Guidance for Industry 

Drug-Induced Liver Injury: 

Premarketing Clinical 

Evaluation 

U.S. Department of Health and Human Services 

Food and Drug Administration 

Center for Drug Evaluation and Research (CDER) 

Center for Biologics Evaluation and Research (CBER) 

July 2009 

Drug Safety 
Guidance for Industry 

Drug-Induced Liver Injury: 

Premarketing Clinical 

Evaluation 

Additional copies are available from: 

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Center for Drug Evaluation and Research 

Food and Drug Administration 

10903 New Hampshire Ave., Bldg. 51, rm. 2201 

Silver Spring, MD 20993-0002 

Tel: 301-796-3400; Fax: 301-847-8714; E-mail: druginfo@fda.hhs.gov 

http://www.fda.gov/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/default.htm 

or 

Office of Communication, Outreach, and Development, HFM-40 

Center for Biologics Evaluation and Research 

Food and Drug Administration 

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Tel: 800-835-4709 or 301-827-1800

http://www.fda.gov/BiologicsBloodVaccines/GuidanceComplianceRegulatoryInformation/Guidances/default.htm 

U.S. Department of Health and Human Services 

Food and Drug Administration 

Center for Drug Evaluation and Research (CDER) 

Center for Biologics Evaluation and Research (CBER) 

July 2009 

Drug Safety 
TABLE OF CONTENTS 

I. INTRODUCTION............................................................................................................. 1

II. BACKGROUND: DILI ................................................................................................... 2

III. SIGNALS OF DILI AND HY’S LAW............................................................................ 3

IV. CLINICAL EVALUATION OF DILI ............................................................................ 7

A. General Considerations.................................................................................................................7

1. Patients with Liver Abnormalities or Disease .................................................................................7

2. Detection of DILI .............................................................................................................................8

3. Confirmation....................................................................................................................................8

4. Close Observation............................................................................................................................9

5. Decision to Stop Drug Administration.............................................................................................9

6. Evaluating Data for Alternative Causes ........................................................................................10

7. Follow-Up to Resolution................................................................................................................11

8. Rechallenge....................................................................................................................................12

B. Research Opportunities...............................................................................................................12

C. Case Report Forms ......................................................................................................................13

D. Interpretation of Signals of DILI or Acute Liver Failure ........................................................14

1. Frequency and Magnitude of Liver AT Abnormalities ..................................................................14

2. Combined Elevations of Aminotransferases and Bilirubin............................................................14

E. Analysis of Signals of DILI..........................................................................................................15

1. Assessment of Drug Metabolism....................................................................................................15

2. Assessment of Liver-Related Adverse Events in Controlled Trials................................................15

3. Assessment of Liver-Related Adverse Events in the Entire Clinical Trials Database ...................16

4. Assessment of Hy’s Law Cases in the Clinical Trials Database....................................................16

5. Overall Assessment of a Drug’s Potential to Cause DILI .............................................................17

REFERENCES............................................................................................................................ 19

APPENDIX A: ILLUSTRATIVE EXAMPLES OF DILI..................................................... 23
Contains Nonbinding Recommendations

Guidance for Industry1

Drug-Induced Liver Injury: 

Premarketing Clinical Evaluation 

This guidance represents the Food and Drug Administration’s (FDA’s) current thinking on this topic. It 

does not create or confer any rights for or on any person and does not operate to bind FDA or the public. 

You can use an alternative approach if the approach satisfies the requirements of the applicable statutes 

and regulations. If you want to discuss an alternative approach, contact the FDA staff responsible for 

implementing this guidance. If you cannot identify the appropriate FDA staff, call the appropriate 

number listed on the title page of this guidance. 

I. INTRODUCTION 

This guidance is intended to assist the pharmaceutical industry and other investigators who are 

conducting new drug development in assessing the potential for a drug2 to cause severe liver 

injury (i.e., irreversible liver failure that is fatal or requires liver transplantation). In particular, 

the guidance addresses how laboratory measurements that signal the potential for such druginduced liver injury (DILI) can be obtained and evaluated during drug development. This 

evaluation is important because most drugs that cause severe DILI do so infrequently; typical 

drug development databases with up to a few thousand subjects exposed to a new drug will not 

show any cases. Databases may, however, show evidence or signals of a drug’s potential for 

severe DILI if the clinical and laboratory data are properly evaluated for evidence of lesser injury 

that may not be severe, but may predict the ability to cause more severe injuries. This guidance 

describes an approach that can be used to distinguish signals of DILI that identify drugs likely to 

cause severe liver injury from signals that do not suggest such a potential. This guidance does 

not address issues of preclinical evaluation for signals of DILI, nor the detection and assessment 

of DILI after drug approval and marketing. 

FDA’s guidance documents, including this guidance, do not establish legally enforceable 

responsibilities. Instead, guidances describe the Agency’s current thinking on a topic and should 

be viewed only as recommendations, unless specific regulatory or statutory requirements are 

 

1

 This guidance has been prepared by the Division of Gastroenterology Products in the Office of New Drugs, the 

Office of Medical Policy, and the Office of Surveillance and Epidemiology in the Center for Drug Evaluation and 

Research (CDER) in cooperation with the Center for Biologics Evaluation and Research (CBER) at the Food and 

Drug Administration (FDA). 

2

 This guidance uses the term drug or product to refer to all products, except whole blood and blood components, 

regulated by CDER and CBER, including vaccines, and uses the term approval to refer to both drug approval and 

biologic licensure. 

1 
Contains Nonbinding Recommendations

cited. The use of the word should in Agency guidances means that something is suggested or 

recommended, but not required. 

II. BACKGROUND: DILI 

DILI has been the most frequent single cause of safety-related drug marketing withdrawals for 

the past 50 years (e.g., iproniazid), continuing to the present (e.g., ticrynafen, benoxaprofen, 

bromfenac, troglitazone, nefazodone). Hepatotoxicity discovered after approval for marketing 

also has limited the use of many drugs, including isoniazid, labetalol, trovafloxacin, tolcapone, 

and felbamate (Temple 2001). Several drugs have not been approved in the United States 

because European marketing experience revealed their hepatotoxicity (e.g., ibufenac, 

perhexiline, alpidem). Finally, some drugs were not approved in the United States because 

premarketing experience provided evidence of the potential for severe DILI (e.g., dilevalol, 

tasosartan, ximelagatran). Although most significant hepatotoxins have caused predominantly 

hepatocellular injury, indicated by leakage of aminotransferase (AT) enzymes from injured liver 

cells without prominent evidence of hepatobiliary obstruction or intrahepatic cholestasis, the 

pattern of injury can vary. Many drugs cause cholestasis, but in general this condition is 

reversible after administration of the offending drug has stopped. Cholestatic injuries are less 

likely to lead to death or transplant, although there have been exceptions. 

Drugs cause liver injuries by many different mechanisms. These injuries resemble almost all 

known liver diseases and there are no pathognomonic findings, even upon liver biopsy, that 

make diagnosis of DILI certain. Therefore, when possible DILI is suspected, it is essential to 

gather additional clinical and laboratory information necessary for differential diagnosis of the 

cause. It is important to observe the time course of the injury, and to seek alternative causes of 

the liver injury, such as acute viral hepatitis A, B, or C; concomitant use of a hepatotoxic drug or 

exposure to hepatotoxins; autoimmune or alcoholic hepatitis; biliary tract disorders; and 

circulatory problems of hypotension or right heart congestive failure that may cause ischemic or 

hypoxic hepatopathy. It is also prudent to assess the subject for previously existing liver disease, 

such as chronic hepatitis C or nonalcoholic steatohepatitis (NASH), that may or may not have 

been recognized before exposure to the experimental drug. It should be recognized that DILI 

may occur also in persons with preexisting liver disease as a superimposed problem. 

