Ganciclovir for Injection - Pharmaceutical Information, Clinical Trials, Detailed Pharmacology, Toxicology
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Ganciclovir for Injection - Scientific Information

Manufacture: Fresenius Kabi USA, LLC
Country: Canada
Condition: Cytomegalovirus Infection, CMV Retinitis
Class: Antiviral agents
Form: Intravenous (IV), Powder
Ingredients: Ganciclovir sodium

Pharmaceutical Information

Drug Substance

Proper name: ganciclovir sodium
Chemical name: 9-[[2-hydroxy-1-(hydroxymethyl)-ethoxy]methyl]guanine, monosodium salt.
Molecular formula and molecular mass: C9H12N5NaO4
277.22
Structural formula:
Physicochemical properties: Ganciclovir sodium is a white or almost white crystalline hygroscopic powder with solubility in water > 50 mg/mL

Clinical Trials

No data available.

Detailed Pharmacology

Animal Pharmacodynamic Studies

Ganciclovir exhibited minimal pharmacological activity in a battery of tests for effects on the central nervous, cardiovascular, and immune systems. No consistent effect was observed when ganciclovir was co-administered with different autonomic drugs.

Human Pharmacology

Absorption

The absolute bioavailability of oral ganciclovir under fasting conditions was approximately 5% (n = 6) and following food was 6 ‒ 9% (n = 32). When ganciclovir was administered orally with food at a total daily dose of 3 g/day (500 mg q3h, 6 times daily and 1000 mg TID), the steady-state absorption as measured by area under the serum concentration vs. time curve (AUC) over 24 hours and maximum serum concentrations (Cmax) were similar following both regimens with an AUC0-24 of 15.9 ± 4.2 (mean ± SD) and 15.4 ± 4.3 μg·hr/mL and Cmax of 1.02 ± 0.24 and 1.18 ± 0.36 μg/mL, respectively (n = 16). At the end of a one-hour intravenous infusion of 5 mg/kg ganciclovir, total AUC ranged between 22.1 ± 3.2 (n = 16) and 26.8 ± 6.1 μg·hr/mL (n = 16) and Cmax ranged between 8.27 ± 1.02 (n = 16) and 9.0 ± 1.4 μg/mL (n = 16).

Distribution

The steady-state volume of distribution of ganciclovir after intravenous administration was 0.74 ± 0.15 L/kg (n = 98). Cerebrospinal fluid concentrations obtained 0.25 and 5.67 hours post-dose in 3 patients who received 2.5 mg/kg ganciclovir intravenously q8h or q12h ranged from 0.31 to 0.68 μg/mL representing 24 to 70% of the respective plasma concentrations. Binding to plasma proteins was only 1 ‒ 2% over ganciclovir concentrations of 0.5 and 51 μg/mL; as such, drug interactions involving binding site displacement are not anticipated.

Metabolism

When administered intravenously, ganciclovir exhibits linear pharmacokinetics over the range of 1.6 to 5.0 mg/kg. Renal excretion of unchanged drug by glomerular filtration and active tubular secretion is the major route of elimination of ganciclovir. In patients with normal renal function, 91.3 ± 5.0% (n = 4) of intravenously administered ganciclovir was recovered unmetabolized in the urine. Systemic clearance of intravenously administered ganciclovir was 3.52 ± 0.80 mL/min/kg (n = 98) while renal clearance was 3.20 ± 0.80 mL/min/kg (n = 47), accounting for 91 ± 11% of the systemic clearance (n = 47).

Special Populations

Renal Impairment

The pharmacokinetics following intravenous administration of ganciclovir sodium for injection were evaluated in 10 immunocompromised patients with renal impairment who received doses ranging from 1.25 to 5 mg/kg (see Table 1).

Table 1
CrCl
(mL/min)
n Dose Clearance
(mL/min)
Mean ± SD
Half-Life
(hours)
Mean ± SD
50 – 79
25 – 49
< 25
4
3
3
3.2 – 5 mg/kg
3 – 5 mg/kg
1.25 – 5 mg/kg
128 ± 63
57 ± 8
30 ± 13
4.6 ± 1.4
4.4 ± 0.4
10.7 ± 5.7

Hemodialysis reduces plasma concentrations of ganciclovir by about 50% after i.v. administration.

