Hepatol Int DOI 10.1007/s12072-013-9482-8
SUPPLEMENT ISSUE: NAFLD
Pharmacological agents for nonalcoholic steatohepatitis Maeva Guillaume • Vlad Ratziu
Received: 8 June 2013 / Accepted: 9 October 2013 Ó Asian Pacific Association for the Study of the Liver 2013
Abstract The rationale for specific pharmacologic therapy in nonalcoholic steatohepatitis (NASH) is determined by the potential for disease progression and the difficulties, in many patients, to successfully implement diet and lifestyle changes in the long term. Because they correct insulin resistance, insulin-sensitizing agents are attractive candidates for the treatment of NASH. However, two randomized studies have shown that vitamin E, despite having no effect on insulin sensitivity, achieves interesting histological and biochemical efficacy. This review provides an insight into the therapeutic efficacy and safety issues of different pharmacological agents tested in human NASH. Keywords Steatohepatitis Fibrosis Steatosis Glitazones Vitamin E Ursodesoxycholic acid Insulin resistance Oxidative stress Insulin sensitizers
M. Guillaume Service d’He´patologie et Gastro-ente´rologie, INSERM 1048 and Universite´ Paul Sabatier, Hoˆpital Purpan, Centre Hospitalier Universitaire de Toulouse, Toulouse, France e-mail: [email protected] M. Guillaume Service d’He´patologie et Gastro-ente´rologie, Hoˆpital Purpan, Pavillon Dieulafoy, 4e`me e´tage, Place du Docteur Baylac, TSA 40031, 31059 Toulouse Cedex 9, France V. Ratziu (&) INSERM U938, Hospital Pitie´ Salpeˆtrie`re, CdR Saint-Antoine and Universite´ Pierre et Marie Curie, Paris, France e-mail: [email protected] V. Ratziu Service d’He´patologie et Gastro-ente´rologie, Hoˆpital Pitie´Salpeˆtrie`re, 47-83, Boulevard de l’hoˆpital, 75651 Paris Cedex 13, France
Introduction Nonalcoholic fatty liver disease (NAFLD) encompasses the entire spectrum of fatty liver disease in individuals without significant alcohol consumption, ranging from nonalcoholic fatty liver (NAFL) to nonalcoholic steatohepatitis (NASH) and cirrhosis. NAFL is defined as the presence of hepatic steatosis with minimal, non-specific inflammation that does not qualify for steatohepatitis; NASH is defined as the presence of lobular inflammation with hepatocyte injury (ballooning) [1]. The risk of progression to cirrhosis, liver failure and liver cancer is minimal for NAFL and real for NASH [2]. In addition, there is growing evidence to support an association between NAFLD and an increased risk of HCC [3]. Patients with NASH have increased liverrelated mortality as well as cardiovascular mortality [4]. The recommended first-line therapy for all NAFLD patients is non-pharmacological: weight reduction through diet, changes in lifestyle, physical exercise and proscribing sedentarity [5, 6]. Additionally, concurrent metabolic disorders such as type 2 diabetes (T2DM), hyperlipidemia or arterial hypertension, when present, should be well controlled, although their specific impact on improving the hepatic disease has not yet been determined. Occasionally, these measures result in a 7–10 % weight loss, a threshold that is possibly associated with histological improvement [5, 6]. Moreover, as insulin resistance, inflammation, an unbalanced adipocytokine ratio and angiogenic factors associated with NAFLD could favor the development of HCC [7], these traditional therapeutic approaches in NAFLD may theoretically reduce the risk of HCC by acting on common pathophysiological pathways shared by NAFLD and HCC. In patients who failed these measures or in those with advanced disease (NASH with significant fibrosis), pharmacological treatments specifically directed at
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improving hepatic inflammation, fibrosis and/or clearing steatohepatitis might be necessary [8]. In some carefully selected patients with morbid obesity and without cirrhosis, bariatric surgery may be proposed [9]. Here we will provide an overview of data obtained so far with pharmacological agents in NASH mostly focusing on controlled studies with histological documentation of NASH. The therapeutic area in NASH is still in its infancy; therefore, there is an urgent need for well-conducted randomized controlled trials with relevant therapeutic endpoints [10].
