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Troglitazone (TGZ) (Rezulin; Pfizer, Madison, NJ) was withdrawn from the market by the U.S. Food and Drug Administration (FDA) in March 2000, because it was associated with the development of acute liver failure.1 Approximately 35,000 TGZ injury claims are pending within the United States.2 However, even 4 years later, the mechanisms of TGZ-induced liver injury remain highly controversial and poorly understood. A review of the topic proposed that TGZ hepatotoxicity was mediated through the induction of steatosis and apoptosis and that the injury could be silent (without increased serum aminotransferases, bilirubin, or alkaline phosphatase).3 In addition, it has been suggested that TGZ may induce liver injury as a consequence of mitochondrial damage, oxidative stress, or by inhibiting the bile salt excretory protein (Bsep).3 Furthermore, these effects have been attributed, at least in part, to the accumulation of TGZ in the liver, particularly among patients with liver disease, as well as to the peculiar metabolism of the drug.3 Finally, it has been postulated that liver fibrosis and cancer may result from TGZ hepatotoxicity.3 These presumed chronic complications would constitute an extraordinary medical burden to the approximately 1.92 million patients treated with TGZ. Because these alleged long-term complications of TGZ-induced liver injury have significant medical, social, legal, and financial repercussions, this review evaluates the scientific merits of the proposed mechanisms and suggest alternative explanations.

Troglitazone-Associated Liver Injury Is Idiosyncratic

  1. Top of page
  2. Troglitazone-Associated Liver Injury Is Idiosyncratic
  3. Epidemiology of Troglitazone-Associated Liver Injury
  4. Troglitazone Pharmacokinetics
  5. Oxidative Stress and Mitochondrial Damage
  6. Bile Salt Export Proteins and Cholestasis
  7. Apoptosis in Cell Cultures
  8. Liver Apoptosis and Mitochondrial Abnormalities in Patients
  9. Liver Fibrosis and Cancer
  10. Potential Genetic and Acquired Susceptibility to TGZ
  11. Conclusions
  12. Acknowledgements
  13. References

Insulin resistance can result from an inherited defect in mitochondrial oxidative phosphorylation in children of patients with type 2 diabetes4 or from acquired defects in mitochondrial biogenesis,5 which leads to reductions in mitochondrial function, in elderly individuals.6

TGZ, a peroxisomal proliferator-activated receptor (PPAR)-γ agonist that improved insulin resistance, was used for the treatment of type 2 diabetes7, 8 in the United States by approximately 1.92 million patients from March 1997 to March 2000.9 As a consequence of the improved insulin sensitivity,8 TGZ had beneficial cardiovascular effects,10–13 protected pancreatic β-cell function,14 and corrected metabolic abnormalities associated with polycystic ovarian syndrome15 and lipodystrophy.16 In addition, TGZ antagonized the corticosteroid induction of insulin resistance and abnormal glucose tolerance.17

In clinical trials, 1.9% of the subjects who received TGZ and 0.6% of the subjects who received placebo had elevations (≥3 times the upper limit of normal) of serum alanine aminotransferase (ALT) concentrations.18 In this cohort of 2,510 subjects, TGZ induced overt liver injury and jaundice in two individuals.18 A retrospective analysis also concluded that approximately 2% of the patients developed TGZ-induced liver injury judging by the serum ALT elevations.19 By comparison, serum ALT elevations (≥3 times the upper limit of normal) occurred in approximately 25% of subjects taking tacrine,20 a drug that has not been associated with acute liver failure; however, probably fewer than 100,000 patients have been treated with this drug. Unfortunately, 94 of the 1.92 million patients developed liver failure while taking TGZ.1 Although only 49 of these liver failure cases were considered to be possibly or probably related to TGZ,9 the numbers may be higher because of incompleteness of reporting.1

TGZ treatment was associated with a characteristic hepatocellular injury,18, 21–35 with rare instances of either a mixed hepatocellular/cholestatic injury or a predominant cholestatic reaction.36–38 The liver injury associated with TGZ is idiosyncratic39; it is unpredictable, neither time- nor dose-dependent,40 and cannot be reproduced in animals.41–43 Although the mechanism of liver injury induced by TGZ is believed to be a metabolic idiosyncrasy (nonimmunological),39 some of the case reports had histological evidences suggestive of an immunological reaction16, 25, 27, 36 or responded favorably to corticosteroids.28, 38 One of these patients promptly developed a similar destructive, granulomatous cholangitis when treated subsequently with rosiglitazone, a related PPAR-γ agonist,38 supporting the diagnosis of an immunological reaction and suggesting a class effect.

According to Zimmerman and Ishak44 and Klatskin and Conn,45 the occurrence of numerous eosinophils or granulomatous inflammatory infiltrates in the liver is sufficiently compelling, albeit circumstantial, evidence that hypersensitivity may play an important role in the injury provoked by TGZ in some patients.

Epidemiology of Troglitazone-Associated Liver Injury

  1. Top of page
  2. Troglitazone-Associated Liver Injury Is Idiosyncratic
  3. Epidemiology of Troglitazone-Associated Liver Injury
  4. Troglitazone Pharmacokinetics
  5. Oxidative Stress and Mitochondrial Damage
  6. Bile Salt Export Proteins and Cholestasis
  7. Apoptosis in Cell Cultures
  8. Liver Apoptosis and Mitochondrial Abnormalities in Patients
  9. Liver Fibrosis and Cancer
  10. Potential Genetic and Acquired Susceptibility to TGZ
  11. Conclusions
  12. Acknowledgements
  13. References

Although valuable to the medical and scientific community, neither the MedWatch Reports to the FDA nor the published case reports provide statistical support for the causal association between TGZ and acute liver failure. In addition, and except in rare instances (e.g., rechallenge38), presumed idiosyncratic liver injuries cannot be attributed unequivocally to TGZ or any other drug.40, 46, 47 As evidence of the diagnostic uncertainties, in the United States, the cases of acute liver failure of undetermined causes (17%) exceed those attributed as probable to all medications combined (13%), excluding acetaminophen.48 Nonetheless, in a prospective study of acute liver failure between 1998 and 2001 at 17 tertiary care centers in the United States, TGZ was prominently implicated, contributing 10% to 15% of the idiosyncratic drug reactions.48 In a small cohort of type 2 diabetic patients and based on a single case of acute liver failure, the incident rate of acute liver failure attributed to TGZ was 240/million person-years,49 compared with the estimated idiopathic acute liver failure of 1 to 2/million person-years.49, 50

Although a study suggested that type 2 diabetes may increase the risk of acute liver failure independently of underlying chronic liver disease or viral hepatitis,51 the study did not eliminate the possible contribution of decompensated chronic liver disease to liver failure. However, in a cohort of diabetic patients treated with hypoglycemic drugs, the incidence of serious liver injury was similar among those treated with sulfonylureas, insulin, metformin. or TGZ (approximately 100/million person-years).52 The estimated risk of unexplained acute liver failure (with coagulopathy and encephalopathy) in type 2 diabetes was approximately 40/million person-years,53 substantially higher than that of 1 to 2/million person-years estimated in the general population.49, 50 It is possible that the retrospective analysis could have introduced some biases,52 because rosiglitazone and pioglitazone have rarely been associated with serious liver injury.49, 50 Another confounding epidemiological variable is the development of subacute liver failure, in the absence of any known cause or TGZ treatment, in approximately 3% of patients with nonalcoholic steatohepatitis (NASH) and unrecognized cirrhosis.54

Thus, TGZ induced serious liver injury in a large number of patients, but the background incidence rate of acute liver failure among type 2 diabetic patients remains to be determined in prospective studies.

Troglitazone Pharmacokinetics

  1. Top of page
  2. Troglitazone-Associated Liver Injury Is Idiosyncratic
  3. Epidemiology of Troglitazone-Associated Liver Injury
  4. Troglitazone Pharmacokinetics
  5. Oxidative Stress and Mitochondrial Damage
  6. Bile Salt Export Proteins and Cholestasis
  7. Apoptosis in Cell Cultures
  8. Liver Apoptosis and Mitochondrial Abnormalities in Patients
  9. Liver Fibrosis and Cancer
  10. Potential Genetic and Acquired Susceptibility to TGZ
  11. Conclusions
  12. Acknowledgements
  13. References

TGZ is metabolized predominantly by sulfation to metabolite 1 (M1) and, to a lesser extent, by glucuronidation to M2 and by oxidation through cytochrome P450 (CYP)3A4 to a quinone M3.42, 55, 56 TGZ concentration in the liver is essentially the same as that in plasma because these pools are in equilibrium.57–61 In rats, the maximal concentration (CMax) of TGZ in plasma and in the liver was approximately 2.6 μmol/L, whereas the concentration of M1+M2+M3 in the liver was roughly 52 μmol/L.58 Thus, TGZ does not accumulate in the liver, as has been suggested,3 and the actual liver TGZ concentration should be used as a reference for cell culture toxicity studies rather than the combined CMax in the liver for TGZ and its metabolites (approximately 18.5 μg/g, approximately 55 μmol/La).57 Virtually all of the excreted dose in bile was represented by M1, M2, and M3. Indeed, TGZ was rapidly metabolized in the rat to M1 and M2. Thirty minutes after the intravenous administration of TGZ (at a dose approximately 3-fold greater than that given to patients), which is expected to result in much higher plasma TGZ concentrations than when administered orally,62 the concentration of TGZ in the liver was undetectable in male and was approximately 10 μmol/L in female rats.60 The diabetic KK mice had a faster metabolism of TGZ than normal mice, with approximately 66% of the dose being metabolized after 1 hour.57 Experiments in diabetic KK mice showed that the combined CMax for TGZ and its metabolites in the liver was lower (approximately 23 μmol/L) than in normal animals.57

