Iron digluconate

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

There are no toxicity to reproduction studies on iron gluconate. Results of a studies conducted with a structurally similar compounds: Iron Sucrose, Ferric Carboxymaltose and iron (II) chloride are reported and used for read across.

Iron (II) Chloride is a good read across material for evaluating the reproductive toxicity potential of iron gluconate because of similarities in their phys/chem properties and similar systemic exposures absorption, distribution, and elimination properties by the oral route of administration. Via read across Iron Gluconate is not classified as toxic to reproduction

Link to relevant study records

Reference

Endpoint:

one-generation reproductive toxicity

Remarks:

based on test type (migrated information)

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

2004

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Reason / purpose for cross-reference:

reference to same study

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Sam: Strain of TacN(SD), Samtako Bio Korea, 77-1 Seorang-dong, Osan-si, Gyeonggido.

- Age at study initiation: 8 weeks old.

- Weight at study initiation: Male: 269.23 - 302.18 g., Female: 191.34 - 221.60 g.

- Fasting period before study: The day before the necropsy.

- Housing:
Facility used:Stainless steel wired cage (260W X 350L X 210H (mm), Daejong Lab), polycarbonate cage (260W X 420L X 180H (mm), MJLTD).
Pre-mating period: 1 - 2 individuals (separate female and male rats), in stainless steel wire cage.
Mating period: One female and one male in one cage, in stainless steel wire cage.
Gestation and nursing period (females): Females individually housed in polycarbonate cage.
Post-mating period (males): 1 - 2 individuals, in stainless steel wire cage.
recovery group: 1 - 2 individuals, in stainless steel wire cage.

- Diet : The lab rat feed 5057 (Agri Purina Korea, 627 Jangdang-dong, Pyeongtaek, Gyeonggido), ad libitum.Analysis of the diet met the criteria of the protocol.

- Water (e.g. ad libitum): Mains water, filtred and sterilised, ad libitum. Analysis of the water met the criteria of the drinking water quality (Ministry of Environment).

- Acclimation period: 7 days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17.7 - 23.0ºC.
- Humidity (%): 40.8 ± 69.1%.
- Air changes (per hr): 10 - 15 changes per hour, front ventilation.
- Photoperiod (hrs dark / hrs light): 12 hrs light (7am - 7pm) / 12 hrs dark. 150 - 300 lux was used.

IN-LIFE DATES:
- Males: From Day 0 To: Day 43.
- Female: From: Day 0 Up To: Day 5 post partum, (female mated but did not show gestation signs) Up To: gestation Day 27.
(Recovery group)
- Males: From: Day 0 To: Day 57.
- Females. From: Day 0 To: Day 69.

Route of administration:

oral: gavage

Type of inhalation exposure (if applicable):

other: not applicable

Vehicle:

water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: The substance was suspended in the water for injection by dose and formulated just before the dosing.

VEHICLE (Water for injection)
- Amount of vehicle : 10 ml/kg/bw.
- Lot/batch no. : AAW4AB, Choongwae Pharma Corporation.

Details on mating procedure:

- M/F ratio per cage: 1:1.

- Length of cohabitation: 2 weeks.

- Proof of pregnancy: Not defined. The copulation was confirmed in the morning and afternoon by the fall of vaginal plug. The date when copulation was confirmed was set to gestation Day 0.

- After successful mating each pregnant female was caged : Individually housed in polycarbonate cage.

- Others: Individuals which did not deliver up to gestation Day 26 and showed the signs of gestation such as inflated mammary glands and abdominal enlargement were regarded as not pregnant. The final gestation was decided by the implantation signs in the uterus in the necropsy. Delivery signs were observed every day. When the delivery was confirmed up to the end of observation (5pm), the day was set on post partum Day 0. When delivery was made after the observation was over, the day after delivery day was set on post partum Day 0.

Analytical verification of doses or concentrations:

no

Details on analytical verification of doses or concentrations:

Not applicable.

Duration of treatment / exposure:

(Males) From Day 0 To: Day 42.
(Female) From: Day 0 Up To: Day 4 post partum, (female mated but did not show gestation signs) Up To: gestation Day 26.

Frequency of treatment:

Once daily.

Details on study schedule:

- Age at mating of the mated animals in the study: At least 10 weeks.

Dose / conc.:

125 mg/kg bw/day (nominal)

Remarks:

nominal in water

Dose / conc.:

250 mg/kg bw/day (nominal)

Remarks:

nominal in water

Dose / conc.:

500 mg/kg bw/day (nominal)

Remarks:

nominal in water

No. of animals per sex per dose:

15 animals per sex per dose and the control group.
(Recovery group; 5 animals per sex at 500 mg/kg/day and control group.)