Insulin sensitizers Insulin resistance is believed to be the main factor underlying liver fat buildup and possibly, through lipotoxic pathways, hepatic inflammation and fibrogenesis; improvement of insulin sensitivity is therefore a legitimate therapeutic target. Conversely, the presence of NAFLD in people with T2DM often makes it difficult to control diabetes: indeed, the intrahepatic triglyceride content is, in some studies, the major determinant of the daily requirements of insulin [11]. Moreover, NAFLD is independently associated with a twofold increased risk of T2DM. Consequently, treatment strategies decreasing intrahepatic triglyceride content and improving insulin sensitivity might contribute to better glycemic control [2]. In addition, as insulin resistance could favor the development of HCC, recent studies suggest that at least some antidiabetic drugs may reduce the risk of developing HCC [12]. This claim was made for metformin, although the data available so far are largely inconclusive from a methodological standpoint [13, 14]. Thiazolidinediones Glitazones are peroxisome proliferator-activated receptor gamma (PPARc) agonists with an insulin-sensitizing activity, currently used for treating T2DM. It has been suggested [15] but not confirmed [16] that glitazones strongly improve adipose tissue insulin resistance, which correlates with the reduction in steatosis and necroinflammation [15]. In controlled clinical trials, pioglitazone improved aminotransferase levels and histological features of liver injury (including resolution of steatohepatitis), except for fibrosis, in NASH patients with insulin resistance or T2DM [17], but also in nondiabetic NASH patients [18, 19]. The largest multicenter, randomized, controlled trial (RCT) (PIVENS) of pioglitazone (at the lower dose of 30 mg/day) in nondiabetic patients with NASH, lasting 2 years, failed to achieve statistically significant improvement of a complex composite primary endpoint that included a 2-point reduction in the NAFLD
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activity score (NAS)[19]. This could be explained by a higher level of statistical significance used in this three-arm trial or by methodological issues ultimately restricting the statistical power [20]. Despite a significant antisteatogenic effect and maintained reduction in alanine transaminase (ALT) levels, rosiglitazone, surprisingly, failed to improve steatohepatitis and its defining histological lesions [21, 22]. A recent meta-analysis of glitazones [23] concluded a histological benefit across the spectrum of histological lesions despite significant and non-reversible weight gain [24]. The use of both glitazones has been severely restricted by black-box warnings based on an increase in cardiovascular events [20], congestive heart failure [25], bone fractures in women [26] and risk of bladder cancer for pioglitazone [27] that justified its market withdrawal in France. While rosiglitazone has been largely withdrawn from the market, current guidelines recommend the use of pioglitazone for the treatment of steatohepatitis in patients with biopsy-proven NASH. However, the long-term safety and benefit is unknown, and its use requires caution in diabetic patients and in those with impaired cardiac function [6, 8]. Metformin Metformin is a safe antidiabetic drug that reduces hepatic glucose production and can induce weight loss. Earlier studies showed encouraging results on aminotransferases [28–30]. However, this drug has no clear benefit on liver histology [30–32], probably because of its limited antisteatogenic effect and its inability to restore adiponectin levels [33]. Possibly, weight loss on therapy might account for some of the discordant findings. Based on data from trials conducted so far, metformin is not recommended specifically for the treatment of NASH [6, 8]. Preclinical studies of glucagon-like peptide-1 agonists suggested these could be promising agents for the treatment of NAFLD [34–36].
Antioxidants and hepatoprotectants Improving insulin sensitivity may not be sufficient for improving liver injury [20]; therefore, specific hepatoprotective agents with antioxidant, antiinflammatory, antiapoptotic or other cytoprotective properties may be required either alone or in addition to insulin sensitizers. Vitamin E A few early studies have suggested that vitamin E (atocopherol), a natural antioxidant, improved liver enzymes, steatosis and histological features of NASH, except for
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fibrosis, when combined with pioglitazone [37] or ursodeoxycholic acid [38]. In the largest RCT to date, the PIVENS trial [19], vitamin E at 800 IU/day improved hepatocyte ballooning and inflammation and cleared steatohepatitis at a higher rate than placebo. These results were partly confirmed in a pediatric trial where vitamin E only improved hepatocyte ballooning and increased clearance of NASH but did not improve inflammation or ALT levels [39]. Interestingly, a 2-year treatment with vitamin E in combination with vitamin C did not result in histological improvement in another pediatric trial [40], thus dampening the enthusiasm for a universal benefit of this molecule in NASH. Another reason for concern is persisting doubt about the long-term safety of vitamin E supplements. Some studies suggest vitamin E might increase overall mortality [41] and the risks of hemorrhagic stroke [42] and prostate cancer [43]. While recommended as a first-line pharmacotherapy in non-diabetic adults with biopsy-proven NASH without cirrhosis [6], clearly more data on immediate efficacy and long-term safety are needed for more confident recommendations. Ursodeoxycholic acid (UDCA) A natural bile acid with hepatoprotective properties induced biochemical and histological improvement in two small RCTs at the dose of 13–15 mg/kg combined with vitamin E [38] or alone [44]. Unfortunately, UDCA alone failed to improve liver enzymes or hepatic histology in three other studies at low (13–15 mg/kg) [38, 45] or higher doses (23–28 mg/kg) [46]. These studies had, however, significant methodological shortcomings [47]. A French multicentric RCT has recently tested the effects of higher doses (28–35 mg/kg) of UDCA for 1 year in 126 NASH patients versus placebo [48]. There was an early, sustained and significant improvement of ALT levels and some suggestion of improvement of insulin resistance surrogate markers and fibrosis scores [48]. In the absence of positive studies with histological outcomes, UDCA is not universally recommended as a treatment for patients with NASH [6, 8]. Pentoxifylline It reduces TNF-a expression in animal models of NASH and has shown biochemical efficacy and histological improvement in a few uncontrolled trials of very small sample size [49]. A recent, 1-year, randomized controlled trial of 55 patients treated with 1,200 mg of pentoxifylline per day [50] has shown efficacy on liver enzymes and histological features of NASH, with even a marginal effect on fibrosis [50]. This beneficial effect could be correlated with decreased oxidized lipid products [51]. These
promising results from an early, small-size, monocentric RCT should be further validated.