Race and Gender

The effects of race and gender were examined in pharmacokinetic studies, among 50 patients receiving an oral dose of 1000 mg every 8 hours. Although the numbers of blacks (n = 8; 16%) and hispanics (n = 10; 20%) were small, there appeared to be a trend towards a lower steady-state Cmax and AUC0-8 in these subpopulations as compared to caucasians. No definitive conclusions regarding gender differences could be made because of the small number of females (n = 6; 12%); however, no differences between males and females were observed.

Pediatrics

Ganciclovir pharmacokinetics were studied in 27 neonates, aged 2 to 49 days. At an intravenous dose of 4 mg/kg (n = 14) or 6 mg/kg (n = 13), the pharmacokinetic parameters were, respectively, Cmax of 5.5 ± 1.6 and 7.0 ± 1.6 μg/mL, systemic clearance of 3.14 ± 1.75 and 3.56 ± 1.27 mL/min/kg, and t1/2 of 2.4 hours (harmonic mean) for both.

Ganciclovir pharmacokinetics were also studied in 10 children, aged 9 months to 12 years. The pharmacokinetic characteristics of ganciclovir were the same after single and multiple (q12h) intravenous doses (5 mg/kg). The steady state volume of distribution was 0.64 ± 2.2 L/kg, Cmax was 7.9 ± 3.9 μg/mL, systemic clearance was 4.7 ± 2.2 mL/min/kg, and t2 was 2.4 ± 0.7 hours. The pharmacokinetics of intravenous ganciclovir in neonates and children are similar to those observed in adults.

Elderly

No studies have been conducted in adults older than 65 years of age.

Virology

Clinical Antiviral Effect of Ganciclovir

Of 314 immunocompromised patients enrolled in an open-label study of the treatment of life- or sight-threatening CMV disease with ganciclovir sodium for injection, 121 patients were identified who had a positive culture for CMV within 7 days prior to treatment and had sequential viral cultures after treatment with ganciclovir sodium for injection. Post-treatment virologic response was defined as conversion to culture negativity, or a greater than 100-fold decrease in CMV infectious units, as shown in the following table:

Table 2
Culture Source No. Patients Cultured No. (%) Patients
Responding
Median Days to
Response
Urine
Blood
Throat
Semen
107
41
21
6
93 (87%)
34 (83%)
19 (90%)
6 (100%)
8
8
7
15

The antiviral activity of ganciclovir sodium for injection was demonstrated in two separate placebo- controlled studies for the prevention of CMV disease in transplant recipients. One hundred forty-nine heart allograft recipients who were either CMV seropositive or had received seropositive heart allografts were randomized to treatment with ganciclovir sodium for injection (5 mg/kg BID for 14 days followed by 6 mg/kg once daily for 5 days/week for an additional 14 days) or placebo. Seventy-two CMV culture-positive allogeneic bone marrow transplant recipients were randomized to treatment with ganciclovir sodium for injection (5 mg/kg BID for 7 days followed by 5 mg/kg once daily) or placebo until day 100 post-transplant. Ganciclovir sodium for injection suppressed CMV shedding in heart allograft and bone marrow allograft recipients. The antiviral effect of ganciclovir sodium for injection in these patients is summarized in the following table:

Table 3: Patients with Positive CMV Cultures
Time Heart Allograft Bone Marrow Allograft
Ganciclovir Placebo Ganciclovir Placebo
Pre-Treatment 1/67 (2%) 5/64 (8%) 37/37 (100%) 35/35 (100%)
Week 2 2/75 (3%) 11/67 (16%) 2/31 (6%) 19/28 (68%)
Week 4 3/66 (5%) 28/66 (43%) 0/24 (0%) 16/20 (80%)

The antiviral activity of ganciclovir sodium for injection was confirmed in two randomized, controlled trials for the maintenance treatment of CMV retinitis in patients with AIDS. Serial cultures of urine were obtained, and cultures of semen, biopsy specimens, blood, and other sources also were obtained when available. Only a small proportion of patients remained culture- positive during maintenance therapy with ganciclovir. The antiviral effect of ganciclovir sodium for injection in the patients in the two studies is summarized in the following table:

Table 4: Patients with Positive CMV Cultures in Two Controlled Clinical Trials
  Patients with Newly Diagnosed
CMV Retinitis*
Patients with Stable, Previously
Treated CMV Retinitis**
Intravenous Intravenous
Start of Maintenance 5/37 (13.5%) 2/66 (3.0%)
Anytime During Maintenance 3/48 (6.3%) 1/45 (2.2%)