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Based on the results in the preliminary test (mortality took place at the dose level of 1000 mg/kg).

- Rationale for animal assignment (if not random): Based on the body weights measured at the end of the quarantine and purification period, rats were deployed as G1 - G2 - G3 - G4 - G4 - G3 - G2 - G1 - G1 - G1 - G3 - G4, where G1 of control, G2 125 mg/kg, G3 of 250 mg/kg and G4 of 500 mg/kg.

- Post-exposure recovery period in satellite groups: For two weeks.

Positive control:

No

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice a day (30 minutes after dosing, and just before the job was over on the day) during the test period.
- Cage side observations checked: Presence of dead animals or animals in critical condition.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Once a day.

BODY WEIGHT: Yes
- Time schedule for examinations: Once daily except mating period.

FOOD CONSUMPTION : Yes
- Time schedule:
(male) The day before the dosing and once a week after the dosing started. After mating, food was supplied on the day of confirming copulation and the day before necropsy. The remains were measured the day after the food supply. Not measured during mating period.
(female) The day before the dosing and once a week after the dosing started. Food was supplied on gestation days 0, 6, 13 and 20 and on post partum days 0 and 3. The remains were measured the day after the food supply. Not measured during mating period.

FOOD EFFICIENCY: No.

WATER CONSUMPTION : Yes
- Time schedule for examinations:
(male) The day before the dosing and once a week after the dosing started. After mating, water was supplied on the day of confirming copulation and the day before necropsy. The remains were measured the day after the water supply. Not measured during mating period.
(female) The day before the dosing and once a week after the dosing started. Water was supplied on gestation days 0, 6, 13 and 20 and on post partum days 0 and 3. The remains were measured the day after the water supply. Not measured during mating period.

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY / CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: For 18 hours the day before the necropsy.
- Anaesthetic used for blood collection: Yes (ether).
- Animals fasted: Yes
- How many animals: 5 males and 5 females of each group.
- Parameters checked:
- Haematology, paramerers examined: RBC, HGB, HCT, MH, MCV, MCH, MCHC, WBC, PLT, NEU, LYM, MONO, EOS, BASO, PT and APTT
- Clinical chemistry, paramerers examined: ALT, AST, Cs, Glu, BUN, Crea, T-Bil, TG, TP, Alb, A/G, P, Ca, Na, K and Cl.

URINALYSIS: Yes
- Time schedule for collection of urine: For 3 to 4 hours the day before the necropsy
- Metabolism cages used for collection of urine: No data
- Animals fasted: No data
- Parameters checked: colour, pH, protein, erythrocyte, specific gravity, glucose and leukocyte.
A certain amount of fresh urine was collected from five males and five females of each group and the recovery group. Comvur10 Test M (Roche, Germany) was used for analysis.

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: The day before necropsy. (motor functions and autonomic nervous system were observed once a day).
- Battery of functions tested: sensory activity and motor fuctions. The auricle reflex and corneal reflex tests were conducted on five males and five females randomly selected from each group and the recovery group to evaluate sensory functions. The traction test was conducted to evaluate motor function.

Oestrous cyclicity (parental animals):

No reported.

Sperm parameters (parental animals):

As a part of histopathological test of the gonad of male rats, sperm generation was examined.

Litter observations:

PARAMETERS EXAMINED
The following parameters were examined in offspring: birth rate, surviving rate, mortality on Day 0 and 4 post partum, body weight Day 0 and 4 post partum, external appearance and sex ratio were examined.

GROSS EXAMINATION OF DEAD PUPS: Not reported.

Postmortem examinations (parental animals):

GROSS PATHOLOGY: Yes
The day after the dosing was over (two weeks after the dosing was over for the recovery group), five rats were placed under ether anesthesia for external appearance examination. After blood collection from the dorsal aorta, the subjects were placed for bloodletting and killed. The organ tissues were observed with macroscopically; external surface and all orifices / cranial cavity and external surface of the brain / nasal cavity and paranasal sinus / thoracic, abdominal and pelvic cavities and their viscera / cervical tissues and organs.
Necropsy was immediately performed on dead animals or animals in critical condition.

- Organ weights: Organs in tables 12 and 13 were removed from 5 males and 5 females of each group at the necropsy and weighed. Testes and epididymides were examined from all male rats.