Studies of up to 52 weeks in monkeys, the closest animal model to humans for the study of TGZ,42, 43 showed that at doses 160-fold higher than that given to patients, TGZ was metabolized efficiently but the sulfate M1 accumulated in plasma.42 In humans, after reaching a steady state, the plasma CMax for TGZ (approximately 3-6 μmol/L) was approximately 12% of the CMax for TGZ + M1 + M3.63 This indicates a very efficient metabolism of TGZ by the liver, supporting the notion that hepatic TGZ in humans is in equilibrium with plasma TGZ.59 The peak level occurs approximately 4 hours after taking TGZ, and the plasma concentration drops progressively from 4 to 24 hours, which would also decrease the liver concentration concomitantly.64, 65 Moreover, because TGZ is tightly bound to albumin in plasma (95%-99.8%), only 0.2% to 5% is available as free plasma TGZ capable of a steady-state equilibrium with hepatocytes.59, 63

It has been speculated that TGZ levels in the livers of patients with alcohol-induced liver disease and NASH should be higher than in normal individuals, leading to greater rates of liver injury in these patients.3 However, this hypothesis is not supported by the available experimental data. In patients with type 2 diabetes, the pharmacokinetics of TGZ were normal, and the plasma CMax (and consequently, the liver CMax) for TGZ, M1, and M3 were not different from those of healthy normal individuals.66 The CMax for TGZ was 16% and 39% lower in cirrhotic patients with moderate and severe liver impairment, respectively.64 The CMax of the quinone M3 was normal, whereas the CMax for the sulfate M1 was increased in cirrhotic patients.64 However, the M1 metabolite showed no toxicity in cultured hepatocytes,55 in HepG2 cells,56 or in monkeys, even at much higher plasma concentrations.42, 43 Indeed, the total integrated plasma concentration for M1 among cirrhotic patients was less than 10% of that in monkeys receiving an extremely high TGZ dose for 52 weeks.43 Likewise, the quinone M3 was not toxic to cultured rat hepatocytes even at concentrations of 100 μmol/L (approximately 50-fold the liver concentration).67

Oxidative Stress and Mitochondrial Damage

  1. Top of page
  2. Troglitazone-Associated Liver Injury Is Idiosyncratic
  3. Epidemiology of Troglitazone-Associated Liver Injury
  4. Troglitazone Pharmacokinetics
  5. Oxidative Stress and Mitochondrial Damage
  6. Bile Salt Export Proteins and Cholestasis
  7. Apoptosis in Cell Cultures
  8. Liver Apoptosis and Mitochondrial Abnormalities in Patients
  9. Liver Fibrosis and Cancer
  10. Potential Genetic and Acquired Susceptibility to TGZ
  11. Conclusions
  12. Acknowledgements
  13. References

AlthoughTGZ has been proposed to exacerbate fatty liver by producing oxidative stress and inflammation in the liver,3 TGZ actually has potent antioxidant and anti-inflammatory properties.

Increased oxidative stress can lead to mitochondrial damage within the cell, and, in turn, decreased mitochondrial oxidation of fatty acids results in the accumulation of triglycerides. Therefore, increased triglyceride concentration in the liver is an indicator of mitochondrial damage.68, 69 TGZ treatment induced fatty liver in both diabetic KK mice, and in hyperlipidemic (NZO × NON) F1 mice,70, 71 but the hepatic triglyceride concentration remained unchanged or was not reported,70, 71 reflecting the accumulation of unidentified lipids, apparently, unrelated to mitochondrial injury. Moreover, similar fatty changes in the liver occurred after the administration of rosiglitazone or pioglitazone.70–72

Therefore, these mouse models are poor predictors of the liver abnormalities associated with TGZ given that they do not reflect the pathological conditions observed in humans and that the current glitazones, which have not been associated with acute liver failure, induce similar abnormalities.

TGZ, at concentrations relevant to human pharmacokinetics, did not induce changes in mitochondrial oxidation in normal cells.73 Although TGZ inhibited fatty acid oxidation and esterification in isolated hepatocytes from starved rats,74 this effect occurred at a concentration approximately 200-fold greater than that achieved therapeutically in the liver.58

Futhermore, TGZ prevented mitochondrial abnormalities and apoptosis, as well as decreased liver and pancreatic islet triglyceride concentration in Zucker diabetic rats and Long-Evans fatty rats.75–77 More importantly, TGZ decreased the liver size and fat accumulation in the livers of patients with insulin resistance and type 2 diabetes or lipodystrophy syndrome, as determined by magnetic resonance imaging and computed tomography scans.16, 78 In addition, TGZ improved mitochondrial oxidation in insulin-resistant patients,16 presumably through PPAR-γ signaling, which is consistent with the stimulation of mitochondrial synthesis and function by PPAR-γ coactivator-1α.5

The activation and induction of PPAR-γ caused by TGZ is also observed with the antioxidant vitamin E79 and anti-inflammatory drugs such as ibuprofen,80, 81 in the absence of cytotoxicity. TGZ is a potent anti-inflammatory drug because it blocks cytokine- and lipopolysaccharide-induced cytotoxicity, activation of macrophages and nitric oxide synthase and chemokine expression.81–87 Like vitamin E, TGZ acts as an antioxidant by inhibiting the oxidation of low-density lipoprotein cholesterol and by blocking the reactive products of oxidative stress.88, 89 These antioxidant and anti-inflammatory effects81, 84, 88, 89 may explain the improved patency of both carotid arteries and coronary artery stents in type 2 diabetic patients who are taking TGZ.90, 91

These beneficial effects of TGZ in patients and animals with insulin resistance are inconsistent with the hypothesis that TGZ induces oxidative stress and mitochondrial abnormalities,3 except possibly in rare, susceptible individuals.

Bile Salt Export Proteins and Cholestasis

  1. Top of page
  2. Troglitazone-Associated Liver Injury Is Idiosyncratic
  3. Epidemiology of Troglitazone-Associated Liver Injury
  4. Troglitazone Pharmacokinetics
  5. Oxidative Stress and Mitochondrial Damage
  6. Bile Salt Export Proteins and Cholestasis
  7. Apoptosis in Cell Cultures
  8. Liver Apoptosis and Mitochondrial Abnormalities in Patients
  9. Liver Fibrosis and Cancer
  10. Potential Genetic and Acquired Susceptibility to TGZ
  11. Conclusions
  12. Acknowledgements
  13. References

The proposal that TGZ and the sulfate M1 may induce intrahepatic cholestasis, contributing indirectly to the development of liver injury in patients taking TGZ,3 is plausible because in isolated rat liver canalicular preparations, TGZ inhibited Bsep.60 Also, impaired biliary excretion of taurocholic acid was observed in isolated perfused livers under albumin-free conditions.60, 92 However, this cholestatic effect of TGZ was prevented by adding albumin to the perfusate.92 This emphasizes the critical role of protein binding in the analysis of TGZ toxicity.59, 65 Although the intravenous administration of TGZ to rats induced acute cholestasis,60 there is no evidence that these effects of TGZ occur after oral administration in chronic studies in animals or in patients.

Comparable experimental results have been observed with other drugs, including cyclosporin A, rifampicin, and glyburide, but there is rarely clinical correlation.93 For example, rifampicin inhibits Bsep in experimental studies94 but ameliorates pruritus (induced by bile acids) in patients with cholestasis.93, 95 This effect is achieved through the stimulation of CYP3A4 activity by rifampicin.96 In turn, 6-α hydroxylation of bile acids by CYP3A4 and subsequent conjugation at C6 by UDP-glucuronosyltransferase increases the renal clearance of bile acids.97, 98 The beneficial effects of rifampicin on bile acid excretion in patients, and the apparent discrepancy between animal and human studies, can be readily explained by the fact that rifampicin is a potent activator of human, but not mouse, pregnane X receptor, which induces CYP3A4.96

In other instances, the correlation between experimental inhibition of Bsep and clinical cholestasis is higher. Because bosentan, an endothelin-1 receptor antagonist that inhibits Bsep in experimental models, can induce cholestasis and liver injury in humans,99 it is used with strict monitoring requirements in patients with pulmonary arterial hypertension.100

The cholestatic effects of TGZ have not been confirmed with chronic studies in animals and humans. Increased serum alkaline phosphatase is a sensitive indicator of cholestasis,101 because bile acids induce the expression of alkaline phosphatase in the liver by enhancing mRNA translation.102 TGZ treatment in monkeys (60- to 120-fold the therapeutic dose) for up to 52 weeks did not increase serum alkaline phosphatase levels.42, 43 In addition, administration of TGZ to monkeys (approximately 20-fold the therapeutic dose) for 4 weeks did not affect sulfobromophthalein plasma clearance, suggesting that TGZ did not impair the hepatic uptake, transport, and biliary excretion of this organic anion.42 Similarly, the serum alkaline phosphatase levels were not increased in the cohort of subjects given TGZ in the clinical trials.103

Alternatively, compensatory mechanisms for the inhibition of Bsep activity by TGZ could exist. These may include the induction of CYP3A4 by TGZ and bile acids,104, 105 which would result in an increased renal excretion of C6-glucuronidated bile acids95–98 and the prevention of hepatocellular damage and inflammation, such as that associated with cholestasis in bile duct–ligated animals.106 PPAR-γ activation is important because it inhibits tumor necrosis factor alpha (TNF-α) production, a contributor to liver inflammation,107 and blocks lipopolysaccharide signaling, a mediator of hepatic injury in cholestasis.108

In brief, if TGZ induced liver injury in humans by affecting Bsep-dependent bile acid excretion or by any other cholestatic mechanism, the event has been rare because cholestasis was unusual among the case reports.36–38

Apoptosis in Cell Cultures

  1. Top of page
  2. Troglitazone-Associated Liver Injury Is Idiosyncratic
  3. Epidemiology of Troglitazone-Associated Liver Injury
  4. Troglitazone Pharmacokinetics
  5. Oxidative Stress and Mitochondrial Damage
  6. Bile Salt Export Proteins and Cholestasis
  7. Apoptosis in Cell Cultures
  8. Liver Apoptosis and Mitochondrial Abnormalities in Patients
  9. Liver Fibrosis and Cancer
  10. Potential Genetic and Acquired Susceptibility to TGZ
  11. Conclusions
  12. Acknowledgements
  13. References

Cell studies are important to understand the effects of drugs on molecular pathways, but the FDA does not require cell studies and considers animal studies sufficient for preclinical assessment of drug toxicity [http://www.fda.gov/cder/guidance/index.htm].