* Study ICM1653: 3 weeks of treatment with intravenous ganciclovir before start of maintenance

** Study ICM1774: 4 weeks to 4 months treatment with intravenous ganciclovir before start of maintenance

Viral Resistance

The current working definition of CMV resistance to ganciclovir in in vitro assays is IC50 > 1.5 μg/mL (6.0 μM). CMV resistance to ganciclovir in individuals with AIDS and CMV retinitis who have not previously been treated with ganciclovir does occur. Viral resistance has been observed in patients receiving prolonged treatment with ganciclovir sodium for injection for CMV retinitis. The possibility of viral resistance should be considered in patients who show poor clinical response or experience persistent viral excretion during therapy.

The principal mechanism of resistance to ganciclovir in CMV is the decreased ability to form the active triphosphate moiety; resistant viruses have been described which contain mutations in the UL97 gene of CMV which controls the phosphorylation of ganciclovir. Mutations in the viral DNA polymerase have also been reported to confer viral resistance to ganciclovir, and may show cross-resistance to other antivirals with a similar mechanism of action.

In Vitro Studies

Ganciclovir is an inhibitor of viral replication in vitro (see Table 5). The relationship between in vitro sensitivity of CMV to antiviral drugs and clinical response has not been established.

Table 5
IN VITRO ACTIVITY OF GANCICLOVIR
Virus IC50 (μM)*
Herpes simplex virus (HSV) 2.4a
Human cytomegalovirus (HCMV) 0.4 – 11.0
Varicella Zoster Virus (VZV) 32.0
Epstein-Barr virus (EBV) 1.0
Murine cytomegalovirus (MCMV) 15.0
Guinea pig cytomegalovirus (GPCMV) 70.0

* μM = 10-6M

a Plaque Reduction Assay

The antiviral activity of ganciclovir against a number of strains of human CMV, in cell culture, is shown in Table 6.

Table 6
IN VITRO ANTIVIRAL ACTIVITY OF GANCICLOVIR AGAINST HUMAN CMV STRAINS
Human CMV Strain Cell Culture IC50 (μM)aganciclovir
AD 169 MRC-5 7
AD 169 MRC-5 1.5 – 6.2
Towne MRC-5 0.4 – 6.2
Towne WI-38 1.0
Major WI-38 4.8
BT 1943 WI-38 1.1
AD 169 HET 7
Davis MRC-5 7
Davis HET 5
Towne MRC-5 2.0
Davis MRC-5 3.1
AD 16 MRC-5 3.1
Eisenhardt-CID9 HEL 2.0
AIDS-O.L. HEL 0.8
AIDS-O.C. HEL 5.5
CHMC-CID HEL 5.9
CMV mononucleosis patients HEL 1.0 – 11.0
Renal transplant patients HEL 0.5 – 9.5
Male homosexual subjects HEL 1.0 – 5.0

a IC50: Median inhibitory dose (µM)

The IC50 of ganciclovir for a variety of cultured mammalian cells in shown in Table 7.

Table 7
EFFECTS of GANCICLOVIR ON HOST CELL PROLIFERATION
Cell Type Ganciclovir IC50 (FM S.D.)
Human Bone Marrow Colony-forming Cells 39 ± 73
Human Embryonic Lung (MRC-5) 110 ± 50
Human Embryonic Tonsil (HET) 250 ± 80
Squirrel Monkey Lung (SML) 1500 ± 95
Guinea Pig Embryo (GPE) 2900 ± 844
Mouse Embryo Fibroblast (MEF) 210 ± 80

a Results eliminate one marrow resistant to ganciclovir

In Vivo Studies

In three animal models of CMV infection ganciclovir has shown in vivo activity. These models are acute MCMV infection, MCMV lung infection and interstitial pneumonia, and acute GPCMV.

As shown in Table 8, various doses of ganciclovir were tested for efficacy against mice infected with CMV. A statistically significant increase in numbers of survivors were observed at doses of 10 mg/kg or more. A dose of 25 mg/kg of acyclovir was required to induce a statistically significant effect in the number of surviving mice.