HISTOPATHOLOGY: Yes
After a uniformly 3 mm thick tissues were removed from fixed tissues of 5 males and 5 females of the control and the dose group of 500 mg/kg, dead bodies of 500 mg/kg group, organs observed macroscopically for abnormality that showed abnormality in organ weight measurement, 1 4 µm histopathological section was stained with haematoxylin and eosin and observed wuith the optic microscope (Olympus BX50, Olympus Optical Co., Japan) for histopathological test. The gonads of male rats were examined for sperm generation and interstitial cells. See Tables 16-1 and 16-2 for the organs examined.

Postmortem examinations (offspring):

GROSS NECROPSY
- Gross necropsy consisted of external appearance including measurement of crown rump length.

HISTOPATHOLOGY / ORGAN WEIGTHS
Not performed.

Statistics:

Levene's test. If significant in the one-way ANOVA due to homogeneous distribution, Dunnett's t-test was conducted. If the distribution was not homogeneous, the proper data transformation was conducted and the the Levene's test was repeated. If the distribution was homogeneous, the one-way ANOVA was conducted. In case of significance, Dunnett's t-test was repeated.

Reproductive indices:

Copulation rate (%) = (number of female and male rats confirmed for copulation ÷ number of male rats used for mating)x100

Male impregnation rate (%) = (number of pregnant female rats ÷ number of female rats used for mating)x100

Female impregnation rate (%) = (number of pregnant female rats ÷ number of female rats confirmed for copulation)x100

Delivery rate (%) = (number of female rats that delivered live neonatas ÷ number of female rats confirmed for copulation)x100

- Measurement of corpora lutea and implantation

% pre – implantation loss = [1 – (number of implantation ÷ number of corpora lutea)] x 100

% post – implantation loss = [1 - (number of neonates – number of corpora lutea)] x 100

Offspring viability indices:

Birth rate (%) = (Number of live neonates during delivery ÷ total number of neonates) x 100

Survival rate on Day 0post partum(
%) = (Number of live neonates on delivery date ÷Number of live neonates during delivery) x 100

Survival rate on Day 4post partum(%) = (Number of live neonates on Day 4post partum÷Number of live neonates on Day 0post partum) x 100

Sex ratio on Day 0post partum=(Number of survived male neonates on Day 0post partum÷ number of survived female neonates on Day 0post partum) x 100

Sex ratio on Day 4post partum=(Number of survived male neonates on Day 4post partum÷ number of survived female neonates on Day 4post partum) x 100

Clinical signs:

no effects observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

males only

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

males only

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Other effects:

not examined

Reproductive function: oestrous cycle:

not specified

Reproductive function: sperm measures:

not specified

Reproductive performance:

no effects observed

Unless stated, there were no effects in recovery group.

MORTALITY
There was no male mortality. Three females were found dead at 500 mg/kg; one in the main group took place on Day 38 and two in the recovery group on Day 46 and 51, respectively.

CLINICAL SIGNS
See Table 2-1 for the results. Blackish stool was found in all animals at all dose groups. Salivation was observed in 13 males and females, and all animals at the dose of 250 mg/kg and above. Soft stool was observed in male at all dose groups and in female at the dose of 500 mg/kg. Diarrhea was found in one female at the dose of 125 mg/kg, three females at 500 mg/kg and ten males at 500 mg/kg. Decrease in locomotion activity was observed in one female at the dose of 250 mg/kg. Paleness was observed in one male and three females, emaciation was observed in one male and one female, soiled perineal region in one female at the dose of 500 mg/kg.

BODY WEIGHT
See Tables 2-1 for the results. A significant decrease in body weight was observed in male at the dose of 250 mg/kg and above during the dosing period. This was observed in the recovery group at the dose of 500 mg/kg. Females showed a significant decrease in body weights at the dose groups of 125 and 500 mg/kg.

FOOD CONSUMPTION
There were no treatment-related effects on food consumption in male, however, there was a significant decrease in food consumption in female at the dose of 125 and 500 mg/kg on gestation day 0. Males in the recovery group at 500 mg/kg showed significant increase in food consumption on Day 3.

WATER CONSUMPTION
See Table 5-1 for the results.There was significant increase in water consumption at 500 mg/kg in both sexes.

HAEMATOLOGY
In males, there was a significant increase of MCV at the dose of 500 mg/kg. There was a significant increase of eosinophil values at the dose of 500 mg/kg recovery group.
In females, there was a significant increase in the number of platelet at the dose of 500 mg/kg in recovery group.

CLINICAL CHEMISTRY
In males, there was a significant decrease of cholinesterase at the dose of 250 mg/kg and above. In the male recovery group, a significant increase of triglyceride was observed at the dose of 500 mg/kg.