It has been argued that apoptosis may be a mechanism responsible for TGZ-induced liver injury.3 Although TGZ has been shown to induce cell death in various cancer cell lines and tumors in animals, it did not affect the corresponding normal primary cells or tissues.109–114 Moreover, these studies with tumoral cells cannot be reliably extrapolated to humans because cancer cells have abnormal mitochondrial morphology, protein content, DNA, and metabolism.115 In addition, TGZ-induced mitochondrial abnormalities in cancer or transformed cell lines were not associated with either apoptosis or cytochrome c release from mitochondria,116, 117 an activator of caspase 9 (a mitochondrial apoptotic pathway).118 Rather, TGZ facilitated apoptosis initiated by TNF-related apoptosis-inducing ligand, at least in some tumor cell lines, by increasing degradation of FLICE-like inhibitory protein long form (FLIPL),109 an inhibitor of caspase 8 (a non-mitochondrial apoptotic pathway).119

Induction of toxicity and cell death by TGZ in normal cells also has been demonstrated, but either with concentrations higher than those achieved in animals and patients given therapeutic doses of TGZ55, 120–122 or in cells that were cultured in the absence or with very low concentrations of albumin (or fetal calf serum).120, 122–124 (Table 1). By contrast, when normal cells, including primary hepatocytes, endothelial cells, peripheral blood monocytes, bone marrow, lymphocytes, and breast cells, were cultured with the appropriate albumin and TGZ concentrations reflective of TGZ pharmacokinetics, there was no cellular toxicity,81, 110, 121 even in the presence of TNF-related apoptosis-inducing ligand,109 a death receptor agonist that activates the extrinsic apoptotic pathway119 (see Table 1). Furthermore, TGZ toxicity to hepatocytes cultured without albumin, which artificially enhances the TGZ bioavailability by up to approximately 500-fold,59 was markedly reduced when albumin was added to the media.123

Table 1. Hepatocyte Studies Assessing Troglitazone-Induced Apoptosis
 Hepatocyte Studies
SpeciesTGZ (μmol/L)Albumin (g/100 mL)Apoptosis
  1. NOTE. Higher troglitazone (TGZ) concentrations than those achieved therapeutically in the liver were used in these hepatocyte studies.

Haskins et al.121Rat200-6007.5No
 Human50-1007.5No
Kostrubsky et al.55Human and pig500No
Toyoda et al.123Rat250.1Yes
 Rat501.0No
Ramachandran et al.122Human500No
Kim et al.109Monkey200.4No

TGZ was cytotoxic to cryopreserved human hepatocytes, judging by the decrease in cellular adenosine triphosphate,125 but the validity of these results is unclear because of the following issues: i) cryopreservation is associated with a loss of hepatocyte viability126, 127; ii) TGZ induced hepatocyte toxicity at concentrations approximately 20-fold greater than that achieved therapeutically in the liver58, 63, 64; iii) the absence of albumin in the culture media increases TGZ bioavailability 20- to 500-fold59, 64; and iv) rosiglitazone also induced cytotoxicity, albeit with a higher median effective concentration

Predictive model systems for TGZ-induced liver toxicity could include highly differentiated human hepatocytes, cocultured with other liver cells (endothelial, macrophages, or stellate cells), given their modulatory role on hepatocyte injury and survival,128, 129 as well as mice expressing humanized xenobiotic responses.96

Liver Apoptosis and Mitochondrial Abnormalities in Patients

  1. Top of page
  2. Troglitazone-Associated Liver Injury Is Idiosyncratic
  3. Epidemiology of Troglitazone-Associated Liver Injury
  4. Troglitazone Pharmacokinetics
  5. Oxidative Stress and Mitochondrial Damage
  6. Bile Salt Export Proteins and Cholestasis
  7. Apoptosis in Cell Cultures
  8. Liver Apoptosis and Mitochondrial Abnormalities in Patients
  9. Liver Fibrosis and Cancer
  10. Potential Genetic and Acquired Susceptibility to TGZ
  11. Conclusions
  12. Acknowledgements
  13. References

Although apoptosis occurs rarely in normal livers, at least one apoptotic body was found in the proximity of 30% of the terminal hepatic veins in normal human and rat livers.130 Apoptosis in all tissues, including the liver, can be easily detected by microscopy because of the characteristic pathological features such as pyknotic and fragmented nuclei and shrunken cytoplasm.131 Hepatocellular apoptosis was detected only in one case report without specifying whether it was increased from background levels, and with normal mitochondria by electron microscopy.16 None of the apoptotic features have been reported in the liver or other tissues, after TGZ treatment, in the other 19 case reports,21–34, 36–38 or in animal studies.41, 43, 70, 76, 77, 110 The presence of acidophilic bodies, in the absence of any typical pathological evidence of apoptosis, is mentioned in a few case reports in association with severe necrosis and inflammation.25, 27, 35

The speculation that TGZ-induced liver injury without increased serum aminotransferases, bilirubin, or alkaline phosphatase3 is not supported by the following findings: i) TGZ did not induce liver apoptosis in animals,43, 70, 76, 77, 110 and ii) stimulation of hepatocyte apoptosis by bile acids, TNF-α/actinomycin or concanavalin A in mice is not silent because it resulted in substantial increases in serum ALT.107, 132, 133

When TGZ was given to patients with NASH, a statistically significant improvement in serum ALT occurred, and a one-grade improvement in the degree of necrosis and inflammation was observed in 5 of the 7 liver biopsies.134 The other PPAR-γ agonists, rosiglitazone and pioglitazone, also improve hepatic inflammation in patients with NASH.135, 136 After the 6-month TGZ treatment, patients did not have evidences of liver apoptosis by electron microscopy or TUNEL assay.134 Mitochondria structural changes (enlarged mitochondria and crystalline inclusions), which are present only in approximately 1% of liver mitochondria among NASH patients,137 were not altered by TGZ treatment.134 However, the small number of patients could have affected the statistical analysis. The significance of these structural mitochondrial findings is uncertain. They may reflect adaptive changes because they also occurred after estrogen/progesterone treatment or pregnancy in rats, associated with decreased mitochondrial oxidation.138, 139

Thus, pathological liver apoptosis in patients treated with TGZ does not seem to be a key mechanism for TGZ-induced liver injury.

Liver Fibrosis and Cancer

  1. Top of page
  2. Troglitazone-Associated Liver Injury Is Idiosyncratic
  3. Epidemiology of Troglitazone-Associated Liver Injury
  4. Troglitazone Pharmacokinetics
  5. Oxidative Stress and Mitochondrial Damage
  6. Bile Salt Export Proteins and Cholestasis
  7. Apoptosis in Cell Cultures
  8. Liver Apoptosis and Mitochondrial Abnormalities in Patients
  9. Liver Fibrosis and Cancer
  10. Potential Genetic and Acquired Susceptibility to TGZ
  11. Conclusions
  12. Acknowledgements
  13. References

The hallmark of chronic liver injury is fibrosis, which eventually evolves into cirrhosis.140, 141 An apparent increase in fibrosis is expected in biopsies from patients with submassive liver necrosis caused by structural collapse. Therefore, the diagnosis of cirrhosis in patients with TGZ-associated submassive liver necrosis cannot be established. In a pilot study, only minor changes occurred in the liver fibrosis score in NASH patients treated with TGZ,134 most likely reflecting sampling error in the biopsies.142

After their activation, hepatic stellate cells are responsible for the production of excess extracellular matrix.143, 144 Thus, regardless of the type, severity, or chronicity of a liver injury, activation of quiescent stellate cells into myofibroblastic stellate cells is indispensable for the development of liver fibrosis/cirrhosis.141 Indeed, this is the basis of potential therapeutic approaches to prevent liver fibrosis by blocking stellate cell activation by using PPAR-γ agonists.106

It has been suggested that hepatocyte apoptosis, associated with increased serum ALT and inflammation, may be a link to liver fibrosis.145 Moreover, apoptotic body engulfment by a human stellate cell line induced expression of collagen type 1.146 Whether apoptotic bodies would be engulfed by stellate cells in the liver and stimulate fibrogenesis, in the presence of competing macrophages and in the absence of an inflammatory response, remains to be determined.147