Only the most overt hepatotoxins can be expected to show cases of severe DILI in the 1,000 to 

3,000 subjects typically studied and described in a new drug application (NDA). Overtly 

hepatotoxic agents (e.g., carbon tetrachloride, chloroform, methylene chloride) are toxic to 

anyone receiving a large enough dose, and drugs that cause such predictable and dose-related 

injury generally are discovered and rejected in preclinical testing. More difficult to detect is 

toxicity that is not predictable or clearly dose-related that occurs at doses well tolerated by most 

people, but seems to depend on individual susceptibilities that have not as yet been characterized. 

Most of the drugs withdrawn from the market for hepatotoxicity have caused death or 

transplantation at frequencies in the range of ≤1 per 10,000, so that a single case of such an event 

rarely would be found even if several thousand subjects were studied. Severe DILI cases rarely 

have been seen in drug development programs of significantly hepatotoxic drugs. 

2 
Contains Nonbinding Recommendations

What are often seen during drug development are mild elevations of serum aminotransferases, 

usually without any symptoms. The problem is that these types of signals can be generated by 

drugs that are capable of causing severe DILI as well as by drugs that have a low potential for 

causing severe injury (e.g., aspirin, tacrine, heparin, hydroxyl-methylglutaryl coenzyme Areductase inhibitors (statins)). Therefore, an approach is needed that can distinguish drugs likely 

to cause severe DILI from drugs unlikely to do so. 

In general, the type of liver injury that leads to severe DILI is a predominantly hepatocellular 

injury. Hepatocellular injury is indicated by rises in AT activities in serum reflecting release of 

alanine or aspartate aminotransferase (ALT or AST) from injured liver cells. The ability to cause 

some hepatocellular injury, however, is not a reliable predictor of a drug’s potential for severe 

DILI. Many drugs that cause transient rises in serum AT activity do not cause progressive or 

severe DILI, even if drug administration is continued. It is only those drugs that can cause 

hepatocellular injury extensive enough to reduce the liver’s functional ability to clear bilirubin 

from the plasma or to synthesize prothrombin and other coagulation factors that cause severe 

DILI. It is important to identify those drugs as early as possible. 

The drugs that have caused severe DILI in humans have not shown clear hepatotoxicity in 

animals, generally have not shown dose-related toxicity, and, as noted, generally have caused 

low rates of severe injury in humans (1 in 5,000 to 10,000 or less). One of the few exceptions to 

these findings is acetaminophen, whose toxicity can be shown in animal models and whose 

toxicity is clearly dose-related. These reactions thus appear to reflect host factors and individual 

susceptibility. Consequently, they have been termed idiosyncratic, meaning dependent upon the 

individual person’s particular constitution. Whether they are the result of genetic and/or 

acquired differences has not yet been established, and to date no genetic, metabolic, or other 

characteristic has been found to reliably predict severe DILI in an individual. 

Some severe DILI examples have presented differently from the more commonly seen 

hepatocellular idiosyncratic type. Perhexiline, an anti-anginal drug marketed in Europe, 

produced toxicity within months of starting the drug that had the histological appearance of 

alcoholic cirrhosis (Pessayre and Biachara et al. 1979). Fialuridine caused modest acute liver 

injury, but most strikingly led to severe metabolic acidosis and multiorgan failure as 

mitochondrial oxidative capacity was obliterated over a period of months (Kleiner and Gaffey et 

al. 1997; Semino-Mora and Leon-Monzon et al. 1997). Valproic acid causes hyperammonemic 

encephalopathy even without notable rises in serum AT activities. Benoxaprofen (Oraflex) 

induced intrahepatic cholestasis that over many months led to significant, sometimes fatal, liver 

injury, especially in elderly patients (Taggart and Alderdice 1982). 

Past experience indicates that appropriate testing and analysis in premarketing trials can detect 

drugs that can cause severe hepatocellular injury. 

III. SIGNALS OF DILI AND HY’S LAW 

Hepatocellular injury (usually detected by serum AT elevations) can be caused by drugs that 

rarely, if ever, cause severe DILI (e.g., aspirin, tacrine, statins, and heparin) as well as by drugs 

3 
Contains Nonbinding Recommendations

that do cause such injury. Evidence of hepatocellular injury is thus a necessary, but not 

sufficient, signal of the potential to cause severe DILI (note, however, that the drugs causing 

hepatic injury through mitochondrial toxicity may not cause early hepatotoxicity). The 

frequency of serum AT elevations also is not a good indicator of a potential for severe DILI, 

because drugs such as tacrine (not a cause of severe DILI) can cause AT elevations in as many as 

50 percent of patients. Very high levels of observed ATs may be a somewhat better indicator of 

potential for severe DILI, but the most specific indicator is evidence of altered liver function

accompanying or promptly following evidence of hepatocellular injury (see below). 

As noted, a typical NDA or biologics license application (BLA) database usually will not show 

any cases of severe DILI, even for a drug that can cause such injury, because the rate of severe 

injury is usually relatively low (1/10,000 or less). Many drugs, however, including both 

significant hepatotoxins and drugs that do not cause severe liver injury, cause laboratory 

evidence of mild, transient hepatic injury, with leakage of liver enzymes and the appearance in 

serum of elevations in AT activities to levels of 3, 5, and sometimes greater than 5 times the 

upper limits of normal (ULN). Generally, ALT is considered a somewhat more liver-specific 

aminotransferase enzyme than AST, although it also occurs in many tissues (Green and Flamm 

2002). The finding of a higher rate of such elevations in drug-treated subjects than in a control 

group is a sensitive signal of a potential to cause severe DILI, but it is not a specific signal. 

A more specific signal of such potential is a higher rate of more marked peak AT elevations 

(10x-, 15xULN), with cases of increases to >1,000 U/L causing increased concern. The single 

clearest (most specific) predictor found to date of a drug’s potential for severe hepatotoxicity, 

however, is the occurrence of a small number of cases of hepatocellular injury (aminotransferase 

elevation) accompanied by increased serum total bilirubin (TBL), not explained by any other 

cause, such as viral hepatitis or exposure to other hepatotoxins, and without evidence of 

cholestasis, together with an increased incidence of AT elevations in the overall trial population 

compared to control. Increased plasma prothrombin time, or its international normalized ratio 

(INR), a consequence of reduced hepatic production of Vitamin K-dependent clotting factors, is 

another potentially useful measure of liver function that might suggest the potential for severe 

liver injury. 

Recognition of the importance of altered liver function, in addition to liver injury, began with 

Zimmerman’s observation that drug-induced hepatocellular injury (i.e., aminotransferase 

elevation) accompanied by jaundice had a poor prognosis, with a 10 to 50 percent mortality from 

acute liver failure (in pretransplantation days) (Zimmerman 1978, 1999). The reason for this 

now seems clear. Because the liver has a large excess of bilirubin-excreting capacity, injury to 

hepatocytes sufficient to cause jaundice or even mild hyperbilirubinemia (i.e., a bilirubin 

>2xULN) represents an extent of liver injury so great that recovery may not be possible in some 

patients. Zimmerman’s observation that hepatocellular injury sufficient to impair bilirubin 

excretion was ominous has been used at the Food and Drug Administration (FDA) over the years 

to identify drugs likely to be capable of causing severe liver injury. The observation of the 

critical importance of altered liver function has been referred to informally as Hy’s Law (Temple 

2001; Reuben 2004). 