URINALYSIS
No treatment-related changes were detected in the urinary parameters examined.

NEUROBEHAVIOUR

ORGAN WEIGHTS
See Tables 12 and 13 for the results.In males, there was a significant increase in absolute and relative liver weight at the doses of 250 mg/kg and above. There was a significant increase in absolute and relative adrenal weight at 500 mg/kg, relative adrenal weight at 250 mg/kg and decrease in relative adrenal weight at 125 mg/kg.
In females, there was a significant increase in absolute and relative liver weight at the dose of 500 mg/kg and a significant decrease in absolute and relative thymus weight at the dose of 500 mg/kg, and absolute thymus weight at 125 mg/kg.
No significant effects of treatment were detected in the male sex organ weights.

GROSS PATHOLOGY
Diaphragmatic nodule in the liver was observed in both sexes in the dose groups and the control group (in males only). Diffuse black coloured liver was observed in all males, haemorrhage with diffuse black pigmentation in the stomach in 12 males, and unilateral atrophy in the testes in one male at the dose of 500 mg/kg. Mass of mesenteric lymphoid nodule was observed in one female at the dose of 500 mg/kg.
(Recovery group) Diaphragmatic nodule was observed in one male at the dose of 500 mg/kg, and in two females in the control group.
(Necropsy of three females found dead) A severe diffuse haemorrhagic glandular stomach was observed in the necropsy in one female found dead on Day 38. Severe distension of stomach were observed in two females dead in recovery group, but during the dosing period, therefore these findings were not considered as of recovery period.


HISTOPATHOLOGY: NON-NEOPLASTIC
See Tables 16-1 and 16-2 for the results.
(Males)
Adrenals: The following effects were observed in male adrenals at the dose of 500 mg/kg; unilateral hyperplasia of zone fasciculate in the adrenal cortex in 4 males, bilateral hyperplasia in hyperplasia in 4 males, and unilateral focal necrosis of adrenal cortex in one male.
Liver: Minimal diffuse haemosiderin deposit of liver parenchymal in 2 males, mild diffuse haemosiderin deposit in 8 males, moderate diffuse haemosiderin deposit in 4 males with one severe case were observed at the dose of 500 mg/kg.
Stomach: Moderate diffuse haemosiderin deposit in the granular in 9 males with 2 severe cases, moderate diffuse cellular infiltration (neutrophil) in submucosa in 3 males, mild hyperkeratosis of forestomach in 7 males, moderate hyperkeratosis of forestomach in one male with severe cases in 3 males.
(Females)
Minimal diffuse haemosiderin deposite in liver parenchymal was observed in one female at the dose of 250 mg/kg and above.
The following non treatment-related findings were observed; bilateral hyperplasia of zone fasciculate in the adrenal cortex in 4 females, bilateral hyperplasia of zone reticulosa in one female, minimal focal haemosiderin deposit of liver parenchymal in one female, mild diffuse haemosiderin deposit of liver parenchymal in one female, moderate multifocal haemosiderin deposit of liver parenchymal in mesenteric lymphnode in one female.
(Recovery group) Diffuse congestion of the adrenal were observed in 2 females. Hyperkeratosis of the forestomach, atrophy of the granularminimally focal haemosiderin deposit of liver parenchymal and focal oval cell proliferation were observed in one male, respectively.

CORPORA LUTEA AND IMPLANTATION
Pre-implantation rates were 14.4% in the control group, 9.4% at 125 mg/kg, 14.3% at 250 mg/kg and 9.8% at 500 mg/kg. Post-implantation loss rates were 6.0% for both the control group and the dose group of 125 mg/kg, 3.1% at 250 mg/kg and 7.0% at 500 mg/kg.

Dose descriptor:

NOAEL

Remarks:

systemic toxicity

Effect level:

125 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: changes in body weight, water consumption, organ weight and histopathology at the dose of 500 mg/kg and changes in organ weight at the dose of 250 mg/kg

Dose descriptor:

NOAEL

Remarks:

systemic toxicity

Effect level:

250 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: changes in organ weights and histopathology were observed at the dose of 500 mg/kg

Dose descriptor:

NOAEL

Remarks:

reproductive and developmental toxicity

Effect level:

500 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Due to the absence of systemic toxicity at any doses tested.

