PERMISSIBLE DAILY EXPOSURE (PDE) DERIVATION DATA

Drug Substance Name Phenytoin Sodium

Company Name: Toxserve Consultancy

Company Address: Maninagar, Ahmedabad - 380008

Name of Author and Reviewer: Kunal Shah, MS (Pharm), Pharmacology and Toxicology,
Sr. Manager, Toxserve Consultancy

Signature and date:

Assessment Review Date:

Sponsor Medical Reviewer Name, Sign and Date:

I. PDE SUMMARY:

Upon extensive review of literature and available non-clinical and human safety data, following PDE for
systemic toxicity was derived for Phenytoin:

Drug Name Phenytoin

Injectable (IV): 33 µg/day (equivalent to 36 µg/day for
Phenytoin Sodium)
Permitted Daily Exposure (PDE) Oral: 59 µg/day (equivalent to 64 µg/day for Phenytoin
Sodium)

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 PERMISSIBLE DAILY EXPOSURE (PDE) DERIVATION DATA
Drug Substance Name Phenytoin Sodium

Table of Contents
I. PDE SUMMARY: ............................................................................................................................... 1
II. INTRODUCTION ................................................................................................................................ 3
III. BASIC DRUG SUBSTANCE INFORMATION .......................................................................... 4
IV. CLINICAL PHARMACOLOGY .................................................................................................... 5
Mechanism of Action: ................................................................................................................. 5
Indication: .................................................................................................................................. 5
Therapeutically Recommended Dose: ........................................................................................... 5
Adverse Health effect: ................................................................................................................. 6
Pharmacokinetic Property: ..................................................................................................... 7
V. HAZARDS IDENTIFIED .................................................................................................................... 8
VI. HAZARD CHARECTERISATION ............................................................................................... 8
Acute Toxicity: ............................................................................................................................ 8
Repeated Dose Toxicity: .............................................................................................................. 9
Reproductive/developmental toxicity:.......................................................................................... 10
Genotoxicity: ............................................................................................................................ 11
Carcinogenicity: ........................................................................................................................ 11
Sensitizing potential: ................................................................................................................. 12
Other Toxicity Studies: .............................................................................................................. 12
VII. BASIS OF PDE: ........................................................................................................................... 12
VIII. SUMMARY AND CONCLUSION: ............................................................................................. 16
IX. SUMMARY OF THE AUTHOR CV ........................................................................................... 19

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Drug Substance Name Phenytoin Sodium

II. INTRODUCTION

The procedure for the determination of the PDE is based on the health effects of the residual active
substances in a manufactured product. For this we used the method for establishing the so-called
Permitted Daily Exposure (PDE) as described in Appendix 3 of ICH Q3C (R6) “Impurities: Guideline for
residual solvents”, Guidance on aspects of cleaning validation In active pharmaceutical ingredient plants
by Active Pharmaceutical Ingredients Committee (APIC), and Guideline on setting health based exposure
limits for use in risk identification in the manufacture of different medicinal products in shared facilities.
EMA/CHMP/CVMP/ SWP/169430/2012 (20 November 2014). The PDE represents a substance specific
dose that is unlikely to cause adverse effects if an individual is exposed at or below this dose every day
for lifetime. (1) (2) (3)

The determination of PDE involves:

1. Hazard identification by reviewing all relevant data

2. Identification of “critical effects”

3. Determination of the no-observed-adverse-effects level (NOAEL) of the findings that are

considered to be critical effects

4. Use of several adjustment factors to account various uncertainties.

As per European Medicines Agency (EMA) guideline EMA/CHMP/CVMP/SWP/169430/2012, following

equation is used for the derivation of the PDE:

NOAEL x Weight Adjustment

PDE =
F1 x F2 x F3 x F4 x F5

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III. BASIC DRUG SUBSTANCE INFORMATION

 Dilantin
Full Chemical Name /  Phenytoin Sodium
Synonyms  5,5-Diphenylhydantoin
 5,5-diphenylimidazolidine-2,4-dione

CAS Number 630-93-3

Molecular Formula C15H11N2NaO2

Structural Formula

Molecular Weight  274.25 g/mol

Phenytoin appears as fine white or almost white crystalline

Appearance and Solubility powder. Odorless or almost odorless. Tasteless. Soluble in
acetic acid; slightly soluble in ethyl ether, benzene, chloroform
(4).

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IV. CLINICAL PHARMACOLOGY

Mechanism of Action:
Phenytoin is a hydantoin derivative and a non-sedative antiepileptic agent with anticonvulsant activity.
Phenytoin potentially acts by promoting sodium efflux from neurons located in the motor cortex reducing
post-tetanic potentiation at synapses. The reduction of potentiation prevents cortical seizure foci
spreading to adjacent areas, stabilizing the threshold against hyperexcitability. In addition, this agent
appears to reduce sensitivity of muscle spindles to stretch causing muscle relaxation (4).

Indication:
Oral: Phenytoin is used for control of tonic-clonic seizures (grand mal epilepsy), partial seizures (focal
including temporal lobe) or a combination of these, and for the prevention and treatment of seizures
occurring during or following neurosurgery and/or severe head injury. Phenytoin has also been employed
in the treatment of trigeminal neuralgia but it should only be used as second line therapy if
carbamazepine is ineffective or patients are intolerant to carbamazepine (5).
Parenteral: Phenytoin is used for control of status epilepticus and the prevention of seizures occurring
during or following neurosurgery. It is also used for treatment of certain cardiac dysrhythmias, particularly
those unresponsive to conventional antidysrhythmic agents or to cardioversion. (6).

