Guidance for Industry 

Clozapine Tablets: In Vivo 

Bioequivalence and In Vitro 

Dissolution Testing 

U.S. Department of Health and Human Services 

Food and Drug Administration 

Center for Drug Evaluation and Research (CDER)

June 2005 

BP 

Revision I 
Guidance for Industry 

Clozapine Tablets: In Vivo 

Bioequivalence and In Vitro 

Dissolution Testing 

Additional copies are available from: 

Office of Training and Communication 

Division of Drug Information, HFD-240 

Center for Drug Evaluation and Research 

Food and Drug Administration 

5600 Fishers Lane 

Rockville, MD 20857 

(Tel) 301-827-4573 

http://www.fda.gov/cder/guidance/index.htm

U.S. Department of Health and Human Services 

Food and Drug Administration 

Center for Drug Evaluation and Research (CDER)

June 2005 

BP 

Revision 1 
TABLE OF CONTENTS 

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

II. BACKGROUND ................................................................................................................... 1

III. IN VIVO STUDIES........................................................................................................... 2

A. Product Information......................................................................................................... 2

B. Steady-State Bioequivalence Study ................................................................................. 3

C. Other Recommendations.................................................................................................. 6

IV. IN VITRO TESTING CRITERIA .................................................................................. 7

A. Dissolution Testing............................................................................................................ 7

B. Content Uniformity Test .................................................................................................. 7

V. WAIVER REQUIREMENTS.............................................................................................. 7
Contains Nonbinding Recommendations 

Guidance for Industry1 1 

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Clozapine Tablets: In Vivo Bioequivalence and In Vitro Dissolution 

Testing 

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

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

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

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

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

11 number listed on the title page of this guidance. 

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I. INTRODUCTION 

This guidance provides recommendations for sponsors of abbreviated new drug applications 

(ANDAs) designing bioequivalence studies for generic clozapine products. This document 

revises the recommendations provided in a guidance on the same topic issued in November 

1996. In the 1996 guidance, the Agency recommended that doses of clozapine tablets be 

administered to healthy subjects as well as to the appropriate patient population in 

bioequivalence studies for generic clozapine products. Because a high number of healthy 

subjects experienced serious adverse effects such as hypotension, bradycardia, syncope, and 

asystole during clozapine bioequivalence studies, FDA is recommending that studies not be 

conducted using healthy subjects. In addition, a single-dose study using a 12.5 mg dose is no 

longer recommended. Instead, this guidance recommends a multiple-dose bioequivalence study 

conducted in patients using the highest dosage strengths (e.g., 100 mg tablets). 

The protocols described in this guidance are designed to reduce the likelihood of adverse events 

or, if adverse events should occur, to ensure that adequate treatment is available. 

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 

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

recommended, but not required. 

II. BACKGROUND 

Clozapine, a dibenzodiazepine derivative with potent antipsychotic properties, is indicated for 

the management of patients with severe schizophrenia who fail to respond adequately to standard 

1

 This guidance has been prepared by the Office of Generic Drugs (OGD) in the Office of Pharmaceutical Science, 

Center for Drug Evaluation and Research, Food and Drug Administration. 
Contains Nonbinding Recommendations 

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antipsychotic drug treatment. A significant risk of agranulocytosis and seizures associated with 

its use is a major factor restricting wide use of clozapine in psychiatric practice. 

The FDA recommends that treatment with clozapine begin with one-half of a 25 milligram (mg) 

tablet (12.5 mg) once or twice daily and that treatment be continued with daily dosage 

increments of 25-50 mg per day, if well tolerated, to achieve a target dose of 300 to 400 mg per 

day by the end of 2 weeks. While many patients respond adequately at doses between 300 and 

600 mg per day, it may be necessary to raise the daily dose to between 600 and 900 mg to obtain 

an acceptable response. Dosing should not exceed 900 mg per day. 

In humans, clozapine from 25 mg and 100 mg tablets is equally bioavailable relative to a 

clozapine solution. Following a dosage of 100 mg twice a day, the average steady-state peak 

plasma concentration occurs at an average of 2.5 hours (range 1-6 hours) after dosing. Food 

does not appear to affect clozapine systemic bioavailability. The mean elimination half-life of

clozapine after a single 75 mg dose is 8 hours (range 4-12 hours), compared to a mean steadystate half-life of 12 hours (range 4-66 hours) following 100 mg twice a day dosing. The 

elimination half-life increases significantly upon multiple dosing relative to single-dose 

administration, raising the possibility of concentration dependent pharmacokinetics. However, 

at steady-state, linearly dose-proportional changes have been observed in AUC, peak, and 

minimum clozapine plasma concentrations after administration of 37.5 mg, 75 mg, and 150 mg

(twice daily). 

Orthostatic hypotension with or without syncope can occur with clozapine treatment. 