Therapies with potential but yet unproven benefit in NASH Angiotensin II receptor antagonists (sartans) Experimental evidence suggests that losartan may have antifibrotic actions, possibly by inhibiting hepatic stellate cell activation [52]. In a rat model of NAFLD, losartan and telmisartan improved fatty liver [53]. While telmisartan, which also acts as a partial agonist of PPARc, improved insulin resistance, losartan could, in addition, reduce the expression of hepatic plasminogen activator inhibitor-1 (PAI-1) [53]. One pilot study in hypertensive patients with NASH has shown that 1 year of treatment with losartan reduced ALT levels, hepatic necroinflammation and fibrosis, but not steatosis [52]. The only randomized openlabel trial in NASH patients that tested 48 weeks of rosiglitazone combined with losartan, compared to rosiglitazone alone, showed that the combination therapy conferred no greater benefit with respect to histopathology [54]. Based on findings that a gene variant (A1166C) in the angiotensin II type 1 receptor predicts the therapeutic response to losartan [55], the possibility to predict response in NASH patient is appealing and should be further tested. Lipid-lowering drugs Statins There is no evidence for a beneficial effect of statins on liver histology in patients with NASH, except, possibly, for a marginal improvement in steatosis [56]. Hence, statins are not recommended for NASH treatment [6]. However, concurrent metabolic comorbidities clearly dictate the use of statins for cardiovascular endpoints in NAFLD patients. Altered liver function tests should not preclude the use of statins in these patients. Numerous studies have shown the safety of statins in regard to hepatotoxicity [57], as these drugs actually lower aminotransferases in patients with the metabolic syndrome [58]. In addition, experimental studies and preclinical observations suggest that statins might prevent portal hypertension and the development of HCC [59–62]. Indeed, by acting on both liver stem cells and endothelial cells, statins might specifically affect some of the main molecular pathways of HCC, such as inhibition of cell proliferation, induction of apoptosis and inhibition of angiogenesis [63]. One meta-analysis using a propensity score analysis has shown that statin use is associated with a
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reduced risk of HCC, most strongly in Asian but also in Western populations [64].
at designing potent peripheral CB1R blockers, but early studies in humans are not yet available.
Fibrates
Uric acid-lowering agents
These conventional triglyceride-lowering agents had no effect on aminotransferases or liver histology in patients with NAFLD [44].
The serum uric acid level is strongly associated with individual components of the metabolic syndrome, although a causal link is controversial [71–73]. Several observational studies have demonstrated a relationship between hyperuricemia and NAFLD [74]. In a large population-based study of nondiabetic adults from the USA, elevated serum uric acid was independently associated with the presence of elevated serum liver enzymes, with ultrasound-diagnosed NAFLD [75] and with a higher risk of cirrhosis-related hospitalization or death [76, 77]. These findings warrant further studies on the role of uric acid in NAFLD, but at this point it is entirely conjectural whether uric acid-lowering drugs could carry therapeutic potential for NASH.
Probucol It is a lipid-lowering agent that also has antioxidative properties. Merat et al. [65] showed that probucol (500 mg/ day) could significantly improve liver enzymes and even hepatic histology, except for fibrosis [66], in NASH patients. Larger RCTs are now required. Omega-3 fatty acids Omega-3 fatty acids are essential polyunsaturated fatty acids (PUFA), unable to be synthesised de novo, that have antiinflammatory and antioxidant properties. Patients with NAFLD have a lower dietary intake of these omega-3 fatty acids than healthy controls, leading to an increased omega6/omega-3 ratio. Downstream products of omega-6 are proinflammatory, increase lipogenesis and hepatic uptake of circulating free fatty acids (FFAs), and reduce hepatocyte fatty acid oxidation and synthesis of fatty acid transport proteins (i.e., VLDL), thus leading to steatosis. Therefore, dietary supplementation of omega-3 PUFAs has been suggested as a treatment of NAFLD. Currently approved in the USA to treat hypertriglyceridemia, omega3 fatty acids improve aminotransferases and steatosis in patients with NAFLD [67]. Two recent meta-analyses found experimental evidence to support their use and require well-conducted RCTs to confirm these results [68, 69]. To date, it is premature to recommend their use for the specific treatment of NAFLD, but they can be considered as the first-line agents to treat hypertriglyceridemia in patients with NAFLD [6]. The results of a large multicentric US study of eicosapentaenoic acid are awaited. Weight-loss agents Orlistat, an enteric lipase inhibitor, did not enhance weight loss or improve liver enzymes and histology [70], and this drug is not recommended for the treatment of NASH [6]. Rimonabant, a non-selective cannabinoid type 1 receptor blocker, was tested in two large international phase 3 trials in NASH patients. Unfortunately, because of severe psychiatric side effects, the marketing authorization was withdrawn, and the studies were halted. Current work aims
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Iron depletion While not a pharmacological agent in a strict sense, phlebotomy is a widely available and inexpensive therapy. Hyperferritinemia, which reflects hyperinsulinemia, is observed in about 20–30 % of patients with NAFLD [78] and is associated with the severity of liver damage [79]. Excessive body iron may play a causal role in insulin resistance through mechanisms that involve a reduced ability to oxidize carbohydrates and altered release of adipokines and beta-cell function [78]. Furthermore, it may facilitate the progression of cardiovascular disease by contributing to the recruitment and activation of macrophages within arterial lesions. Iron might also induce oxidative stress in hepatocytes, activation of HSCs and malignant transformation by promotion of cell growth and DNA damage [78]. In rats with fatty liver, iron depletion by deferoxamine upregulates glucose uptake, and increases insulin receptor activity and insulin signaling in hepatocytes [80, 81]. In humans, iron depletion by phlebotomy improved insulin resistance [81–85], liver iron content, NAS [82, 86, 87] and liver enzymes [83, 85]. It is unclear at this point whether and how iron depletion could be incorporated in the treatment strategies of patients with NASH [88]. Modulation of gut microbiota Gut microbiota is associated with obesity and the metabolic syndrome and could also play a role in the development of NASH, probably because of increased endotoxemia and activation of the Toll-like receptor-4 signaling cascade [89, 90]. Prebiotic modulation of gut microbiota in ob/ob mice
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lowers intestinal permeability and increases the endogenous glucagon-like peptide 2 (GLP-2) [91]. Ampicillin and neomycin improved insulin resistance in animals [92–94]. Based on these data, treatments aiming at manipulating enteric flora by using intestinally focused antibiotics, probiotics or prebiotics are proposed. In healthy [95] and in overweight and obese adults [96], it improves glucose tolerance compared to placebo. This effect was associated with weight loss and with an increased plasma GLP-1 and peptide YY concentrations. [95, 96]. Probiotics, which exhibit immunoregulatory and antiinflammatory activity, showed beneficial effects on liver fibrosis [90] in animal models of NASH. It could also improve steatosis and insulin resistance [97, 98], and preliminary data from two pilot studies suggest that they may be well tolerated and may improve conventional liver function tests [99, 100]. Randomized controlled clinical trials are now awaited [89, 101–103].