4 
Contains Nonbinding Recommendations

Hy’s Law is essentially a translation of Zimmerman’s observation that pure hepatocellular injury 

sufficient to cause hyperbilirubinemia is an ominous indicator of the potential for a drug to cause 

serious liver injury. Thus, a finding of ALT elevation, usually substantial, seen concurrently 

with bilirubin >2xULN, identifies a drug likely to cause severe DILI (fatal or requiring 

transplant) at a rate roughly 1/10 the rate of Hy’s Law cases. It is critical to rule out other causes 

of injury (e.g., other drugs or viral hepatitis) and to rule out an obstructive basis for the elevated 

bilirubin, so that alkaline phosphatase (ALP) should not be substantially elevated. In all cases to 

date, the small number of Hy’s Law cases has arisen on a background of an increased incidence 

of more modest signs of hepatocellular injury (e.g., greater incidence of 3xULN elevations in AT 

than seen in a control group). 

Briefly, Hy’s Law cases have the following three components: 

1. The drug causes hepatocellular injury, generally shown by a higher incidence of 3-fold or 

greater elevations above the ULN of ALT or AST than the (nonhepatotoxic) control drug 

or placebo 

2. Among trial subjects showing such AT elevations, often with ATs much greater than 

3xULN, one or more also show elevation of serum TBL to >2xULN, without initial 

findings of cholestasis (elevated serum ALP) 

3. No other reason can be found to explain the combination of increased AT and TBL, such 

as viral hepatitis A, B, or C; preexisting or acute liver disease; or another drug capable of 

causing the observed injury 

Finding one Hy’s Law case in the clinical trial database is worrisome; finding two is considered 

highly predictive that the drug has the potential to cause severe DILI when given to a larger 

population. Clinical trials of the beta blocker dilevalol (enantiomer of labetalol, a 

diastereoisomeric mixture) showed two such cases in about 1,000 exposures. The drug was not 

approved in the United States, and examination of a postmarketing study in Portugal revealed 

fatal liver injury. Clinical trials of tasosartan, an angiotensin II blocking agent, showed a single 

Hy’s Law case. This led to a request for a much larger premarketing database and the drug was 

abandoned. 

Severe DILI can be estimated to occur at a rate of at least one-tenth the rate of the so-called Hy’s 

Law cases (Temple 2001). This observation was recently confirmed in large studies of DILI in 

Spain (Andrade and Lucena et al. 2005) and in Sweden (Björnsson and Olsson 2005) in which 

approximately 10 percent of subjects with hyperbilirubinemia or jaundice died or needed liver 

transplants. 

Recent examples of some drugs causing idiosyncratic hepatotoxicity (e.g., bromfenac, 

troglitazone, ximelagatran) further illustrate the predictive value of Hy’s Law, where findings 

during clinical trials were noted and severe DILI occurred after marketing. These examples are 

described in detail in Appendix A. 

Hy’s Law cases represent one end of a spectrum of laboratory abnormalities that indicate liver 

injury. Each of these cases has different sensitivity and specificity as a predictor for the potential 

for severe liver injury. Although it is not possible to provide precise specificity and sensitivity 

5 
Contains Nonbinding Recommendations

estimates for the various signals, guidance can be provided on use of these major indicators of a 

potential for severe DILI, as follows: 

• An excess of AT elevations to >3xULN compared to a control group

AT elevations to >3xULN are relatively common and may be seen in all groups, but an 

excess of these elevations compared to a control group is nearly always seen for drugs 

that ultimately prove severely hepatotoxic at relatively high rates (1/10,000). Therefore, 

the sensitivity of a significantly increased incidence compared to control (e.g., of 

>3xULN AT elevations) as an indicator of a potential for liver injury is high. But many 

drugs show this signal without conferring a risk of severe injury (e.g., tacrine, statins, 

aspirin, heparin), indicating low specificity for an excess of AT elevations alone. There 

are no good data to predict how great this excess incidence of AT elevations should be 

compared to controls to suggest an increased risk of DILI. Such an excess may not be 

apparent for drugs with a potential to cause idiosyncratic DILI that are used for short 

treatment courses, such as many antibiotics. 

• Marked elevations of AT to 5x-, 10x-, or 20xULN in modest numbers of subjects in 

the test drug group and not seen (or seen much less frequently) in the control group 

Many, but not all, severely hepatotoxic drugs show such elevations, indicating high 

sensitivity for predicting severe DILI; again, however, some drugs, such as tacrine and 

others that are not severely hepatotoxic, also can cause AT elevations to this degree, so 

that specificity of this finding is suboptimal. 

• One or more cases of newly elevated total serum bilirubin to >2xULN in a setting of 

pure hepatocellular injury (no evidence of obstruction, such as elevated ALP typical 

of gall bladder or bile duct disease, or malignancy, or impaired glucuronidation 

capacity caused by genetic (Gilbert syndrome) or pharmacologic (treatment with 

atazanavir or other drugs) factors), with no other explanation (viral hepatitis, 

alcoholic or autoimmune hepatitis, other hepatotoxic drugs), accompanied by an 

overall increased incidence of AT elevations >3xULN in the test drug group 

compared to placebo3

 

The sensitivity of this observation appears high for any given incidence rate of severe 

DILI if enough people are exposed to the drug. For example, if the true incidence of 

severe injury is 1/10,000, and the rate of Hy’s Law cases is 1/1,000, about 3,000 exposed 

subjects (Rule of 3) would be needed to have a 95 percent probability of observing at 

least one Hy’s Law case in the treated population (Rosner 1995).4

 The specificity of this 

 

3

 This constellation of findings is the hallmark of a Hy’s Law case. The predictive value of these three findings for a 

drug’s potential to cause DILI may be different if these findings are identified in patients with preexisting liver 

disease, fatty liver disease such as NASH, chronic hepatitis C or B, or bilirubin metabolism abnormalities (Gilbert 

syndrome), or in patients on drugs that treat liver disease or that inhibit bilirubin glucuronidation, such as indinavir 

or atazanavir (Zhang and Chando et al. 2005). 

4

 The Rule of 3 is derived from simple binomial calculation. There will be at least a 95 percent chance of seeing one 

or more cases of DILI in 3n trial subjects if its true incidence is 1 in n subjects, and the group is well observed. 

6 
Contains Nonbinding Recommendations

finding appears very high if two or more cases are seen (e.g., dilevalol, bromfenac, 

troglitazone, ximelagatran). We are not aware of the occurrence of false positive Hy’s 

Law findings for a drug that was subsequently found not to cause severe DILI in a larger 

treatment population. Therefore, the finding of two Hy’s Law cases, and probably even 

one, is a strong predictor of a significant risk of severe liver injury. Failure to find a case, 

however, does not imply that a drug with AT elevations is free of a risk of severe DILI. 

The degree of assurance depends on the population exposed for a long enough time, the 

discontinuation rules used in the protocols, and the true incidence rate of severe DILI. 

IV. CLINICAL EVALUATION OF DILI 

A. General Considerations 

For most drugs in development that reach phase 3 testing, the chances of encountering severe 

DILI are low. An increased incidence of mild hepatotoxicity (AT elevations) in early trials 

usually results in heightened screening to detect and evaluate liver injury during phase 3 testing. 

It is critical, however, to determine whether mild hepatotoxicity reflects a potential for severe 

DILI or reflects a capacity for only limited injury. To make this distinction, it is important to 

detect any cases of more severe injury and to examine such cases closely, observing the course 

and outcome of the injury, and seeking additional information that might identify other causes. 