Therapeutically Recommended Dose:

Oral
adult dosage for seizure: Initially 3 to 4 mg/kg/day with subsequent dosage adjustment if necessary. For
most adults a satisfactory maintenance dose will be 200 mg to 500 mg daily in single or divided doses.
Exceptionally, a daily dose outside this range may be indicated. Dosage should normally be adjusted
according to serum levels where assay facilities exist (5).
Adult dosage for trigeminal neuralgia: The clinically effective dose has not been established in clinical
trials. In adults, 300-500 mg daily given in divided doses has been reported in the literature. Dosing
should be adjusted based on clinical response. Determination of serum phenytoin levels is advised.
Levels of total phenytoin should not exceed 20 µg/ml (5).
Paediatric population dosage for seizures: Initially, 5 mg/kg/day in two divided doses, with subsequent
dosage individualised to a maximum of 300 mg daily. A recommended daily maintenance dosage is
usually 4 mg/kg - 8 mg/kg (5).
Parenteral:
Status Epilepticus: Following the use of diazepam in patients having continuous seizures and in the initial
management of serial epilepsy a loading dose of Phenytoin 10 mg/kg - 15 mg/kg should be injected
slowly intravenously, at a rate not exceeding 50 mg per minute in adults (this will require approximately
20 minutes in a 70 kg patient). The loading dose should be followed by maintenance doses of 100 mg
orally or intravenously every 6 to 8 hours.
Cardiac Arrhythmias: Dosage 3.5 mg/kg - 5 mg/kg of bodyweight intravenously initially, repeated once if
necessary.
Neurosurgery: In a patient who has not previously received the drug, Parenteral phenytoin 100 mg - 200
mg (2-4 ml) may be administered intramuscularly at approximately 4-hours intervals prophylactically
during neurosurgery and continued during the post-operative period for 48-72 hrs. The dosage should
then be reduced to a maintenance dose of 300 mg and adjusted according to serum level estimations (6).

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Dosage is individualized because of the great variation of response among patients and the relatively
narrow therapeutic serum concentration range. A therapeutic serum concentration of phenytoin is 10 to
20 µg/mL, which results from approximately 4 to 8 mg/kg-BW/day (14 to 164 mg/d) for infants to age 6, to
3 to 15 mg/kg-BW/day (minimum adult dose is 300 mg/d) for children over age 6 to adults (7).

Adverse Health effect:

The most common adverse reactions are nervous system reactions, including nystagmus, ataxia, slurred
speech, decreased coordination, somnolence, and mental confusion (8).

The following adverse reactions associated with the use of Phenytoin were identified in clinical studies or
post marketing reports. Because these reactions are reported voluntarily from a population of uncertain
size, it is not always possible to reliably estimate their frequency or establish a causal relationship to drug
exposure. The most notable signs of toxicity associated with the intravenous use of this drug are
cardiovascular collapse and/or CNS depression. Hypotension does occur when the drug is administered
rapidly by the intravenous route. The rate of administration is very important; it should not exceed 50 mg
per minute in adults, and 1 to 3 mg/kg/min (or 50 mg per minute, whichever is slower) in pediatric patients

Body As a Whole: Allergic reactions in the form of rash and rarely more serious forms and DRESS (Drug
Reaction with Eosinophilia and Systemic Symptoms) have been observed. Anaphylaxis has also been
reported. There have also been reports of coarsening of facial features, systemic lupus erythematosus,
periarteritis nodosa, and immunoglobulin abnormalities.
Cardiovascular: Severe cardiovascular events and fatalities have been reported with atrial and ventricular
conduction depression and ventricular fibrillation. Severe complications are most commonly encountered
in elderly or critically ill patients.
Digestive System: Acute hepatic failure, toxic hepatitis, liver damage, nausea, vomiting, constipation,
enlargement of the lips, and gingival hyperplasia.
Hematologic and Lymphatic System: Hematopoietic complications, some fatal, have occasionally been
reported in association with administration of phenytoin. These have included thrombocytopenia,
leukopenia, granulocytopenia, agranulocytosis, and pancytopenia with or without bone marrow
suppression. While macrocytosis and megaloblastic anemia have occurred, these conditions usually
respond to folic acid therapy. Lymphadenopathy, including benign lymph node hyperplasia,
pseudolymphoma, lymphoma, and Hodgkin's Disease have been reported.
Laboratory Test Abnormality: Phenytoin may decrease serum concentrations of thyroid hormone (T4 and
T3), sometimes with an accompanying increase in thyroid-stimulating hormone (TSH), but usually in the
absence of clinical hypothyroidism. Phenytoin may also produce lower than normal values for
dexamethasone or metyrapone tests. Phenytoin may also cause increased serum levels of glucose,
alkaline phosphatase, and gamma glutamyl transpeptidase (GGT).
Nervous System: The most common adverse reactions encountered with phenytoin therapy are nervous
system reactions and are usually dose-related. Reactions include nystagmus, ataxia, slurred speech,
decreased coordination, somnolence, and mental confusion. Dizziness, vertigo, insomnia, transient
nervousness, motor twitchings, paresthesia, and headaches have also been observed. There have also
been rare reports of phenytoin induced dyskinesias, including chorea, dystonia, tremor and asterixis,
similar to those induced by phenothiazine and other neuroleptic drugs. Cerebellar atrophy has been
reported, and appears more likely in settings of elevated phenytoin levels and/or long-term phenytoin use.

