinger et al. 1982; Teschke et al. 1983; Yamauchi et al. 1984). This result is presumed partly due to the specific hepatic tissue components analyzed, and contributions from extrahepatic tissues to blood GGT activity (Ikeda and Taniguchi 2005; Whitfield 2001). Still, the contrast between circulating and hepatic GGT activity suggests that serum values with alcohol consumption may reflect enzyme release into circulation in excess of retention in hepatic tissue (Hauge et al. 1998; Seitz et al. 1985), consistent with the extracellular GGT-mediated catabolism of GSH (Chikhi, et al. 1999; Ikeda and Taniguchi 2005). In summary, serum or tissue GGT increases with administration of DME-inducing xenobiotics in humans is now generally considered an adaptive response that is only indirectly linked with DME induction (Zhang et al. 2005).

Hormone Effects Associated with Prototypic Hepatic Enzyme-inducing Agents in Humans

The potential for DME induction to cause endocrine disruption has long been a focus of clinical studies. The CYP3A and CYP2 enzymes are particularly responsible for metabolism of multiple steroid hormones, and CYP3A4 has been specifically implicated in causing human endocrine disruption because of its role in metabolizing progesterone, estradiol, testosterone, and cortisol (Anderson 2004; Sarlis and Gourgiotis 2005). Phase II enzymes additionally conjugate several nonsteroidal and some steroidal hormones; the UGT1A family is particularly responsible for conjugation of thyroid and estrogenic hormones (Kato et al. 2008; Mackenzie et al. 2000; Takahashi et al. 2008), and the UGT2B family has a significant role in glucuronidation of human androgens (Barbier and Bélanger 2003). Drug-metabolizing enzyme induction has also been suspected of altering other endogenous human products, including circulating insulin, glucose, lipids, bilirubin, and vitamins D, K, B12, and folate (Benedetti et al. 2005; Lahtela 1986; Moreau et al. 2008; Podvinec et al. 2004; Putignano et al. 1998; Roth et al. 2008; Xie et al. 2003; You 2004). In this section, only clinical pathology linking DME induction to human thyroid and reproductive hormone levels will be discussed owing to limited comparable information on these other effects with laboratory animals.

Endogenous Thyroid Hormone Effects Associated with Prototypic Hepatic Enzyme-inducing Agents in Humans

Effects of DME-inducing agents on circulating thyroid hormone levels have been reported in a few human studies. In particular, the UGT1A enzymes facilitate T₄ excretion and are induced by a wide variety of agents that activate human PXR, CAR, and/or AhR (Xie et al. 2003; Kato et al. 2008). Accordingly, decreased serum T₄ and free T₄ (fT₄) in volunteers given rifampicin alone or with PB or antipyrine for fourteen days were attributed to increased UGT-mediated T₄ turnover (Ohnhaus and Studer 1983). Similarly, decreased fT₄ and increased thyroid gland volume were reported in volunteers after one month of dosing with rifampicin (Christensen et al. 1989). Comparable effects on serum thyroid hormone profiles have also been described in patients given DPH, PB, CBZ, and/or oxcarbazepine (Chin and Schussler 1968; Connel et al. 1984; Finucane and Griffiths 1976; Hansen et al. 1974; Hirfanoglu et al. 2007; Liewendahl et al. 1985; Tanaka et al. 1987; Vainionpää et al. 2004). Notably, T₃ was not comparably affected by UGT-inducing agents in these studies, a finding which has been attributed to primary metabolism of T₃ through sulfation and deiodination rather than glucuronidation in humans, in contrast with rats (Benedetti et al. 2005; Findlay et al. 2000; Visser et al. 1993). More significantly, no notable increase in TSH was reported in most of these clinical studies (Apeland et al. 2006; Benedetti et al. 2005; Isojärvi et al. 2001; Verrotti et al. 2009). Because of this lack of TSH change or clinical thyroid illness, the collective human literature suggests that little to no effect on thyroid function occurs in otherwise healthy subjects given DME-inducing agents. However, in persons with underlying thyroid conditions, increased incidence of hypothyroidism has been infrequently reported in association with phase II enzyme-inducing therapies (Takasu et al. 2006). As noted previously, prominent differences in thyroid hormone biology and induction of specific UGTs potentially accounts for much of the difference between rodent and human susceptibility to thyroid hormone disruption associated with DME-inducing agents (Kato et al. 2005; Kester et al. 2003; Wu and Farelly 2006).

Endogenous Reproductive Hormone Effects Associated with Prototypic Hepatic Enzyme-inducing Agents in Humans

In contrast with preclinical drug safety studies, in published clinical studies, reproductive hormone effects have received greater attention than thyroid hormone changes in association with DME induction. Induction of phase I oxidative hydroxylation and/or phase II conjugation reactions may facilitate steroid hormone inactivation or excretion (You 2004). In addition to the well-studied example of rifampin on estradiol-containing contraceptive metabolism in women (Isojärvi 2008; Mohutsky et al. 2009; Thorneycroft et al. 2006), increased endogenous estradiol metabolism has been a proposed clinical outcome of CYP3A4 induction, in general (Yu et al. 2005). Clinical antiepileptic regimens have been most often linked with reproductive hormone disruption through co-induction of aromatase (CYP19), responsible for conversion of testosterone to estradiol (Isojärvi, 2008; Herzog et al. 1991). Reproductive hormone alterations have also been attributed to DME induction via AhR- and PPAR-γ–activating agents (Nakanishi 2008). Animal models are variable in their relevance for study of these reproductive effects (Taubøll et al. 2008). Rationale for the use of nonhuman primates, specifically cynomolgus monkeys, as models for the effects of DME induction on steroid reproductive hormone phase II metabolism has been presented (Barbier and Bélanger 2003; Nishimura et al. 2008).

Conclusions on DME Induction and Clinical Pathology Biomarkers in Humans

As in preclinical species, toxicity associated with DME induction in man has been based largely on the effects of a few prototypic inducing agents. In general, administration of CYP3A4- or CYP2E1-inducing agents has been associated with increased serum or plasma activity of GGT most commonly, and ALP, AST, and ALT less often. However, as noted in animal studies, these findings have been inconsistent between studies and generally show no or only poor association with direct measures of DME induction. Rather, downstream effects of ROS formation, as well as idiosyncratic or pharmacologically mediated hepatotoxicity, more likely influence these blood chemistry changes. Increases in serum GGT may particularly represent an adaptive response to xenobiotic-related oxidative stress. Drug-metabolizing enzyme–inducing agents have also been reported to influence human endocrine patterns. Although the overall mechanisms for DME induction are similar between humans and laboratory animals, the CYP induction profile and response to specific inducers in preclinical species are limited in predictability of the response in humans.

Overall Summary

For both laboratory animals and man, collective preclinical and clinical study data generally do not support a consistent effect of DME on traditional clinical laboratory parameters. However, this review paper does support the following findings: in the rat, in the absence of findings other than hepatocellular hypertrophy, DME induction alone is not expected to be associated with increases in serum or plasma activities of ALT, ALP, or GGT. In the dog and the monkey, there is also no consistent relationship between administration of DME-inducing agents and changes in these clinical pathology parameters in the preclinical literature. However, in dogs given phenobarbital or corticosteroids, increased liver ALP or GGT activities suggest that direct or indirect induction of select hepatobiliary injury markers can occur both in the absence of liver injury and independent of DME induction. Correlations between tissue and serum activities of ALT, ALP, or GGT are limited and inconsistent, and increases in serum/plasma activities that are substantial or involve changes in other hepatobiliary markers generally reflect hepatobiliary insult rather than DME induction. Most notably, hepatic DME induction and associated changes in preclinical species are not necessarily predictive of the occurrence, magnitude, or enzyme induction profile in humans.