The following general recommendations for evaluating and monitoring potential drug-induced 

hepatotoxicity may not be suitable for all situations and should be modified for special 

populations, such as people with preexisting liver disease or malignancies, and in light of 

accumulating data. In addition, clinical trials of cellular and gene therapies and of vaccines pose 

specific challenges related to trial size and design, biodistribution and persistence of vectors, the 

function and anatomic location of cellular products, and other factors. Applicants are 

encouraged to discuss these issues with the relevant review division. 

1. Patients with Liver Abnormalities or Disease 

Patients are sometimes excluded from clinical trials because of baseline liver test abnormalities 

or a history of liver disease, but there is no well-established reason to do this, except perhaps to 

avoid confusion between the previous disease and an effect of the test drug. Patients with acute 

viral, autoimmune, alcoholic, or other types of hepatitis are unstable and generally not 

appropriate subjects for clinical trials other than trials of treatments for their acute illness. 

Patients with stable underlying liver disease can be included cautiously in late-stage clinical 

trials, but probably not if bilirubin excretory or protein synthetic functions are impaired, unless 

there is a strong need that they be treated. This implies that diagnostic screening for liver test 

abnormalities should be conducted before enrolling subjects into trials. Patients with stable liver 

disease generally should be included in at least some phase 3 trials if they are likely to be treated 

with the drug if it is marketed. Preexisting liver disease has not been thought to make patients 

more susceptible to DILI (Zimmerman 1978, 1999), but it may be that a diminished liver reserve

or the ability to recover could make the consequences of injury worse. This appears to be the 

case with highly active antiretroviral therapy in patients with chronic viral hepatitis. If the drug 

7 
Contains Nonbinding Recommendations

is intended to be prescribed or marketed to such patients after approval, they should be enrolled 

in controlled trials. 

2. Detection of DILI 

Depending on the mechanism underlying DILI, different drugs can be associated with different 

treatment time/hazard profiles. In many cases, there is a delay of at least a few weeks between 

initiation of treatment and onset of liver injury. However, for some drugs, rapid onset of injury 

may occur, sometimes in the presence of a systemic hypersensitivity reaction that can be 

associated with multi-organ involvement, fever, eosinophilia, and/or rash. In general, early trials 

of a drug in trial subjects with presumably normal liver function should involve obtaining liver 

enzyme (ALT, AST, ALP) and bilirubin tests every 2 to 4 weeks, at least for a few months. For 

drugs being studied with short treatment courses, both baseline and post-treatment liver enzyme 

testing should be performed, since there may be a gap between the end of treatment and the onset 

of liver injury. In circumstances when there is a high likelihood that such a drug will be 

chronically used in an off-label fashion, long-term treatment trials to measure risk for DILI may 

be warranted. 

It is uncertain whether early and nonspecific symptoms (e.g., anorexia, nausea, fatigue, right 

upper abdominal discomfort, vomiting) precede or follow the first laboratory signs of hepatic 

injury (rising ALT, AST, or ALP), and the pattern of clinical and laboratory changes may vary 

with different drugs and recipients. In most cases, however, the first evidence of a problem is the 

discovery of elevated AT or ALP during routine serial measurements. In longer trials, if there is 

no sign of liver injury after a reasonable length of exposure (e.g., 3 months), the monitoring 

interval can be increased to once every 2 to 3 months. Later trials also can use less frequent liver 

chemistry monitoring if there is no indication of hepatotoxicity in earlier trials. 

As previously noted, if symptoms compatible with DILI precede knowledge of serum chemical 

test abnormalities, liver enzyme measurements should be made immediately, regardless of when 

the next visit or monitoring interval is scheduled. In some cases, symptoms may be an early sign 

of injury and although typically less sensitive than serum enzyme elevations, they may indicate a 

need for prompt serum testing. Reliance on early symptoms, rather than serum enzyme 

monitoring, has become the standard for monitoring isoniazid therapy for prophylaxis of 

tuberculosis and seems to prevent severe liver injury if acted upon promptly by discontinuation 

of isoniazid (Nolan and Goldberg et al. 1999). Attention to symptoms does not supplant routine 

periodic assessment of AT, TBL, and ALP in trials of investigational drugs. 

3. Confirmation 

In general, an increase of serum AT to >3xULN should be followed by repeat testing within 48 

to 72 hours of all four of the usual serum measures (ALT, AST, ALP, and TBL) to confirm the 

abnormalities and to determine if they are increasing or decreasing. There also should be inquiry 

made about symptoms. Serum AT may rise and fall quite rapidly, and waiting a week or two 

before obtaining confirmation of elevations may lead to a false conclusion that the initially 

observed abnormality was spurious. Of greater concern, delay in retesting may allow 

progression to severe worsening if the initial abnormality was the herald of a severe reaction to 

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Contains Nonbinding Recommendations

follow. The need for prompt repeat testing is especially great if AT is much greater than 3xULN 

and/or TBL is greater than 2xULN. For outpatient trials, or trials in which subjects are far away 

from the trial site, it may be difficult for the subjects to return to the trial site promptly. In this 

case, the subjects should be retested locally, but normal laboratory ranges should be recorded, 

results should be made available to trial investigators immediately, and the data should be 

included in the case reports. If symptoms persist or repeat testing shows AT >3xULN for 

subjects with normal baseline measures or 2-fold increases above baseline values for subjects 

with elevated values before drug exposure, it is appropriate to initiate close observation to 

determine whether the abnormalities are improving or worsening (see below). If close 

monitoring is not possible, the drug should be discontinued. 

4. Close Observation 

It is critical to initiate close observation immediately upon detection and confirmation of early 

signals of possible DILI, and not to wait until the next scheduled visit or monitoring interval. A 

threshold of aminotransferase levels greater than 3xULN seems reasonable, as lesser elevations 

are common and nonspecific. If additional testing, beyond that specified in the trial protocol, is 

carried out, it is important that the subject’s information be added to the case report forms and 

database. 

Close observation includes: 

• Repeating liver enzyme and serum bilirubin tests two or three times weekly. Frequency 

of retesting can decrease to once a week or less if abnormalities stabilize or the trial drug 

has been discontinued and the subject is asymptomatic. 

• Obtaining a more detailed history of symptoms and prior or concurrent diseases. 

• Obtaining a history of concomitant drug use (including nonprescription medications and 

herbal and dietary supplement preparations), alcohol use, recreational drug use, and 

special diets. 

• Ruling out acute viral hepatitis types A, B, C, D, and E; autoimmune or alcoholic 

hepatitis; NASH; hypoxic/ischemic hepatopathy; and biliary tract disease. 

• Obtaining a history of exposure to environmental chemical agents. 

• Obtaining additional tests to evaluate liver function, as appropriate (e.g., INR, direct 

bilirubin). 

• Considering gastroenterology or hepatology consultations. 

5. Decision to Stop Drug Administration 

It has been observed that dechallenge (stopping drug administration) does not always result in 

immediate improvement in abnormal lab values. Abnormal test values and symptoms may 

progress for several days or even weeks after discontinuation of the drug that caused the 

abnormality. For example, rising TBL usually follows serum AT increases by a few days to 

weeks. The primary goal of close observation is to determine as quickly as possible whether 

observed abnormal findings are transient and will resolve spontaneously or will progress. For 

most DILI, no specific antidotes are available (except N-acetylcysteine for acute acetaminophen 

9 
Contains Nonbinding Recommendations

overdose if given promptly, and, possibly, intravenous carnitine for valproic acid hepatotoxicity). 

Promptly stopping the offending drug usually is the only potentially effective therapy. 