Active fibrogenesis is common in many tumors, and collapse of the architecture after cell death would enhance the fibrotic appearance. For example, the combination of TGZ with all-trans-retinoic acid induced apoptosis and fibrosis in breast tumors implanted into mice.110 Treatment with TGZ alone (at a dose approximately 200-fold greater than the therapeutic dose) was not associated with either apoptosis or fibrosis of these tumors.110

Some of the signaling pathways critical for the induction of collagen type 1 gene expression and stellate cell activation can be blocked with antioxidants.144, 148, 149 TGZ and related PPAR-γ agonists also block the activation of hepatic stellate cells, thereby inhibiting the production of the extracellular matrix proteins that constitutes the wound healing fibrotic response of the liver.83, 150, 151

The suggestion that TGZ could have induced cancer in patients3 is not supported by the experimental data. Increased incidence of liver hemangiosarcomas and hepatocellular carcinomas occurred with TGZ in mice at 20 and 40 times the human therapeutic dose.41 No tumors of any type were induced by TGZ in rats at 40 times or in monkeys at 160 times the human therapeutic dose.41–43 Furthermore, TGZ was neither mutagenic in bacteria nor clastogenic in bone marrow of mice or rats, indicating no genotoxic risk.152 In addition, none of the analyzed 165 tumors in mice had inactivating p53 mutations, and fewer than 5% had mutations of Ki- and Ha- ras oncogenes.153, 154

Thus, there is no evidence that TGZ facilitates the development of liver fibrosis and cirrhosis140, 141 or the induction of liver tumors in humans.41

Potential Genetic and Acquired Susceptibility to TGZ

  1. Top of page
  2. Troglitazone-Associated Liver Injury Is Idiosyncratic
  3. Epidemiology of Troglitazone-Associated Liver Injury
  4. Troglitazone Pharmacokinetics
  5. Oxidative Stress and Mitochondrial Damage
  6. Bile Salt Export Proteins and Cholestasis
  7. Apoptosis in Cell Cultures
  8. Liver Apoptosis and Mitochondrial Abnormalities in Patients
  9. Liver Fibrosis and Cancer
  10. Potential Genetic and Acquired Susceptibility to TGZ
  11. Conclusions
  12. Acknowledgements
  13. References

The metabolism of TGZ by the CYP3A4 in hepatocytes, is a common pathway for approximately 50% of all medications, as well as estrogen, testosterone, and bile acids.155 Corticosteroids, St. John's Wort, and some antibiotics, sedatives, and cardiovascular medications,155 for example, share the induction of CYP3A4 with TGZ,105 which would increase TGZ metabolism into M3.

Relevant to the understanding of the variable individual susceptibility to TGZ hepatotoxicity, single-nucleotide polymorphisms have been found in three genes critical for the overall activity of CYP3A.156 This is important because the hepatic expression of CYP3A4 varies approximately 50-fold, and the in vivo CYP3A4 enzymatic function (drug clearance) varies at least 20-fold among individuals.157 Depending on the drug studied, 60% to 90% of the individual variability in CYP3A function is caused by genetic factors.157, 158 Single-nucleotide polymorphisms of members of the CYP3A gene family affect various ethnic groups differently and could have contributed to the individual susceptibility to TGZ.156, 159 CYP3A activity, which would predict TGZ metabolism into M3, can be accurately determined in vivo by a breath test based on N-demethylation of erythromycin.160

The quinone M3, which has a chemical structure similar to that of vitamin E,55 has not been found to be toxic after prolonged treatment of monkeys with up to 320-fold the therapeutic TGZ dose.42, 43 However, in susceptible individuals, M3 conceivably could react with proteins, RNA, or DNA to induce liver injury. Also, M3-derived reactive intermediates are covalently bound to microsomal protein and glutathione,67, 161 but the significance of these adducts in the pathogenesis of TGZ-associated hepatoxicity is unknown.

In a cohort of 4,079 patients, combined genetic polymorphisms involving specific genes were associated with increased susceptibility for TGZ-induced liver injury.162 They included heterozygous single point mutations for CYP1A1 (increased drug interactions), NQO1 (reduced metabolic activity), GLUT-1 (increased risk for type 2 diabetes), PPARγ-892 (increased insulin sensitivity), and PPARγ-1431 (increased leptin levels).162 In a small group of patients, a strong correlation was also observed between TGZ-induced liver injury and the combined glutathione-S-transferase GSTT1-GSTM1 null genotype.163

Conclusions

  1. Top of page
  2. Troglitazone-Associated Liver Injury Is Idiosyncratic
  3. Epidemiology of Troglitazone-Associated Liver Injury
  4. Troglitazone Pharmacokinetics
  5. Oxidative Stress and Mitochondrial Damage
  6. Bile Salt Export Proteins and Cholestasis
  7. Apoptosis in Cell Cultures
  8. Liver Apoptosis and Mitochondrial Abnormalities in Patients
  9. Liver Fibrosis and Cancer
  10. Potential Genetic and Acquired Susceptibility to TGZ
  11. Conclusions
  12. Acknowledgements
  13. References

The mechanisms by which TGZ caused severe liver injury remain unknown, but they may include genetic or acquired susceptibilities involving CYP3A, CYP1A1, NQO1, GLUT-1, PPARγ, Bsep, or GST genes. No evidence exists that TGZ induced pathological apoptosis of hepatocytes in culture (under appropriate experimental conditions), in animals or in patients.

In addition, no scientific evidence supports the following notions about TGZ: (i) that it accumulates in the liver; (ii) that it stimulates oxidative stress in the liver; (iii) that it injures the mitochondria in hepatocytes (except in transformed or cancer cells); (iv) that it induces cholestasis in patients (except in rare cases); (v) that it provokes steatosis in the liver; (vi) that it causes silent liver injury; and (vii) that exposed patients are at risk of developing cirrhosis and liver cancer.

The liver injury induced by TGZ was mainly hepatocellular and attributable to a metabolic idiosyncrasy. Although in some cases evidence exists for an immunological reaction, this mechanism cannot explain most of the cases. Identification of the mechanisms responsible for TGZ-induced liver injury, using tissues from well-characterized cases, might help to minimize the risk of similar toxicities with other drugs.

Acknowledgements

  1. Top of page
  2. Troglitazone-Associated Liver Injury Is Idiosyncratic
  3. Epidemiology of Troglitazone-Associated Liver Injury
  4. Troglitazone Pharmacokinetics
  5. Oxidative Stress and Mitochondrial Damage
  6. Bile Salt Export Proteins and Cholestasis
  7. Apoptosis in Cell Cultures
  8. Liver Apoptosis and Mitochondrial Abnormalities in Patients
  9. Liver Fibrosis and Cancer
  10. Potential Genetic and Acquired Susceptibility to TGZ
  11. Conclusions
  12. Acknowledgements
  13. References

The author thanks Dr. Martina Buck (University of California, San Diego) for her critical review of the manuscript, and to Drs. Urs A. Boelsterli (National University of Singapore), D. Montgomery Bissell (University of California, San Francisco), Joe V. Selby (Kaiser Permanente), Hashem El-Serag (University of Texas, Houston), James Everhart (National Institutes of Health), and Abigail Jacobs (Food and Drug Administration) for their valuable comments. The author thanks Lauren de los Santos for the preparation of the manuscript. Space limitations prevented the citation of many important publications.

  • a

    The molecular weight of TGZ is 445D and the hepatic water content was estimated to be 75% (μg/g × 1000/445 × 100/95 = μmol/L).

