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Administrative data

Key value for chemical safety assessment

Effects on fertility

Link to relevant study records
Reference
Endpoint:
toxicity to reproduction
Remarks:
other: Effect on the fertility of male rats
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
2000
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Information is available from the pre-approval review of Aczote (a cream containing 5% dapsone for use against acne) by U.S. FDA.
Principles of method if other than guideline:
Treatment of males only. Fertility effects on females have been studied separately
GLP compliance:
yes (incl. QA statement)
Limit test:
no
Species:
rat
Strain:
other: Rat/Crl:CD(SD)IGS BR VAF/Plus
Sex:
male
Route of administration:
oral: gavage
Vehicle:
CMC (carboxymethyl cellulose)
Remarks:
0.5%
Details on exposure:
The study was conducted in two stages, with an initial study that involved dosages of 0, 12, 30 and 75 mg/kg/day, and a second study that involved exposures at 0, 0.5, 2 and 12 mg/kg/day. Therefore there were two groups which received 0 mg/kg/day (vehicle control group) and two groups which received 12 mg/kg/day.
Details on mating procedure:
Only males were treated. Untreated females were used to confirm male reproductive potential.
Analytical verification of doses or concentrations:
yes
Duration of treatment / exposure:
F0 males were dosed with the test material 63 days prior to cohabitation with untreated females and continued through the day prior to sacrifice
Frequency of treatment:
daily
Remarks:
Doses / Concentrations:
0
Basis:
actual ingested
mg/kg bw/day
Remarks:
Doses / Concentrations:
0.5
Basis:
actual ingested
mg/kg bw/day
Remarks:
Doses / Concentrations:
3
Basis:
actual ingested
mg/kg bw/day
Remarks:
Doses / Concentrations:
12
Basis:
actual ingested
mg/kg bw/day
Remarks:
Doses / Concentrations:
30
Basis:
actual ingested
mg/kg bw/day
No. of animals per sex per dose:
25 males (treated) and 25 females (untreated)
Control animals:
yes, concurrent vehicle
Positive control:
none
Parental animals: Observations and examinations:
F0 male rats observed for viability and clinical signes twice daily. Body weight was recorded daily. F0 females were nonitred for body weight and clinical signs on gestation days 0, 7, 10 and 13. All F0 females were sacrificed on gestation day 13, cesarian sectioned, gross necropsied, and the numbers of corpora lutea, implantation sites, and viable and non-viable embryos was recorded. Following Completion of the cohabitation period, F1 males wer subjected to gross necropsy, the reproductive organs (each testis, each epididymidis, seminal vesicles and prostate) weighted, and sperm concentration and motility were evaluated.
Oestrous cyclicity (parental animals):
Females were not treated
Sperm parameters (parental animals):
Concentration and motility
Postmortem examinations (parental animals):
Gross pathology of males
Postmortem examinations (offspring):
not performed in this study
Statistics:
yes
Reproductive indices:
yes
Offspring viability indices:
yes
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Pale or blue extremities, salvation, poor grooming. Incidence proportional to exposure
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
BW gain significantly reduced > 75 mg/kg/day.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
BW gain significantly reduced > 75 mg/kg/day.
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Other effects:
not examined
Description (incidence and severity):
Test substance intake: gavage
Reproductive function: oestrous cycle:
not examined
Description (incidence and severity):
only males treated
Reproductive function: sperm measures:
effects observed, treatment-related
Description (incidence and severity):
motility: Effects above 75 mg/kg/day; sperm number: no effect
Number of implantation sites in females and viable embryos significantly reduced if males were treated >12 mg/kg/day, presumably due to the numbers or effectiveness of sperm.
Key result
Dose descriptor:
NOAEL
Effect level:
>= 0.5 - < 3 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
male
Basis for effect level:
other: Effect is on the number of motile sperm
Clinical signs:
not examined
Mortality / viability:
no mortality observed
Body weight and weight changes:
not examined
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not examined
Histopathological findings:
not examined
Only male fertility tested.
Remarks on result:
not measured/tested
Reproductive effects observed:
not specified
Conclusions:
Dapsone impaired fertility in male rats at high doses. This was evident from reductions in fertility index (numbers of rats pregnant/number of rats mated), reduced sperm motility (percentage of observed sperm that were motile), and reduced numbers of implantations and viable embryos in the females that did become pregnant. Statistically significant reductions in percentage of motile sperm were observed at exposures of 3 mg/kg/day and above. 0.5 mg/kg/day was the apparent NOAEL in this study, although a non-significant trend toward a reduction in the percentage of motile sperm may have been apparent.
Executive summary:

In this study, male rats were treated for 63 days prior cohabitation with untreated females. Dapsone at high doses impaired fertility in male rats. This was evident from reductions in fertility index (numbers of rats pregnant/number of rats mated), reduced sperm motility (percentage of observed sperm that were motile), and reduced numbers of implantations and viable embryos in the females that did become pregnant. Statistically significant reductions in percentage of motile sperm were observed at exposures of 3 mg/kg/day and above. 0.5 mg/kg/day was the apparent NOAEL in this study, although a non-significant trend toward a reduction in the percentage of motile sperm may have been apparent.

Effect on fertility: via oral route
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
0.5 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
Good; The studies are not OECD-type studies, howerver designed to investigate the fertility and teratogenicity endpoint similar to a regular study. In addition, because dapsone is used as a pharmaceutical for > 60 years with hundred thousands of treated patients and thousands of people protected from malaria by dapsone (including the well surveilled US and AUS military personnel who served in Vietnam) the effects of dapsone are well reported in literature, a selection of which has been included in the present dossier.
Additional information

The main toxic effect of dapsone is methemoglobin formation. This effects occurs in humans rarely at a daily dose of 100 mg/person/day and becomes more frequent in patients with a metabolic disease. The effect is frequent and clinically relevant at about 400 mg/person/day (or ~ 6 mg/kg/day). Fertility effects have been noted in male rats at 12 mg/kg/day and in female rats at 30 mg/kg/day. The effects in male rats are a reduction of sperm number and motility and in female rats an increased number of early resorptions and derived effects (e.g. number of offsprings).


Short description of key information:
Male rats: Reduction of sperm number and motility (males). Effect occurs at toxic doses comparable with therapeutic clinical doses and above.
Female rats: Increased number of early resorptions. Effect occurs at clearly toxic doses for the female.

Justification for selection of Effect on fertility via oral route:
Effects on both male and female fertility were noted. The NOAEL for male fertility effects is lower than the NOAEL for female fertility effects.