Remarks on result:

other: Generation: parents and offspring (migrated information)

Clinical signs:

no effects observed

Mortality / viability:

no mortality observed

Body weight and weight changes:

no effects observed

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Histopathological findings:

not examined

OBSERVATION OF NEONATES
No treatment-related effects were observed on mean live neonates, birth rates, survival rates and sex ratios on days 0 and 4 post partum. The only abnormalilty found in the external appearance examinations is an acaudate was observed in one neonate at 500 mg/kg. Crown Rump Length (CRL) of female neonates showed a significant decrease at 125 mg/kg on Day 4 post partum.

Key result

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

ca. 500 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Due to the absence of systemic toxicity at any doses tested.

Reproductive effects observed:

not specified

Conclusions:

The oral administration of the test material to rats by gavage resulted in changes in body weight, water consumption, organ weight and histopathology in males at the dose of 500 mg/kg and changes in organ weight at the dose of 250 mg/kg. Therefore 125 mg/kg/day can be considered the No Observed Adverse Effect Level (NOAEL) in male rats. In females, changes in organ weights and histopathology were observed at the dose of 500 mg/kg. Therefore 250 mg/kg/day can be considered the NOAEL in female rats. The changes except body weight changes in males seemed to be reversible.
There was no treatment-related effects on reproductive functions in parental animals and development of neonates at any doses tested. the NOAEL for reproduction and developmental toxicity was considered to be 500 mg/kg/day.

Executive summary:

The test was intended to evaluate NOAEL (No Observed Adverse Effect Level) on reproduction process such as mating, conception, gestation, childbirth and development of neonates and the effect on the whole body including nerve and immune systems when female rats were orally dosed with Iron dichloride (CAS No.7758-94-3) once a day up topost partumday 4 from two weeks prior to the mating and male rats were orally dosed once a day with iron dichloride till two weeks before and after the mating.

Male and female SD rats were dosed with the test substance (0 (Control group), 125, 250 and 500 mg/kg/day) from two weeks before mating. Male SD rates were dosed once a day till two weeks after mating while female SD rats were dosed once a day up topost partumday 4. A total of 42 doses were provided for male rats while female rates had 42 to 54 dosages depending on mating and delivery of individuals. Clinical signs and mortality were observed and body weight and food and water consumption were measured. In the necropsy, gross examination of organs and tests on corpus luteum graviditatis and implantation rates were conducted. In addition, tests for sensory and motor functions, urinalysis and hematological and blood chemical tests were given and organ weights were measured for five individuals randomly selected from each group. External abnormalities, sex ratio, body weights, CRL (Crown Rump Length) and survival rate were observed onpost partumdays 0 and 4.

During the observation period, the main group dosed with the substance showed signs such as melaena (black stool) and salivation but these signs were observed to disappear after dosing in the recovery group. There was no mortality in male SD rats, but three mortalities took place in female individuals at 500 mg/kg. The cause for mortalities was presumably the gastrointestinal damage by the substance. It was found that male individuals were more sensitive to body weight and food consumption than female counterparts. The change by the test substance was not recognized in mating data, sensory functions, motor functions, urine analysis and blood test. Gastric hemorrhage with blackened liver and black pigmentation of liver discovered in the necropsy findings was presumed to be caused by the test substance, but it was found to improve for the recovery period of two weeks. Weight changes in the liver and adrenal were observed in the absolute and relative organ weights of male individuals at 250 and 500 mg/kg and female individuals at 500 mg/kg.The histopathological test found parenchymal hemosiderosis and hyperplasia of adrenocortical zona fasciculate as well. It was found that the substance had no effect on birth rate, survival rate, body weight and CRL of neonates.

As a result of the test using iron dichloride (CAS No.7758-94-3), the NOAEL of repeated doses to male and female SD rats were 125 and 250 mg/kg/day, respectively.As there was no difference observed in reproductive functions of male and female SD rates and development of neonates between the control group and main group, NOAEL was thought to be 500 mg/kg/day. The effect of the test substance was none on reproductive functions, sensory functions, motor functions, urinalysis, and hematological and blood chemical findings which showed no differences between the control group and main group. Except the body weight change of male SD rats, clinical signs, water consumption, organ weights, necropsy findings and histopathological findings were reversible as they recovered after the dosing was over.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

500 mg/kg bw/day

Species:

rat

Effects on developmental toxicity

Description of key information

There are no effects on developmental toxicity studies on ferrous gluconate. Results of studies conducted with a structurally similar compounds are reported and used for read across. 
Via read across Iron Gluconate is not classified as toxic to development