A difficult question is when should the investigational drug be stopped? Because transient 

fluctuations of ALT or AST are common, and progression to severe DILI or acute liver failure is 

uncommon, automatic discontinuation of trial drug upon finding a greater than 3xULN elevation 

of ALT or AST may be unnecessary. For most people, the liver appears capable of adapting to 

injury by foreign chemical substances, which may render a person tolerant to the drug despite 

continued exposure. Stopping a drug at the first hint of mild injury does not permit learning 

whether adaptation will occur, as it does for drugs such as tacrine, which cause liver injury but 

do not cause severe DILI. On the other hand, continuing drug appears unacceptably dangerous if 

there is marked serum aminotransferase elevation or evidence of functional impairment, as 

indicated by rising bilirubin or INR, which represent substantial liver injury. Although there is 

no published consensus on exactly when to stop a drug in the face of laboratory abnormalities 

and the decision will be affected by information on related drugs, the accumulating clinical 

experience, the clinical status of the patient, and many other factors, the following can be 

considered a basic guide. Discontinuation of treatment should be considered if: 

• ALT or AST >8xULN 

• ALT or AST >5xULN for more than 2 weeks 

• ALT or AST >3xULN and (TBL >2xULN or INR >1.5) 

• ALT or AST >3xULN with the appearance of fatigue, nausea, vomiting, right upper 

quadrant pain or tenderness, fever, rash, and/or eosinophilia (>5%) 

It should be noted that although these guidelines have not been evaluated systematically in a 

prospective fashion, they represent an approach that is similar to current practice. 

6. Evaluating Data for Alternative Causes 

An important purpose of close observation is to gather additional clinical information to seek 

other possible causes of the observed liver test abnormalities, such as one of the following 

common causes: 

• Acute viral hepatitis. The usual onset of hepatocellular DILI is indistinguishable from 

acute viral hepatitis A or B. Hepatitis C is much less often acute in its onset and tends to 

be insidious, but it sometimes can resemble acute DILI. The presence of acute viral 

hepatitis A, B, and C should be evaluated by serological markers. Viral hepatitis D 

(requires concomitant hepatitis B infection) and E are relatively rare in the United States. 

Hepatitis E is more common in developing countries, including Southeast Asia, and 

should be considered in recent travelers to those countries and in patients in trials 

conducted in those countries. Also rare are hepatocellular liver injuries caused by 

Epstein-Barr virus, cytomegalovirus, herpes simplex virus, toxoplasmosis, varicella, and 

parvovirus, although these infections are seen more typically in immuno-suppressed 

individuals. Adolescent and young adult patients with possible DILI should be tested for 

Epstein-Barr virus. Hepatitis is common among transplant patients with cytomegalovirus 

disease. 

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• Alcoholic and autoimmune hepatitis. Acute alcoholic hepatitis usually is recurrent, 

with a history of binging exposure to alcohol preceding episodes, and it has some 

characteristic features, such as associated fever, leukocytosis, right upper quadrant pain 

and tenderness, hepatomegaly, and AST >ALT, that may help distinguish it from other 

causes of liver injury. Other features of the physical examination may include the 

presence of stigmata of cirrhosis, such as spider nevi, palmar erythema, estrogenic 

changes in males, and Dupuytren’s contractures. Alcoholic and autoimmune hepatitis 

should be assessed by history, physical examination, and laboratory testing, including 

serologic testing (e.g., antinuclear or other antibodies). 

• Hepatobiliary disorders. Biliary tract disease, such as migration of gallstones or 

intrahepatic lesions, more often causes cholestatic injury initially and should be 

investigated with gall bladder and ductal imaging studies, especially if ALP is increased. 

Malignant interruption of the biliary tract also should be considered. 

• NASH. NASH may be seen in obese, hyperlipoproteinemic, and/or diabetic patients and 

may be associated with fluctuating aminotransferase levels, and hepatic and sometimes 

splenic enlargement. It is sometimes associated with cirrhosis and portal hypertension. 

• Cardiovascular causes. Cardiovascular disease, especially right heart failure and 

hypotension or any cause of impaired oxygenation of the liver, may cause acute 

centrilobular hypoxic cell necrosis (ischemic hepatitis) with rapid and sometimes 

spectacular increases of serum AT (e.g., AT >10,000 U/L). Cardiovascular dysfunction 

or impaired liver oxygenation, including hypotension or right heart failure, should be 

assessed by physical examination and history. 

• Concomitant treatments. It is critical to discover concomitant treatments, including 

exposure to nonprescription and dietary supplement products that might be responsible 

for injury. Many people take multiple drugs, perhaps less often in controlled clinical 

trials because of exclusion criteria, but subjects may not report taking disallowed drugs or 

other agents. The possible exposure to potentially toxic herbal or dietary supplement 

mixtures (sometimes of unknown composition), nonprescription medications such as 

acetaminophen, or to occupational chemical agents may not be volunteered unless 

subjects are specifically questioned. 

7. Follow-Up to Resolution 

All trial subjects showing possible DILI should be followed until all abnormalities return to 

normal or to the baseline state. DILI may develop or progress even after the causative drug has 

been stopped. Results should be recorded on the case report form and in the database. Note that 

longer follow-up can sometimes reveal an off-drug repetition of what had appeared to be DILI, 

indicating that liver injury was related to underlying liver disease. 

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8. Rechallenge 

Whether or not to rechallenge a subject who showed mild DILI is a difficult decision. 

Reexposure may initiate a sometimes explosive and more severe reaction, as was observed with 

halothane several decades ago. Some cases of DILI show indicators of immunological reaction 

such as eosinophilia, rash, fever, or other symptoms or findings, and it is possible that such cases 

are more prone to recur with reexposure. Rechallenge may not be considered negative unless the 

subject is exposed to and tolerates the same dose and treatment duration that preceded the 

original reaction. A negative rechallenge does not necessarily allow a conclusion that the drug 

did not cause the injury. Most people can adapt to xenobiotic substances, including new drugs, 

and develop tolerance for them. This has been observed even for drugs that can cause severe 

injury, such as isoniazid. The large majority of people showing hepatocellular injury while 

taking isoniazid recover fully or recover while continuing to take the drug, and some, but not all, 

can resume or continue taking the drug without further adverse consequence. If such tolerance 

has developed, the use of rechallenge to verify drug causation would give a false negative result. 

Generally, rechallenge of subjects with significant AT elevations (>5xULN) should not be 

attempted. If such subjects are rechallenged, they should be followed closely. Rechallenge can 

be considered if the subject has shown important benefit from the drug and other options are not 

available or if substantial accumulated data with the test drug do not show a potential for severe 

injury. The subject should be made aware of the potential risk, and consent to the rechallenge, 

and the institutional review board consulted. 

B. Research Opportunities 

It is not known why only a few people show severe DILI in response to a hepatotoxic drug while 

others show nothing or seem to adapt. The current thinking is that both genetic and acquired 

factors may be important in determining the susceptibility to injury. Close observation provides 

a major opportunity to gather and store serial samples of blood and urine, to investigate 

characteristics of subjects who show evidence of mild or severe DILI, and to see how they differ 

from each other and from people who do not show any effects despite being similar in age, sex, 

and drug exposure. These serial samples can be studied by genomic, proteomic, and 

metabolomic methods to determine how subjects differ, and to seek biomarkers that identify the 

susceptible persons. 

As part of the Critical Path Initiative,5

 the FDA is working with industry, academia, and other 

experts to broaden its understanding of the biochemical and genetic bases of DILI. It is hoped 

that predictive bioassays and biomarkers can be identified through analysis of systematically 

collected biospecimens that will help determine which patients are most likely to suffer liver 

injury from specific compounds. If tests that identify people susceptible to severe DILI can be 

developed, a drug that is hepatotoxic to them could remain available to other people who are not 

susceptible to severe DILI. 