Effects on developmental toxicity

Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2004
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: NTP study
Qualifier:
according to guideline
Guideline:
EPA OPP 83-3 (Prenatal Developmental Toxicity Study)
Deviations:
not applicable
GLP compliance:
not specified
Limit test:
no
Species:
mouse
Strain:
CD-1
Details on test animals or test system and environmental conditions:
Time - mated mice (CD-1) Swiss albino were dosed by gavage.
Mice were kept in groups in stainless-steel containers at 22 degree C
Route of administration:
oral: gavage
Vehicle:
CMC (carboxymethyl cellulose)
Remarks:
0.5 %
Details on exposure:
Doses of 0, 50, 100, 200 mg/kg bw were applied by gavage divided in two portions: one half in the morning, one half in the evening. as for Humans.
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:
Gestation days 6-15 , necropsy at day 17
Frequency of treatment:
daily (morning and evening)
Duration of test:
17 days
Remarks:
Doses / Concentrations:
200
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
100
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
50
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
0
Basis:
nominal conc.
No. of animals per sex per dose:
20-21 only females
Control animals:
yes, concurrent vehicle
Details on study design:
Timed-mated mice (20-21 per group) were monitored at regular intervals for clinical signs of toxicity, feed consumption, and body weight. At necropsy (gd 17), the following observations were made: clinical condition; maternal body, and gravid uterine weights; pregnancy status; and number of corpora lutea. In the gravid uterus, the numbers of prenatal deaths (resorptions and/or dead fetuses) and live fetuses were recorded. Live fetuses were weighed, sexed, and examined for morphological anomalies (external, visceral, and skeletal).
The study was performed in two replicates.
Maternal examinations:
Timed-mated mice (20-21 per group) were monitored at regular intervals for clinical signs of toxicity, feed consumption, and body weight. At necropsy (gd 17), the following observations were made: clinical condition; maternal body, and gravid uterine weights; pregnancy status; and number of corpora lutea. In the gravid uterus, the numbers of prenatal deaths (resorptions and/or dead fetuses) and live fetuses were recorded. Live fetuses were weighed, sexed, and examined for morphological anomalies (external, visceral, and skeletal). The study was performed in two replicates.
Ovaries and uterine content:
yes
Fetal examinations:
yes
Statistics:
no data
Indices:
yes
Historical control data:
no data
Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
For dapsone alone, maternal mortality (10%) was noted at the high dose, but otherwise clinical observations were unremarkable. At scheduled necropsy (gd 17), gross findings included enlarged spleens in 1, 12, 13 or 17 dams in the control through high dose groups, respectively. Maternal body weight, weight change, and gravid uterine weight were each reduced at the highest dose of dapsone (200 mg/kg/day). As noted during the pre-treatment period, there was a tendency for dapsone groups to consume more feed than the controls. Consistent with this observation, maternal feed consumption was usually at or above controls for subsequent measurement periods. The exception to this pattern was a significant reduction in maternal relative feed consumption at the highest dose of dapsone at the beginning of the treatment period (gd 6 to 9).
Dose descriptor:
NOAEL
Effect level:
ca. 200 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
Indices of prenatal mortality appeared to be elevated at the highest dose of dapsone (e.g., 16% resorbed implantation sites vs. 2% for controls). Some indices of prenatal mortality reached statistical significance and effects tended to be more severe in the second replicate. Fetal body weights (male, female or both) were significantly reduced by 25% at the highest dose of dapsone. Altered incidences of fetal morphological anomalies (malformations or variations) were noted, but patterns of effects varied across replicates.
Remarks on result:
not measured/tested
Abnormalities:
not specified
Developmental effects observed:
not specified

Variations and malformations in Dapsone treated embryos were not statistically significant as they were distributed unevenly over all dose groups.

Conclusions:
Dapsone is not teratogenic in CD-1 Swiss albino mice.
Executive summary:

Based on the NTP-study conducted, Dapsone is not teratogenic in CD-1 Swiss albino mice.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
200 mg/kg bw/day
Study duration:
subacute
Species:
mouse
Quality of whole database:
The two studies together (KS.developemtal toxicity and female.fertiltiy.01, which includes the evaluation of teratogenicity) indicate that no teratogenciity occurs.
Additional information

Justification for selection of Effect on developmental toxicity: via oral route:
Based on the NTP-study conducted, Dapsone is not teratogenic in CD-1 Swiss albino mice. The result is confirmed in the combined study on female fertility and teratogenicity in the rat (Female fertility.01).

Toxicity to reproduction: other studies

Additional information

The fertility effect in the rat (sperm motility and number) is relevant at doses significantly above human exposure during therapy or workplace exposure. Clinical experience shows that in the course of high dapsone therapy methemoglobinaemia is very frequent at a comparable dose as in the rat study. In some studies rats showed also pale or blue extremities and other effects. There is a clear evidence that the fertility effects occur at doses which provoke clear signs of toxicity.The effect is therefore considered secondary to parental toxicity.