model of NAFLD, FXR agonist decreased ALT levels and significantly reduced inflammatory cell infiltration and hepatic fibrosis without changes in steatosis [111]. A large multicentric RCT of obeticholic acid, an FXR agonist with insulin-sensitizing properties, has completed enrollment in the US. Final results are expected in 2014. Caspase inhibitors GS-9450 is a selective caspase inhibitor that improved liver injury in murine models of NASH in particular in the methionine–choline deficient diet. In human trials, it reduced aminotransferases in patients with biopsy-proven NASH in a short-term phase 2 RCT [112], but development is currently halted because of concerns about possible hepatotoxicity. GFT505
L-carnitine
This is a modulator of mitochondrial FFA transport and boxidation, with antioxidative and antiinflammatory properties [104]. In an RCT performed on 74 NASH patients, Lcarnitine (2 g/day) supplementation to the diet for 24 weeks improved ALT, triglycerides and the lipid profile, HOMA-IR, TNFa, steatosis and NAS, compared with the placebo plus diet group [105]. These results are interesting but have not been reproduced so far. Thyromimetics Hypothyroidism is associated with the risk of developing the metabolic syndrome, and high-though-normal TSH values are independently associated with NASH in euthyroid patients [106]. Thyroid hormones such as T3 might have a prosteatogenic effect by increasing plasma FFA flux from the periphery to the liver and increasing hepatic lipogenesis [107, 108]. However, a specific agonist of the b isoform of the thyroid hormone receptor induced hepatic beta-oxidation of fatty acids resulting in a reduction of triacyl and diacyl glycerol in rodent models of hepatic steatosis [109]. This reduction in hepatic fat, especially if accompanied by an improvement of insulin resistance, could prove a useful pharmacological option to test in humans [110].
It is a dual PPAR alpha-delta agonist that improves the lipid profile and glycemic control in patients with atherogenic dyslipidemia and pre-diabetes [113]. It also has potent antiinflammatory and antifibrotic activities in several models of chronic murine liver injury. Contrary to PPAR gamma agonists, which can induce weight gain and cardiovascular events, its safety profile seems unremarkable so far. Based on these preclinical results and the results of several phase 2a trials in humans, a large international study of GFT-505 has been initiated in patients with NASH. Pre-clinical studies Pre-clinical studies have tested other therapeutic agents in mouse models of NAFLD. Administration of retinoids could improve insulin sensitivity through upregulation of hepatic leptin receptor expression [114]. Pharmacological inhibition of adipocyte fatty acid binding protein, a key mediator of inflammatory response in Kupffer cells, ameliorated hepatic steatosis, macrophage infiltration and hepatocyte ballooning [115]. Carbamazepine or rapamycin, which both enhance macroautophagy, could improve steatosis and insulin sensitivity by increasing lipid degradation [116]. Conclusion and perspectives
Agents in development The farnesoid X receptor (FXR) This is a member of the nuclear hormone receptor superfamily, which plays an essential role in the regulation of enterohepatic circulation and lipid homeostasis. In a mouse
Therapeutic research and drug development in NASH has only started, but will experience considerable traction because of the clear unmet medical need. While diet and lifestyle changes should always be the first-line therapy, current practice guidelines recommend the use of hepatic pharmacological therapy in patients with NASH and
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advanced fibrosis, without cirrhosis [6]. Despite limited results on efficacy and safety, vitamin E and pioglitazone are the recommended pharmacological agents, both of them being cost-effective in this indication [23]. Future therapeutic research in NASH should focus on exploring innovative molecules with better efficacy and a better safety profile. Because of the numerous comorbidities and heavy drug intake in patients with NASH, any new agent should not worsen, through off-target effects or drug-drug interactions, the concurrent diseases or therapeutic regimens. An individualized, dual approach of improving insulin resistance and conferring hepatoprotective, antiinflammatory and antifibrotic actions might be necessary for effective treatment of this complex condition. Compliance with Ethical Requirements This article is a review article and does not contain any studies with human or animal subjects. Conflict of interest V. R. served as a consultant for Axcan, Astellas, Enterome, GalMed, Genfit, Gilead and Roche. M. G. declared no conflict of interest.