In addition, identification of common genotypic characteristics among patients experiencing 

DILI in response to one or more class-related hepatotoxic drugs might permit the development of 

 

5

 See http://www.fda.gov/ScienceResearch/SpecialTopics/CriticalPathInitiative/default.htm. 

12 
Contains Nonbinding Recommendations

in vitro or ex vivo tests or genetically altered animal strains that can be used to better predict 

serious hepatotoxic potential, or the lack thereof, of new drugs belonging to the same or closely 

related classes. 

C. Case Report Forms 

Because DILI has resulted in the marketing withdrawal or cessation of development of many 

drugs, every clinical trial should include case report form pages specifically designed to capture 

information pertinent to the evaluation of treatment-emergent liver abnormalities. In addition to 

collecting information on laboratory abnormalities, clinical symptoms, and the potential cause of 

any hepatic illness, case report forms and narratives should include the following information for 

cases in which liver injury is found (including control subjects with such injury): 

• Time and date from start of drug administration to start of illness. 

• Time and date of cessation of drug, or interruption of drug administration. 

• Complete description of the injury, including systemic symptoms, other organ 

involvement, rash, fever, and eosinophilia. 

• Outcomes such as death, liver transplant, hospitalization, recovery, and treatment for 

DILI. 

• Free text describing the course of illness, including pertinent physical examination 

findings, such as hepatomegaly, splenomegaly, right-upper quadrant tenderness, the time 

course of abnormalities of aminotransferases, ALP, TBL with dates of testing, normal 

ranges, and results for tests done in addition to those specified in the original protocol, 

and tests done during any unscheduled visits. These additional laboratory test results, 

including reference ranges, should also become part of the overall database. Supportive 

tabular and/or graphical display of serial laboratory data is often desirable in addition to 

narrative information. Pre-study AT values should be sought, which may suggest chronic 

liver disease and/or an acute process that may have preceded exposure to the 

investigational drug. 

• Risk factors, especially history of alcohol use; risk factors for NASH such as diabetes, 

obesity, and hypertriglyceridemia, which may prompt ultrasound examination of the liver 

to detect steatosis. 

• All concomitant drugs (dose, start and stop dates, whether they are known to be 

hepatotoxic, information on rechallenge or dechallenge with drugs with the same or 

similar structure). 

• Evaluation of nondrug causes: recent hepatitis A, B, and C serology; evidence for biliary 

obstruction; imaging study results; acute alcoholic hepatitis (recent drinking and AST 

>2xALT are supportive); recent history of severe hypotension or congestive heart failure; 

other underlying viral disease. 

• All supplemental information, including consultation reports, narrative information, and 

special studies. 

Any potential Hy’s Law case should be handled as a serious unexpected adverse event associated 

with the use of the drug and reported to the FDA promptly (i.e., even before all other possible 

causes of liver injury have been excluded). It should be promptly reported to the FDA before 

fully working up the patient to rule out other etiologies. Reporting should include all available 

13 
Contains Nonbinding Recommendations

information, especially that needed for evaluating the severity and likelihood that the drug 

caused the reaction, and should initiate a close follow-up until complete resolution of the 

problem and completion of all attempts to obtain supplementary data. 

D. Interpretation of Signals of DILI or Acute Liver Failure 

1. Frequency and Magnitude of Liver AT Abnormalities 

The presence of even a single case of severe liver injury resulting from treatment in the 

premarketing clinical trials database is a signal of a high level of hepatotoxic risk. More 

commonly, however, there will be no identifiable cases of severe liver injury, but rather varying 

degrees of serum AT abnormalities that need to be interpreted. As previously noted, slight 

abnormalities of this kind (to <3xULN) are common in untreated and placebo-treated subjects 

and are not informative about the potential for the development of severe DILI. Subjects with 

such abnormalities should be watched. 

Therefore, it has become standard practice to look at greater deviations, such as AT values ≥3x-, 

5x-, or 10xULN. Because these abnormalities are often associated with other causes, such as 

NASH or hepatitis C, they can occur in placebo-treated groups, and it is important to compare 

their incidence in drug-exposed subject groups to that observed in control groups (i.e., placebo or 

treatment with products that do not cause elevation of aminotransferases). A significantly 

increased incidence of AT abnormalities >3xULN is a signal of a potential for severe DILI, but, 

even though it has high sensitivity, it is not specific. Abnormalities of greater magnitude (e.g., 

>10xULN) are rarely seen spontaneously in placebo arms of clinical trials in most settings. 

Therefore, greater magnitude AT elevations can be examined in the entire clinical trials database, 

not just in the controlled trials. Serum AT activity is a relatively volatile measurement, often 

rising and falling within days. It cannot be concluded from one measurement that a peak value 

has been seen, so detection of an abnormal rise calls for serial measures to determine which way 

the abnormality is moving, whether increasing or decreasing. 

A number of factors may confound interpretation of AT abnormalities seen in NDA or BLA 

databases. Although the more extreme AT elevations may be better predictors of toxicity than 

smaller elevations, close monitoring can affect the magnitude of abnormalities seen if it leads to 

earlier cessation of drug treatment. In addition, the contribution of drug treatment to an 

exacerbation of preexisting liver disease or the effects of concomitant hepatotoxic drugs may be 

difficult to determine. Finally, normalization of abnormalities on continued treatment is not 

proof that the abnormality was not drug-caused, as it can result from liver adaptation to the drug. 

2. Combined Elevations of Aminotransferases and Bilirubin 

When AT abnormalities indicating hepatocellular injury are accompanied by evidence of 

impaired hepatic function (bilirubin elevation >2xULN), in the absence of evidence of biliary 

obstruction (i.e., significant elevation of ALP) or some other explanation of the injury (e.g., viral 

hepatitis, alcohol hepatitis), the combined finding (i.e., Hy’s Law cases) represents a signal of a 

potential for the drug to cause severe DILI. Experience has indicated that the occurrence of even 

14 
Contains Nonbinding Recommendations

one or two well-documented cases of this combination is ominous, indicating a likelihood that 

the drug will cause severe liver injury. 

The absence of Hy’s Law cases in an NDA or BLA database may allow an estimate of an upper 

limit of the rate for severe DILI, using the Rule of 3 derived from simple binomial calculation. 

There will be at least a 95 percent chance of seeing one or more cases of DILI in 3n trial subjects 

if its true incidence is 1 in n subjects, and the group is well observed. Thus, if no cases of AT 

and bilirubin elevations are seen in 3,000 well-observed subjects, it can be concluded with 95 

percent confidence that the true rate of such occurrences is not more than 1 per 1,000. This 

calculation would then suggest a rate of expected severe liver injury ≤1 per 10,000 exposed 

patients, assuming that the rate of severe injury among patients with concomitant AT and TBL 

elevations is about 10 percent (Andrade and Lucena et al. 2005; Björnsson and Olsson 2005). 

E. Analysis of Signals of DILI 

Based on the FDA’s experience, the following analyses related to liver injury potential should be 

carried out and included in an NDA or BLA, or included in an investigational new drug 

application when DILI is suspected and being evaluated. 

1. Assessment of Drug Metabolism 

The metabolism of a drug can markedly affect the safety profile of the drug. A drug may be 

metabolized to a hepatotoxic metabolite (e.g., acetaminophen, halothane, isoniazid). Most 

hepatotoxic drugs have been oxidatively metabolized by the CYP450 system. 