References

  1. Top of page
  2. Troglitazone-Associated Liver Injury Is Idiosyncratic
  3. Epidemiology of Troglitazone-Associated Liver Injury
  4. Troglitazone Pharmacokinetics
  5. Oxidative Stress and Mitochondrial Damage
  6. Bile Salt Export Proteins and Cholestasis
  7. Apoptosis in Cell Cultures
  8. Liver Apoptosis and Mitochondrial Abnormalities in Patients
  9. Liver Fibrosis and Cancer
  10. Potential Genetic and Acquired Susceptibility to TGZ
  11. Conclusions
  12. Acknowledgements
  13. References
  • 1
    Graham DJ, Green L, Senior JR, Nourjah P. Troglitazone-induced liver failure: a case study. Am J Med 2002; 114: 299306.
  • 2
    Company News; Pfizer wins diabetes drug liability case. New York Times 2004 May 28;C:4.
  • 3
    Smith MT. Mechanisms of troglitazone hepatotoxicity. Chem Res Toxicol 2003; 16: 679687.
  • 4
    Petersen K, Dufour S, Befroy D, Garcia R, Shulman G. Impaired mitochondrial activity in the insulin-resistant offspring of patients with type 2 diabetes. N Engl J Med 2004; 350: 664671.
  • 5
    Nisoli E, Clementi E, Paolucci C, Cozzi V, Tonello C, Sciorati C, et al. Mitochondrial biogenesis in mammals: the role of endogenous nitric oxide. Science 2003; 299: 896899.
  • 6
    Petersen K, Befroy D, Dufour S, Dziura J, Ariyan C, Rothman D, et al. Mitochondrial dysfunction in the elderly: possible role in insulin resistance. Science 2003; 300: 11401142.
  • 7
    Inzucchi SE, Maggs DG, Spollett GR, Page SL, Rife FS, Walton V, et al. Efficacy and metabolic effects of metformin and troglitazone in type II diabetes mellitus. N Engl J Med 1998; 338: 867872.
  • 8
    Taylor R. Causation of type 2 diabetes—the Gordian knot unravels. N Engl J Med 2004; 350: 639641.
  • 9
    Faich GA, Moseley RH. Troglitazone (Rezulin) and hepatic injury. Pharmacoepidemiol Drug Saf 2001; 10: 537547.
  • 10
    Schwart S, Raskin P, Fonseca V, Graveline JF. Effect of troglitazone in insulin-treated patients with type II diabetes mellitus. Troglitazone and Exogenous Insulin Study Group. N Engl J Med 1998; 338: 861866.
  • 11
    Ghazzi MN, Perez JE, Antonucci TK, Driscoll JH, Huang SM, Faja BW, et al. Cardiac and glycemic benefits of troglitazone treatment in NIDDM. The Troglitazone Study Group. Diabetes 1997; 46: 433439.
  • 12
    Sung BH, Izzo JL Jr, Dandona P, Wilson MF. Vasodilatory effects of troglitazone improve blood pressure at rest and during mental stress in type 2 diabetes mellitus. Hypertension 1999; 34: 8388.
  • 13
    Kruszynska YT, Yu JG, Olefsky JM, Sobel BE. Effects of troglitazone on blood concentrations of plasminogen activator inhibitor 1 in patients with type 2 diabetes and in lean and obese normal subjects. Diabetes 2000; 49: 633639.
  • 14
    Buchanan TA, Xiang AH, Peters RK, Kjos SL, Marroquin A, Goico J, et al. Preservation of pancreatic beta-cell function and prevention of type 2 diabetes by pharmacological treatment of insulin resistance in high-risk hispanic women. Diabetes 2002; 51: 27962803.
  • 15
    Azziz R, Ehrmann D, Legro RS, Whitcomb RW, Hanley R, Fereshetian AG, et al. Troglitazone improves ovulation and hirsutism in the polycystic ovary syndrome: a multicenter, double blind, placebo-controlled trial. J Clin Endocrinol Metab 2001; 86: 16261632.
  • 16
    Arioglu E, Duncan-Morin J, Sebring N, Rother KI, Gottlieb N, Lieberman J, et al. Efficacy and safety of troglitazone in the treatment of lipodystrophy syndromes. Ann Intern Med 2000; 133: 263274.
  • 17
    Willi SM, Kennedy A, Wallace P, Ganaway E, Rogers NL, Garvey WT. Troglitazone antagonizes metabolic effects of glucocorticoids in humans: effects on glucose tolerance, insulin sensitivity, suppression of free fatty acids, and leptin. Diabetes 2002; 51: 28952902.
  • 18
    Watkins PB, Whitcomb RW. Hepatic dysfunction associated with troglitazone. N Engl J Med 2001; 338: 916917.
  • 19
    St. Peter JV, Neafus KL, Khan MA, Vessey JT, Lockhart MS. Factors associated with the risk of liver enzyme elevation in patients with type 2 diabetes treated with a thiazolidinedione. Pharmacotherapy 2001; 21: 183188.
  • 20
    Watkins PB, Zimmerman HJ, Knapp MJ, Gracon SI, Lewis KW. Hepatotoxic effects of tacrine administration in patients with Alzheimer's disease. JAMA 1994; 271: 10231024.
  • 21
    Booth AM, Caldwell SH, Iezzoni JC. Troglitazone-associated hepatic failure. Am J Gastroenterol 2000; 95: 557558.
  • 22
    Caldwell SH, Hespenheide EE, von Borstel RW. Myositis, microvesicular hepatitis, and progression to cirrhosis from troglitazone added to simvastatin. Dig Dis Sci 2001; 46: 376378.
  • 23
    Vella A, de Groen PC, Dinneen SF. Fatal hepatotoxicity associated with troglitazone. Ann Intern Med 1998; 129: 1080.
  • 24
    Iwase M, Yamaguchi M, Yoshinari M, Okamura C, Hirahashi T, Tsuji H, et al. A Japanese case of liver dysfunction after 19 months of troglitazone treatment. Diabetes Care 1999; 22: 13821384.
  • 25
    Kohlroser J, Mathai J, Reichheld J, Banner BF, Bonkovsky HL. Hepatotoxicity due to troglitazone: report of two cases and review of adverse events reported to the United States Food and Drug Administration. Am J Gastroenterol 2000; 95: 272276.
    Direct Link:
  • 26
    Fukano M, Amano S, Sato J, Yamamota K, Adachi H, Okabe H, et al. Subacute hepatic failure associated with a new antidiabetic agent, troglitazone: a case report with autopsy examination. Hum Pathol 2000; 31: 250253.
  • 27
    Murphy EJ, Davern T, Shakil A, Shick L, Masharani U, Chow H, et al. Troglitazone-induced hepatic failure. Acute Liver Failure Study Group. Dig Dis Sci 2000; 45: 549553.
  • 28
    Prendergast KA, Berg CL, Wisniewski R. Troglitazone-associated hepatotoxicity treated successfully with steroids. Ann Intern Med 2000; 133: 751.
  • 29
    Neuschwander-Tetri BA, Isley WL, Oki JC, Ramrakhiani S, Quiason SG, Phillips NJ, et al. Troglitazone-induced hepatic failure leading to liver transplantation: a case report. Ann Intern Med 1998; 129: 3841.
  • 30
    Li H, Heller DS, Leevy CB, Zierer KG, Klein KM. Troglitazone-induced fulminant hepatitis: report of a case with autopsy findings. J Diabetes Complications 2000; 14: 175177.
  • 31
    Jagannath S, Rai R. Rapid-onset subfulminant liver failure associated with troglitazone. Ann Intern Med 2000; 132: 677.
  • 32
    Malik AH, Prasad P, Saboorian MH, Thiele DL, Malet PF. Hepatic injury due to troglitazone. Dig Dis Sci 2000; 45: 210214.
  • 33
    Schiano T, Dolehide K, Hart J, Baker AL. Severe but reversible hepatitis induced by troglitazone. Dig Dis Sci 2000; 45: 10391042.
  • 34
    Shibuya A, Watanabe M, Fujita Y, Saigenji K, Kuwao S, Takahashi H, et al. An autopsy case of troglitazone-induced fulminant hepatitis. Diabetes Care 1998; 21: 21402143.
  • 35
    Herrine SK, Choudary C. Severe hepatotoxicity associated with troglitazone. Ann Intern Med 1999; 130: 163164.
  • 36
    Gitlin N, Julie NL, Spurr CL, Lim KN, Juarbe HM. Two cases of severe clinical and histologic hepatotoxicity associated with troglitazone. Ann Intern Med 2000; 129: 3638.
  • 37
    Menon KVN, Angulo P, Lindor KD. Severe cholestatic hepatitis from troglitazone in a patient with nonalcoholic steatohepatitis and diabetes mellitus. Am J Gastroenterol 2001; 96: 16311634.
    Direct Link:
  • 38
    Bonkovsky HL, Azar R, Bird S, Szabo G, Banner B. Severe cholestatic hepatitis caused by thiazolidinediones: risks associated with substituting rosiglitazone for troglitazone. Dig Dis Sci 2002; 47: 16321637.
  • 39
    Bissell D, Gores GJ, Laskin DL, Hoofnagle JH. Drug-induced liver injury: mechanisms and test systems. HEPATOLOGY 2001; 33: 10091013.
  • 40
    Kaplowitz N. Drug-induced liver disorders: implications for drug development and regulation. Drug Safety 2001; 24: 483490.
  • 41
    Herman JR, Dethloff LA, McGuire EJ, Parker RF, Walsh KM, Gough AW, et al. Rodent carcinogenicity with the thiazolidinedione antidiabetic agent troglitazone. Toxicol Sci 2002; 68: 226236.
  • 42
    Herman JR, Brodfuehrer JI, Koup JR, McGuire EJ. Troglitazone: Studies in nonrodents and relevance to preclinical safety assessment. Food and Drug Administration 1994; RR: 74502299.
  • 43