Table 8
EFFECTS OF GANCICLOVIR ON MCMV (SMITH STRAIN) INDUCED MORTALITY WHEN TREATMENT WAS STARTED 6 HOURS AFTER INFECTION
Drug (mg/kg)a Survivors/Total Mean Survival Time (days)b
Saline 2/20 (10)c 4.4 ± 0.78d
Ganciclovir    
1 2/20 (10) 6.2 ± 1.8f
5 2/20 (10) 6.3 ± 1.4f
10 8/20 (40)e 7.7 ± 1.8f
25 15/20 (75)e 6.4 ± 0.55f
50 19/20 (95)e 7.0 ± 0.0f

a Half-daily doses were administered s.c. at 9am and 3pm for 5 days
b Of the mice that died

c Percent survival

d Standard deviation

e Statistically significant (p < 0.05) by Fisher exact test

f Statistically significant (p < 0.05) by Mann-Whitney U-test

In another experiment, treatment began either 6, 24, 48, 72 or 96 hours after the infection. A statistically significant increase in the number of survivors was observed when ganciclovir therapy was started 48 hours or less after inoculation (Table 9).

Table 9
EFFECTS of 50 mg/kg GANCICLOVIR ON MCMV (SMITH STRAIN) INDUCED MORTALITY WHEN TREATMENT WAS STARTED 6, 24, 48, 72 or 96 HOURS AFTER INFECTION
Drug (hours after infection)a Survivors/Total Mean Survival Time (days)b
Saline 2/19 (11)c 5.2 ± 1.2d
Ganciclovir    
6 18/20 (90)e 6.5 ± 0.71
24 15/19 (79)e 10.7 ± 3.8
48 9/19 (47)e 8.0 ± 2.5
72 6/20 (30)e 6.1 ± 1.6
96 1/20 (5)e 4.8 ± 0.71

a Half-daily doses were administered s.c. at 9am and 3pm for 5 days
b Of the mice that died

c Percent survival

d Standard deviation

e Statistically significant (p < 0.05) by Fisher exact test

Doses of ganciclovir ranging from approximately 40 ‒ 300 mg/kg (ad libitum in drinking water starting 24 hours post-infection) reduced salivary titers of MCMV by 84 ‒ 99 percent and lung titers by 97 ‒ 99 percent. Ganciclovir treatment of diffuse interstitial pneumonitis also reduced the replication of MCMV in both lung and salivary glands but did not block pneumonitis development.

Guinea pig CMV is very insensitive to ganciclovir (Table 5), nevertheless, ganciclovir (25 mg/kg intraperitoneally, BID for 7 days) reduces GPCMV titers in the salivary glands. Histopathology showed that the lesions in the kidney and salivary gland from ganciclovir treated animals were significantly less severe than controls.

Toxicology

Tables 10 to 14 summarize the toxicological studies conducted with ganciclovir.

The most sensitive target organ for the primary toxic effects of ganciclovir was the testis. Other systems affected by ganciclovir treatment, but less sensitive than the male reproductive system, were the hematopoietic, integumentary, female reproductive, gastrointestinal, and urinary systems and the developing embryo/fetus. Except for the effects on the male reproductive system, and for some effects on the hematopoietic system and the skin, the changes induced by the administration of the drug occurred at dosages greater than the proposed clinical dose. The adverse effects due to ganciclovir treatment were generally reversible on withdrawal of drug treatment unless the doses used were exceptionally high and except for certain effects on the developing embryo/fetus.

Table 10
ACUTE TOXICOLOGY
SPECIES
STRAIN
SEX (N)
AGE
ROUTE
PROCEDURE
VOLUME
PRETEST
CONDITIONING
DOSE
(MG/KG)
MORTALITY LD50 SIGNS OF TOXICITY
        STUDY
DAY
ANIMALS
(N)
   
Mouse
Swiss-Webster
Male (12)
Female (12)
8 – 12 weeks
Oral
Stomach tube
0.2 mL/10 g body
weight
1 mo. acclim.
Period; 3 hr. fast
prior to dosing
900
2000
-
-
0
0
LD50 > 2000 mg/kg Clinical condition normal except for
occasional inactivity and/or rough coat. No
treatment related effects for body weight
intakes.
Mouse
Swiss-Webster
Male (18)
Female (18)
8 – 12 weeks
Intravenous tail-
vein inject.
0.2 mL/10 g body
weight
1 mo. accilm.
period
0
900