Link to relevant study records

Reference

Endpoint:

developmental toxicity

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

Date of study initiation: August 5, 2000

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

Study is reported by US-FDA in the drug approval process for the product Ferinject (NDA 22-054 Ferinject) as iron replacement product indicated for the treatment of iron deficiency anemia, the original study report is not available but study and test item summaries can be accessed via: Drug Approval Package NJECTAFER (ferric carboxymaltose) Injection Company: Luitpold Pharmaceuticals, Inc. Application No.: 203565 Approval Date: 07/25/2013 http://www.accessdata.fda.gov/drugsatfda_docs/nda/2013/203565Orig1s000TOC.cfm Pharmacology Review(s), Chemistry Review(s) and Other Review(s)

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

not specified

Principles of method if other than guideline:

To study the potential teratogenic effects of VIT-45 in pregnant rabbits, VIT-45 was given by intravenous infusion to rabbits (22/group) at 0, 4.5, 9, 13.5, and 18 mg/kg/day from gestation days 6 to 19.

Study is reported by US-FDA in the drug approval process for the product Ferinject (NDA 22-054 Ferinject) as iron replacement product indicated for the treatment of iron deficiency anemia, the original study report is not available but study and test item summaries can be accessed via: Drug Approval Package NJECTAFER (ferric carboxymaltose) Injection Company: Luitpold Pharmaceuticals, Inc. Application No.: 203565 Approval Date: 07/25/2013 http://www.accessdata.fda.gov/drugsatfda_docs/nda/2013/203565Orig1s000TOC.cfm
Pharmacology Review(s), Chemistry Review(s) and Other Review(s)

GLP compliance:

yes

Limit test:

no

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: no data
- Age at study initiation: 19-27 weeks
- Weight at study initiation: 3.45 - 5.55 kg
- Fasting period before study: no data
- Housing: no data
- Diet (e.g. ad libitum): no data

Study initiation date: 2000-08-05

Route of administration:

intravenous

Vehicle:

other: watery solution of sodium chloride 0.9 %

Details on exposure:

no data

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

no data

Details on mating procedure:

no data

Duration of treatment / exposure:

treatment from gestation day 6 to 19

Frequency of treatment:

once daily i.v.

No. of animals per sex per dose:

22 female rabbits/group, for dose group 18 only 10 animals/group

Control animals:

yes, concurrent vehicle

Details on study design:

no data

Maternal examinations:

CAGE SIDE OBSERVATIONS: no data

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: daily

FOOD CONSUMPTION ): Yes , daily

WATER CONSUMPTION: No data


POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day: no data
- Organs examined: yes, corpora lutea, gravid uterus

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of resorptions: Yes

Fetal examinations:

- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: all per litter
- Skeletal examinations: Yes
- Head examinations: Yes: all per litter

Statistics:

no data

Details on maternal toxic effects:

Maternal toxic effects:yes

Details on maternal toxic effects:
Two dams per dose group died or were sacrificed. One dam in dose group 4.5, 9 and 18 mg/kg and two dams of group 13.5 mg/kg aborted. Prior to death/sacrifice dams in group 13.5 and 18 mg/kg had weight loss and reduced food intake. Most of these dose groups dams had liver tinged orange with accenturated lobular pattern. Brown/dark pinna was noted in the rabbits at 9 and 13.5 mg/kg and orange/brown eyelids in rabbits at 18 mg/kg. At necropsy orange/brown discoloration in various organs were noted it dams of group 9, 13.5 and 18 mg/kg/day. Preimplantation loss was marked in high dose group (33%) compared to control (18%).

Dose descriptor:

LOAEL

Effect level:

4.5 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

other: maternal toxicity

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:yes. Remark: at maternal toxic dose

Details on embryotoxic / teratogenic effects:
Three fetuses at high dose and two fetuses in 13.5 mg/kg group had marked domed craniums. Both 13.5 mg fetuses had additionally internal hydrocephaly or suspected hydrocephaly (major malformations). One fetus in 9 mg group had moderately domed cranium (minor malformation). The domed cranium was in incidence and severity dose related.

Key result

Dose descriptor:

NOEL

Effect level:

4.5 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

other: Basis for effect level / Remarks: head malformation

Abnormalities:

not specified

Developmental effects observed:

not specified

Table 1: Litter value – group value

Live young

Resorption

Implantation Loss %

Group

No. pregnant dams

Corpora lutea

Implant-ations

Males

Females

Total

Early

Late

Total

Pre-

Post-

1(control)

22

Means/ SD

12.5/3.4

10.1/2.9

4.1/1.7

4.1/2.29

8.2/2.3

0.9/0.9

1.0/1.0

1.9/1.3

18.1

17.4

2(4.5 mg/kg)

16

Means/ SD

13.1/1.9

11.5/2.8

4.9/1.7

4.7/1.8

9.6/2.5

0.5/0.7

1.4/1.2

1.9/1.3

13.1

15.0

3(9 mg/kg)