    Rothwell C, McGuire EJ, Altrogge DM, Masuda H, de la Iglesia FA. Chronic toxicity in monkeys with the thiazolidinedione antidiabetic agent troglitazone. J Toxicol Sci 2002; 27: 3547.
  • 44
    Zimmerman HJ, Ishak KG. Hepatic injury due to drugs and toxins. In: MacsweenRNM, AnthonyPP, ScheuerPJ, BurtAD, PortmannBC, eds. Pathology of the Liver. London, UK: Churchill Livingstone, 1994: 563634.
  • 45
    Klatskin G, Conn H. Toxic and drug-induced hepatic injury. In: KlatskinG, ConnH, eds. Histopathology of the Liver. New York: Oxford University Press, 1993: 111140.
  • 46
    Danan G, Benichou C. Causality assessment of adverse reactions to drugs—I. A novel method based on the conclusions of international consensus meetings: application to drug-induced liver injuries. J Clin Epidemiol 1993; 46: 13231330.
  • 47
    Benichou C, Danan G, Flahault A. Causality assessment of adverse reactions to drugs—II. An original model for validation of drug causality assessment methods: case reports with positive rechallenge. J Clin Epidemiol 1993; 46: 13311336.
  • 48
    Ostapowicz G, Fontana R, Schiodt F, Larson A, Davern T, Han S, et al. Results of a prospective study of acute liver failure at 17 tertiary care centers in the United States. Ann Intern Med 2002; 137: 947954.
  • 49
    Graham DJ, Drinkard CR, Shatin D. Incidence of idiopathic acute liver failure and hospitalized liver injury in patients treated with troglitazone. Am J Gastroenterol 2003; 98: 175179.
    Direct Link:
  • 50
    Kaplowitz N, Lewis JH, Watkins PB. Did this drug cause my patient's hepatitis? Ann Intern Med 2003; 138: 159.
  • 51
    El-Serag HB, Everhart JE. Diabetes increases the risk of acute hepatic failure. Gastroenterology 2002; 122: 18221828.
  • 52
    Chan KA, Truman A, Gurwitz JH, Hurley JS, Martinson B, Platt R, et al. A cohort study of the incidence of serious acute liver injury in diabetic patients treated with hypoglycemic agents. Arch Intern Med 2003; 163: 728734.
  • 53
    Chan KA, Truman A, Gurwitz JH, Hurley JS, Martinson B, Platt R, et al. Risk of unexplained acute liver failure in diabetes mellitus—Reply. Arch Intern Med 2003; 163: 26502651.
  • 54
    Caldwell SH, Hespenheide EE. Subacute liver failure in obese women. Am J Gastroenterol 2002; 97: 20582062.
    Direct Link:
  • 55
    Kostrubsky VE, Sinclair JF, Ramachandran V, Venkataramanan R, Wen YH, Kindt E, et al. The role of conjugation in hepatotoxicity of troglitazone in human and porcine hepatocyte cultures. Drug Metab Dispos 2000; 28: 11921197.
  • 56
    Yamamoto Y, Nakajima M, Yamazaki H, Yokoi T. Cytotoxicity and apoptosis produced by troglitazone in human hepatoma cells. Life Sci 2001; 70: 471482.
  • 57
    Kawai K, Kawasaki-Tokui Y, Odaka T, Tsurata F, Kazui M, Iwabuchi H, et al. Disposition and metabolism of the new oral antidiabetic drug troglitazone in rats, mice and dogs. Arzneimittelforschung 1997; 47: 356368.
  • 58
    Kawai K, Kawasaki Y, Tokui T, Kuroiwa C, Shigehara E, Shinozaki H, et al. Metabolism of CI-991: III. Distribution in rat tissues. Food and Drug Administration 1991; RR: 76401690.
  • 59
    Izumi T, Enomoto S, Hosiyama K, Sasahara K, Shibukawa A, Nakagawa T, et al. Prediction of the human pharmacokinetics of troglitazone, a new and extensively metabolized antidiabetic agent, after oral administration, with an animal scale-up approach. J Pharm Exp Ther 1996; 277: 16301641.
  • 60
    Funk C, Ponelle C, Scheuermann G, Pantze M. Troglitazone-induced intrahepatic cholestasis by an interference with the hepatobiliary export of bile acids in male and female rats. Toxicology 2001; 167: 8398.
  • 61
    Funk C, Ponelle C, Scheuermann G, Pantze M. Cholestatic potential of troglitazone as a possible factor contributing to troglitazone-induced hepatotoxicity: in vivo and in vitro interaction at the canalicular bile salt export pump (Bsep) in the rat. Mol Pharmacol 2001; 59: 627635.
  • 62
    Izumi T, Maeda E, Sasahara K, Semmes R, Koup J. Biopharmaceutical studies on a new antidiabetic, CI-991, VIII. Pharmacokinetic study of dose proportionality of CI-991 and its metabolites after a single-intravenous and oral administration of CI-991. Food and Drug Adminstration 1992; RR: 76401353.
  • 63
    Loi CM, Young M, Randinitis E, Vassos A, Koup JR. Clinical pharmacokinetics of troglitazone. Clin Pharmacokinet 1999; 37: 91104.
  • 64
    Ott P, Ranek L, Young MA. Pharmacokinetics of Troglitazone, a PPAR-gamma agonist, in patients with hepatic insufficiency. Eur J Clin Pharmacol 1998; 54: 567571.
  • 65
    Loi CM, Alvey CW, Vassos A, Randinitis E, Sedman AJ, Koup JR. Steady-state pharmacokinetics and dose proportionality of troglitazone and its metabolites. J Clin Pharmacol 1999; 39: 920926.
  • 66
    Loi CM, Randinitis E, Vassos A, Kazierad DJ, Koup JR, Sedman AJ. Lack of effect of type II diabetes on the pharmacokinetics of troglitazone in a multiple-dose study. J Clin Pharmacol 1997; 37: 11141120.
  • 67
    Tettey JN, Maggs JL, Rapeport WG, Pirmohamed M, Park BK. Enzyme-induction dependent bioactivation of troglitazone and troglitazone quinone in vivo. Chem Res Toxicol 2001; 14: 965974.
  • 68
    Jaeschke H, Gores GJ, Cederbaum AI, Hinson JA, Pessayre D, Lemasters JJ. Mechanisms of hepatotoxicity. Toxicol Sci 2002; 65: 166176.
  • 69
    Fromenty B, Pessayre D. Inhibition of mitochondrial beta-oxidation as a mechanism of hepatotoxicity. Pharmacol Ther 1995; 67: 101154.
  • 70
    Bedoucha M, Atzpodien E, Boelsterli UA. Diabetic KKAγ mice exhibit increased hepatic PPARγ-1gene expression and develop hepatic steatosis upon chronic treatment with antidiabetic thiazolidinediones. J Hepatol 2001; 35: 1723.
  • 71
    Watkins SM, Reifsnyder PR, Pan HJ, German JB, Leiter EH. Lipid metabolome-wide effects of the PPARgamma agonist rosiglitazone. J Lipid Res 2002; 43: 18091817.
  • 72
    Weinstock RS, Murray FT, Diani A, Sangani GA, Wachowski MB, Messina JL. Pioglitazone: in vitro effects on rat hepatoma cells and in vivo liver hypertrophy in KKAy mice. Pharmacology 1997; 54: 169178.
  • 73
    Lenhard JM, Kliewer SA, Paulik MA, Plunket KD, Lehmann JM, Weiel JE. Effects of troglitazone and metformin on glucose and lipid metabolism: alterations of two distinct molecular pathways. Biochem Pharmacol 1997; 54: 801808.
  • 74
    Fulgencio JP, Kohl C, Girard J, Pegorier JP. Troglitazone inhibits fatty acid oxidation and esterification, and gluconeogenesis in isolated hepatocytes from starved rats. Diabetes 1996; 45: 15561562.
  • 75
    Higa M, Zhou YT, Ravazzola M, Baetens D, Orci L, Unger RH. Troglitazone prevents mitochondrial alterations, beta cell destruction, and diabetes in obese prediabetic rats. Proc Natl Acad Sci U S A 1999; 96: 1151311518.
  • 76
    Jia DM, Tabaru A, Nakamura H, Fukumitsu KI, Akiyama T, Otsuki M. Troglitazone prevents fatty changes of the liver in obese diabetic rats. J Gastroenterol Hepatol 2000; 15: 11831191.
  • 77
    Shimabukuro M, Zhou YT, Lee Y, Unger RH. Troglitazone lowers islet fat and restores beta cell function of Zucker diabetic fatty rats. J Biol Chem 1998; 273: 35473550.
  • 78
    Katoh S, Hata S, Matsushima M, Ikemoto S, Inoue Y, Yokoyama J, et al. Troglitazone prevents the rise in visceral adiposity and improves fatty liver associated with sulfonylurea therapy: a randomized controlled trial. Metabolism 2001; 50: 414417.
  • 79
    Davies GF, McFie PJ, Khandelwal RL, Roesler WJ. Unique ability of troglitazone to up-regulate peroxisome proliferator-activated receptor-gamma expression in hepatocytes. J Pharm Exp Ther 2002; 300: 7277.
  • 80
    Lehmann JM, Lenhard JM, Oliver BB, Ringold GM, Kliewer SA. Peroxisome proliferator-activated receptors alpha and gamma are activated by indomethacin and other non-steroidal anti-inflammatory drugs. J Biochem 1997; 272: 34063410.
  • 81
    Jiang C, Ting AT, Seed B. PPAR-gamma agonists inhibit production of monocyte inflammatory cytokines. Nature 1998; 391: 8286.
  • 82
    Maggi LB Jr, Sadeghi H, Weigand C, Scarim AL, Heitmeier MR, Corbett JA. Anti-inflammatory actions of 15-deoxy-delta 12,14-prostaglandin J2 and troglitazone: evidence for heat shock-dependent and -independent inhibition of cytokine-induced inducible nitric oxide synthase expression. Diabetes 2000; 49: 346355.
  • 83