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SUPPLEMENT ISSUE: NAFLD
Pharmacological agents for nonalcoholic steatohepatitis Maeva Guillaume • Vlad Ratziu
Received: 8 June 2013 / Accepted: 9 October 2013 Ó Asian Pacific Association for the Study of the Liver 2013
Abstract The rationale for specific pharmacologic therapy in nonalcoholic steatohepatitis (NASH) is determined by the potential for disease progression and the difficulties, in many patients, to successfully implement diet and lifestyle changes in the long term. Because they correct insulin resistance, insulin-sensitizing agents are attractive candidates for the treatment of NASH. However, two randomized studies have shown that vitamin E, despite having no effect on insulin sensitivity, achieves interesting histological and biochemical efficacy. This review provides an insight into the therapeutic efficacy and safety issues of different pharmacological agents tested in human NASH. Keywords Steatohepatitis Fibrosis Steatosis Glitazones Vitamin E Ursodesoxycholic acid Insulin resistance Oxidative stress Insulin sensitizers
M. Guillaume Service d’He´patologie et Gastro-ente´rologie, INSERM 1048 and Universite´ Paul Sabatier, Hoˆpital Purpan, Centre Hospitalier Universitaire de Toulouse, Toulouse, France e-mail: [email protected] M. Guillaume Service d’He´patologie et Gastro-ente´rologie, Hoˆpital Purpan, Pavillon Dieulafoy, 4e`me e´tage, Place du Docteur Baylac, TSA 40031, 31059 Toulouse Cedex 9, France V. Ratziu (&) INSERM U938, Hospital Pitie´ Salpeˆtrie`re, CdR Saint-Antoine and Universite´ Pierre et Marie Curie, Paris, France e-mail: [email protected] V. Ratziu Service d’He´patologie et Gastro-ente´rologie, Hoˆpital Pitie´Salpeˆtrie`re, 47-83, Boulevard de l’hoˆpital, 75651 Paris Cedex 13, France
Introduction Nonalcoholic fatty liver disease (NAFLD) encompasses the entire spectrum of fatty liver disease in individuals without significant alcohol consumption, ranging from nonalcoholic fatty liver (NAFL) to nonalcoholic steatohepatitis (NASH) and cirrhosis. NAFL is defined as the presence of hepatic steatosis with minimal, non-specific inflammation that does not qualify for steatohepatitis; NASH is defined as the presence of lobular inflammation with hepatocyte injury (ballooning) [1]. The risk of progression to cirrhosis, liver failure and liver cancer is minimal for NAFL and real for NASH [2]. In addition, there is growing evidence to support an association between NAFLD and an increased risk of HCC [3]. Patients with NASH have increased liverrelated mortality as well as cardiovascular mortality [4]. The recommended first-line therapy for all NAFLD patients is non-pharmacological: weight reduction through diet, changes in lifestyle, physical exercise and proscribing sedentarity [5, 6]. Additionally, concurrent metabolic disorders such as type 2 diabetes (T2DM), hyperlipidemia or arterial hypertension, when present, should be well controlled, although their specific impact on improving the hepatic disease has not yet been determined. Occasionally, these measures result in a 7–10 % weight loss, a threshold that is possibly associated with histological improvement [5, 6]. Moreover, as insulin resistance, inflammation, an unbalanced adipocytokine ratio and angiogenic factors associated with NAFLD could favor the development of HCC [7], these traditional therapeutic approaches in NAFLD may theoretically reduce the risk of HCC by acting on common pathophysiological pathways shared by NAFLD and HCC. In patients who failed these measures or in those with advanced disease (NASH with significant fibrosis), pharmacological treatments specifically directed at
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improving hepatic inflammation, fibrosis and/or clearing steatohepatitis might be necessary [8]. In some carefully selected patients with morbid obesity and without cirrhosis, bariatric surgery may be proposed [9]. Here we will provide an overview of data obtained so far with pharmacological agents in NASH mostly focusing on controlled studies with histological documentation of NASH. The therapeutic area in NASH is still in its infancy; therefore, there is an urgent need for well-conducted randomized controlled trials with relevant therapeutic endpoints [10].