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Dermatologic: Dermatological manifestations sometimes accompanied by fever have included

scarlatiniform or morbilliform rashes. A morbilliform rash (measles-like) is the most common; other types
of dermatitis are seen more rarely. Serious and sometimes fatal dermatologic reactions, including toxic
epidermal necrolysis (TEN) and StevensJohnson syndrome (SJS), have been reported with phenytoin
treatment. The onset of symptoms is usually within 28 days, but can occur later. DILANTIN should be
discontinued at the first sign of a rash, unless the rash is clearly not drug-related. If signs or symptoms
suggest SJS/TEN, use of this drug should not be resumed and alternative therapy should be considered.
Studies in patients of Chinese ancestry have found a strong association between the risk of developing
SJS/TEN and the presence of HLA-B*1502, an inherited allelic variant of the HLA B gene, in patients
using carbamazepine. Limited evidence suggests that HLA-B*1502 may be a risk factor for the
development of SJS/TEN in patients of Asian ancestry taking other antiepileptic drugs associated with
SJS/TEN, including phenytoin. Consideration should be given to avoiding phenytoin as an alternative for
carbamazepine in patients positive for HLA-B*1502. If a rash occurs, the patient should be evaluated for
signs and symptoms of Drug Reaction with Eosinophilia and Systemic Symptoms (DRESS). DRESS, also
known as Multiorgan hypersensitivity, has been reported in patients taking antiepileptic drugs, including
Phenytoin. Some of these events have been fatal or life-threatening. DRESS typically, although not
exclusively, presents with fever, rash, lymphadenopathy, and/or facial swelling, in association with other
organ system involvement, such as hepatitis, nephritis, hematological abnormalities, myocarditis, or
myositis sometimes resembling an acute viral infection. Eosinophilia is often present. Because this
disorder is variable in its expression, other organ systems not noted here may be involved. It is important
to note that early manifestations of hypersensitivity, such as fever or lymphadenopathy, may be present
even though rash is not evident. If such signs or symptoms are present, the patient should be evaluated
immediately. Phenytoin should be discontinued if an alternative etiology for the signs or symptoms cannot
be established. There have also been reports of hypertrichosis.
Local toxicity: Soft tissue irritation and inflammation has occurred at the site of injection with and without
extravasation of intravenous phenytoin. Edema, discoloration, and pain distal to the site of injection
(described as “purple glove syndrome”) have also been reported following peripheral intravenous
phenytoin injection. Soft tissue irritation may vary from slight tenderness to extensive necrosis, and
sloughing. The syndrome may not develop for several days after injection. Although resolution of
symptoms may be spontaneous, skin necrosis and limb ischemia have occurred and required such
interventions as fasciotomies, skin grafting, and, in rare cases, amputation. Because of the risk of local
toxicity, intravenous Phenytoin should be administered directly into a large peripheral or central vein
through a large-gauge catheter. Prior to the administration, the patency of the IV catheter should be
tested with a flush of sterile saline. Each injection of parenteral Phenytoin should then be followed by a
flush of sterile saline through the same catheter to avoid local venous irritation caused by the alkalinity of
the solution. Intramuscular Phenytoin administration may cause pain, necrosis, and abscess formation at
the injection site.
Special Senses: Altered taste sensation including metallic taste.
Urogenital: Peyronie’s disease. (8)

Pharmacokinetic Property:
Absorption:
Studies using Dilantin have shown that phenytoin and its sodium salt are usually completely absorbed
from the GI tract. Absorption of phenytoin is slow and variable among products (poor in neonates) for oral
administration, immediate for iv administration, and very slow but complete (92%) for intramuscular
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Drug Substance Name Phenytoin Sodium

administration (4).The absolute bioavailability of an oral dosage form (capsules) varied from 57.7 to
85.6% (9).

Distribution:
Phenytoin’s volume of distribution has been estimated to be between 0.52 and 1.19 litres/kg, and it is
highly protein bound (usually 90% in adults). In serum, phenytoin binds rapidly and reversibly to proteins.
About 90% of phenytoin in plasma is bound to albumin. The plasma half-life of phenytoin in man
averages 22 hours with a range of 7 to 42 hours (6).

Metabolism:
Phenytoin is hydroxylated in the liver by an enzyme system which is saturable. Small incremental doses
may produce very substantial increases in serum levels when these are in the upper range of therapeutic
concentrations (6).
A small percentage of individuals who have been treated with phenytoin have been shown to metabolize
the drug slowly. Slow metabolism may be caused by limited enzyme availability and lack of induction; it
appears to be genetically determined. If early signs of dose-related central nervous system (CNS) toxicity
develop, serum levels should be checked immediately. Further, Phenytoin is a potent inducer of hepatic
drug-metabolizing enzymes (8).

Excretion:
The parameters controlling elimination are also subject to wide interpatient variation. The serum level
achieved by a given dose is therefore also subject to wide variation (6).

V. HAZARDS IDENTIFIED

A thorough and comprehensive literature search was performed for acute dose toxicity, repeated dose
toxicity, reproductive and developmental toxicity, genotoxicity and carcinogenicity endpoints. The
available relevant information has been included in the section below:

Hazards Yes No Unknown

Genotoxicity X

Reproductive/developmental toxicity X

Carcinogenicity X

Highly sensitizing potential X

VI. HAZARD CHARECTERISATION

Acute Toxicity:
Acute toxicity (LD50) of Phenytoin is reported as below (10):

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Mouse Oral: 150 mg/kg

Rat Oral 1635mg/kg
Rat Intravenous 96mg/kg
Rat IM >337mg/kg
Rabbit Oral >3000mg/kg

In humans, the lethal dose in pediatric patients is not known. The lethal dose in adults is estimated to be
2 to 5 grams. The initial symptoms are nystagmus, ataxia, and dysarthria. Other signs are tremor,
hyperreflexia, lethargy, slurred speech, blurred vision, nausea, and vomiting. The patient may become
comatose and hypotensive. Death is caused by respiratory and circulatory depression. There are marked
variations among individuals with respect to phenytoin serum levels where toxicity may occur.
Nystagmus, on lateral gaze, usually appears at 20 mcg/mL, ataxia at 30 mcg/mL, dysarthria and lethargy
appear when the serum concentration is over 40 mcg/mL, but as high a concentration as 50 mcg/mL has
been reported without evidence of toxicity. As much as 25 times the therapeutic dose has been taken to
result in a serum concentration over 100 mcg/mL with complete recovery. Irreversible cerebellar
dysfunction and atrophy have been reported (8).