There are Human data (see e.g. the review by Wolf et al (2002) from pregnant leprosy patients exposed to high doses of dapsone. The effects on the foetus were not teratogenc, but sometimes embryotoxic/lethal (likely by low availability of oxygen during development).

Justification for classification or non-classification

The main toxic effect of dapsone is methaemoglobin formation. This effect occurs in humans rarely at a daily dose of 100 mg/person/day and becomes more frequent in patients with a metabolic disease. The effect is frequent and clinically relevant at about 400 mg/person/day (or ~ 6 mg/kg/day).

 

Effects on the rat male and female reproductive systems have been noted in several experimental studies. A large amount of data is available in published literature, but the Extended One Generation Study as requested by ECHA is summarised as follows:

 

The test item was administered to the rats daily, via oral gavage. Cohorts of F1 animals were used to assess the potential for systemic toxicity, immunotoxicity and potential effects on the sexual maturation, estrous cycles and reproductive performance

In the F0 generation, three groups of 25 male and 25 female rats received dapsone at dose levels of 7.5, 15 or 30 mg/kg/day at a volume dose of 5 mL/kg/day.  Males were treated for ten weeks before pairing, up to necropsy after litters were weaned.  Females were treated for ten weeks before pairing, throughout pairing up to necropsy on Day 28 post partum.  In the F1 generation, 40 males and 40 females were treated from weaning to their scheduled termination (relevant to each cohort) at the same dose levels and volume-dose as the F0 generation.  A similarly constituted Control group received the vehicle, CMC (carboxymethylcellulose) (0.5%) at the same volume dose.

Clinical observations, body weight, food consumption and macropathology examinations were performed on all animals for signs of toxicity, with special emphasis on the integrity and performance of the male and female reproductive system and the health (including thyroid hormone analysis), growth, development and function of the offspring. 

 

Effects noted on this study are summarised in tabular format, in the attachment available in Chapter 13.

 

Based on the results of this study,  it is concluded that a No-Observed-Adverse-Effect-Level (NOAEL) for systemic toxicity in Sprague Dawley rats was not established and lies below 7.5 mg/kg/day since test item related germ cell degeneration in the testes was detected at all doses in Dapsone F0 and F1A males.

The NOAEL for Reproductive toxicity also lies below 7.5 mg/kg/day, since reduced F1 and F2B litter size was seen at all doses of Dapsone.

The NOAEL for Developmental immunotoxicity was 30 mg/kg/day.

 

In the extended one generation study, the degenerative changes observed in the testes were of a minimal to mild (slight) nature, and at this level likely did not result in a significant reduction in sperm production, at least not at a histopathological level, and so no reduced luminal sperm in the epididymides was observed, except in two animals (a control and a Group 2), which was associated with spontaneous degenerative changes (in the control) and a tumour (in the Group 2 animal) in the testes in these animals, and wasn’t treatment-related. The minimal cell debris observed in the epididymides in some males correlated with the minimal to mild testicular degeneration.

 

The sperm analysis conducted in the F0 animals shows no true dose response due to high inter-animal variability, though is statistically different from control for % motility and several mobility parameters (VAP, VSL, VCL, ALH, BCF. Only the highest and control dose levels were analysed for their morphology, but significant abnormalities and decapitation was observed at the high dose. However, a true dose response is seen in the litter sizes of the F1 animals, all treated groups had statistically significantly lower litter size than control.

The sperm analysis conduced in the F1 Cohort 1A showed a monotonic dose response for (%) motility, but sperm in the 7.5 mg/kg bw/d group were motile and fast moving. Though sperm assessments from the 15 mg/kg bw/d treatment group showed most sperm as motile, they were slow moving and statistically different from the control group. Morphology was only assessed for the high dose group (30 mg/kg bw/d) similar abnormalities were seen as in the F0 animals.