Footnotes

The views expressed in this article are the personal views of the authors and do not necessarily represent the policies, positions, or opinions of their respective organizations.

References

Acheampong-Mensah

-1976-. Activity of gamma-glutamyl transpeptidase in serum of patients receiving anticonvulsant or anticoagulant therapy. Clin Biochem 9, 67–70.

Aiges

H. W.

Daum

Olson

Kahn

Teichberg

-1980-. The effects of phenobarbital and diphenylhydantoin on liver function and morphology. J Pediatr 97, 22–26.

Alatalo

P. I.

Koivisto

H. M.

Hietala

J. P.

Puukka

K. S.

Bloigu

Niemelä

O. J.

-2008-. Effect of moderate alcohol consumption on liver enzymes increases with increasing body mass index. Am J Clin Nutr 88, 1097–103.

Alatalo

Koivisto

Puukka

Hietala

Anttila

Bloigu

Niemelä

-2009-. Biomarkers of liver status in heavy drinkers, moderate drinkers and abstainers. Alcohol Alcohol 44, 199–203.

Aldenhövel

H. G.

-1988-. The influence of long-term anticonvulsant therapy with diphenylhydantoin and carbamazepine on serum gamma-glutamyltransferase, aspartate aminotransferase, alanine aminotransferase and alkaline phosphatase. Eur Arch Psychiatry Neurol Sci 237, 312–16.

Allaway

S. L.

Ritchie

C. D.

Robinson

Seear

Reznek

Fry

I. K.

Thompson

G. R.

-1988-. Detection of alcohol-induced fatty liver by computerized tomography. J R Soc Med 81, 149–51.

Amacher

D. E.

Schomaker

S. J.

Burkhardt

J. E.

-1998-. The relationship among microsomal enzyme induction, liver weight and histological change in rat toxicology studies. Food Chem Toxicol 36, 831–39.

Amacher

D. E.

Schomaker

S. J.

Boldt

S. E.

Mirsky

-2006-. The relationship among microsomal enzyme induction, liver weight, and histological change in cynomolgus monkey toxicology studies. Food Chem Toxicol 44, 528–37.

Amacher

D. E.

Schomaker

S. J.

Burkhardt

J. E

-2001-. The relationship among microsomal enzyme induction, liver weight and histological change in beagle dog toxicology studies. Food Chem Toxicol 39, 817–25.

10.

Anderson

G. D.

-2008-. Gender differences in pharmacological response. Int Rev Neurobiol 83, 1–10.

11.

Anderson

G. D.

-2004-. Pharmacogenetics and enzyme induction/inhibition properties of antiepileptic drugs. Neurology 63, S3–8.

12.

Apeland

Kristensen

Strandjord

R. E.

Mansoor

M. A.

-2006-. Thyroid function during B-vitamin supplementation of patients on antiepileptic drugs. Clin Biochem 39, 282–86.

13.

Aycicek

Iscan

-2007-. The effects of carbamazepine, valproic acid and phenobarbital on the oxidative and antioxidative balance in epileptic children. Eur Neurol 57, 65–69.

14.

Bain

P. J.

-2003-. Liver. In Duncan and Prasse’s Veterinary Laboratory Medicine: Clinical Pathology - Latimer

K. S.

Mahaffey

E. A.

Prasse

K. W.

, eds.-, pp. 193–214, Iowa State University Press, Ames, IA.

15.

Balazs

Farber

T. M.

Feuer

-1978-. Drug-induced changes in serum alkaline phosphatase and alanine aminotransferase activities not related to hepatic injuries. Arch Toxicol Suppl 1, 159–63.

16.

Banciu

Weidenfeld

Marcoane

Berinde

-1983-. Serum gamma-glutamyltranspeptidase assay in the detection of alcohol consumers and in the early and stadial diagnosis of alcoholic liver disease. Med Interne 21, 23–29.

17.

Barbier

Bélanger

-2003-. The cynomolgus monkey -Macaca fascicularis- is the best animal model for the study of steroid glucuronidation. J Steroid Biochem Mol Biol 85, 235–45.

18.

Bartels

Hauck

Vogel

-1975-. Aminopyrine - an effective modifier of liver and serum gamma glutamyl transpeptidase. J Pediatr 86, 298–301.

19.

Barter

R. A.

Klaassen

C. D.

-1994-. Reduction of thyroid hormone levels and alteration of thyroid function by four representative UDP-glucuronosyltransferase inducers in rats. Toxicol Appl Pharmacol. 128, 9–17.

20.

Benedetti

M. S.

Whomsley

Baltes

Tonner

-2005-. Alteration of thyroid hormone homeostasis by antiepileptic drugs in humans: involvement of glucuronosyltransferase induction. Eur J Clin Pharmacol 61, 863–72.

21.

Boll

Weber

L. W.

Font

Stampfl

-1998-. The enzyme inducers 3-methylcholanthrene and phenobarbital affect the activities of glucocorticoid hormone-regulated enzymes in rat liver and kidney. Toxicology 126, 127–36.

22.

Boone

L. I.

Meyer

D. J.

Cusick

Ennulat

Provencher Bollinger

Everds

N. E.

Meador

Elliott

Honor

Bounous

Jordan

-2005-. Selection and interpretation of clinical pathology indicators of hepatic injury in preclinical studies. Vet Clin Pathol 34, 182–88.

23.

Borg

-1996-. Treatment of alcohol dependence: experiences of using biological markers in monitoring and prevention of relapse. Alcohol Alcohol 31, 621–24.

24.

Botts

-2010-. Introduction to Hepatic Drug Metabolizing Enzyme Induction in Drug Safety Evaluation Studies. Tox Path 38, 794–96..

25.

Boyd

J. W.

-1983-. The mechanisms relating to increases in plasma enzymes and isoenzymes in diseases of animals. Vet Clin Pathol 12, 9–24.

26.

Braide

S. A.

Davies

T. J.

-1987-. Factors that affect the induction of gamma glutamyltransferase in epileptic patients receiving anti-convulsant drugs. Ann Clin Biochem 24, 391–99.

27.

Bunch

S. E.

Castleman

W. L.

Baldwin

B. H.

Hornbuckle

W. E.

Tennant

B. C.

-1985-. Effects of long-term primidone and phenytoin administration on canine hepatic function and morphology. Am J Vet Res 46, 105–15.

28.

Callaghan

Majeed

O’Connell

Oliveira

D. B. G.

-1994-. A comparative study of serum F protein and other liver function tests as an index of hepatocellular damage in epileptic patients. Acta Neurol Scand, 89, 237–41.

29.

Capen

C. C.

-1997-. Mechanistic data and risk assessment of selected toxic end points of the thyroid gland. Toxicol Pathol 25, 39–48.