Repeated Dose Toxicity:

Human:
A decreased/detrimental cognitive effect (alertness, attention, and memory) were observed following one
day, seven day, and one month, respectively, of phenytoin exposure. The one-month exposure also
detected mood effects. Phenytoin dosage was 10 mg/kg BW-d, 200 to 250 mg/d (average 3.3 mg/kg BW-
d for 21.7 ± 2.32 year age, and 200 to 600 mg/d (mean 404 mg/d and average 5.6 mg/kg BW-d for 30
year mean age, respectively, administered to adolescents, adults, and/or mature adults (7).

In another human study. cerebellar volume was measured using magnetic resonance imaging scans of
23 male and 33 female epilepsy patients aged 4 to 65 years (mean 33.6 years) exposed to phenytoin for
two or more months. Mean daily dose of phenytoin was 301 mg (range 100 to 650 mg) (mean 4.2 mg/kg
BW-d for 33.6 years). Blood serum levels of phenytoin were available in 18 patients, with toxic levels
found in 9 patients. History of alcohol consumption was positive in 12 (21.5 percent) patients. Smoking
was not included as a risk factor in this study. Cerebellar volumes were transformed into Z-scores and
volumes below minus 2 SD from the mean of the control group were considered abnormal. Abnormal
(decreased volume) cerebellar atrophy in this group was noted when volumes were compared to a
healthy human control group. The atrophy correlated with duration of epilepsy (p = 0.01) and duration of
treatment with phenytoin (p = 0.001) but, not with age, age at seizure onset, maximum dosage used, or
mean daily dosage of phenytoin (7).

Animals:
In a 13-week toxicity study rats were dosed 300, 600, 1200, 2400 and 4,800 ppm Phenytoin in the diet.
The final mean body weights of males and females exposed to 2,400 or 4,800 ppm were significantly
decreased. No chemical-related gross lesions were present in the tissues of exposed rats.
Microscopically, centrilobular hypertrophy of hepatocytes was observed in the liver of rats in the 4,800
ppm groups. NOAEL = 1200 ppm (60 mg/kg/day) (11).

In 13-week toxicity study, mice were exposed at 75, 150, 300, 600 and 1,200 ppm Phenytoin in the diet.
With the exception of one male, all mice exposed to 1,200 ppm died before the end of the study. No other
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chemical-related deaths occurred. All groups of mice except the 1,200 ppm groups gained weight over
the 13-week period; however, an exposure related decrease in body weight gain was seen in males and
females. Feed consumption by exposed and control groups was generally similar. Chemical related
histomorphology lesions were present in the liver of exposed mice, particularly 600 ppm males, and
consisted of centrilobular hypertrophy of hepatocytes. Females appeared to be less sensitive than males
to the effects of drug on growth and on histomorphology liver lesions. LOAEL = 75 ppm (11.25
mg/kg/day) (11)

Reproductive/developmental toxicity:
Human:
In humans, prenatal exposure to phenytoin may increase the risks for congenital malformations and other
adverse developmental outcomes. Prenatal phenytoin exposure is associated with an increased
incidence of major malformations, including orofacial clefts and cardiac defects. In addition, the fetal
hydantoin syndrome, a pattern of abnormalities including dysmorphic skull and facial features, nail and
digit hypoplasia, growth abnormalities (including microcephaly), and cognitive deficits has been reported
among children born to epileptic women who took phenytoin alone or in combination with other
antiepileptic drugs during pregnancy. There have been several reported cases of malignancies, including
neuroblastoma, in children whose mothers received phenytoin during pregnancy. Meta-analyses using
data from published observational studies and registries have estimated an approximately 2.4-fold
increased risk for any major malformation in children with prenatal phenytoin exposure compared to
controls. An increased risk of heart defects, facial clefts, and digital hypoplasia has been reported (8).

A potentially life-threatening bleeding disorder related to decreased levels of vitamin K-dependent clotting
factors may occur in newborns exposed to phenytoin in utero. This drug-induced condition can be
prevented with vitamin K administration to the mother before delivery and to the neonate after birth (8).

Animals:
There are several studies available in literature evaluating reproductive and developmental toxicity of
Phenytoin (the effect dose level in mg/kg/day indicated in {} bracket):

Decreased fetal weight was observed in mice {250} and decreased whole brain {100} and hindbrain
weights {150} were detected in rats. Body weight differences were detected in neonatal mice {19},
pubertal rats {200}, and sexually mature rats {150}. Additionally, neonatal survival was decreased in rats
{150, 200}. Consistent with the symptoms of fetal hydantoin syndrome in humans, the teratogenicity {cleft
palate} of phenytoin was documented in mice {45}. A possible correlation with this observation is a
detection of altered craniofacial gene expression in embryonic mice {60}. (7)