2. Assessment of Liver-Related Adverse Events in Controlled Trials 

Applicants should provide an analysis of the incidence of abnormalities in AT, bilirubin, and 

ALP levels seen in subjects in controlled trials with at least one dose of drug exposure. 

Generally, the analysis should be for pooled data, although trial-to-trial differences may be of 

interest. Incidence can be given as the number of events per number of subjects exposed, or can 

incorporate treatment exposure, as the number of events per subject-years of exposure, 

preferably both. Changes in mean values for groups are not informative. For many drugs, it 

appears that a minimum duration of exposure is needed before DILI occurs. Therefore, it is 

useful to describe liver-related adverse events for subjects who have had the minimum duration 

of exposure (e.g., subjects with at least 1-month exposure). For some drugs, patterns of early 

injury after initiation of treatment may occur, and for these patients testing intervals should be 

modified appropriately. Incidences for pooled data should include, but are not limited to: 

• 3x-, 5x-, 10x-, and 20xULN elevations of AST, ALT, and either ALT or AST. 

• Any elevations of bilirubin; elevated TBL to >2xULN. 

• Any elevations of ALP >1.5xULN. 

• Elevation of AT (>3xULN) accompanied by elevated bilirubin (>1.5xULN, >2xULN). 

• Elevation of AT in temporal association with nausea, vomiting, anorexia, abdominal 

pain, or fatigue. 

15 
Contains Nonbinding Recommendations

• Possibly liver-related deaths and liver-related treatment discontinuations. These cases 

should be described and time-to-event analyses should be performed. Follow-up status 

also should be provided. There should be a description of any histologic and rechallenge 

data. 

All incidences should be calculated separately for drug-, placebo-, and active-controlled groups. 

Normal ranges for all tests should be provided. Time-to-event analyses for events occurring with 

increased incidence should be provided (e.g., elevated AT, bilirubin). The contribution of sex, 

age, risk factors, and drug dose or regimen to the abnormalities seen should be explored. 

3. Assessment of Liver-Related Adverse Events in the Entire Clinical Trials 

Database 

Applicants should provide an analysis of the incidence of abnormalities in AT, bilirubin, and 

ALP levels for the entire clinical trials database, including subjects with exposure of at least one 

dose of trial drug in phase 1 or phase 2 trials, or in uncontrolled, open label, extension trials. We 

recommend the same evaluation as for the controlled trials database discussed in section IV.D.2. 

Time-to-event analyses of events occurring at increased incidence, and rates of death and trial 

withdrawal in subjects with abnormalities, should be provided. The contribution of sex, age, 

drug dose or regimen, use of concomitant drugs, and underlying disease to the abnormalities seen 

should be explored. 

4. Assessment of Hy’s Law Cases in the Clinical Trials Database 

NDA and BLA submissions should include a listing of possible Hy’s Law cases identified by 

treatment group (e.g., subjects with any elevated AT of >3xULN, ALP <2xULN, and associated 

with an increase in bilirubin ≥2xULN). A narrative summary for each Hy’s Law case should be 

provided. Narrative summaries should not only provide, in text format, the data that are already 

presented in the case report tabulation, but also should provide a complete synthesis of all 

available clinical data and an informed discussion of the case, allowing for a better 

understanding of what the subject experienced. For a narrative summary to be useful, it should 

contain the following information: 

• Subject’s age, sex, weight, and height 

• Discussion of signs and symptoms related to hepatotoxicity: type and timing to exposure 

• Relationship of exposure duration and dose to the development of the liver injury 

• Pertinent medical history 

• Concomitant drugs with dates and doses 

• Pertinent physical exam findings 

• Test results (e.g., laboratory data, biopsy data and reports, with dates and normal ranges) 

• Time course of serum enzyme and bilirubin elevations (consider tabular and/or graphical 

display of serial laboratory data) 

• A summary of all available clinical information including, if known: 

− Prior or current history of ethanol use 

− Presence of risk factors for NASH (e.g., obesity, diabetes, marked 

hypertriglyceridemia) 

16 
Contains Nonbinding Recommendations

− Evidence for pre- or co-existing viral hepatitis, or other forms of liver disease, prestudy AT values, if available 

− Symptoms and clinical course including follow-up to resolution 

− Special studies (i.e., ultrasound, radiologic examinations, liver biopsy results) 

− Presence or absence of possible confounders, including concomitant illness, use of 

concomitant drugs that are known hepatotoxins, such as acetaminophen 

• Discussion of hepatotoxicity as supported by available clinical data and overall 

assessment of the treating physician, consultants, and applicants as to the likelihood of 

DILI 

• Treatment provided 

• Dechallenge and rechallenge results, if done 

• Outcomes and follow-up information 

• Copies of hospital discharge summaries, pathology and autopsy reports 

The availability of liver biopsy, explant, or autopsy slides for pathology review by review staff 

or external expert consultants has been helpful in the FDA’s assessment of Hy’s Law cases. 

Reports of external consultant opinions solicited by the applicant should be provided to the FDA. 

Applicants also should provide complete narrative summaries that include the components 

previously listed for all subjects who died of hepatic illness, or who discontinued trial drugs for 

hepatotoxicity, including subjects with abnormalities consistent with protocol-specific stopping 

rules. 

In some cases, a drug under consideration in the United States will have been marketed in other 

countries. In these cases it is important for the applicant to provide a synopsis of the global 

safety experience and level of usage and to describe in detail all cases of hepatotoxicity observed 

or suspected. 

5. Overall Assessment of a Drug’s Potential to Cause DILI 

The overall assessment should characterize a drug’s potential for DILI and should consider at 

least the following questions: 

• Was liver monitoring sufficiently frequent and thorough to characterize DILI risk? 

• Were there any cases of probable severe DILI? 

• Were there signals of a potential for DILI (e.g., AT elevations, Hy’s Law cases) and how 

were these signals assessed? 

• What doses and durations of exposure were associated with hepatotoxicity signals? 

• What approximate incidence of mild, moderate, and severe DILI can be expected 

postmarketing? 

• Is the trial information sufficient to inform an overall risk-benefit assessment? 

• Was there sufficient drug exposure (i.e., number of trial subjects and duration of 

treatment of each trial subject) and adequate liver test monitoring to reliably set an upper 

boundary for risk of severe DILI after marketing? 

17 
Contains Nonbinding Recommendations

18 

• What rate of severe injury (assuming Hy’s Law cases occur at about 10 times the rate of 

severe injury) has been suggested or has been ruled out (e.g., no Hy’s Law cases in 3,000 

subjects implies a rate of such cases of <1/1,000 and thus a rate of severe DILI of 

<1/10,000)? This consideration should reflect the presence or absence of other signals, 

such as marked elevations of AT. 

• Will some form of monitoring, by symptoms or serum testing, be needed? Usually, this 

would be considered only if there was evidence of severe liver injury or the potential for 

it. If so, effectiveness of monitoring, whether by symptoms or laboratory tests, and at 

what intervals should be discussed, and whether the results justify a monitoring 

recommendation in product labeling at the time of marketing approval. 
Contains Nonbinding Recommendations

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Contains Nonbinding Recommendations

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20 
Contains Nonbinding Recommendations

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21 
Contains Nonbinding Recommendations

22 

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Wilkins, Philadelphia. 
Contains Nonbinding Recommendations

APPENDIX A: ILLUSTRATIVE EXAMPLES OF DILI 

Duract (bromfenac) 

Bromfenac was a nonsteroidal anti-inflammatory drug (NSAID) studied for both shortterm analgesia and long-term arthritis treatment. There was little evidence of 

hepatotoxicity in the short-term analgesic trials, but during longer term clinical trials in 

arthritis, ALT elevations >3xULN were seen in 2.8 percent of patients on bromfenac, 

compared to none in placebo group. Among 1,195 exposed patients, there were two 

cases in which there was elevated TBL as well as AT elevation in the clinical trial data 

submitted for review in the NDA. Concerns about possible liver toxicity led to the 

approval of bromfenac in July 1997 for short-term use only and not for osteoarthritis or 

rheumatoid arthritis. As an NSAID, however, it was prescribed long-term off-label in 

arthritic patients, and severe hepatotoxicity emerged. Within 6 months of approval, 

reports of severe hepatic failure, including two cases requiring liver transplant, were 

received. All severe cases involved the use of bromfenac for more than 10 days, the 

maximum duration of treatment recommended in the labeling. 