30.

Center

S. A.

-2007-. Interpretation of liver enzymes. Vet Clin Small Anim 37, 297–333.

31.

Cepelak

Lipovac

Juretić

Birek

-1986-. Effects of neuroleptic phenothiazines on the activities of aminotransferases and gamma-glutamyltransferase in serum and liver. J Clin Chem Clin Biochem 24, 583–87.

32.

Chikhi

Holic

Guellaen

Laperche

-1999-. Gamma-glutamyl transpeptidase gene organization and expression: a comparative analysis in rat, mouse, pig and human species. Comp Biochem Physiol B Biochem Mol Biol 122, 367–80.

33.

Chin

Schussler

G. C.

-1968- Decreased serum free thyroxine concentration in patients treated with diphenylhydantoin. J Clin Endocrinol. 28, 181–86.

34.

Christensen

H. R.

Simonsen

Hegedüs

Hansen

B. M.

Døssing

Kampmann

J. P.

Hansen

J. M.

-1989-. Influence of rifampicin on thyroid gland volume, thyroid hormones, and antipyrine metabolism. Acta Endocrinol 121, 406–10.

35.

Conigrave

K. M.

Degenhardt

L. J.

Whitfield

J. B.

Saunders

J. B.

Helander

Tabakoff

; WHO/ISBRA Study Group -2002-. CDT, GGT, and AST as markers of alcohol use: the WHO/ISBRA collaborative project. Alcohol Clin Exp Res 26, 332–39.

36.

Connell

J. M.

Rapeport

W. G.

Brodie

M. J.

-1984-. Changes in circulating thyroid hormones during short-term hepatic enzyme induction with carbamazepine. Eur J Clin Pharmacol. 26, 453–456.

37.

Cooper

B. W.

Cho

T. M.

Thompson

P. M.

Wallace

A. D.

-2008-. Phthalate induction of CYP3A4 is dependent on glucocorticoid regulation of PXR expression. Toxicol Sci 10, 268–77.

38.

Daeppen

J. B.

Schoenfeld-Smith

Smith

T. L.

Schuckit

M. A.

-1999-. Characteristics of alcohol dependent subjects with very elevated levels of gamma-glutamyltransferase -GGT-. J Stud Alcohol 60, 589–94.

39.

de Lédinghen

Combes

Trouette

Winnock

Amouretti

de Mascarel

Couzigou

-2004-. Should a liver biopsy be done in patients with subclinical chronically elevated transaminases? Eur J Gastroenterol Hepatol 16, 879–83.

40.

DeSandro

Catinot

Kriszt

Cordier

Richert

-1992-. Male rat hepatic UDP-glucuronosyltransferase activity toward thyroxine. Activation and induction properties—Relation with thyroxine plasma disappearance rate. Biochem Pharmacol 43, 1563–69.

41.

Deutsch

Fritsch

Gölles

Semmelrock

H. J.

-1986-. Effects of anticonvulsive drugs on the activity of gammaglutamyltransferase and aminotransferases in serum. J Pediatr Gastroenterol Nutr 5, 542–48.

42.

Devi

S. L.

Viswanathan

Anuradha

C. V.

-2009-. Taurine enhances the metabolism and detoxification of ethanol hepatic fibrosis in rats treated with iron and alcohol. Environ Toxicol Pharm 27, 120–26.

43.

de Wolff

F. A.

Peters

A. C.

van Kempen

G. M.

-1982-. Serum concentrations and enzyme induction in epileptic children treated with phenytoin and valproate. Neuropediatrics 13, 10–13.

44.

Dickins

-2004-. Induction of cytochrome P450. Curr Top Med Chem 4, 1745–66.

45.

Dresser

G. K.

Spence

J. D.

Bailey

D. G.

-2000-. Pharmacokinetic-pharmacodynamic consequences and clinical relevance of cytochrome P450 3A4 inhibition. Clin Pharmacokinet 38, 41–57.

46.

Ellis

Worthy

Goldberg

D. M.

-1979-. Lack of value of serum gamma-glutamyl transferase in the diagnosis of hepatobiliary disease. Clin Biochem 12, 142–45.

47.

Ennulat

Magid-Slav

Rehm

Tatsuoka

-2010-. Diagnostic performance of traditional hepatobiliary markers of drug-induced liver injury in the rat. Toxicol Sci; doi: 10.1093/toxsci/kfq144.

48.

Evans

W. E.

Relling

M. V.

-1999-. Pharmacogenomics: translating functional genomics into rational therapeutics. Science 286, 487–91.

49.

Faucette

S. R.

Sueyoshi

Smith

C. M.

Negishi

Lecluyse

E. L.

Wang

-2006-. Differential regulation of hepatic CYP2B6 and CYP3A4 genes by constitutive androstane receptor but not pregnane X receptor. J Pharmacol Exp Ther 317, 1200–9.

50.

Findlay

K. A.

Kaptein

Visser

T. J.

Burchell

-2000-. Characterization of the uridine diphosphate-glucuronosyltransferase-catalyzing thyroid hormone glucuronidation in man. J Clin Endocrinol Metab 85, 2879–83.

51.

Finucane

J. F.

Griffiths

R. S.

-1976-. Effect of phenytoin therapy on thyroid function. Br J Clin Pharmacol 3, 1041–44.

52.

Fittschen

Bellamy

J. E.

-1984-. Prednisone-induced morphologic and chemical changes in the liver of dogs. Vet Pathol 21, 399–406.

53.

Frey

Braegger

C. P.

Ghelfi

-2002-. Neonatal cholestatic hepatitis from carbamazepine exposure during pregnancy and breast feeding. Ann Pharmacother 36, 644–47.

54.

Frezza

Pozzato

Chiesa

Terpin

Barbone

Di Padova

-1989-. Abnormal serum gamma-glutamyltranspeptidase in alcoholics. Clues to its explanation. Neth J Med 34, 22–28.

55.

Gaskill

C. L.

Burton

S. A.

Gelens

H. C.

Ihle

S. L.

Miller

J. B.

Shaw

D. H.

Brimacombe

M. B.

Cribb

A. E.

-1999-. Effects of phenobarbital treatment on serum thyroxine and thyroid-stimulating hormone concentrations in epileptic dogs. J Am Vet Med Assoc 215, 489–96.

56.

Gaskill

C. L.

Miller

L. M.

Mattoon

J. S.

Hoffmann

W. E.

Burton

S.A.

Gelens

H. C.

Ihle

S. L.

Miller

J. B.

Shaw

D. H.

Cribb

A. E.

-2005-. Liver histopathology and liver and serum alanine aminotransferase and alkaline phosphatase activities in epileptic dogs receiving phenobarbital. Vet Pathol 42, 147–60.

57.

Gheno

Mazzei

-1983-. Biochemical indicators in chronic alcoholism. Comparison between the enzymes glutamate dehydrogenase and gamma glutamyl transferase. Minerva Med. 74, 1441–47.

58.

Giffen

P. S.

Pick

C. R.

Price

M. A.

Williams

York

M. J.

-2002-. Alpha-glutathione S-transferase in the assessment of hepatotoxicity—its diagnostic utility in comparison with other recognized markers in the Wistar Han rat. Toxicol Pathol 30, 365–72.

59.