2000
-
1**
6
7
8
1**
4
8
0
1M, 1F
1F
1M, 1F
1M, 1F
3M
1M, 5F
1F
Estimated LD50
900 mg/kg
Rough coat recorded for vehicle-treated
mice. Drug related effects: pallor,
unthriftiness, hypothermia, inactivity,
labored/increased respiration, necrosis and
ulceration at injection site. Decreased body
weight and food intakes observed during
week 1 postdosing.
Dog
Beagle
Male (1)
Female (1)
9 – 16 months
Oral
Stomach tube
10 mL/kg body
weight
5 – 10 mo. acclim.
period; overnight
fast prior to dosing
1000 - 0 LD50 > 1000 mg/kg No treatment-related changes observed in
body weight, gross pathology, haematology
or clinical chemistry.
Dog
Beagle
Male (1)
Female (1)
18 – 29 months
Intravenous
cephalic vein
injection
5 mL/kg
13 – 22 mo acclim.
period
500 5
7
1M
1F
Estimated LD50 
< 500 mg/kg
Anorexia, diarrhea, hypothermia, vomiting,
collapse, salivation, unthriftiness. Body
weight loss of 15
‒ 20% Sanguinous changes
in stomach and intestines. Male: leukopenia;
increased BUN, GOT, GPT
Dog
Beagle
Male (1)
Female (1)
10 Months
Intravenous
cephalic vein
injection
1.5 mL/kg
6 mo. acclim
period
150 - 0 Estimated
> 150 mg/kg
No treatment-related effects on clinical
condition or body weight. Male (day after
dosing): slight increase in RBC count,
hemoglobin hematocrit, total protein and
albumin.

* M = MALE; F = FEMALE

** Deaths for all animals occurred during or within 1 minute of dosing and were therefore not considered drug-related.

Table 11
MULTIDOSE TOXICITY STUDIES
SPECIES
STRAIN
DURATION
OF DOSING
ROUTE
DOSE
(MG/KG/DAY)
MORTALITY HEMATOLOGY CLINICAL
CHEMISTRY
ORGAN
WEIGHTS
PATHOLOGY COMMENTS
Mouse
Swiss-Webster
3 months
Gavage (Only
male dosed)
           
  0 1/45 - - - - -
  10 1/45 NDE NDE Testes
decreased
Testicular atrophy &
hypospermatogenesis,
with complete recovery
by 130 days postdosing.
Decreased fertility & increased
abnormal sperm morphology, with
recovery between 30
‒ 130 days
postdosing. No dominant lethal
effect.
  100 0/45 NDE NDE Testes
decreased
Testicular atrophy &
aspermatogenesis
Infertility & increased abnormal
sperm morphology.
  1000 4/45 NDE NDE. No
treatment-
related changes
in plasma FSH,
LH, or
testosterone
Testes
decreased
Testicular atrophy &
aspermatogenesis
Infertility & increased abnormal
sperm morphology. Decreased food
intake & body weight.
Mouse
Swiss-Webster
3 months
Gavage (Only
female dose)
           
  0 1/85 - - - - -
  100 0/85 NDE NDE NDE NDE NDE
  300 1/85 NDE NDE NDE NDE NDE
  1000 2/85 NDE NDE. No
treatment-
related changes
in plasma FSH
and LH
NDE NDE NDE

NDE = no drug-related effect

ND = not done

Table 11 (cont’d)
SPECIES
STRAIN
DURATION
OF DOSING
ROUTE
DOSE
(MG/KG/DAY)
MORTALITY HEMATOLOGY CLINICAL
CHEMISTRY
ORGAN
WEIGHTS
PATHOLOGY COMMENTS
Mouse
Swiss-Webster
1 month
Intravenous
(males &
females dosed)
           
  0 0/25(M)
0/25 (F)
- - - - -
  15 0/25(M)
0/25 (F)
NDE NDE NDE Testes decreased
Spleen increased
(females)
Aspermatogenesis and reproductive
organ atrophy (males). Partial
recovery at 1 month postdosing.
  45 5/25(M)
3/25 (F)
Hypothermia,
inactivity, pallor,
rough coat, wasting,
unthriftness.
Recovery 1 week
post-dosing.
NDE NDE Testes decreased
Spleen increased
Aspermatogenesis and reproductive
organ atrophy (males). Renal
cortical damage.
Atrophy of skin adnexal tissue.
Little evidence of recovery at 1
month postdosing.
  135 8/25(M)
7/25 (F)
Decreased body
weight during first
&/or second week
of treatment.
Hypothermia
inactivity, pallor,
rough coat, wasting,
unthriftiness.
Recovery 1 week
postdosing.
Decreased
erythrocyte count,
hemoglobin &
haematocrit.
Abnormal
erythrocyte
morphology in some
mice. Complete
recovery at 1 month
postdosing.
Increased
GOT, GPT, &
BUN in
females.
Complete
recovery at 1
month
postdosing.
Males: testes,
prostate gland &
seminal vesicles
decreased; spleen
increased.