In response, the FDA and the manufacturer strengthened the warnings in the package 

insert with a boxed warning, and issued a Dear Health Care Professional Letter. Despite 

these efforts, the manufacturer and the FDA continued to receive reports of severe 

injuries, including reports of death or need for liver transplantation (Moses and Schroeder 

et al. 1999; Hunter and Johnston et al. 1999; Rabkin and Smith et al. 1999; Fontana and 

McCashland et al. 1999). Given the availability of other effective NSAIDs, bromfenac 

was withdrawn from the market in June 1998. The two Hy’s Law cases in the long-termexposed population of about 1,000 subjects during drug development predicted an 

occurrence of severe hepatotoxicity during chronic use at a rate of about 1/5,000 to 

10,000 people. Following approval, rates of acute liver failure for bromfenac were 

estimated to be in the range of 1/10,000 (Goldkind and Laine 2006). 

Rezulin (troglitazone) 

Troglitazone was approved by the FDA in January 1997 for the treatment of Type 2 

diabetes mellitus. In reviews of the clinical trials of troglitazone conducted before 

approval there were no cases of liver failure among 2,510 subjects exposed to the drug in 

the NDA database, but 1.9 percent of troglitazone-treated subjects had ALT >3xULN 

compared to 0.3 percent of placebo-treated subjects, 1.7 percent had ALT >5xULN, and 

0.2 percent (5 subjects) had ALT >30xULN (2 subjects in the last group also experienced 

jaundice). The median duration of troglitazone therapy before peak ALT elevation was 

121 days. In the Diabetes Prevention Trial at the National Institutes of Health (NIH) 

performed after approval, 4.3 percent of 585 troglitazone-treated subjects had ALT 

≥3xULN, 1.5 percent had ALT >8xULN, and 2 subjects had ALT >30xULN, compared 

to 3.6 percent of subjects with ALT ≥3xULN in the placebo group (Knowler and 

Hamman et al. 2005). One of the subjects in the Diabetes Prevention Trial with ALT 

>30xULN developed liver failure and died, despite receiving a liver transplant. The 

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second subject recovered. These data suggest that the rate of severe liver injury would be 

about 1 in 3,000 to 10,000. 

After marketing, there were numerous reports of acute liver failure associated with 

troglitazone use (Gitlin and Julie et al. 1998; Vella and deGroen et al. 1998; Herrine and 

Choudary 1999), and four letters were sent to practicing physicians between 1997 and 

1999, urging monthly monitoring and careful use. These letters did not significantly 

affect the monitoring done by physicians, and AT monitoring recommended in the Dear 

Health Care Professional Letters and in the package insert was not regularly performed 

(Graham and Drinkard et al. 2001). Moreover, an analysis of 94 cases of liver failure 

reported spontaneously to the FDA showed that the progression from normal hepatic test 

results to irreversible liver injury occurred in less than a month (the recommended 

monitoring interval) in 19 patients. The onset of injury began after 3 days to more than 2 

years of troglitazone use (Graham and Green et al. 2003; Graham and Drinkard et al. 

2003). Time from jaundice to hepatic encephalopathy, liver transplantation, or death 

usually was rapid, averaging 24 days. Troglitazone was withdrawn from the U.S. market 

in March 2000, when other drugs in the same class with similar efficacy but little or no 

evidence of hepatotoxicity became available (i.e., rosiglitazone, pioglitazone). 

Apart from constituting another example of the predictive value of evidence of 

hepatocellular injury accompanied by even two cases of elevated bilirubin, there were 

other lessons learned from the troglitazone experience: 1) monitoring recommendations 

may not be well followed by physicians, even after warning letters are sent to all 

practicing physicians; and 2) some cases of severe hepatotoxicity occur rapidly, within 

less than a reasonable and practical recommended interval for monitoring, indicating that 

monitoring would provide at best only partial protection, even if recommendations were 

followed. 

Exanta (ximelagatran) 

Exanta (ximelagatran), an oral anticoagulant (antithrombin), was not marketed in the 

United States because of hepatotoxicity and other concerns discovered during clinical 

trials. Issues related to potential liver toxicity of ximelagatran were presented and 

discussed at an FDA advisory committee meeting in September 2004 (He 2004). During 

short-term clinical trials of the drug for prevention of thromboembolic complications 

after joint replacement surgical procedures, there was no increased rate of transaminase 

elevations in the ximelagatran group compared to the enoxaparin-warfarin group, and no 

serious hepatotoxicity was seen. But in longer term trials (more than 35 days) in patients 

with chronic atrial fibrillation to prevent embolic or thrombotic strokes, an increase in 

ALT >3xULN occurred in 7.6 percent of 6,948 patients compared to 1.1 percent of 

patients receiving warfarin treatment; and 1.5 percent of ximelagatran-treated patients 

had ALT >10xULN. 

Increases in AT typically occurred 1 to 6 months after the initiation of ximelagatran 

administration with peak levels within 2 to 3 months postrandomization. Among the 531 

ximelagatran patients with ALT >3xULN, 39 percent completed the trial on treatment, 

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25 

while 61 percent discontinued the drug. Almost all patients with ALT >3xULN returned 

to <2xULN whether the drug was stopped or not, although the return to normal was faster 

if ximelagatran was stopped. Of 18 patients who resumed drug after ALT returned to 

normal, only 2 had elevations recur. Concomitant elevations of ALT >3xULN and 

bilirubin >2xULN were observed in 37 of about 7,000 patients with ximelagatran and 5 

of 6,230 patients with comparator. At least 13 of 37 patients in the ximelagatran group 

had no alternative explanation for the concomitant ALT and bilirubin elevation. Nine of 

the 37 patients died, but in most cases the deaths were not clearly hepatotoxicity-related. 

Only one autopsy was done and it showed a small, friable and diffusely mottled liver 

suggestive of severe diffuse hepatic necrosis, but liver failure from ximelagatran might 

have contributed to some of the other deaths (He 2004; Lewis 2006; Kaplowitz 2006; 

Senior 2006; Temple 2006). Because severe hepatotoxicity was observed in an 

orthopedic surgery trial in an extended treatment of 35 days, Exanta was withdrawn in 

February 2006 from the 22 countries in which it had been approved, and further 

development in the United States was abandoned. 

Again, short-term tolerance of ximelagatran, with resolution of even substantial 

elevations of ALT in most cases, did not predict long-term safety. The relatively high 

rate of Hy’s Law cases, about 0.2 percent or 1/500 (13 cases among 7,000 exposed 

patients), predicted the occurrence of severe hepatotoxicity, at a rate of about 1/5,000 (10 

percent of the rate of Hy’s Law cases). In fact, at least one death occurred among the 

7,000 exposed patients from subsequent liver toxicity, further supporting such an 

estimate.