Ginsberg

Slikker

Jr Bruckner

Sonawane

-2004-. Incorporating children’s toxicokinetics into a risk framework. Environ Health Perspect 112, 272–83.

60.

Glinghammar

Rafter

Lindström

A. K.

Hedberg

J. J.

Andersson

H. B.

Lindblom

Berg

A. L.

Cotgreave

-2009-. Detection of the mitochondrial and catalytically active alanine aminotransferase in human tissues and plasma. Int J Mol Med 23, 621–31.

61.

Goldberg

D. M.

Roomi

M. W.

Roncari

D. A.

-1981-. Triacylglycerol metabolism in the phenobarbital-treated rat. Biochem J 196, 337–46.

62.

Goldstein

Yang

Reagan

W. J

Nettleton

Rajamohan

Lawton

Nelms

Park

Gong

-2007-. Characterization or rat alanine aminotransferase isoenzymes ALT1 and ALT2. Toxicologist 96, 227.

63.

Gomes de Sá Ribeiro

Mde. F.

da Costa Gayotto

L. C.

de Alencar Fischer Chamone

Strauss

-2005-. Alcoholic intake predisposes to more interface hepatitis in chronic hepatitis C. Ann Hepatol 4, 176–83.

64.

Graham

M. J.

Lake

B. G.

-2008-. Induction of drug metabolism: species differences and toxicological relevance. Toxicology 254, 184–91.

65.

Greaves

-2007-. Liver and pancreas. In Histopathology of Preclinical Toxicity Studies, 3rd ed, pp. 457–504., Elsevier, London, UK.

66.

Haidukewych

John

-1986-. Chronic valproic acid and coantiepileptic drug therapy and incidence of increases in serum liver enzymes. Ther Drug Monit 8, 407–10.

67.

Haining

J. L.

-1970-. Kinetics of induction of rat liver enzymes by glucocorticoids. Mol Pharmacol 6, 444–47.

68.

Handschin

Meyer

U. A.

-2003-. Induction of drug metabolism: the role of nuclear receptors. Pharmacol Rev 55, 649–73.

69.

Hansen

J. M.

Skovsted

Lauridsen

U. B.

Kirkegaard

Siersbaek-Nielsen

-1974-. The effect of diphenylhydantoin on thyroid function. J Clin Endocrinol Metab 39, 785–89.

70.

Hauge

Nilsson

Persson

Hultberg

-1998-. Gamma-glutamyl transferase, intestinal alkaline phosphatase and beta-hexosaminidase activity in duodenal biopsies from chronic alcoholics. Hepatogastroenterology 45, 985–89.

71.

Heinemeyer

Roots

Lestau

Klaiber

H. R.

Dennhardt

-1986-. D-glucaric acid excretion in critical care patients—comparison with 6 beta-hydroxycortisol excretion and serum gamma-glutamyltranspeptidase activity and relation to multiple drug therapy. Br J Clin Pharmacol 21, 9–18.

72.

Hermida

Fernández

M. P.

Tutor

J. C.

-2002-. Relationship between changes in drug score, D-glucaric acid excretion, and gamma-glutamyltransferase and beta-glucuronidase serum activities during anticonvulsant treatment. Clin Lab 48, 415–19.

73.

Herzog

A. G.

Levesque

L. A.

Drislane

F. W.

Ronthal

Schomer

D. L.

-1991-. Phenytoin-induced elevation of serum estradiol and reproductive dysfunction in men with epilepsy. Epilepsia 32, 550–53.

74.

Hewitt

N. J.

Lecluyse

E. L.

Ferguson

S. S.

-2007-. Induction of hepatic cytochrome P450 enzymes: methods, mechanisms, recommendations, and in vitro-in vivo correlations. Xenobiotica 10–11, 1196–224.

75.

Hildebrandt

A. G.

Roots

Speck

Saalfrank

Kewitz

-1975-. Evaluation of in vivo parameters of drug metabolizing enzyme activity in man after administration of clemastine, phenobarbital or placebo. Eur J Clin Pharmacol 8, 327–36.

76.

Hirfanoglu

Serdaroglu

Camurdan

Cansu

Bideci

Cinaz

Gucuyener

-2007-. Thyroid function and volume in epileptic children using carbamazepine, oxcarbazepine and valproate. Pediatr Int 49, 822–26.

77.

Hoffmann

W. E.

Wilson

B. W.

Solter

P. F.

-1999-. Clinical enzymology. In The Clinical Chemistry of Laboratory Animals - Loeb

W. F.

Quimby

F. W.

, eds.-, pp. 399–454, Taylor and Francis, Philadelphia, PA.

78.

Hood

Liu

Klaassen

C. D.

-1999-. Effects of phenobarbital, pregnenolone-16alpha-carbonitrile, and propylthiouracil on thyroid follicular cell proliferation. Toxicol Sci 50, 45–53.

79.

Hood

Hashmi

Klaassen

C. D.

-1999-. Effects of microsomal enzyme inducers on thyroid-follicular cell proliferation, hyperplasia, and hypertrophy. Toxicol Appl Pharmacol 160, 163–70.

80.

Hood

Allen

M. L.

Liu

Klaassen

C. D.

-2003-. Induction of T-4- UDP-GT activity, serum thyroid stimulating hormone, and thyroid follicular cell proliferation in mice treated with microsomal enzyme inducers. Toxicol Appl Pharmacol 188, 6–13.

81.

Huseby

N. E.

Kindberg

G. M.

Grøstad

Berg

-1992-. Clearance of purified human liver gamma-glutamyltransferase after intravenous injection in the rat. Clin Chim Acta 205, 197–203.

82.

Ikeda

Taniguchi

-2005-. Gene expression of gamma-glutamyltranspeptidase. Methods Enzymol 401, 408–25.

83.

Ishii

Ebihara

Okuno

Munakata

Takagi

Arai

Shigeta

Tsuchiya

-1986-. Gamma-Glutamyl transpeptidase activity in liver of alcoholics and its localization. Alcohol Clin Exp Res 10, 81–5.

84.

Isojärvi

-2008-. Disorders of reproduction in patients with epilepsy: antiepileptic drug related mechanisms. Seizure 17, 111–19.

85.

Isojärvi

J. I.

Turkka

Pakarinen

A. J.

Kotila

Rättyä

Myllylä

V. V.

-2001-. Thyroid function in men taking carbamazepine, oxcarbazepine, or valproate for epilepsy. Epilepsia 42, 930–34.

86.

Ivanov

Adjarov

Etarska

Stankushev

Brumbarov

Kerimova

-1980-. Elevated liver gamma-glutamyl transferase in chronic alcoholics: Enzyme 25, 304–8.

87.

Jackson

E. R.

Kilroy

Joslin

D. L.

Schomaker

S. J.

Pruimboom-Brees

Amacher

D. E.

-2008-. The early effects of short-term dexamethasone administration on hepatic and serum alanine aminotransferase in the rat. Drug Chem Toxicol 31, 427–45.

88.

Jadhao

S. B.

Yang

R. Z.

Lin

Anania

F. A.

Shuldiner

A. R.

Gong

D. W.

-2004-. Murine alanine aminotransferase: cDNA cloning, functional expression, and differential gene regulation in mouse fatty liver. Hepatology 39, 1297–302.