Females: uterus
decreased; spleen
liver and kidney
increased.
Aspermatogenesis. Reproductive
organ atrophy (males & females).
Inhibition of ovarian cycling. Renal
cortical damage. Atrophy of skin
adnexal tissue. Little evidence of
recovery at 1 month postdosing.
Rat
Sprague-Dawley
3 months
Oral (in feed)
(males and
females dosed)
           
  0 0/6 (M)
0/6 (F)
- - - - -
  100 0/6 (M)
0/6 (F)
NDE NDE NDE Testes decreased Testicular atrophy and
aspermatogenic tubules.
  500 0/6 (M)
0/6 (F)
NDE NDE NDE Testes decreased Testicular atrophy and
aspermatogenic tubules.
  1400 0/6 (M)
0/6 (F)
NDE NDE NDE Testes decreased Testicular atrophy and
aspermatogenic tubules.
  5000 1/6 (M)
0/6 (F)
8-12% decrease in
body weight gain
NDE NDE Testes decreased Testicular atrophy and
aspermatogenic tubules.
Dog
Beagle
1 month
Intravenous            
  0 0/3 (M)
0/2 (F)
- - - - -
  0.4 0/3 (M) NDE NDE NDE   Testicular atrophy and
hypospermatogenesis after a one
month recovery.
  1.2 0/3 (M)
0/2 (F)
NDE NDE NDE   Testicular atrophy and
hypospermatogenesis after a one
month recovery.
  3.6 0/3 (M)
0/2 (F)
NDE NDE NDE   Testicular atrophy and
hypospermatogenesis. Recovery
expected based on presence of
spermatogonia and primary
spermatocytes one month
postdosing.
Dog
Beagle
3 months
Oral tube
(stomach)
           
  Males            
  0 0/6 - - - - -
  0.2 0/6 NDE NDE NDE NDE Minimal-to-slight testicular atrophy
and hypospermatogenesis.
Sebaceous gland atrophy. Complete
recovery from all lesions by 65 days
postdosing.
  2.0 0/6 Decr. body weight
& testes vol.
NDE NDE Testes dec. Testicular atrophy &
aspermatogenesis. Decr.
epididymal sperm. Sebaceous gland
atrophy. Complete recovery from
all lesions by 4 mths postdosing.
  20.0 0/6 Decr. body wgt.
and testes vol.
lacrimation
Decr. hemoglobin
& hematocrit. Decr.
erythrocyte,
leukocyte & platelet
counts.
No treatment
related changes
in FSH, LH or
testosterone
Testes dec. Testicular atrophy &
aspermatogenesis. Decr. epididymal
sperm. Bone marrow
hypocellularity. Sebaceous gland &
hair follicle atrophy. Complete
recovery from all lesions by
4 months postdosing.
  Females            
  0 0/6 - - - - -
  2.0 0/6 NDE NDE NDE NDE NDE
  6.0 0/6 Decr. body weight
Lacrimination
Decr. leukocyte
and platelet
counts.
NDE NDE Bone marrow hypocellularity.
Sebaceous gland atrophy. Complete
recovery from all lesions by 65 days
postdosing.
  20.0 1/6 Decr. body wgt.
Lacrimation
Decr. hemoglobin
& hematocrit.
Decr. erythrocyte,
leukocyte &
platelet counts.
No treatment
related changes
in FSH, LH or
testosterone.
NDE Bone marrow hypocellularity.
Sebaceous gland and hair follicle
atrophy. Complete
recovery from all lesions by
4 months postdosing
Dog
Beagle
1 month
i.v.            
  0 0/3 (M)
0/3 (F)
- - - - -
  10 0/3 (M)
0/3 (F)
Clear ocular
discharge
Leukocyte count
slightly decreased
NDE Testes decr. Decr. bone marrow cellularity.
Atrophic testes. Sebaceous
gland atrophy.
  30 0/3 (M)
1/3 (F)
Clear ocular
discharge in all
dogs. Decr. body
wgt, anorexia
emesis hypothermia
& bloody diarrhea
in female that died.
Decr. leukocytes,
platelets, and/or
reticulocytes in all
dogs partial
recovery at 2 wks
postdosing. Decr.
erythrocytes,
hemoglobin &
hematocrit in female
that died.
Effects similar
to high-dose
dogs were seen
in female that
died.
Testes decr. Decr. bone marrow cellularity.
Partial recovery at 2 wks.
postdosing. Atrophic testes.
Epidermal, sebaceous and hair
follicle atrophy.
  90 3/3 (M)
3/3 (F)
Decr. body weight
anorexia emesis,
hypothermia bloody
diarrhea
dehydration and
clear ocular
discharge in all
dogs.
Decr. leukocytes,
reticulocytes &
platelets. Incr.
erythrocytes
hemoglobin and
hematocrit.
Decr. Na & Cl.
Incr. BUN
creatinine
alkaline
Phosphatase,
phosphorous,
cholesterol
& triglycerides.
Testes normal
dogs died too
early to show
effects.
Decr. bone marrow cellularity.
Severe gastrointestinal degeneration
& atrophy. Renal tubular dilation.
Epidermal sebaceous and hair
follicle atrophy. Thymic involution.
Ovarian suppression and lymphoid
atrophy, possibly secondary to
stress of systemic illness.