Insulin sensitizers Insulin resistance is believed to be the main factor underlying liver fat buildup and possibly, through lipotoxic pathways, hepatic inflammation and fibrogenesis; improvement of insulin sensitivity is therefore a legitimate therapeutic target. Conversely, the presence of NAFLD in people with T2DM often makes it difficult to control diabetes: indeed, the intrahepatic triglyceride content is, in some studies, the major determinant of the daily requirements of insulin [11]. Moreover, NAFLD is independently associated with a twofold increased risk of T2DM. Consequently, treatment strategies decreasing intrahepatic triglyceride content and improving insulin sensitivity might contribute to better glycemic control [2]. In addition, as insulin resistance could favor the development of HCC, recent studies suggest that at least some antidiabetic drugs may reduce the risk of developing HCC [12]. This claim was made for metformin, although the data available so far are largely inconclusive from a methodological standpoint [13, 14]. Thiazolidinediones Glitazones are peroxisome proliferator-activated receptor gamma (PPARc) agonists with an insulin-sensitizing activity, currently used for treating T2DM. It has been suggested [15] but not confirmed [16] that glitazones strongly improve adipose tissue insulin resistance, which correlates with the reduction in steatosis and necroinflammation [15]. In controlled clinical trials, pioglitazone improved aminotransferase levels and histological features of liver injury (including resolution of steatohepatitis), except for fibrosis, in NASH patients with insulin resistance or T2DM [17], but also in nondiabetic NASH patients [18, 19]. The largest multicenter, randomized, controlled trial (RCT) (PIVENS) of pioglitazone (at the lower dose of 30 mg/day) in nondiabetic patients with NASH, lasting 2 years, failed to achieve statistically significant improvement of a complex composite primary endpoint that included a 2-point reduction in the NAFLD
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activity score (NAS)[19]. This could be explained by a higher level of statistical significance used in this three-arm trial or by methodological issues ultimately restricting the statistical power [20]. Despite a significant antisteatogenic effect and maintained reduction in alanine transaminase (ALT) levels, rosiglitazone, surprisingly, failed to improve steatohepatitis and its defining histological lesions [21, 22]. A recent meta-analysis of glitazones [23] concluded a histological benefit across the spectrum of histological lesions despite significant and non-reversible weight gain [24]. The use of both glitazones has been severely restricted by black-box warnings based on an increase in cardiovascular events [20], congestive heart failure [25], bone fractures in women [26] and risk of bladder cancer for pioglitazone [27] that justified its market withdrawal in France. While rosiglitazone has been largely withdrawn from the market, current guidelines recommend the use of pioglitazone for the treatment of steatohepatitis in patients with biopsy-proven NASH. However, the long-term safety and benefit is unknown, and its use requires caution in diabetic patients and in those with impaired cardiac function [6, 8]. Metformin Metformin is a safe antidiabetic drug that reduces hepatic glucose production and can induce weight loss. Earlier studies showed encouraging results on aminotransferases [28–30]. However, this drug has no clear benefit on liver histology [30–32], probably because of its limited antisteatogenic effect and its inability to restore adiponectin levels [33]. Possibly, weight loss on therapy might account for some of the discordant findings. Based on data from trials conducted so far, metformin is not recommended specifically for the treatment of NASH [6, 8]. Preclinical studies of glucagon-like peptide-1 agonists suggested these could be promising agents for the treatment of NAFLD [34–36].
Antioxidants and hepatoprotectants Improving insulin sensitivity may not be sufficient for improving liver injury [20]; therefore, specific hepatoprotective agents with antioxidant, antiinflammatory, antiapoptotic or other cytoprotective properties may be required either alone or in addition to insulin sensitizers. Vitamin E A few early studies have suggested that vitamin E (atocopherol), a natural antioxidant, improved liver enzymes, steatosis and histological features of NASH, except for
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fibrosis, when combined with pioglitazone [37] or ursodeoxycholic acid [38]. In the largest RCT to date, the PIVENS trial [19], vitamin E at 800 IU/day improved hepatocyte ballooning and inflammation and cleared steatohepatitis at a higher rate than placebo. These results were partly confirmed in a pediatric trial where vitamin E only improved hepatocyte ballooning and increased clearance of NASH but did not improve inflammation or ALT levels [39]. Interestingly, a 2-year treatment with vitamin E in combination with vitamin C did not result in histological improvement in another pediatric trial [40], thus dampening the enthusiasm for a universal benefit of this molecule in NASH. Another reason for concern is persisting doubt about the long-term safety of vitamin E supplements. Some studies suggest vitamin E might increase overall mortality [41] and the risks of hemorrhagic stroke [42] and prostate cancer [43]. While recommended as a first-line pharmacotherapy in non-diabetic adults with biopsy-proven NASH without cirrhosis [6], clearly more data on immediate efficacy and long-term safety are needed for more confident recommendations. Ursodeoxycholic acid (UDCA) A natural bile acid with hepatoprotective properties induced biochemical and histological improvement in two small RCTs at the dose of 13–15 mg/kg combined with vitamin E [38] or alone [44]. Unfortunately, UDCA alone failed to improve liver enzymes or hepatic histology in three other studies at low (13–15 mg/kg) [38, 45] or higher doses (23–28 mg/kg) [46]. These studies had, however, significant methodological shortcomings [47]. A French multicentric RCT has recently tested the effects of higher doses (28–35 mg/kg) of UDCA for 1 year in 126 NASH patients versus placebo [48]. There was an early, sustained and significant improvement of ALT levels and some suggestion of improvement of insulin resistance surrogate markers and fibrosis scores [48]. In the absence of positive studies with histological outcomes, UDCA is not universally recommended as a treatment for patients with NASH [6, 8]. Pentoxifylline It reduces TNF-a expression in animal models of NASH and has shown biochemical efficacy and histological improvement in a few uncontrolled trials of very small sample size [49]. A recent, 1-year, randomized controlled trial of 55 patients treated with 1,200 mg of pentoxifylline per day [50] has shown efficacy on liver enzymes and histological features of NASH, with even a marginal effect on fibrosis [50]. This beneficial effect could be correlated with decreased oxidized lipid products [51]. These
promising results from an early, small-size, monocentric RCT should be further validated.