89.

Jayyosi

Muc

Erick

Thomas

P. E.

Kelley

-1996-. Catalytic and immunochemical characterization of cytochrome P450 isozyme induction in dog liver. Fundam Appl Toxicol 31, 95–102.

90.

Jin

J. Y.

DuBois

D. C.

Almon

R. R.

Jusko

W. J.

-2004-. Receptor/gene-mediated pharmacodynamic effects of methylprednisolone on phosphoenolpyruvate carboxykinase regulation in rat liver. J Pharmacol Exp Ther 309, 328–39.

91.

Jones

S. A.

Moore

L. B.

Shenk

J. L.

Wisely

G. B.

Hamilton

G. A.

McKee

D. D.

Tomkinson

N. C.

LeCluyse

E. L.

Lambert

M. H.

Willson

T. M.

Kliewer

S. A.

Moore

J. T.

-2000-. The pregnane X receptor: a promiscuous xenobiotic receptor that has diverged during evolution. Mol Endocrinol 14, 27–39.

92.

Jover

Bort

Gómez-Lechón

M. J.

Castell

J. V.

-2002-. Down-regulation of human CYP3A4 by the inflammatory signal interleukin-6: molecular mechanism and transcription factors involved. FASEB J 16, 1799–801.

93.

Kalra

B. S.

-2007-. Cytochrome P450 enzymes and their therapeutic implications: an update. Indian J Med Sci 61, 102–16.

94.

Kato

Ikushiro

Emi

Tamaki

Suzuki

Sakaki

Yamada

Degawa

-2008-. Hepatic UDP-glucuronosyltransferases responsible for glucuronidation of thyroxine in humans. Drug Metab Dispos 36, 51–55.

95.

Kato

Suzuki

Ikushiro

Yamada

Degawa

-2005-. Decrease in serum thyroxine level by phenobarbital in rats is not necessarily dependent on increase in hepatic UDP-glucuronosyltransferase. Drug Metab Dispos 33, 1608–12.

96.

Kazemi-Shirazi

Veloso

M. P.

Frommlet

Steindl-Munda

Wrba

Zehetmayer

Marsik

Ferenci

-2008-. Differentiation of nonalcoholic from alcoholic steatohepatitis: are routine laboratory markers useful? Wien Klin Wochenschr 120, 25–30.

97.

Kennedy

-2008-. Hormonal regulation of hepatic drug-metabolizing enzyme activity during adolescence. Clin Pharmacol Ther 84, 662–73.

98.

Kester

M. H.

Kaptein

Roest

T. J.

van Dijk

C. H.

Tibboel

Meinl

Glatt

Coughtrie

M. W.

Visser

T. J.

-2003-. Characterization of rat iodothyronine sulfotransferases. Am J Physiol Endocrinol Metab 285, 592–8.

99.

Klaunig

J. E.

Isenberg

Bachowski

K. L.

Kolaja

K. L.

Jiang

Stevenson

D. E.

Walborg

E. F.

Jr. -1998-. The role of oxidative stress in chemical carcinogenesis. Environ. Health Perspect 106 Suppl 1, 289–295.

100.

Kotlyar

Carson

S. W.

-1999-. Effects of obesity on the cytochrome P450 enzyme system. Int J Clin Pharmacol Ther 37, 8–19.

101.

Kramer

J. A.

Blomme

E. A.

Bunch

R. T.

Davila

J. C.

Jackson

C. J.

Jones

P. F.

Kolaja

K. L.

Curtiss

S. W.

-2003-. Transcription profiling distinguishes dose-dependent effects in the livers of rats treated with clofibrate. Toxicol Pathol 31, 417–31.

102.

Lahrichi

Ratanasavanh

Galteau

M. M.

Siest

-1982-. Effect of chronic ethanol administration on gamma-glutamyltransferase activities in plasma and in hepatic plasma membranes of male and female rats. Enzyme 28, 251–57.

103.

Lahtela

J. T.

-1986-. Effect of long-term anticonvulsant therapy on glucose metabolism in humans. Epilepsia. 27, 711–16.

104.

Lake

B. G.

Renwick

A. B.

Cunninghame

M. E.

Price

R. J.

Surry

Evans

D. C.

-1998-. Comparison of the effects of some CYP3A and other enzyme inducers on replicative DNA synthesis and cytochrome P450 isoforms in rat liver. Toxicology 131, 9–20.

105.

Landi

M. S.

Kissinger

J. T.

Campbell

S. A.

Kenney

C. A.

Jenkins

E. L.

-1990-. The effects of four types of restraint on serum alanine aminotransferase and aspartate animotransferase in the Macaca fascicularis. J Am Coll Toxicol 9, 517–23.

106.

Lassen

E. D.

-2004-. Laboratory Evaluation of the Liver. In Veterinary hematology and clinical chemistry - Thrall

M. A.

Baker

D. C.

, eds.-, pp. 355–77., Blackwell Publishing, Ames, IA.

107.

Lecureux

Dieter

M. Z.

Nelson

D. M.

Watson

Wong

Gemzik

Klaassen

C. D.

Lehman-McKeeman

L. D.

-2009-. Hepatobiliary disposition of thyroid hormone in Mrp2-deficient TR- rats: reduced biliary excretion of thyroxine glucuronide does not prevent xenobiotic-induced hypothyroidism. Toxicol Sci 108, 482–91.

108.

Lemasters

J. J.

Gores

G. J.

Nieminen

A. L.

Dawson

T. L.

Wray

B. E.

Herman

-1990-. Multiparameter digitized video microscopy of toxic and hypoxic injury in single cells. Environ Health Perspect 84, 83–94.

109.

Leonard

T. B.

Neptun

D. A.

Popp

J. A.

-1984-. Serum gamma glutamyl transferase as a specific indicator of bile duct lesions in the rat liver. Am J Pathol 116, 262–69.

110.

Liewendahl

Tikanoja

Helenius

Majuri

-1985-. Free thyroxin and free triiodothyronine as measured by equilibrium dialysis and analog radioimmunoassay in serum of patients taking phenytoin and carbamazepine. Clin Chem 31,1993–96.

111.

Lindblom

Rafter

Copley

Andersson

Hedberg

J. J.

Berg

A. L.

Samuelsson

Hellmold

Cotgreave

Glinghammar

-2007-. Isoforms of alanine aminotransferases in human tissues and serum—differential tissue expression using novel antibodies. Arch Biochem Biophys 466, 66–77.

112.

Litchfield

M. H.

Conning

D. M.

-1972-. Effect of phenobarbitone on plasma and hepatic alkaline phosphatase activity in the dog. Naunyn Schmiedebergs Arch Pharmacol 272, 358–62.

113.

Liu

Pan

Whitington

P. F.

-2009-. Increased hepatic expression is a major determinant of serum alanine aminotransferase elevation in mice with nonalcoholic steatohepatitis. Liver Int 29, 337–43.

114.

Loeb

W. F.

-1999-. The rat. In The Clinical Chemistry of Laboratory Animals - Loeb

W. F.

, ed.-, pp. 33–48, Taylor and Francis, Philadelphia, PA.

115.

Cederbaum

A. I.

-2008-. CYP2E1 and oxidative liver injury by alcohol. Free Radic Biol Med 44, 723–38.