Phenytoin effected changes in developmental assessments in rats and included accelerated eye opening
and olfactory orientation {50}, delayed reflex function {50}, and decreased startle response {50}. Memory
and learning in rats was also affected by phenytoin and included decreased radial maze and decreased
nonmatching-to-sample abilities {50}, spatial reference memory-based learning deficit {200}, and complex
maze deficit {100}. Brain concentrations of neuropeptides were affected by phenytoin in rats and found to
be decreased in the mesolimbic cortex for somatostatin and increased in the hippocampus and amygdala
for neuropeptide Y {100}. Phenytoin also caused increased adrenaline and noradrenaline concentrations
in response to stress in rats {150} and increased hyperexcitability in monkeys {20}. (7)

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To examine correlations between dose levels of phenytoin (PHT) and neurotoxic effects on cerebellar
development, 10, 17.5, 25, and 35 mg/kg PHT suspended in sesame oil administered orally to newborn
Jcl:ICR mice once a day during postnatal days 2–4 and determined plasma PHT concentrations during
the administration period. Mortality rates were 12.5% and 35.2% in males and 15.3% and 33.3% in
females for the 25 and 35 mg/kg PHT-treated groups during the PHT treatment, respectively. In the 25
and 35 mg/kg PHT-treated groups, total brain weight, the size of the cerebellum, and cerebellar weight
were significantly reduced on postnatal day 21. However, in the 10 and 17.5 mg/kg PHT-treated groups,
total brain weight and the size and weight of the cerebellum did not differ from those of the control group.
Histologically, the number of pyknotic cells in the external granular layer (EGL) in the 25 and 35 mg/kg
PHT-treated groups was increased on postnatal day 5, and the EGL was thicker than in the control group
on postnatal day 14. Some of the Purkinje cells in the 35 mg/kg PHT-treated group showed degeneration.
Plasma PHT levels were 10.7 ± 2.2 and 24.6 ± 2.6 μg/ml in the 25 and 35 mg/kg PHT groups on the third
day of PHT treatment, respectively. In the 25 mg/kg PHT group, plasma PHT level was found to be in the
therapeutic range for humans, 10–20 μg/mL (12).

The neurotoxic effects of PHT was examined on postnatal brain development, PHT was administered at
doses of 10, 17.5, 25, or 35 mg/kg to newborn mice once a day during postnatal days (PD) 2-4. Total
brain weight, cerebral weight, and cerebellar weight in the group treated with 25 or 35 mg/kg were
significantly reduced compared to controls from PD 5 to 21. Mice treated with PHT at 25 or 35 mg/kg
showed decreased locomotor abilities and righting reflex on PD 5. In all phenytoin treatment groups,
phenytoin levels in the brain were higher than those in the plasma on the third day of PHT treatment.
Decreased early motor function, motor coordination, locomotor activity, and learning were also observed.
This study reveals neurotoxic effects of PHT on postnatal brain development in mice. LOAEL (post-natal
development) = 10 mg/kg/day (13).

Genotoxicity:
Phenytoin was negative in in-vivo genotoxicity assays of Ames Salmonella test, and mouse lymphoma
and chromosome aberrations tests in CHO cells. Further, in the in-vivo tests of bone marrow
micronucleus, bone marrow chromosome aberrations and Drosophila tests, phenytoin was negative.
However, it produced positive result in sister chromatid exchange (SCE) assay in CHO cells in-vitro and
equivocal results bone marrow SCE in-vivo. Based on this result, a mutagenic MOA for phenytoin can
neither be ruled out nor accepted (7).

Carcinogenicity:
Two well-conducted lifetime (chronic) oral exposure cancer studies in rats and mice are available: NTP
and Dethloff et al.

The NTP study included two-year feed (oral diet) experiments for F344/N rats and B6C3F1 mice. This
study found that chronic feeding of phenytoin did not increase tumor incidences in female F344 rats or
male B6C3F1 mice. The male rats that were fed a diet containing 2,400 ppm (122 mg/kg) were found to
have marginally increased incidence (p = 0.054) of liver neoplasms when compared to controls. There
was clear evidence of carcinogenicity in female mice, with increased incidence (p < 0.001) of
hepatocellular neoplasms. The adult female mice (F1 generation) that were fed diets containing 0, 200, or
600 ppm (0, 50, or 160 mg/kg) provide this evidence. The combined liver neoplasms (hepatocellular

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adenoma, hepatocellular carcinomas, and hepatoblastomas) incidences were 5, 14, and 30, respectively,
for each dose. The adjusted combined liver neoplasms incidence rates were reported as 13.3, 34.8, and
66.4 percent, respectively, for each dose (7) (11).

The Dethloff study is a second chronic oral exposure cancer study of phenytoin in rats (Wistar) and mice
(B6C3F1). This study found that feeding of phenytoin did not increase tumor incidences in the female
rats. The male rats were found to have an increased incidence (p < 0.01) of skin pilomatricoma at the
highest phenytoin dose (100 mg/kg). No other statistically significant increases in any tumor type,
including all liver tumors singly or combined, were found for the rats. Phenytoin was administered in the
diet at doses of 0, 10, 25, and 45 mg/kg for mice. Increased incidences (p < 0.01) of hepatocellular
adenomas were noted in male mice; however, the control group had many of these tumors, which
reduces the effectiveness of Benchmark Dose runs for developing a cancer slope factor. The female mice
had increased incidences (p < 0.01) of hepatocellular adenomas and combined liver tumors that included
hemangioma, hemangiosarcoma, hepatocellular adenoma, hepatocellular carcinoma, and Kupffer cell
sarcoma. This study did not give details in order to determine time to first tumor or how many animals
were at risk. Therefore, it is necessary to assume all 50 animals/group at the start of the study were at
risk (7).

There are other carcinogenicity studies as well evaluated in Carcinogenicity Potency Database which
derives TD50 values on each species and sex from the most potent target site in each positive
experiment. TD50, is the daily dose rate in mg/kg body weight/day to induce tumors in half of test animals
that would have remained tumor-free at zero dose. For phenytoin lowest derived TD50 value is 59.1
mg/kg/day (14).