Therapies with potential but yet unproven benefit in NASH Angiotensin II receptor antagonists (sartans) Experimental evidence suggests that losartan may have antifibrotic actions, possibly by inhibiting hepatic stellate cell activation [52]. In a rat model of NAFLD, losartan and telmisartan improved fatty liver [53]. While telmisartan, which also acts as a partial agonist of PPARc, improved insulin resistance, losartan could, in addition, reduce the expression of hepatic plasminogen activator inhibitor-1 (PAI-1) [53]. One pilot study in hypertensive patients with NASH has shown that 1 year of treatment with losartan reduced ALT levels, hepatic necroinflammation and fibrosis, but not steatosis [52]. The only randomized openlabel trial in NASH patients that tested 48 weeks of rosiglitazone combined with losartan, compared to rosiglitazone alone, showed that the combination therapy conferred no greater benefit with respect to histopathology [54]. Based on findings that a gene variant (A1166C) in the angiotensin II type 1 receptor predicts the therapeutic response to losartan [55], the possibility to predict response in NASH patient is appealing and should be further tested. Lipid-lowering drugs Statins There is no evidence for a beneficial effect of statins on liver histology in patients with NASH, except, possibly, for a marginal improvement in steatosis [56]. Hence, statins are not recommended for NASH treatment [6]. However, concurrent metabolic comorbidities clearly dictate the use of statins for cardiovascular endpoints in NAFLD patients. Altered liver function tests should not preclude the use of statins in these patients. Numerous studies have shown the safety of statins in regard to hepatotoxicity [57], as these drugs actually lower aminotransferases in patients with the metabolic syndrome [58]. In addition, experimental studies and preclinical observations suggest that statins might prevent portal hypertension and the development of HCC [59–62]. Indeed, by acting on both liver stem cells and endothelial cells, statins might specifically affect some of the main molecular pathways of HCC, such as inhibition of cell proliferation, induction of apoptosis and inhibition of angiogenesis [63]. One meta-analysis using a propensity score analysis has shown that statin use is associated with a
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reduced risk of HCC, most strongly in Asian but also in Western populations [64].
at designing potent peripheral CB1R blockers, but early studies in humans are not yet available.
Fibrates
Uric acid-lowering agents
These conventional triglyceride-lowering agents had no effect on aminotransferases or liver histology in patients with NAFLD [44].
The serum uric acid level is strongly associated with individual components of the metabolic syndrome, although a causal link is controversial [71–73]. Several observational studies have demonstrated a relationship between hyperuricemia and NAFLD [74]. In a large population-based study of nondiabetic adults from the USA, elevated serum uric acid was independently associated with the presence of elevated serum liver enzymes, with ultrasound-diagnosed NAFLD [75] and with a higher risk of cirrhosis-related hospitalization or death [76, 77]. These findings warrant further studies on the role of uric acid in NAFLD, but at this point it is entirely conjectural whether uric acid-lowering drugs could carry therapeutic potential for NASH.
Probucol It is a lipid-lowering agent that also has antioxidative properties. Merat et al. [65] showed that probucol (500 mg/ day) could significantly improve liver enzymes and even hepatic histology, except for fibrosis [66], in NASH patients. Larger RCTs are now required. Omega-3 fatty acids Omega-3 fatty acids are essential polyunsaturated fatty acids (PUFA), unable to be synthesised de novo, that have antiinflammatory and antioxidant properties. Patients with NAFLD have a lower dietary intake of these omega-3 fatty acids than healthy controls, leading to an increased omega6/omega-3 ratio. Downstream products of omega-6 are proinflammatory, increase lipogenesis and hepatic uptake of circulating free fatty acids (FFAs), and reduce hepatocyte fatty acid oxidation and synthesis of fatty acid transport proteins (i.e., VLDL), thus leading to steatosis. Therefore, dietary supplementation of omega-3 PUFAs has been suggested as a treatment of NAFLD. Currently approved in the USA to treat hypertriglyceridemia, omega3 fatty acids improve aminotransferases and steatosis in patients with NAFLD [67]. Two recent meta-analyses found experimental evidence to support their use and require well-conducted RCTs to confirm these results [68, 69]. To date, it is premature to recommend their use for the specific treatment of NAFLD, but they can be considered as the first-line agents to treat hypertriglyceridemia in patients with NAFLD [6]. The results of a large multicentric US study of eicosapentaenoic acid are awaited. Weight-loss agents Orlistat, an enteric lipase inhibitor, did not enhance weight loss or improve liver enzymes and histology [70], and this drug is not recommended for the treatment of NASH [6]. Rimonabant, a non-selective cannabinoid type 1 receptor blocker, was tested in two large international phase 3 trials in NASH patients. Unfortunately, because of severe psychiatric side effects, the marketing authorization was withdrawn, and the studies were halted. Current work aims
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Iron depletion While not a pharmacological agent in a strict sense, phlebotomy is a widely available and inexpensive therapy. Hyperferritinemia, which reflects hyperinsulinemia, is observed in about 20–30 % of patients with NAFLD [78] and is associated with the severity of liver damage [79]. Excessive body iron may play a causal role in insulin resistance through mechanisms that involve a reduced ability to oxidize carbohydrates and altered release of adipokines and beta-cell function [78]. Furthermore, it may facilitate the progression of cardiovascular disease by contributing to the recruitment and activation of macrophages within arterial lesions. Iron might also induce oxidative stress in hepatocytes, activation of HSCs and malignant transformation by promotion of cell growth and DNA damage [78]. In rats with fatty liver, iron depletion by deferoxamine upregulates glucose uptake, and increases insulin receptor activity and insulin signaling in hepatocytes [80, 81]. In humans, iron depletion by phlebotomy improved insulin resistance [81–85], liver iron content, NAS [82, 86, 87] and liver enzymes [83, 85]. It is unclear at this point whether and how iron depletion could be incorporated in the treatment strategies of patients with NASH [88]. Modulation of gut microbiota Gut microbiota is associated with obesity and the metabolic syndrome and could also play a role in the development of NASH, probably because of increased endotoxemia and activation of the Toll-like receptor-4 signaling cascade [89, 90]. Prebiotic modulation of gut microbiota in ob/ob mice
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lowers intestinal permeability and increases the endogenous glucagon-like peptide 2 (GLP-2) [91]. Ampicillin and neomycin improved insulin resistance in animals [92–94]. Based on these data, treatments aiming at manipulating enteric flora by using intestinally focused antibiotics, probiotics or prebiotics are proposed. In healthy [95] and in overweight and obese adults [96], it improves glucose tolerance compared to placebo. This effect was associated with weight loss and with an increased plasma GLP-1 and peptide YY concentrations. [95, 96]. Probiotics, which exhibit immunoregulatory and antiinflammatory activity, showed beneficial effects on liver fibrosis [90] in animal models of NASH. It could also improve steatosis and insulin resistance [97, 98], and preliminary data from two pilot studies suggest that they may be well tolerated and may improve conventional liver function tests [99, 100]. Randomized controlled clinical trials are now awaited [89, 101–103].