19

Means/ SD

11.6/2.0

9.6/3.1

3.9/2.0

4.4/2.1

8.3/3.5

0.6/0.8

0.7/0.9

1.3/1.1

16.3

14.0

5(13.5mg/kg)

16

Means/ SD

12.1/2.0

10.6/2.7

4.1/2.2

5.1/2.4

9.2/2.7

0.7/0.8

0.8/1.1

1.4/1.5

12.2

13.7

4(18 mg/kg)

7

Means/ SD

12.1/1.9

8.4/3.4

3.1/2.8

3.7/2.2

6.9/3.1

0.1/0.4

1.4/1.2

1.6/1.3

32.7

17.0

Table 2: Fetal examination – major abnormalities – group incidences

Gross Observation
	Fetus				Litters
Group	1	2	3	5	1	2	3	5
Dose (mg Fe/kg/day)	0	4.5	9	13.5	0	4.5	9	13.5
Number examined	181#	154	158	147	22	16	19	16
Number affected	2	1	2	3	2	1	2	3
Malformations
Marked internal hydrocephaly/suspected, marked domed cranium	-	-	-	2	-	-	-	2
Partially fused bilateral frontal, small left orbital socket	1	-	-	-	1	-	-	-
Fused bilateral parietal to interparietal, partially fused bilateral frontal, dilated pulmonary trunk, narrow ascending aorta, retroesophageal aortic arch, large/small atria, misshapen ventricle	-	1	-	-	-	1	-	-
Palatine irregularity, protruding tongue, incompletely ossified cranial bones and vertebral elements, irregularly ossified, kinked and medially thickened ribs, irregularly ossified long bones	-	-	-	1	-	-	-	1
Dilated ascending aorta and aortic arch, transposition of ascending aorta and dorsally displaced pulmonary trunk, displaced ductus arteriosus, ventricular septal defect	-	-	1	-	-	-	1	-
Lumbar scoliosis	-	-	1	-	-	-	1	-
Lumbosacral vertebral irregularities	1	-	-	-	1	-	-	-

  # Includes 1 early birth

Table 3: Fetal examinations – placental abnormalities – group incidences

Gross Observation
	Fetus				Litters
Group	1	2	3	5	1	2	3	5
Dose (mg Fe/kg/day)	0	4.5	9	13.5	0	4.5	9	13.5
Number examined	181#	154	158	147	22	16	19	16
Malformations
Amniotic sac, milky fluid in	1	-	-	-	1	-	-	-
Amniotic sac, tinged orange	-	-	-	55	-	-	-	6
Placenta, milky fluid around	1	1	-	-	1	1	-	-
Placenta, maternal potion tinged orange	-	-	-	95	-	-	-	10
Placenta, pale and swollen	-	-	-	9	-	-	-	1
Number of fetuses affected	1	1	-	122	1	1	-	13

  # Includes 1 early birth

Conclusions:

The test substance does only show fetal effects at maternal toxic dose. Therefore, it is not considered as teratogenic.

Executive summary:

Pregnant rabbits were treated intravenously with ferric carboxymaltose (CAS 9007-72-1) at the doses of 4.5, 9, 13.5 and 18 mg Fe/kg /day from day 6 to 19 of pregnancy. After sacrifice or premature death section was performed on dams and fetuses.

Two dams per dose group died or were sacrificed. Prior to death/sacrifice dams in group 13.5 and 18 mg/kg suffered from weight loss and reduced food intake and showed a liver tinged orange with accentuated lobular pattern. Orange/brown discoloration in variuos organs were noted it dose group 9 mg/kg upwards. Several dams starting at dose group 4.5 mg/kg onwards aborted. Preimplantation loss was marked in high dose group (33 %) compared to control (18 %).

Three fetuses at high dose and two fetuses in 13.5 mg/kg developed marked domed craniums. Both 13.5 mg fetuses had additionally internal hydrocephaly or suspected hydrocephaly (major malformations). One fetus in 9 mg group had moderately domed cranium (minor malformation). The domed cranium was in incidence and severity dose related. All fetal effects appeared at maternal toxic dose. Therefore, ferric carboxymaltose is assumed to be not teratogenic under the conditions of this test.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

4.5 mg/kg bw/day

Species:

rabbit

Additional information

Justification for selection of Effect on developmental toxicity: via oral route:
There are no effects on developmental toxicity studies on ferrous gluconate. Results of a studies conducted with a structurally similar compounds (Ferric carboxymaltose, iron sucrose and ferrous sulphate) are reported and used for read across.