116.

A. P.

-2001-. Species comparison in P450 induction: effects of dexamethasone, omeprazole, and rifampin on P450 isoforms 1A and 3A in primary cultured hepatocytes from man, Sprague-Dawley rat, minipig, and beagle dog. Chem Biol Interact 134, 271–81.

117.

Lucas

Ménez

Girre

Berthou

Bodénez

Joannet

Hispard

Bardou

L. G.

Ménez

J. F.

-1995-. Cytochrome P450 2E1 genotype and chlorzoxazone metabolism in healthy and alcoholic Caucasian subjects. Pharmacogenetics 5, 298–304.

118.

Mackenzie

P. I.

Miners

J. O,.

McKinnon

R. A.

-2000-. Polymorphisms in UDP glucuronosyltransferase genes: functional consequences and clinical relevance. Clin Chem Lab Med 38, 889–92.

119.

Maronpot

-In press-. Hepatic enzyme induction: Microscopic and ultrastructural appearance. Toxicol Pathol

120.

Martignoni

Groothuis

G. M.

de Kanter

-2006-. Species differences between mouse, rat, dog, monkey and human CYP-mediated drug metabolism, inhibition and induction. Expert Opin Drug Metab Toxicol 2, 875–94.

121.

Martignoni

de Kanter

Grossi

Mahnke

Saturno

Monshouwer

-2004-. An in vivo and in vitro comparison of CYP induction in rat liver and intestine using slices and quantitative RT-PCR. Chem Biol Interact 151, 1–11.

122.

McClain

R. M.

-1995-. Mechanistic considerations for the relevance of animal data on thyroid neoplasia to human risk assessment. Mut Res 333, 131–42.

123.

McClain

R. M.

Levin

A. A.

Posch

Downing

J. C

-1989-. The effect of phenobarbital on the metabolism and excretion of thyroxine in rats. Toxicol Appl Pharmacol 99, 216–28.

124.

McClain

R. M.

Posch

R. C.

Bosakowski

Armstrong

J. M.

-1988-. Studies on the mode of action for thyroid gland tumor promotion in rats by phenobarbital. Toxicol Appl Pharmacol 94, 254–65.

125.

Meek

M. E.

Bucher

J. R.

Cohen

S. M

Dellarco

Hill

R. N.

Lehman-McKeeman

L. D.

Longfellow

D. G.

Pastoor

Seed

Patton

D. E.

-2003-. A framework for human relevance analysis of information on carcinogenic modes of action. Crit Rev Toxicol 33, 591–653.

126.

Meijer

E. P.

Coolen

S. A.

Bast

Westerterp

K. R.

-2001-. Exercise-induced oxidative stress in older adults as measured by antipyrine oxidation. Metabolism 50, 1484–88.

127.

Mendis

G. P.

Gibberd

F. B.

Hunt

H. A.

-1993-. Plasma activities of hepatic enzymes in patients on anticonvulsant therapy. Seizure 2, 319–23.

128.

Merlob

Mor

Litwin

-1992-. Transient hepatic dysfunction in an infant of an epileptic mother treated with carbamazepine during pregnancy and breastfeeding. Ann Pharmacother 26, 1563–65.

129.

Meskar

Plee-Gautier

Amet

Berthou

Lucas

-2001-. Alcohol-xenobiotic interactions. Role of cytochrome P450 2E1. Pathol Biol 49, 696–702.

130.

Meyer

D. J.

Harvey

J. W.

-2004-. Hepatobiliary and skeletal muscle enzymes and liver function tests. In Veterinary Laboratory Medicine. Interpretation and Diagnosis, pp. 174–75., Saunders, Philadelphia, PA.

131.

Misslbeck

N. G.

Campbell

T. C.

Roe

D. A.

-1986-. Increase in hepatic gamma-glutamyltransferase -GGT- activity following chronic ethanol intake in combination with a high fat diet. Biochem Pharmacol 35, 399–404.

132.

Moh

M. E.

Watson

R. R.

-1989-. Changes in nutritional status associated with alcohol abuse. In Diagnosis of Alcohol Abuse - Watson

R. R.

, ed-, pp. 125–46, CRC Press, Boca Raton, FL.

133.

Mohutsky

M. A.

-In press-. Hepatic drug-metabolizing enzyme induction and implications for preclinical and clinical risk assessment. Toxicol Pathol

134.

Moreau

Vilarem

M. J.

Maurel

Pascussi

J. M.

-2008-. Xenoreceptors CAR and PXR activation and consequences on lipid metabolism, glucose homeostasis, and inflammatory response. Mol Pharmaceutics 5, 35–41.

135.

Moreland

T. A.

Park

B. K.

Rylance

G. W.

-1982-. Microsomal enzyme induction in children: the influence of carbamazepine treatment on antipyrine kinetics, 6 beta-hydroxycortisol excretion and plasma gamma glutamyltranspeptidase activity. Br J Clin Pharmacol 14, 861–65.

136.

Nakanishi

-2008-. Endocrine disruption induced by organotin compounds; organotins function as a powerful agonist for nuclear receptors rather than an aromatase inhibitor. J Toxicol Sci 33, 269–76.

137.

Nishimura

Stein

Berges

Teschke

-1981-. Gamma-glutamyltransferase activity of liver plasma membrane: Induction following chronic alcohol consumption. Biochem. Biophys Res Comm 99, 142–48.

138.

Nishimura

Teschke

-1982-. Effect if chronic alcohol consumption on the activities of liver plasma membrane enzymes: gamma-glutamyltransferase, alkaline phosphatase and 5’-nucleotidase. Biochem. Pharmacol. 31, 377–81.

139.

Nishimura

Koeda

Shimizu

Nakayama

Satoh

Narimatsu

Naito

-2008-. Comparison of inducibility of sulfotransferase and UDP-glucuronosyltransferase mRNAs by prototypical microsomal enzyme inducers in primary cultures of human and cynomolgus monkey hepatocytes. Drug Metab Pharmacokinet 23, 45–53.

140.

O’Brien

P. J.

Slaughter

M. R.

Polley

S. R.

Kramer

-2002-. Advantages of glutamate dehydrogenase as a blood biomarker of acute hepatic injury in rats. Lab Anim 36, 313–21.

141.

O’Connor

J. C.

Frame

S. R.

Davis

L. G.

Cook

J. C.

-1999-. Detection of thyroid toxicants in a tier I screening battery and alterations in thyroid endpoints over 28 days of exposure. Toxicol Sci 51, 54–70.

142.

S. I.

Kim

C. I.

Chun

H. J.

Park

S. C.

-1998-. Chronic ethanol consumption affects glutathione status in rat liver. J Nutr 128, 758–63.

143.

Ohnhaus

E. E.

Studer

-1983-. A link between liver microsomal enzyme activity and thyroid hormone metabolism in man. Br J Clin Pharmacol 15, 71–76.

144.

Perucca

Grimaldi

Frigo

G. M.

Sardi

Mönig

Ohnhaus

E. E.

-1988-. Comparative effects of rifabutin and rifampicin on hepatic microsomal enzyme activity in normal subjects. Eur J Clin Pharmacol 34, 595–99.

145.