NDE = no drug-related effect; Observations apply to both male and females, unless specified otherwise.

Table 12
Reproduction Studies
SPECIES
STRAIN &
SEX/GROUP
DOSE ROUTE
BREEDING/
SACRIFICE SCHEDULE
RESULTS/CONCLUSIONS
FERTILITY AND
REPRODUCTION
   
Mouse Swiss-Webster
Males: 20/group,
dosed for 60 days before mating plus
9 days during mating.
Females: 38 ‒ 40/group, undosed
0, 0.4, 2.0, and 10.0
mg/kg/day intravenous
breeding postdosing.
2 months and 6 months.
Necropsy (males): 2 months
and 7 months.
Decreased fertility in mid- and high-dose
males at the end of treatment, associated
with atrophy and decreased sperm in
testes and epididymis. Mid-dose fertility
completely recovered after 2 months,
high-dose fertility minimally recovered
after 6 months. No treatment-related
effects on mating behavior, litter size and
survival indices. No dominant lethal
effects.
Mouse Swiss-Webster
Males: Undosed
Females: 40/group,
dosed from 14 days before mating
through weaning.
0, 5, 20 and 90 mg/kg/day
intravenous breeding after 14
days dosing (females allowed
to litter) and at 2 months
postdosing. Necropsy
(females): at both mating
periods.
Decreased receptivity to mating,
decreased pregnancy rate and increased
resorption rate in high-dose females at
first mating period; offspring showed
hypoplastic testes and seminal vesicles,
and increased incidence of epithelial
hyperplasia and hyperkeratosis of the
non-glandular stomach. Complete
recovery at 2 months from treatment
related changes in mating, fertility and
embryolethality. Offspring of low and
mid-dose females showed no treatment-
related changes, and had normal mating
behavior, fertility, and offspring viability
for second-generation mating cycle.
TERATOLOGY    
Mouse Swiss-Webster
Females: 25/group
dosed days 7 ‒ 16 gestation
0, 12, 36 and 108 mg/kg/day
Necropsy day 18 gestation
Decreased body weight gain in high-dose
dams, associated with increased
resorptions, decreased live-litter size and
decreased livefetus weights. Growth
retardation in high-dose fetuses, but no
treatment-related teratologic changes
noted after external, skeletal and visceral
examination.
Rabbit Dutch-Belted
Females: 21/group
dosed days 7 ‒ 19 gestation
0, 6, 20 and 60 mg/kg/day
Necropsy day 29 gestation
High-dose dams showed clinical signs of
toxicosis, decreased body weights and
resorption of 12/14 litters. Decreased fetal
weight and fetal growth retardation in
both mid- and high-dose groups. Fetal
malformations observed in two mid-dose
and two high-dose dams, including cleft
palate, hydrocephaly and microphthalmia;
teratogenic effects did not appear to be
secondary to maternal toxicity.