IARC Classification: possibly carcinogenic to humans (Group 2B) (15).

Sensitizing potential:
Dermatological manifestations sometimes accompanied by fever have included scarlatiniform or
morbilliform rashes. A morbilliform rash (measles-like) is the most common; other types of dermatitis are
seen more rarely. Serious and sometimes fatal dermatologic reactions, including toxic epidermal
necrolysis (TEN) and Stevens Johnson syndrome (SJS), have been reported with phenytoin treatment
(8).

Other Toxicity Studies:

No further data available on other toxicological investigation on Phenytoin.

VII. BASIS OF PDE:

The first step in developing PDE is to identify the most relevant critical endpoint. The dose associated
with this endpoint or lack thereof, is then used as a point-of-departure (POD).

The adverse effects reported in the human studies (lower IQ, decreased/detrimental cognitive effects,
mood effects, and cerebellar atrophy) are consistent with reported phenytoin side effects (mood or mental
changes and cognitive impairment). Therefore, the low end of the therapeutic dose range (3 mg/kg BW-d)
is identified as the lowest-observed-adverse-effect-level (LOAEL) (7).

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Evaluation of the phenytoin doses and toxic effects reveals that effects due to neonatal exposure are the
most sensitive. In the mice study of neonatal exposure adverse effects includes include decreases in the
following: total brain/brainstem/cerebral/cerebellar weights, early motor function, motor coordination,
locomotor activity, and learning. These effects maybe interrelated, since motor
function/coordination/activity is controlled by the brainstem, cerebrum, and cerebellum, while learning is
centered in the cerebrum (7).

Phenytoin produced carcinogenicity in animal studies. If the mode of action (MOA) for a carcinogenic
substance is anticipated to be mutagenic, a linear (non-threshold) approach is appropriate for risk
assessment of carcinogenic substance. In the genotoxicity test batteries, six tests were negative, one
positive and one equivocal a mutagenic MOA for phenytoin can neither be ruled out nor accepted.
Therefore, the MOA for phenytoin has not been established and the default linear (non-threshold)
extrapolation is used for the cancer evaluation (7).

ICH M7 guideline recommends that linear extrapolation to a probability of 1 in 100,000 (i.e., the accepted
lifetime risk level used) is achieved by dividing the TD50 (doses giving a 50% tumor incidence equivalent
to a cancer risk probability level of 1:2). by 50,000 (16).

The lowest derived PDE value out of these above POD’s for different critical effects will be considered as
final POD.

Scenario 1: Lowest human therapeutic dose

Lowest human therapeutic dose of 3 mg/kg/day is considered as LOAEL and selected as one of the POD.

Derived PDE:

POD Critical Effects

LOAEL of 3 mg/kg/day lower IQ, decreased/detrimental cognitive effects, mood

effects, and cerebellar atrophy

APPLICATION OF ADJUSTMENT FACTORS

Factor IV/IM Oral Rationale or Comment
F1 (extrapolation between 1 1 For extrapolation from humans to humans
species)
F2 (Inter-individual variability) 10 10 A default factor of 10 for account of variability between
individuals
F3 (Duration of exposure) 5 5 The therapeutic dose represents mainly acute effects
hence uncertainty factor of 5 applied for repeated daily
administration.
F4 (Severity of toxicity) 10 10 Adverse effects (neurotoxicity) are considered to be
extension of pharmacological effects however
therapeutic dose range is very narrow with steep

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dose-response.
F5 (NOAEL/NOEL vs 3 3 POD is considered LOAEL.
LOAEL/LOEL)
Additional modifying factors (MF) 1 1 No additional factor required.
α (Correction for Bioavailability) 1.75 1 IV: Bioavailability of oral route considered to be 57%
conservatively and hence correction factor of 1.75
applied (100/57=1.75) for IV PDE.
Oral: The POD value was based on oral therapeutic
dose and the intended PDE route is oral. Hence, no
correction factor for bioavailability required.

PDE (mg/day) ( ) ( )
(Parenteral route)

= 0.057 mg/day (57 µg/day)

PDE (mg/day) ( ) ( )
(Oral route)

= 0.1 mg/day (100 µg/day)

Scenario 2: Post-natal developmental toxicity study in Mice

In a postnatal developmental toxicity in mice, neonates exposed orally to phenytoin at 10 to 35 mg/kg/day

produced neurotoxicity. Therefore, LOAEL in this study was considered to be 10 mg/kg/day.

Derived PDE:

POD Critical Effects

LOAEL of 10 mg/kg/day decreases in total brain/brainstem/cerebral/cerebellar weights,

early motor function, motor coordination, locomotor activity,
and learning.

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APPLICATION OF ADJUSTMENT FACTORS

Factor IV/IM Oral Rationale or Comment
F1 (extrapolation between 12 12 For extrapolation from mice to humans
species)
F2 (Inter-individual variability) 10 10 A default factor of 10 for account of variability
between individuals
F3 (Duration of exposure) 1 1 Based on available developmental studies neonatal
exposure are the most sensitive
F4 (Severity of toxicity) 10 10 Developmental toxicity in absence of maternal
toxicity
F5 (NOAEL/NOEL vs 3 3 POD is LOAEL.
LOAEL/LOEL)
Additional modifying factors (MF) 1 1 No additional uncertainty factor warranted.
α (Correction for Bioavailability) 1.75 1 IV: Bioavailability of oral route considered to be 57%
conservatively and hence correction factor of 1.75
applied (100/57=1.75) for IV PDE.
Oral: The POD value was based on oral therapeutic
dose and the intended PDE route is oral. Hence, no
correction factor for bioavailability required