Orthostatic hypotension is more likely to occur during initial titration in association with rapid 

dose escalation and may even occur with the first dose. Due to the hypotensive effects 

associated with administration of clozapine to healthy subjects, the original recommendations in 

a guidance on clozapine tablets published in November 1996 are being changed. This document 

revises and supersedes the previous version of the guidance. The Agency currently recommends 

that steady-state studies to evaluate the bioequivalence of clozapine products be performed only 

on patients who are already receiving an established maintenance dose of an approved clozapine 

product and have failed to respond adequately to standard antipsychotic drug treatment. The 

Agency believes that the previously recommended study design using half tablets in healthy 

subjects was adequate to establish bioequivalence of generic clozapine products; however, the 

safety concerns associated with the use of clozapine in healthy subjects are significant, and it is 

recommended that this practice not be continued. 

III. IN VIVO STUDIES 

A. Product Information 

1. FDA Designated Reference Product 

Applicants may consult FDA’s Approved Drug Products with Therapeutic Equivalence 

Evaluations (Orange Book) for the appropriate reference product. 

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Contains Nonbinding Recommendations 

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The test batch or lot should be manufactured under production conditions and should be 

at least 10% of the size of the largest lot planned for full production, or a minimum of 

100,000 units, whichever is larger. 

3. Potency 

The assayed potency of the reference product should not differ from that of the test 

product by more than 5%. 

B. Steady-State Bioequivalence Study 

The objective of this steady-state bioequivalence study is to compare the rate and extent 

of absorption of a generic formulation with a reference formulation when administered at 

equal doses, as labeled. 

Potential sponsors should consider the following study design. This study is appropriate 

for institutionalized or noninstitutionalized patients. Procedures should be in place to 

ensure medication compliance in either setting. 

1. Steady-State Study in Patients Receiving a Stable Dose of Clozapine 

The study would be conducted in patients who are receiving a stable daily dose of 

clozapine administered in equally divided doses at 12-hour intervals. Patients who are 

receiving multiples of 100 mg every 12 hours would be eligible to participate in the study 

of the 100 mg strength by continuing their established maintenance dose. According to 

the randomization schedule, an equal number of patients would receive either the generic 

formulation (Treatment A) or the reference formulation (Treatment B) in the same dose 

as administered prior to the study every 12 hours for 10 days. 

Patients would then be switched to the other product for a second period of 10 days. No 

washout period is necessary between the two treatment periods. After the study is 

completed, patients could be continued on their current dose of clozapine using an 

approved clozapine product as prescribed by their clinicians. 

2. Procedures for the Study 

Before the study begins, the proposed protocol must be approved by an institutional 

review board (IRB).2

 

The FDA recommends that applicants enroll a sufficient number of patients to ensure 

adequate statistical power. 

2

 See 21 CFR 314.94(a)(7)(iii). 

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Contains Nonbinding Recommendations 

Patients should receive study treatment A or B with 240 milliliters (ml) of water at fixed 

12-hour intervals for 10 days, using multiples of the 100 mg strength. 

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Blood samples should be collected over a dosing interval on day 10, following 

preliminary sampling on days 7, 8, and 9 to confirm steady-state conditions. The last 

dose of clozapine to be taken before blood sampling for each period should be 

administered at the clinical site to assure exact timing of sampling. 

3. Patient Entry Criteria and Facilities 

To enter into this study, patients should be appropriate candidates for clozapine therapy 

(as stated in product labeling) and have been taking a stable dose of clozapine for at least 

three months. Patients should be otherwise healthy as determined by physical 

examination, medical history, and routine hematologic and biochemical tests. 

Outpatients should be hospitalized for at least 2 days during the collection of each set of 

pharmacokinetic samples. The clinical and analytical laboratories used for the study 

should be identified in the study report, along with the names, titles, and curriculum vitae 

of the medical and scientific/analytical directors. 

4. Safety Monitoring 

White blood cell (WBC) counts should be monitored and clozapine treatment modified, if 

necessary, in accordance with the agranulocytosis warning in the labeling of the 

reference listed drug product. Patients requiring modification of clozapine treatment 

should be dropped from the study and provided with prompt medical care. Blood 

pressure, heart rate, and body temperature should be monitored during the study and 

immediate medical care provided for any significant abnormalities. 

 

5. Restrictions 

Patients should fast for at least 8 hours prior to and 4 hours after the administration of the 

morning dose of the test or reference treatment on day 10 of each period (i.e., the days on 

which blood samples are to be collected to assess the concentration-time curve). All 

meals on day 10 should be standardized during the study. 

Water may be allowed, except for 1 hour before and 1 hour after drug administration, 

when no liquid should be permitted other than that needed for drug dosing. 