Perucca

Hedges

Makkilt

K. A.

Ruprah

Wilson

J. F.

Richens

-1984-. A comparative study of the relative enzyme inducing properties of anticonvulsant drugs in epileptic patients. Br J Clin Pharmacol 18, 401–10.

146.

Peterson

R. L.

Casciotti

Block

Goad

M. E.

Tong

Meehan

J. T.

Jordan

R. A.

Vinlove

M. P.

Markiewicz

V. R.

Weed

C. A.

Dorner

A. J.

-2004-. Mechanistic toxicogenomic analysis of WAY-144122 administration in Sprague-Dawley rats. Toxicol Appl Pharmacol 196, 80–94.

147.

Podvinec

Handschin

Looser

Meyer

U. A.

-2004-. Identification of the xenosensors regulating human 5-aminolevulinate synthase. Proc Natl Acad Sci U S A 101, 9127–32.

148.

Putignano

Kaltsas

G. A.

Satta

M. A.

Grossman

A. B.

-1998-. The effects of anti-convulsant drugs on adrenal function. Horm Metab Res 30, 389–97.

149.

Qatanani

Zhang

Moore

D. D.

-2005-. Role of the constitutive androstane receptor in xenobiotic-induced thyroid hormone metabolism. Endocrinology 146, 995–1002.

150.

Rajamohan

Nelms

Joslin

D. L.

Reagan

W. J.

Lawton

-2006-. cDNA cloning, expression, purification, distribution, and characterization of biologically active canine alanine aminotransferase-1. Protein Expr Purif 48, 81–89.

151.

Rambabu

Matsuda

Katunuma

-1986-. Studies on turnover rates of rat γ-glutamyltranspeptidase after chronic ethanol administration in vivo. Biochem Med Metab Biol 35, 335–44.

152.

Ratanasavanh

Tazi

Galteau

M. M.

Siest

-1979-. Localization of gamma-glutamyltransferase in subcellular fractions of rat and rabbit liver: effect of phenobarbital. Biochem Pharmacol 28, 1363–65.

153.

Roberts

B. J.

Shoaf

S. E.

Song

B. J.

-1995-. Rapid changes in cytochrome P4502E1 -CYP2E1- activity and other P450 isozymes following ethanol withdrawal in rats. Biochem Pharmacol 49, 1665–73.

154.

Roomi

M. W.

Goldberg

D. M.

-1981-. Comparison of γ-glutamyl transferase induction by phenobarbital in the rat, guinea pig, and rabbit. Biochem Pharmacol 30, 1563–71.

155.

Rosen

Roberts

N. R.

Budnick

L. E.

Nichol

C. A.

-1959-. Corticosteroids and transaminase activity: The specificity of the glutamic-pyruvic transaminase response. Endocrinology 65, 256–64.

156.

Roth

Looser

Kaufmann

Meyer

U. A.

-2008-. Sterol regulatory element binding protein 1 interacts with pregnane X receptor and constitutive androstane receptor and represses their target genes. Pharmacogenet Genomics 18, 325–37.

157.

Rutgers

H. C.

Batt

R. M.

Vaillant

Riley

J. E.

-1995-. Subcellular pathologic features of glucocorticoid-induced hepatopathy in dogs. Am J Vet Res 56, 898–907.

158.

Sabzevari

Hatcher

Kentish

O’Sullivan

Gibson

G. G.

-1996-. Bifonazole, but not the structurally-related clotrimazole, induces both peroxisome proliferation and members of the cytochrome P4504A sub-family in rat liver. Toxicology 106, 19–26.

159.

Saito

Kaneko

Sato

Yoshitake

Yamada

-1991-. Hepatic UDP-glucuronosyltransferase-s- activity toward thyroid hormones in rats: Induction and effects on serum thyroid levels following treatment with various enzyme inducers. Toxicol Appl Pharmacol 111, 99–106.

160.

Salaspuro

-1987-. Use of enzymes for the diagnosis of alcohol-related organ damage. Enzyme 37, 87–107.

161.

Sano

Kawada

Yamaguchi

Kawakita

Kobayashi

-1981-. Effects of phenytoin on serum gamma-glutamyl transpeptidase activity. Epilepsia 22, 331–38.

162.

Sapolsky

R. M.

Romero

L. M.

Munck

A. U.

-2000-. How do glucocorticoids influence stress responses? Integrating permissive, suppressive, stimulatory, and preparative actions. Endocr Rev 21, 55–89.

163.

Sarlis

N. J.

Gourgiotis

-2005-. Hormonal effects on drug metabolism through the CYP system: perspectives on their potential significance in the era of pharmacogenomics. Curr Drug Targets Immune Endocr Metabol Disord 5, 439–48.

164.

Satoh

Igarashi

Hirota

Kitagawa

-1982-. Induction of hepatic γ-glutamyl transpeptidase in rats by repeated administration of aminopyrine. J Pharmacol Exp Ther 221, 795–800

165.

Schulte-Hermann

-1974-. Induction of liver growth by xenobiotic compounds and other stimuli. Crit Rev Toxicol 3, 97–158.

166.

Seitz

H. K.

Velasquez

Waldherr

Veith

Czygan

Weber

Deutsch-Diescher

O. G.

Kommerell

-1985-. Duodenal gamma-glutamyltransferase activity in human biopsies: effect of chronic ethanol consumption and duodenal morphology. Eur J Clin Invest 15, 192–96.

167.

Selinger

M. J.

Matloff

D. S.

Kaplan

M. M.

-1982-. Gamma-glutamyl transpeptidase activity in liver disease: serum elevation is independent of hepatic GGTP activity. Clin Chim Acta 125, 283–90.

168.

Solter

P. F.

Hoffmann

W. E.

Chambers

M. D.

Schaeffer

D. J.

Kuhlenschmidt

M. S.

-1994-. Hepatic total 3 alpha-hydroxy bile acids concentration and enzyme activities in prednisone-treated dogs. Am J Vet Res 55, 1086–92.

169.

Solter

P. F.

-2005-. Clinical pathology approaches to hepatic injury. Toxicol Pathol 33, 9–16.

170.

Takahashi

Maruo

Mori

Iwai

Sato

Takeuchi

-2008-. Effect of D256N and Y483D on propofol glucuronidation by human uridine 5′-diphosphate glucuronosyltransferase -UGT1A9-. Basic Clin Pharmacol Toxicol 103, 131–36.

171.

Takasu

Kinjou

Kouki

Takara

Ohshiro

Komiya

-2006-. Rifampin-induced hypothyroidism. J Endocrinol Invest 29, 645–49.

172.

Tanaka

-1998-. Clinically important pharmacokinetic drug-drug interactions: role of cytochrome P450 enzymes. J Clin Pharm Ther 23, 403–16.

173.

Tanaka

Kodama

Yokoyama

Komatsu

Konishi

Momota

Matsuo

-1987-. Thyroid function in children with long-term anticonvulsant treatment. Pediatr Neurosci 13, 90–94.

174.

Taubøll

Røste

L. S.

Svalheim

Gjerstad

-2008-. Disorders of reproduction in epilepsy—what can we learn from animal studies? Seizure. 17, 120–26.

175.