model of NAFLD, FXR agonist decreased ALT levels and significantly reduced inflammatory cell infiltration and hepatic fibrosis without changes in steatosis [111]. A large multicentric RCT of obeticholic acid, an FXR agonist with insulin-sensitizing properties, has completed enrollment in the US. Final results are expected in 2014. Caspase inhibitors GS-9450 is a selective caspase inhibitor that improved liver injury in murine models of NASH in particular in the methionine–choline deficient diet. In human trials, it reduced aminotransferases in patients with biopsy-proven NASH in a short-term phase 2 RCT [112], but development is currently halted because of concerns about possible hepatotoxicity. GFT505
L-carnitine
This is a modulator of mitochondrial FFA transport and boxidation, with antioxidative and antiinflammatory properties [104]. In an RCT performed on 74 NASH patients, Lcarnitine (2 g/day) supplementation to the diet for 24 weeks improved ALT, triglycerides and the lipid profile, HOMA-IR, TNFa, steatosis and NAS, compared with the placebo plus diet group [105]. These results are interesting but have not been reproduced so far. Thyromimetics Hypothyroidism is associated with the risk of developing the metabolic syndrome, and high-though-normal TSH values are independently associated with NASH in euthyroid patients [106]. Thyroid hormones such as T3 might have a prosteatogenic effect by increasing plasma FFA flux from the periphery to the liver and increasing hepatic lipogenesis [107, 108]. However, a specific agonist of the b isoform of the thyroid hormone receptor induced hepatic beta-oxidation of fatty acids resulting in a reduction of triacyl and diacyl glycerol in rodent models of hepatic steatosis [109]. This reduction in hepatic fat, especially if accompanied by an improvement of insulin resistance, could prove a useful pharmacological option to test in humans [110].
It is a dual PPAR alpha-delta agonist that improves the lipid profile and glycemic control in patients with atherogenic dyslipidemia and pre-diabetes [113]. It also has potent antiinflammatory and antifibrotic activities in several models of chronic murine liver injury. Contrary to PPAR gamma agonists, which can induce weight gain and cardiovascular events, its safety profile seems unremarkable so far. Based on these preclinical results and the results of several phase 2a trials in humans, a large international study of GFT-505 has been initiated in patients with NASH. Pre-clinical studies Pre-clinical studies have tested other therapeutic agents in mouse models of NAFLD. Administration of retinoids could improve insulin sensitivity through upregulation of hepatic leptin receptor expression [114]. Pharmacological inhibition of adipocyte fatty acid binding protein, a key mediator of inflammatory response in Kupffer cells, ameliorated hepatic steatosis, macrophage infiltration and hepatocyte ballooning [115]. Carbamazepine or rapamycin, which both enhance macroautophagy, could improve steatosis and insulin sensitivity by increasing lipid degradation [116]. Conclusion and perspectives
Agents in development The farnesoid X receptor (FXR) This is a member of the nuclear hormone receptor superfamily, which plays an essential role in the regulation of enterohepatic circulation and lipid homeostasis. In a mouse
Therapeutic research and drug development in NASH has only started, but will experience considerable traction because of the clear unmet medical need. While diet and lifestyle changes should always be the first-line therapy, current practice guidelines recommend the use of hepatic pharmacological therapy in patients with NASH and
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advanced fibrosis, without cirrhosis [6]. Despite limited results on efficacy and safety, vitamin E and pioglitazone are the recommended pharmacological agents, both of them being cost-effective in this indication [23]. Future therapeutic research in NASH should focus on exploring innovative molecules with better efficacy and a better safety profile. Because of the numerous comorbidities and heavy drug intake in patients with NASH, any new agent should not worsen, through off-target effects or drug-drug interactions, the concurrent diseases or therapeutic regimens. An individualized, dual approach of improving insulin resistance and conferring hepatoprotective, antiinflammatory and antifibrotic actions might be necessary for effective treatment of this complex condition. Compliance with Ethical Requirements This article is a review article and does not contain any studies with human or animal subjects. Conflict of interest V. R. served as a consultant for Axcan, Astellas, Enterome, GalMed, Genfit, Gilead and Roche. M. G. declared no conflict of interest.
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