Ferrous Sulphate is a good read across material for evaluating the reproductive toxicity potential of iron gluconate because of similarities in their phys/chem properties and similar systemic exposures absorption, distribution, and elimination properties by the oral route of administration. Via read across Iron Gluconate is not classified as toxic to development

The read-across justification is attached.

Justification for classification or non-classification

The reproductive effects of iron gluconate can be reasonably predicted by read across to other sources of iron that have adequate bioavailability to ensure appropriate systemic exposure. The gluconic acid portion of the molecule is not critical in a reprotoxicity/developmental toxicity read across because of the benign nature of this material. Once absorbed, the gluconate, or the cyclized form, D-glucano-1,5-lactone, would be metabolized in the same way as any other form of glucose as part of normal cellular respiration via the Krebs cycle.

 

Good surrogates for iron gluconate would be any number of ferrous salts with dissociation constants similar to iron gluconate. Iron dichloride (CAS 7758-94-3) and iron sulphate (CAS 7720-78-7) are used in this read–across justification because they will deliver systemic exposure at sufficient levels to adequately test the reproductive and developmental potential of the ferrous ion. 

 

After oral exposure, iron gluconate, iron sulphate and iron dichloride, will dissociate into their ionic forms in a similar manner. The pKa of iron gluconate is estimated to be 3.7 (OECD SIDS 2004) while that of iron dichloride and iron sulphate are approximately 3.1 and 3.05, respectively. Once dissociated, the ferrous ions would be absorbed for systemic distribution in the same way. The amount of delivered ferrous ions would be calculated based on the weight percentage of iron in each of the molecules and the endpoint values expressed in iron equivalents rather than as the weight of the parent molecule. For purposes of these calculations, iron constitutes 12% of the mass of the iron gluconate, 44% of the mass of iron dichloride and 37% of the mass of the iron sulphate.

 

Overall, iron sulphate and iron dichloride are good read across materials for evaluating the reproductive potential of iron gluconate because of similarities in its phys/chem properties and similar toxicokinetics via the oral route of administration.

In a combined repeated dose toxicity study (OECD 422) of ferrous chloride,oral administration of the test material to rats by gavage resulted in no treatment-related effects on reproductive functions in parental animals and development of neonates at any doses tested. The NOAEL for reproduction and developmental toxicity was considered to be 500 mg/kg/day (170 mg Fe/kg/day).

In the REACH dossier for both iron dichloride and iron sulphate (chosen read across substances), iron sucrose (CAS 8047-67-4)

and ferric carboxymaltose (CAS 9007-72-1) were used as read across substances for developmental endpoints, for which data from an OECD 414 test protocol (or similar) were presented. These substances were also used as read-across for other iron salts

 

Data for ferric carboxymaltose (CAS 9007-72-1) was reported by US-FDA as part of a drug approval process for a product to be used as an iron replacement in iron deficiency anaemia. The LOAEL for maternal toxicity was 4.5 mg/kg/day based on the numbers aborting and mortality. A NOAEL could not be derived due to effects being seen at the lowest dose tested. The NOAEL for developmental toxicity was 4.5 mg/kg/day based on head malformations such as hydrocephaly, domed cranium observed at in the 13.5 mg/kg group. The study was deemed reliable with restrictions (Klimisch 2 score) as the original study is not available although study and test item summaries were accessible. Overall, becauseferric carboxymaltoseonly caused developmental effects at maternally toxic doses it is not considered to be teratogenic.

 

Data for iron sucrosewas reported by US-FDA as part of a drug approval process. The NOAEL for maternal toxicity was 3 mg/kg/day based on the dose related abortion and decreased body weight in the 6.5 mg/kg group. The NOAEL for developmental toxicity was also 3 mg/kg/day based on fluid in abdomen seen in the 6.5 mg/kg group. The study was deemed reliable with restrictions (Klimisch 2 score). Iron sucrose was considered not to be teretogenic due to developmental effects ibly being oberved at maternally toxic doses, or there being a lack of dose-dependancy for some effects.

In a study published in 1974 (US FDA 1974) , the US FDA evaluated the teratologic and developmental effects of ferrous sulphate in both rats and mice and concluded the ferrous ion did not cause significant effects in either endpoint up to the levels tested. A NOAEL for reproductive effects was established at the maternal tox level of 160 mg/kg (58 mg Fe/kg/day), and developmental effects at > 1000 mg/kg/day (360 mg Fe/kg/day).  

Via read across Iron Gluconate is classified as non-hazardous for this end point