PDE (mg/day) ( ) ( )
(Parenteral route) α

= 0.079 mg/day (79 µg/day)

PDE (mg/day) ( ) ( )
(Oral route)

= 0.138 mg/day (138 µg/day)

Scenario 3: PDE based on TD50 value for carcinogenicity

Due to positive, negative and equivocal results in genotoxicity studies, phenytoin is conservatively
considered to be genotoxic carcinogen and hence its cancer risk should be assessed using non-
threshold approach as recommended in ICH M7 guideline. A life time exposure value i.e. Acceptable
Intake (AI) can be established for mutagenic carcinogens based on rodent carcinogenicity potency data
such as TD50 values (doses giving a 50% tumour incidence equivalent to a cancer risk probability level

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of 1:2). ICH M7 guideline recommends that linear extrapolation to a probability of 1 in 100,000 (i.e., the
accepted lifetime risk level used) is achieved by dividing the TD50 (doses giving a 50% tumour incidence
equivalent to a cancer risk probability level of 1:2). by 50,000 (16). CPDB assess several carcinogenicity
experiments on phenytoin and derived TD50 value. The lowest TD50 value is 59.1 mg/kg/day based on
oral NTP mice study (14).

Bioavailability of oral route considered to be 57% conservatively and hence for IV route correction factor
of 1.75 applied (100/57=1.75).

PDE (mg/day) ( ) ( )
(Parenteral route) α( )

= 0.033 mg/day (33 µg/day)

PDE (mg/day) ( ) ( )
(Oral route) α( )

= 0.059 mg/day (59 µg/day)

VIII. SUMMARY AND CONCLUSION:

The calculated intravenous PDE for Phenytoin is 33 µg/day and oral PDE is 59 µg/day based upon the
Carcinogenic Potency Database derived TD50 value of 59.1 mg/kg/day in oral mice carcinogenicity study.

In conclusion, a PDE value for Phenytoin was determined based on the conservative scenario amongst
all of three approaches including human therapeutic dose.

The derived PDE value is expected to be protective of the adverse effects observed under clinical
settings, and systemic and developmental toxicity including carcinogenicity observed in animal studies.
The calculated value represents a sufficient margin of safety for patients.

REFERENCES:
1. European Medicines Agency. Guideline on setting health based exposure limits for use in risk
identification in the manufacture of different medicinal products in shared facilities. [Online].; 2014

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Drug Substance Name Phenytoin Sodium

[cited 2020 02 04. Available from: https://www.ema.europa.eu/en/documents/scientific-

guideline/guideline-setting-health-based-exposure-limits-use-risk-identification-manufacture-
different_en.pdf.

2. ICH HARMONISED GUIDELINE. IMPURITIES: GUIDELINE FOR RESIDUAL SOLVENTS Q3C(R6). [Online].;
2016 [cited 2020 02 04. Available from: https://database.ich.org/sites/default/files/Q3C-
R6_Guideline_ErrorCorrection_2019_0410_0.pdf.

3. Active Pharmaceutical Ingredients Committee (APIC). Guidance on aspects of cleaning validation In

active pharmaceutical ingredient plants. [Online].; 2016 [cited 2020 02 04. Available from:
https://apic.cefic.org/pub/APICCleaningValidationGuide-updateSeptember2016-final.pdf.

4. PubChem Database. Phenytoin, CID=1775, and Phenytoin sodium, CID=657302. [cited 2020 03 29.
Available from: https://pubchem.ncbi.nlm.nih.gov/compound/Phenytoin-sodium.

5. SmPC. Epanutin 30mg/5ml oral Suspension. Upjohn UK Limited.. [cited 2022 06 25. Available from:
https://www.medicines.org.uk/emc/product/2257/smpc.

6. SmPC. Phenytoin Hospira 50 mg/ml Injection BP. Hospira UK Ltd. [cited 2022 06 25. Available from:
https://www.medicines.org.uk/emc/product/3794/smpc.

7. Fritz KL. Toxicological Assessment. Phenytoin, CAS #57-41-0. Interim Draft. Michigan Waste and
Hazardous Materials Division.; 2010 [cited 2020 03 29. Available from:
https://www.michigan.gov/documents/deq/deq-rrd-
PhenytoinKLFToxicologicalAssessmentInterimDraft_323614_7.pdf.

8. US FDA Prescribing Information (US PI). DILANTIN® (phenytoin sodium) injection, for intravenous or
intramuscular use. Pfizer Inc.. [cited 2020 03 29. Available from:
https://www.accessdata.fda.gov/drugsatfda_docs/label/2017/010151s047s048lbl.pdf.

9. Guger R. et al. Phenytoin: Pharmacokinetics and bioavailability. Clinical Pharmacology &

Therapeutics. 1976 February; 19(2): 135-142.

10. Safety Data Sheet. Dilantin. Pfizer Inc.. [cited 2020 03 29. Available from: https://pfe-pfizercom-
prod.s3.amazonaws.com/products/material_safety_data/phenytoin_tablets_15-Feb-2018_0.pdf.

11. NTP (National Toxicology Program) (1993). NTP technical report on the perinatal toxicology and
carcinogenesis studies of 5,5-diphenylhydantoin in F344/N rats and B6C3F1 mice. Research Triangle
Park, NC, NIH No. 94-2859.. Available from:

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https://www.who.int/foodsafety/chem/jecfa/en/tox_guidelines.pdf.

12. Ohmori H. Effects of low-dose phenytoin administered to newborn mice on developing cerebellum.
Neurotoxicology and Teratology. 1997; 19(3): 205-211.