Table 13
MUTAGENICITY STUDIES
STUDY TYPE DOSE EFFECTS CONCLUSION
Ames plate test with
Salmonella and mitotic
gene conversion assay
with Saccharomyces (both
assays with and without
activation)
1, 10, 100, 500,
1000, 2500, 5000
and 10000
mcg/plate
Not toxic to bacterial strain TA-
100 or yeast strain D at
10,000 mcg/plate; cytotoxicity
not evaluated in four other
bacterial strains used. Results
of both tests conducted in the
presence or absence of
activation were negative.
DHPG did not demonstrate
mutagenic activity in any
assays conducted in this
evaluation and was considered
not mutagenic under these test
conditions.
Ames suspension test with
Salmonella (with and
without activation)
500, 1000, 1250,
2500, and 5000
mcg/mL
Not toxic to bacterial strain TA-
100 - cytoxicity not evaluated
in four other bacterial strains
used. Results of both tests
conducted in the presence or
absence of activation were
negative.
DHPG did not demonstrate
mutagenic activity in any
assays conducted in this
evaluation and was considered
not mutagenic under these test
conditions.
Sister chromatid exchange
assay with human
lymphocytes (with
activation only)
0.250, 0.500,
0.750, 1.0, 1.5 &
2.0 mg/mL
Delayed cell growth at
concentrations of 1.0 mg/mL
and greater. Results of tests
conducted in the presence of
activation were positive, at all
doses tested.
DHPG was positive at all dose
levels in human lymphoblasts
under the conditions of this
test.
Mouse lymphoma forward
mutation assay in L5178Y
cells, with and without
activation
50, 100, 400, 500,
600, & 800
mcg/mL without
activation;
100, 400, 800, &
1000 mcg/mL
with activation
Moderate to very high toxicity
induced by range of doses used.
Results of all tests conducted
under conditions of moderate
toxicity were positive, with or
without activation.
DHPG was positive over a
wide range of dose levels
under the conditions of this
test.
Cell transformation assay
in BALB/c-3T3 cells,
without activation
1.88, 7.5, 15, 25
and 40 mcg/mL
Complete lethality at
concentrations of 62.5 mcg/mL
and greater. Cell survival
12.8% at 31.3 mcg/mL, near
control levels at concentrations
of 3.91 mcg/mL and less.
Transformation responses to
doses tested not significantly
elevated relative to spontaneous
transformant frequencies; no
evidence of dose-related
increase in transforming
activity.
DHPG was considered
inactive in this assay at the
concentrations tested.
Mouse micronucleus assay
in CD-1(1CR) male and
female mice
50, 150, and
500 mg/kg body
weight,
intravenous
Bone marrow cytotoxicity
observed at high dose. Positive
dose response trends seen in
both sexes at mid and high
doses, with max.
DHPG was considered
negative in this assay at
50 mg/kg and positive at 150
and 500 mg/kg.

Table 14
OTHER TOXICITY STUDIES
PROTOCOL SPECIES
STRAIN
ROUTE
DOSE
NO./SEX PER
GROUP
STUDY
DURATION
RESULTS
Single-Dose
Gonadotoxicity
Mouse
Swiss-
Webster
Intravenous
0, 2, 10, 30,
100 & 300
mg/kg
Males: 15/dose
group
Females: 15/dose
group (30,100 &
300 mg/kg only)
Single dose with
necropsy at 2
wks., 1 mo. and 3
mos. postdosing
Testicular atrophy &
hypospermatogenesis in
males at 2 wks. and 1
mo. postdosing after 30,
100 or 300 mg/kg;
complete or partial
recovery at 3 mos. No
treatment-related
changes seen in
females.
Single-Dose
Gonadotoxicity
Dog
Beagle
Intravenous
0, 1, 6, 30 &
150 mg/kg
Males: 2/dose
group
Single dose with
hemicastration at
1 mo. followed by
necropsy at 2
mos, or
hemicastration at
2 mos. Followed
by necropsy at 4
mos. (1 dog each
per dose group)
Hypospermatogenesis at
1 & 2 mos. post-
treatment after 30 or
150 mg/kg; complete
recovery at 4 mos. No
treatment-related
changes in clinical
condition, body
weight, FSH, LH or
testosterone.
Vein Irritation Rabbit
New
Zealand
Intravenous
45 mg/ml
solution
Females: 2/dose
group
Single dose with
necropsy at 10, 30
& 240 minutes
postdosing.
No gross or microscopic
pathological changes
were observed in the
treated ear veins.