Teschke

Neuefeind

Nishimura

Strohmeyer

-1983-. Hepatic gamma-glutamyltransferase activity in alcoholic fatty liver: comparison with other liver enzymes in man and rats. Gut 24, 625–30.

176.

Thorneycroft

Klein

Simon

-2006-. The impact of antiepileptic drug therapy on steroidal contraceptive efficacy. Epilepsy Behav 9, 31–39.

177.

Thulin

Rafter

Stockling

Tomkiewicz

Norjavaara

Aggerbeck

Hellmold

Ehrenborg

Andersson

Cotgreave

Glinghammar

-2008-. PPARalpha regulates the hepatotoxic biomarker alanine aminotransferase -ALT1- gene expression in human hepatocytes. Toxicol Appl Pharmacol 231, 1–9.

178.

Tutor

J. C.

Alvarez-Prechous

Bernabeu

Pardinas

M. C.

Paz

J. M.

Lareu

-1988-. Urinary D-glucaric acid and serum hepatic enzyme levels in chronic alcoholics. Clin Biochem 21, 193–98.

179.

Unakami

Komoda

Watanabe

Tanimoto

Sakagishi

Ikezawa

-1987-. Molecular nature of three liver alkaline phosphatases detected by drug administration in vivo: differences between soluble and membranous enzymes. Comp Biochem Physiol B 88, 111–18.

180.

Vădan

Gheorghe

Becheanu

Iacob

Gheorghe

-2003-. Predictive factors for the severity of liver fibrosis in patients with chronic hepatitis C and moderate alcohol consumption. Rom J Gastroenterol 12, 183–87.

181.

Vainionpää

L. K.

Mikkonen

Rättyä

Knip

Pakarinen

A. J.

Myllylä

V. V.

Isojärvi

J. I. T.

-2004- Thyroid function in girls with epilepsy with carbamazepine, oxcarbamazepine, or valproate monotherapy and after withdrawal of medication. Epilepsia 45, 197–203.

182.

Valentine

B. A.

Blue

J. T.

Shelley

S. M.

Cooper

B. J.

-1990-. Increased serum alanine aminotransferase activity associated with muscle necrosis in the dog. J Vet Intern Med 4, 140–43.

183.

Verrotti

Laus

Scardapane

Franzoni

Chiarelli

-2009-. Thyroid hormones in children with epilepsy during long-term administration of carbamazepine and valproate. Eur J Endocrinol 160, 81–86.

184.

Viollon-Abadie

Lassere

Debruyne

Nicod

Carmichael

Richert

-1999-. Phenobarbital, beta-naphthoflavone, clofibrate, and pregnenolone-16alpha-carbonitrile do not affect hepatic thyroid hormone UDP-glucuronosyl transferase activity, and thyroid gland function in mice. Toxicol Appl Pharmacol 155, 1–12.

185.

Visser

T. J.

Kaptein

van Raaij

J. A. G. M.

Joe

C. T. T.

Ebner

Burchell

-1993-. Multiple UDP-glucuonosyltransferases for the glucuronidation of thyroid hormones with preference for 3,3′5′-triiodothyronine -reverse T3-. FEBS Lett 315, 65–68.

186.

Weingand

Brown

Hall

Davies

Gossett

Neptun

Waner

Matsuzawa

Salemink

Froelke

Provost

J. P.

Dal Negro

Batchelor

Nomura

Groetsch

Boink

Kimball

Woodman

York

Fabianson-Johnson

Lupart

Melloni

-1996-. Harmonization of animal clinical pathology testing in toxicity and safety studies. The Joint Scientific Committee for International Harmonization of Clinical Pathology Testing. Fundam App Toxicol 29, 198–201.

187.

Whitfield

J. B.

Allen

J. K.

Adena

M. A.

Hensley

W. J.

-1981-. Effect of drinking on correlations between biochemical variables. Ann Clin Biochem 18, 143–45.

188.

Whitfield

J. B.

-2001-. Gamma glutamyl transferase. Crit Rev Clin Lab Sci 38, 263–355.

189.

Wiedmeyer

C. E.

Solter

P. E.

Hoffmann

W. E.

-2002a-. Alkaline phosphatase expression in tissues from glucocorticoid-treated dogs. Am J Vet Res 63, 1083–88.

190.

Wiedmeyer

C. E.

Solter

P. E.

Hoffmann

W. E.

-2002b-. Kinetics of mRNA expression of alkaline phosphatase isoenzymes in hepatic tissues from glucocorticoid-treated dogs. Am J Vet Res 63, 1089–95.

191.

Wilkinson

G. R.

-1996-. Cytochrome P4503A -CYP3A- metabolism: prediction of in vivo activity in humans. J Pharmacokinet Biopharm 24, 475–90.

192.

Williams

G. M.

Iatropoulos

M. J.

-2002- Alteration of Liver Cell Function and Proliferation: Differentiation Between Adaptation and Toxicity. Toxicol Pathol 3, 41–53.

193.

K.-M.

Farrelly

J. G.

-2006-. Preclinical development of new drugs that enhance thyroid hormone metabolism and clearance: inadequacy of using rats as an animal model for predicting human risks in an IND and NDA. Am J Ther 13, 141–144.

194.

Xie

Yeuh

M. F.

Radominska-Pandya

Saini

S. P.

Negishi

Bottroff

B. S.

Cabrera

G. Y.

Tukey

R. H.

Evans

R. M.

-2003-. Control of steroid, heme, and carcinogen metabolism by nuclear pregnane X receptor and constitutive androstane receptor. Proc Natl Acad Sci U S A 100, 4150–55.

195.

C. Y.

Kong

A. N.

-2005-. Induction of phase I, II and III drug metabolism/transport by xenobiotics. Arch Pharm Res 28, 249–68.

196.

Yamada

Wilson

J. S.

Lieber

C. S.

-1985-. The effects of ethanol and diet on hepatic and serum γ-glutamyltranspeptidase activities in rats. J Nutr 115, 1285–90.

197.

Yamauchi

Kimura

Kawase

Watanabe

Kitahara

Ogura

Fujisawa

Kameda

-1984-. Hepatic gamma-glutamyl transferase and hepatic fibrosis in patients with alcoholic liver disease. Enzyme 32, 110–15.

198.

Yang

R. Z.

Park

Reagan

W. J.

Goldstein

Zhong

Lawton

Rajamohan

Qian

Liu

Gong

D. W.

-2009-. Alanine aminotransferase isoenzymes: molecular cloning and quantitative analysis of tissue expression in rats and serum elevation in liver toxicity. Hepatology 49, 598–607.

199.

You

-2004-. Steroid hormone biotransformation and xenobiotic induction of hepatic steroid metabolizing enzymes. Chem Biol Interact 147, 233–46.

200.

A. M.

Fukamachi

Krausz

K. W.

Cheung

Gonzalez

F. J.

-2005-. Potential role for human cytochrome P450 3A4 in estradiol homeostasis. Endocrinology 146, 2911–19.

201.

Zhang

Chando

T. J.

Everett

D. W.

Patten

C. J.

Dehal

S. S.

Humphreys

W. G.

-2005-. In vitro inhibition of UDP glucuronosyltransferases by atazanavir and other HIV protease inhibitors and the relationship of this property to in vivo bilirubin glucuronidation. Drug Metab Dispos 33, 1729–39.