13. Hatta T. Neurotoxic Effects of Phenytoin on Postnatal Mouse Brain Development Following Neonatal
Administration. Neurotoxicology and Teratology. 1999; 21(1): 21-8.

14. The Carcinogenic Potency Database (CPDB). 5,5-Diphenylhydantoin (CAS 57-41-0).. [cited 2020 03
29. Available from: https://files.toxplanet.com/cpdb/chempages/5%2C5-
DIPHENYLHYDANTOIN.html.

15. IARC. Summary of Data Reported and Evaluation. 5,5-Diphenyl-2,4-imidazolidinedione. CAS# 57-41-
0. Vol. 66 (1996) (p. 175). [cited 2020 03 30. Available from:
http://www.inchem.org/documents/iarc/vol66/phenytoin.html.

16. ICH M7: Assessment and Control of DNA Reactive (Mutagenic) Impurities In Pharmaceuticals To
Limit Potential Carcinogenic Risk..; 2017 [cited 2020 03 30. Available from:
https://database.ich.org/sites/default/files/M7_R1_Guideline.pdf.

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IX. SUMMARY OF THE AUTHOR CV

KUNAL U. SHAH

SUMMARY
A competent toxicology professional with close to 7+ years of experience in various product safety and
regulatory affairs activities like ICH-Q3, EU-REACH registration, GHS/CLP classification, EC No
1223/2009, Human health hazard assessment of drug substance/cosmetic ingredients, data mining
through secondary data search and report writing

PROFESSIONAL EXPERIENCE

SENIOR MANAGER
TOXSERVE CONSULTANCY FEB 2015 – PRESENT

 Determination of Health-Based Exposure Limits (HBEL)

o Calculation of a Permitted Daily Exposure (PDE) drug substance and intermediate chemicals.
o Determination of Occupational Hazard Band (OHB) or Occupational Health Limits (OEL)
o Determining appropriate Hazard Notations for drug substance and intermediate chemicals
 Toxicological literature search and Safety Data Sheet (SDS)
o Preparation of Toxicological Profile for chemical ingredients – Endpoint wise summarization of
various animal and human studies through extensive literature review
o Safety Data Sheet SDS/MSDS compilation as per OSHA's Hazard Communication Standard
(HCS), GHS/CLP classification
 Consumer (Cosmetic) Safety Assessment for EU and Latin America market as per Regulation
(EC) No 1223/2009
o Cosmetic Ingredient Monograph/Review
o Calculation of a Margin of safety of ingredient
o Product Information file (PIFs)
o Cosmetic Product Safety Report (CPSR, Part A and Part B)

SENIOR ASSOCIATE
WNS GLOBAL SERVICES, GURGAON, INDIA MAY 2013 – JAN 2015

Supported Global Product Stewardship of the leading FMCG companies as clients of organisation
with the following work experience:
 Chemical hazard characterisation and safety assessment
o Comprehensive searching of chemicals for toxicological data in various public and private online
databases (ScienceDirect, ExPub, PubMed, eChemPortal, ChemView, ChemIDplus, Toxnet,
IPCS, PCPC, NTIS, IRIS, OECD HPV, ACToR etc.)
o Collation and abstract summarisation of identified data
o Writing toxicity profiles and hazard assessments (including analogue and/or metabolite based) for
chemicals, identifying critical study demonstrating the toxic effect of concern, selecting
NOAEL/LOAEL values and deriving safety values (e.g., MACE, RfD) by applying appropriate
safety/uncertainty factors.
o Safety assessment of data deficient chemical using Weight of Evidence (WoE) approach through
Structure Activity Relationship (SAR) based read-across and/or in-silico prediction using OECD
QSAR Toolbox.

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o Worked on wide spectrum of chemicals belonging to categories including fatty acids, fatty
alcohols, PEG, Alkyl PEG ethers (and sulphates), Alkyl sulphates, Sorbitan fatty acid esters,
Perfluoroalkyls, Petroleum products and botanicals
 EU cosmetic regulation
o Preparation of Cosmetic Product Safety Report (CPSR) for products ranging hair dyes, hair
sprays, shampoos, conditioner etc.
o Dermal absorption assessment of cosmetic ingredients
o GHS/CLP based classification and data management
o Extraction and collation of information from European Chemical Agency (ECHA) registered
substances dossier for preparation of MSDS
o Analysis of the supplier information/MSDS sheet of various chemicals and verification of chemical
identity through SciFinder for client’s internal database maintenance

RESEARCH ASSOCIATE
PIRAMAL HEALTHCARE LIMITED, MUMBAI, INDIA SEPT 2011 - MAY 2013

 Accountable for study protocol preparation, data acquisition and analysis, reporting results to
Medicinal chemistry group in oral and written forms
 Assisted in different phases (dosing, clinical pathology and necropsy) of 14 days and 28 days
repeat dose toxicity study in rodents for regulatory submission of NCEs
 Performance of acute toxicity study in rodents under supervision
 Coordinated toxicokinetic study with different functional group
 Supporting in screening of NCEs for genotoxicity; Micronucleus Test and AMES assay
 Preparation, revision and training on SOP for system, instrument, operation and process-based
works for GLP implementation
 Training on different functional group programs for improvement in the scientific aspect as well as
quality system, for the conduct and GLP compliance of the studies

EDUCATIONAL DEVELOPMENT

Masters in Pharmacology & Toxicology, 2009-2011

National Institute of Pharmaceutical Education and Research, NIPER, S.A.S. Nagar, Punjab, India; 8.28
CGPA

Bachelor of Pharmacy, 2005-2009

Gujarat University, Ahmedabad, India; First Class

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