    Marra F, Efsen E, Romanelli RG, Caligiuri A, Pastacaldi S, Batignani G, et al. Ligands of peroxisome proliferator-activated receptor gamma modulate profibrogenic and proinflammatory actions in hepatic stellate cells. Gastroenterology 2000; 119: 466478.
  • 84
    Ricote M, Huang JT, Welch JS, Glass CK. The peroxisome proliferator-activated receptor-gamma is a negative regulator of macrophage activation. Nature 1998; 391: 82.
  • 85
    Combs CK, Johnson DE, Karlo JC, Cannady SB, Landreth GE. Inflammatory mechanisms in Alzheimer's disease: inhibition of β-amyloid-stimulated proinflammatory responses and neurotoxicity by PPARγ agonists. J Neurosci 2000; 20: 558567.
  • 86
    Sigrist S, Bedoucha M, Boelsterli UA. Down-regulation by troglitazone of hepatic tumor necrosis factor-alpha and interleukin-6 mRNA expression in a murine model of non-insulin-dependent diabetes. Biochem Pharmacol 2000; 60: 6775.
  • 87
    Shimabukuro M, Koyama K, Lee Y, Unger RH. Leptin- or troglitazone-induced lipopenia protects islets from interleukin 1beta cytotoxicity. J Clin Invest 1997; 100: 17501754.
  • 88
    Noguchi N, Sakai H, Kato Y, Tsuchiya J, Yamamoto Y, Niki E, et al. Inhibition of oxidation of low density lipoprotein by troglitazone. Atherosclerosis 1996; 123: 227234.
  • 89
    Inoue I, Katayama S, Takahashi K, Negishi K, Mizayaki T, Sonoda M, et al. Troglitazone has a scavenging effect on reactive oxygen species. Bioch Biophys Res Commun 1997; 235: 116.
  • 90
    Minamikawa J, Yamauchi M, Inoue D, Koshiyama H. Another potential use of troglitazone in noninsulin-dependent diabetes mellitus. J Clin Endocrin Metab 1998; 83: 10411042.
  • 91
    Takagi T, Yamamuro A, Tamita K, Yamabe K, Katayama M, Morioka S, et al. Impact of troglitazone on coronary stent implantation using small stents in patients with type 2 diabetes mellitus. Am J Cardiol 2002; 89: 318322.
  • 92
    Preininger K, Stingl H, Englisch R, Furnsinn C, Graf J, Waldhausl W, et al. Acute troglitazone action in isolated perfused rat liver. Br J Pharmacol 1999; 126: 372378.
  • 93
    Bohan A, Boyer JL. Mechanisms of hepatic transport of drugs: implications for cholestatic drug reactions. Semin Liv Dis 2002; 22: 123136.
  • 94
    Stieger B, Fattinger K, Madon J, Kullak-Ublick GA, Meier PJ. Drug- and estrogen-induced cholestasis through inhibition of the hepatocellular bile salt export pump (Bsep) of rat liver. Gastroenterology 2000; 118: 422430.
  • 95
    Wietholtz H, Marschall HU, Sjovall J, Matern S. Stimulation of bile acid 6 alpha-hydroxylation by rifampin. J Hepatol 1996; 24: 713718.
  • 96
    Xie W, Barwick JL, Downes M, Blumberg B, Simon CM, Nelson MC, et al. Humanized xenobiotic response in mice expressing nuclear receptor SXR. Nature 2000; 406: 435439.
  • 97
    Marschall HU, Matern H, Egestad B, Matern S, Sjovall S. 6 Alpha-glucuronidation of hyodeoxycholic acid by human liver, kidney and small bowel microsomes. Biochem Biophys Acta 1987; 921: 392397.
  • 98
    Araya Z, Wikvall K. 6Alpha-hydroxylation of taurochenodeoxycholic acid and lithocholic acid by CYP3A4 in human liver microsomes. Biochem Biophys Acta 1999; 1438: 4754.
  • 99
    Fattinger K, Funk C, Pantze M, Weber C, Reichen J, Stieger B, et al. The endothelin antagonist bosentan inhibits the canalicular bile salt export pump: a potential mechanism for hepatic adverse reactions. Clin Pharmacol Ther 2001; 69: 223231.
  • 100
    Rubin LJ, Badesch DB, Barst RJ, Galie N, Black CM, Keogh A, et al. Bosentan therapy for pulmonary arterial hypertension. N Engl J Med 2002; 346: 896903.
  • 101
    Pratt DS, Kaplan M. Evaluation of abnormal liver-enzyme results in asymptomatic patients. N Engl J Med 2000; 342: 12661271.
  • 102
    Seetharam S, Sussman NL, Komoda T, Alpers DH. The mechanism of elevated alkaline phosphatase activity after bile duct ligation in the rat. HEPATOLOGY 1986; 6: 374380.
  • 103
    Baron B, Valiquett T, Vassos A, Venable T. Integrated summary of the safety of Troglitazone (CI-991). Food and Drug Administration 1996; RR: 72003538.
  • 104
    Xie W, Rodominska-Pandya A, Shi Y, Simon CM, Nelson MC, Ong ES, et al. An essential role for nuclear receptors SXR/PXR in detoxification of cholestatic bile acids. Proc Natl Acad Sci U S A 2001; 98: 33753380.
  • 105
    Sahi J, Hamilton G, Sinz M, Barros S, Huang SM, Lesko LJ, et al. Effect of troglitazone on cytochrome P450 enzymes in primary cultures of human and rat hepatocytes. Xenobiotica 2000; 30: 273284.
  • 106
    Galli A, Crabb DW, Ceni E, Salzano R, Svegliati-Baroni G, Ridolfi F, et al. Antidiabetic thiazolidinediones inhibit collagen synthesis and hepatic stellate cell activation in vivo and in vitro. Gastroenterology 2002; 122: 19241940.
  • 107
    Guicciardi ME, Duessing J, Miyoshi H, Bronk SF, Svingen PA, Peters C, et al. Cathepsin B contributes to TNF-alpha-mediated hepatocyte apoptosis by promoting mitochondrial release of cytochrome c. J Clin Invest 2000; 106: 11271137.
  • 108
    Sewnath ME, van der Poll T, Ten Kate FJ, Van Noorden CJ, Gouma DJ. Interleukin-1 receptor type I gene-deficient bile duct-ligated mice are partially protected against endotoxin. HEPATOLOGY 2002; 35: 149158.
  • 109
    Kim Y, Nanjoo S, Sporn M, Reed J. An inducible pathway for degradation of FLIP protein sensitizes tumor cells to TRAIL-induced apoptosis. J Biol Chem 2002; 277: 2232022329.
  • 110
    Elstner E, Muller C, Koshizuka K, Williamson EA, Park D, Asou H, et al. Ligands for peroxisome proliferator-activated receptor gamma and retinoic acid receptor inhibit growth and induce apoptosis of human breast cancer cells in vitro and in BNX mice. Proc Natl Acad Sci U S A 1998; 95: 88068811.
  • 111
    Kubota T, Koshizuka K, Williamson EA, Asou H, Said JW, Holden S, et al. Ligand for peroxisome proliferator-activated receptor gamma (troglitazone) has potent antitumor effect against human prostate cancer both in vitro and in vivo. Cancer Res 1998; 58: 33443352.
  • 112
    Tsubouchi Y, Sano H, Kawahito Y, Mukai S, Yamada R, Kohno M, et al. Inhibition of human lung cancer cell growth by the peroxisome proliferator-activated receptor-gamma agonists through induction of apoptosis. Bioch Biophys Res Commun 2000; 270: 400405.
  • 113
    Ohta K, Endo T, Haraguchi K, Hershman JM, Onaya T. Ligands for peroxisome proliferator-activated receptor gamma inhibit growth and induce apoptosis of human papillary thyroid carcinoma cells. J Clin Endocrinol Metab 2001; 86: 21702177.
  • 114
    Harris SG, Phipps RP. Prostaglandin D(2), its metabolite 15-d-PGJ(2), and peroxisome proliferator activated receptor-gamma agonists induce apoptosis in transformed, but not normal, human T lineage cells. Immunology 2002; 105: 2334.
  • 115
    Cuezva JM, Krajewska M, Lopez de Heredia M, Krajewski S, Santamaria G, Kim H, et al. The bioenergetic signature of cancer: a marker of tumor progression. Cancer Res 2002; 62: 66746681.
  • 116
    Shishido S, Koga H, Harada M, Kumemura H, Hanada S, Taniguchi E, et al. Hydrogen peroxide overproduction in megamitochondria of troglitazone-treated human hepatocytes. HEPATOLOGY 2003; 37: 136147.
  • 117
    Koga H, Sakisaka S, Harada M, Takagi T, Hanada S, Taniguchi E, et al. Involvement of p21(WAF1/Cip1), p27(Kip1), and p18(INK4c) in troglitazone-induced cell-cycle arrest in human hepatoma cell lines. HEPATOLOGY 2001; 33: 10871097.
  • 118
    Green D, Kroemer G. The pathophysiology of mitochondrial cell death. Science 2004; 305: 626629.
  • 119
    Micheau O, Tschopp J. Induction of TNF receptor I-mediated apoptosis via two sequential signaling complexes. Cell 2003; 114: 181190.
  • 120
    Gouni-Berthold I, Berthold HK, Weber AA, Ko Y, Seul C, Vetter H, et al. Troglitazone and rosiglitazone induce apoptosis of vascular smooth muscle cells through an extracellular signal-regulated kinase-independent pathway. Arch Pharmacol 2001; 363: 215221.
  • 121
    Haskins JR, Rowse P, Rahbari R, de la Iglesia FA. Thiazolidinedione toxicity to isolated hepatocytes revealed by coherent multiprobe fluorescence microscopy and correlated with multiparameter flow cytometry of peripheral leukocytes. Arch Toxicol 2001; 75: 425438.
  • 122