 

Extensive details of the use of dapsone is available from occupational exposure and use in the medical/pharmaceutical industry, including exposure of pregnant women and lactating mothers. Dapsone has been used as an acne treatment and in the prevention of malaria and is still recommended by the WHO for leprosy treatment (WHO, 2021). A selection of the available information has been included in the present dossier, where they include toxicologically relevant information. There is a low and insignificant risk of congenital malformations in patients receiving dapsone during pregnancy.

 

A 2018 study by Akinsomisoye et al, used both in vivo and in vitro methods to investigate dapsone effects on the male reproductive system in the rat. Male rats were treated with dapsone via oral gavage for 5 days or 6 weeks. Body weight, reproductive organ weights, sperm parameters, reproductive histology and hormone levels were recorded. The in vivo study showed significant decrease in body and reproductive organ weights, sperm parameters and reproductive hormone concentrations. These effects were duration dependant and recoverable following cessation of treatment. Further details of the study and the in vitro study are summarised in an additional toxicological information endpoint record, however the results indicate effects on testicular and epididymal structure and function are the drivers of male reproductive stress.

 

In a study conducted by Norman et al. in 2000, male rats were treated for 63 days prior to cohabitation with untreated females. Dapsone at high doses impaired fertility in male rats. This was evident from reductions in fertility index (numbers of rats pregnant/number of rats mated), reduced sperm motility (percentage of observed sperm that were motile), and reduced numbers of implantations and viable embryos in the females that did become pregnant. Statistically significant reductions in percentage of motile sperm were observed at exposures of 3 mg/kg/day and above. 0.5 mg/kg/day was the apparent NOAEL in this study, although a non-significant trend toward a reduction in the percentage of motile sperm may have been apparent.

 

The fertility effect in the rat (sperm motility and number) is relevant at doses significantly above human exposure during workplace exposure. Dapsone’s uses for the present dossier are exclusively at industrial sites, for uses such as pre-preg manufacturing via automated impregnation and curing in autoclaves or compression moulding. A high level of control is in place.

Clinical experience shows that methemoglobinemia occurs at a comparable dose as in the rat studies included in the present dossier. Evidence of methaemoglobin formation is shown in the 2021 Extended One Generation Study conducted to OECD TG 443.

 

There is a clear evidence that the fertility effects occur only at doses which provoke clear signs of systemic toxicity. However, a no observed effect level for fertility or general toxicity could not be identified on the key reproductive toxicity study conducted in rats. The human epidemiological study referenced in section 7.10.2 is considered to represent the most appropriate value for use in risk characterisation as it provides the lowest effect level and therefore the DNELs used for risk assessment have used 0.35 mg/kg bw/day as a starting point. Due to the fact that this data originates from human exposure this should be considered more relevant and species specific rather than estimation from rat models.

 

Dapsone, CAS 80-08-0, is classified as follows due to the effects observed on the EOGRTs study:

 

Category 1, STOT, Testes and Epididymis;

Effects on sperm morphology and motility

Effects on male reproductive system; testis and epididymis

Effects on implantation numbers and litter size

 

Cat 2 STOT, RE, Blood

 

Due to the widely available information on the reprotoxic effects of Dapsone, closed systems and extensive RMMs and OCs are required for the industrial use of Dapsone. The use of the substance relevant to the present dossier, is limited to adhesives, castings, laminates and advanced composites where an extremely high level of control is in place at the industrial sites.

 

 

References

 

Akinsomisoye OS, Gupta G, Raji Y (2019) Experimental dapsone administration induces infertility in male Wistar rats: Mechanisms and clinical implications.https://doi.org/10.1016/j.pathophys.2019.07.004

 

Norman A (2005) Application Number 21-794 Pharmacology Review. Dept of Health and Human Services, Public Health Service, Food and Drug Administration, ATLS-119.

 

World Health Organisation (2021) Leprosy Elimination, WHO recommended MDT regimens. Recoverable from:https://www.who.int/lep/mdt/regimens/en/

 

Additional information