Patients with any of the following should be excluded from the study: 

 

• A history of allergic reactions to clozapine or other chemically related psychotropic 

drugs 

• Concurrent primary psychiatric or neurological diagnosis, including organic mental 

disorder, severe tardive dyskinesia, or idiopathic Parkinson’s disease 

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Contains Nonbinding Recommendations 

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• A total white blood cell count below 4000/ml, or an absolute neutrophil count below 

2000/ml

• A history of granulocytopenia or myeloproliferative disorders (drug-induced or 

idiopathic) 

• Significant orthostatic hypotension (i.e., a drop in systolic blood pressure of 30 mm

Hg or more and/or a drop in diastolic blood pressure of 20 mm Hg or more on 

standing) 

• Concurrent use of antihypertensive medication or any medication that might predispose to orthostatic hypotension 

• A medical or surgical condition that might interfere with the absorption, metabolism,

or excretion of clozapine 

• A history of epilepsy or risk for seizures 

• Concurrent use of other drugs known to suppress bone marrow function 

• Expected changes in concomitant medications during the period of study 

• Positive tests for drug or alcohol abuse at screening or baseline 

• A history of alcohol or drug dependence by Diagnostic and Statistical Manual of 

Mental Disorders IV (DSM-IV) criteria during the 6-month period immediately prior 

to study entry 

• Compliance with outpatient medication schedule not expected 

• History of multiple syncopal episodes 

6. Blood Sampling 

Venous blood samples should be collected after the day 10 morning dose to assess the 

concentration-time curve at predose (0 hours) and at 0.25, 0.5, 1.0, 1.5, 2.0, 2.5, 3.0, 3.5, 

4.0, 5.0, 6.0, 8.0, 10.0, 12.0 hours. The predose blood sampling should include at least 

three successive trough level samples (Cmin). These samples should be collected on the 

last 3 days of dosing in each period to ensure that steady-state blood plasma/serum levels 

are achieved in each study period. 

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Contains Nonbinding Recommendations 

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1. Precautions and Safety Issues 

• Patients should be confined for at least 12 hours after the first dose of the test and 

reference products. 

• Patients should remain in the supine position for the first 6 hours after the first dose, 

even if they were previously on a stable dose of clozapine. 

• Patients should be adequately hydrated. This may be achieved by administering 240 

ml of water before the overnight fast, 240 ml of water one hour before dosing, 240 

ml of water with the study dose, and 240 ml of water every 2 hours for 6 hours postdosing. 

• Patients must be adequately informed of possible cardiovascular adverse effects in 

the consent form.

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2. Statistical Analysis of Pharmacokinetic Data (Blood Plasma/Serum)

The following pharmacokinetic data should be used for the evaluation of bioequivalence 

of the multiple dose study: 

• Individual and mean blood drug concentration levels 

• Individual and mean trough levels (Cmin ss) 

• Individual and mean peak levels (Cmax ss) 

• Calculation of individual and mean steady-state AUCinterdose (AUCinterdose is AUC 

during a dosing interval at steady-state) 

• Individual and mean percent fluctuation [ =100 * (Cmax ss – Cmin ss)/Caverage ss] 

• Individual and mean time to peak concentration 

The log-transformed AUC and Cmax data should be analyzed statistically using analysis of 

variance. The 90% confidence interval for the ratio of the geometric means of the 

pharmacokinetic parameters (AUC and Cmax) should be within 80-125%. Fluctuation 

for the test product should be evaluated for comparability with the fluctuation of the 

reference product. The trough concentration data should also be analyzed statistically to 

verify that steady-state was achieved prior to Period 1 and Period 2 pharmacokinetic 

sampling. 

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 See 21 CFR 50.25. 

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Contains Nonbinding Recommendations 

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3. Clinical Report and Adverse Reactions 

Patient medical histories, physical examination and laboratory reports, and all incidents 

of possible adverse reactions should be reported. 

IV. IN VITRO TESTING CRITERIA 

A. Dissolution Testing 

Dissolution testing on 12 dosage units of the test product versus 12 units of the reference 

product should be conducted for all strengths. The lot used in the biostudy should be 

used for dissolution testing as well. The United States Pharmacopeia (USP) method is 

recommended for this product. Sampling times of 15, 30, 45 and 60 minutes are 

recommended. 

The percent of label claim dissolved at each specified testing interval should be reported 

for each individual dosage unit. The mean percent dissolved, the range (highest, lowest) 

of dissolution, the coefficient of variation (relative standard deviation), and similarity 

comparisons of dissolution profiles (f2 calculations) should be reported. 

B. Content Uniformity Test 

Content uniformity testing on the test product lots should be performed as described in 

the latest edition of the USP. 

V. WAIVER REQUIREMENTS 

Waiver of in vivo bioequivalence study requirements for the lower strengths of a generic product 

can be granted if the following conditions are met:4

1. The in vivo study on the 100 mg tablet is acceptable. 

2. The strengths are proportionally similar in active and inactive ingredients to the 

strength tested in vivo. 

3. All strengths meet an appropriate in vitro dissolution test. 

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 See 21 CFR 320.22(d)(2) 

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