    Ramachandran V, Kostrubsky VE, Komoroski BJ, Zhang S, Dorko K, Esplen JE, et al. Troglitazone increases cytochrome P-450 3A protein and activity in primary cultures of human hepatocytes. Drug Metab Dispos 1999; 27: 11941199.
  • 123
    Toyoda Y, Tsuchida A, Iwami E, Miwa I. Toxic effect of troglitazone on cultured rat hepatocytes. Life Sci 2001; 68: 18671876.
  • 124
    Okura T, Nakamura M, Takata Y, Watanabe S, Kitami Y, Hiwada K. Troglitazone induces apoptosis via the P53 and Gadd45 pathway in vascular smooth muscle cells. Eur J Pharmacol 2000; 407: 227235.
  • 125
    Hewitt NJ, Lloyd S, Hayden M, Butler R, Sakai Y, Springer R, et al. Correlation between troglitazone cytotoxicity and drug metabolic enzyme activities in cryopreserved human hepatocytes. Chem Biol Interact 2002; 142: 7382.
  • 126
    Yagi T, Hardin JA, Valenzuela YM, Miyoshi H, Gores GJ, Nyberg SL. Caspase inhibition reduces apoptotic death of cryopreserved porcine hepatocytes. HEPATOLOGY 2001; 33: 14321440.
  • 127
    Lloyd TD, Orr S, Skett P, Berry DP, Dennison AR. Cryopreservation of hepatocytes: a review of current methods for banking. Cell Tissue Bank 2003; 4: 315.
  • 128
    LeCouter J, Moritz DR, Li B, Phillips GL, Liang XH, Gerber H-P, et al. Angiogenesis-independent endothelial protection of liver: role of VEGFR-1. Science 2003; 299: 890893.
  • 129
    Chojkier M, Fierer J. D-Galactosamine hepatotoxicity is associated with endotoxin-sensitivity and mediated by lymphoreticular cells in mice. Gastroenterology 1985; 88: 115121.
  • 130
    Benedetti A, Jezequel AM, Orlandi F. Preferential distribution of apoptotic bodies in acinar zone 3 of normal human and rat liver. J Hepatol 1988; 7: 319324.
  • 131
    Schulte-Herman R, Bursch W, Grasl-Kraupp B. Active cell death (apoptosis) in liver biology and disease. Prog Liver Dis 1995; 13: 135.
  • 132
    Miyoshi H, Rust C, Roberts PJ, Burgart LJ, Gores GJ. Hepatocyte apoptosis after bile duct ligation in the mouse involves Fas. Gastroenterology 1999; 117: 732736.
  • 133
    Trautwein C, Rakemann T, Malek NP, Plumpe J, Tiegs G, Manns MP. Concanavalin A-induced liver injury triggers hepatocyte proliferation. J Clin Invest 1998; 101: 19601969.
  • 134
    Caldwell SH, Hespenheide EE, Redick JA, Iezzoni JC, Battle EH, Sheppard BL. A pilot study of a thiazolidinedione, troglitazone, in nonalcoholic steatohepatitis. Am J Gastroenterol 2001; 96: 519525.
    Direct Link:
  • 135
    Promrat K, Lutchman G, Uwaifo GI, Freedman RJ, Soza A, Heller T, et al. A pilot study of pioglitazone treatment for nonalcoholic steatohepatitis. HEPATOLOGY 2004; 39: 188196.
  • 136
    Neuschwander-Tetri BA, Brunt EM, Wehmeier KR, Oliver D, Bacon BR. Improved nonalcoholic steatohepatitis after 48 weeks of treatment with the PPAR-gamma ligand rosiglitazone. HEPATOLOGY 2003; 38: 10081017.
  • 137
    Caldwell SH, Swerdlow RH, Khan EM, Iezonni JC, Hespenheide EE, Parks JK, et al. Mitochondrial abnormalities in non-alcoholic steatohepatitis. J Hepatol 1999; 31: 430434.
  • 138
    Grimbert S, Fisch C, Deschamps D, Berson A, Fromenty B, Feldmann G, et al. Effects of female sex hormones on mitochondria: possible role in acute fatty liver of pregnancy. Am J Physiol 1995; 268: 107115.
  • 139
    Grimbert S, Fromenty B, Fisch C, Letteron P, Berson A, Durand-Schneider AM, et al. Decreased mitochondrial oxidation of fatty acids in pregnant mice: possible relevance to development of acute fatty liver of pregnancy. HEPATOLOGY 1993; 17: 628637.
  • 140
    Ankoma-Sey V, Friedman S. Hepatic stellate cells. In: StrainA, DiehlA, eds. Liver Growth and Repair. London: Chapman & Hall, 1998: 512537.
  • 141
    Chojkier M. Regulation of collagen gene expression. In: StrainA, DiehlA, eds. Liver Growth and Repair. London: Chapman & Hall, 1998: 430450.
  • 142
    Regev A, Berho M, Jeffers LJ, Milikowski C, Molina EG, Pyrsopoulos NT, et al. Sampling error and intraobserver variation in liver biopsy in patients with chronic HCV infection. Am J Gastroenterol 2002; 97: 26142618.
    Direct Link:
  • 143
    Friedman S, Roll F, Boyles J, Bissell D. Hepatic lipocytes: the principal collagen-producing cells of the liver. Proc Natl Acad Sci U S A 1985; 82: 86818685.
  • 144
    Lee KS, Buck M, Houglum K, Chojkier M. Activation of hepatic stellate cells by TGFα and collagen type I is mediated by oxidative stress through c-myb expression. J Clin Invest 1995; 96: 24612468.
  • 145
    Canbay A, Higuchi H, Bronk SF, Taniai M, Sebo TJ, Gores GJ. Fas enhances fibrogenesis in the bile duct ligated mouse: a link between apoptosis and fibrosis. Gastroenterology 2002; 123: 13231330.
  • 146
    Canbay A, Taimr P, Torko N, Higuchi H, Friedman S, Gores GJ. Apoptotic body engulfment by a human stellate cell line is profibrogenic. Lab Invest 2003; 83: 655663.
  • 147
    Canbay A, Friedman S, Gores GJ. Apoptosis: the nexus of liver injury and fibrosis. HEPATOLOGY 2004; 39: 273278.
  • 148
    Lee KS, Cottam HB, Houglum K, Wasson DB, Carson D, Chojkier M. Pentoxyfilline blocks stellate cell activation independently of its cAMP phosphodiesterase inhibitory activity. Am J Physiol 1997; 273: G1094G1100.
  • 149
    Chojkier M, Houglum K, Lee KS, Buck M. Long- and short-term D-α-tocopherol supplementation inhibits liver collagen α1(I) gene expression. Am J Physiol 1998; 275: G14801485.
  • 150
    Miyahara T, Schrum L, Rippe R, Xiong S, Yee HF Jr, Motomura K, et al. Peroxisome proliferator-activated receptors and hepatic stellate cell activation. J Biol Chem 2000; 275: 3571535722.
  • 151
    Galli A, Crabb DW, Price D, Ceni E, Salzano R, Surrenti C, et al. Peroxisome proliferator-activated receptor gamma transcriptional regulation is involved in platelet-derived growth factor-induced proliferation of human hepatic stellate cells. HEPATOLOGY 2000; 31: 101108.
  • 152
    Hirano K, Hagiwara T, Umezawa T, Uchida T, Watanabe M, Matsushima H. Mutagenicity studies of CS-045 in bacteria, in mammalian tissue culture cells and in mice. Rinsho Iyaku 1993; 9: 375388.
  • 153
    Duddy SK, Gorospe SM, Bleavins MR, de la Iglesia FA. Spontaneous and thiazolidinedione-induced B6C3F1 mouse hemangiosarcomas exhibit low ras oncogene mutation frequencies. Toxicol Appl Pharmacol 1999; 160: 133140.
  • 154
    Duddy SK, Parker RF, Bleavins MR, Gough AW, Rowse P, Gorospe SM, et al. p53 is not inactivated in B6C3F1 mouse vascular tumors arising spontaneously or associated with long-term administration of the thiazolidinedione troglitazone. Toxicol Appl Pharmacol 1999; 156: 106112.
  • 155
    Gura T. SNP-ing Drugs to Size. Science 2001; 293: 595.
  • 156
    Kuehl P, Zhang L, Lin Y, Lamba J, Assem M, Schuetz J, et al. Sequence diversity in CYP3A promoters and characterization of the genetic basis of polymorphic CYP3A5 expression. Nat Genet 2001; 27: 383391.
  • 157
    Eichelbaum M, Burk O. CYP3A genetics in drug metabolism. Nature Medicine 2001; 7: 285287.
  • 158
    Ozdemir V, Kalowa W, Tang BK, Paterson AD, Walker SE, Endrenyi L, et al. Evaluation of the genetic component of variability in CYP3A4 activity: a repeated drug administration method. Pharmacogenetics 2000; 10: 373388.
  • 159
    Hustert E, Haberl M, Burk O, Wolbold R, He YQ, Klein K, et al. The genetic determinants of the CYP3A5 polymorphism. Pharmacogenetics 2001; 11: 773779.
  • 160
    Schmidt LE, Olsen AK, Stentoft K, Rasmussen A, Kirkegaard P, Dalhoff K. Early postoperative erythromycin breath test correlates with hepatic cytochrome P4503A activity in liver transplant recipients. Clin Pharmacol Ther 2001; 70: 446454.
  • 161
    He K, Talaat RE, Pool WF, Reily MD, Reed JE, Bridges A, et al. Metabolic activation of troglitazone: identification of a reactive metabolite and mechanisms involved. Drug Metab Dispos 2004; 32: 639646.
  • 162
    de la Iglesia FA, Haskins JR, Feuer G. Hepatotoxicity of cardiovascular and antidiabetic drugs. In: KaplowitzN, DeLeveLD, eds. Drug-Induced Liver Disease. New York: Marcel Dekker, 2003: 549592.
  • 163
    Watanabe I, Tomita A, Shimizu M, Sugawara M, Yasumo H, Koishi R, et al. A study to survey susceptible genetic factors responsible for troglitazone-associated hepatotoxicity in Japanese patients with type 2 diabetes mellitus. Clin Pharmacol Ther 2003; 73: 435455.