Disodium 3,3'-dithiobis[propanesulphonate]

Endpoint summary

Description of key information

Repeated Dose Toxicity Oral: Carcinogenicity study, oral: drinking water, 350 mg/kg bw/day, rat (Sprague-Dawley), male, 40/group, similar to OECD 451, RA from Mesna: NO(A)EL = 350 mg/kg bw/day, there were no signs of toxicity in the animals that received Mesna alone.
Repeated Dose Toxicity Oral: 6 month study, oral: gavage, 500, 1000 and 2000 mg/kg bw, rat (BD II and BD IX), m/f, 10/group, RA from Mesna: NOAEL = 2000 mg/kg bw, tolerated without compound linked mortality or morbidity.
Repeated Dose Toxicity Oral: Chronic toxicity study, oral: drinking water, 1000 mg/L drinking water ≙ 10 mg/kg, rat (DA, Lewis), m/f, 15/group, similar to OECD 452, RA from Mesna: NO(A)EL = 10 mg/kg, no effects on mortality, body weight, gross pathology, histology, nonneoplastic and neoplastic lesions compared to vehicle control.
Repeated Dose Toxicity – other routes: 6 weeks, daily iv-application: 1000 mg/kg bw/day, rats (Sprague-Dawley), m/f, 10/group: NO(A)EL = 1000 mg/kg bw/day, tolerated without side effects.
Repeated Dose Toxicity – other routes: 6 weeks, daily iv-application: 316 mg/kg bw/day, dogs (Beagle), m/f, 3/group: LOEL = 316 mg/kg bw/day, the only toxic effects were vomiting and diarrhoea.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

chronic toxicity: oral

Type of information:

read-across based on grouping of substances (category approach)

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Read-across data (CAS 19767-45-4). The publication is well documented and meets basic scientific principles.

Justification for type of information:

REPORTING FORMAT FOR THE CATEGORY APPROACH
For details, please refer to the attached read-across justification. In brief:

1. HYPOTHESIS FOR THE CATEGORY APPROACH (ENDPOINT LEVEL)
There are two category approaches relevant for all human health associated endpoints: Chain-length category and similar metabolic pathway.
Chain-length Category: Both SPS and Dimesna are sodium salts of two sulphonated alkanes connected via a disulphide group. SPS contains two propane moieties, Dimesna ethane ones, hence, these chemicals only differ minor in their hydrocarbon chains in one –CH2– group. The same applies to the carbon chain in MPS and MESNA, connecting the sodium sulfonate with the sulfhydryl moiety. The reactivity and toxicological relevance of this difference in chain length is considered to be minor compared to the chemicals properties triggered by the two remaining respective functional groups. Comparing the actually available information on the substances with regard to their physico-chemical properties, the minor influence of the hydrocarbon chain length becomes obvious. The melting points for the disulphide compounds and the sulfhydryl ones are consistent, and the ones for the ethane derivatives are as expected slightly lower. All compounds are very soluble in water, and similar consistencies are noted with regard to vapour pressure and partition coefficient.
Metabolic pathway: Here it is aimed to justify the read-across from both MPS to SPS and Mensa to Dimesna, and Dimesna to SPS and Mesna to MPS (and vice versa) based on the available information on their metabolism.
Generally, Mesna and Dimesna are considered to be a metabolite of each other. Also, other metabolites of Mesna were identified, besides Mesna-Mesna (i.e., Dimesna), such as Mesna-Cys, Mesna-homocysteine, Mesna-cysteinylglutamate, Mesna-cysteinylglycine, and Mesna-GSH which have been collectively termed “Dimesna” in some studies, while others refer to the mixed disulfides containing a single Mesna moiety as “Mesna”, quantifying Dimesna separately. The relevant functional groups for the enzymatic and non-enzymatic metabolism of Dimesna and Mesna are the disulphide resp. thiol functional groups. Those are both contained in the related substances SPS and MPS, which only differ from the former in their hydrocarbon chains in one –CH2– group, the basic structure and functional groups are however identical. Hence, only taking into account the given functional groups, a similar toxicodynamic behaviour of SPS and MPS compared to Dimesna and Mesna can be expected.

2. CATEGORY APPROACH JUSTIFICATION (ENDPOINT LEVEL)
As shown above, Mesna, MPS, Dimesna and SPS can be used for read-across to each other by grouping of chemicals. Mesna and MPS and Dimesna and SPS share similar physico-chemical properties as well as they exhibit similar toxicological properties, where data is available. Their alkyl side chains differ only in one –CH2- group, so it can be concluded that e.g. absorption, distribution patterns, or excretion from organ systems and body are comparable. Furthermore, Dimesna and Mesna are considered a metabolite of each other, which allows the conclusion that the same also applies for SPS and MPS. Conclusively, data for Mesna, MPS, and Dimesna can be used to cover data gaps for SPS; especially for the required endpoints for human health assessment.
Freely available toxicological information on Dimesna is lacking, so the available information on SPS, MPS and Mesna will be compared in order to obtain contributing information for the read-across justification, as set out in the attachment
The available data indicate that SPS, MPS and Mesna do not need to be classified as acute toxic according to Regulation (EC) No 1272/2008, all available LD50 (oral or dermal) values are greater than 2000 mg/kg bw, clearly indicating the comparability of the substances and the relative harmlessness of all group members including the target chemical SPS with regard to acute toxicity.
All available Ames tests on SPS, MPS and Mesna are consistently negative. Furthermore, the available in vitro micronucleus test on MPS, the SCE assay and in vivo micronucleus test on Mesna do also not give any indication that this group of substances bears any genotoxic properties. Here, gene mutation as well as chromosome mutation and clastogenicity endpoints are covered. In addition, the SCE assay is indicative for an enhanced repair activity upon genotoxic damage, which may result in several outcomes, e.g. point mutations, chromosome breaks etc., which support additionally the hypothesis that this group does not bear genotoxic properties of any kind.
In the available publications on carcinogenicity, both Mesna and its dimer Dimesna did not trigger any signs of toxicity or carcinogenic activity up to the highest dose tested, i.e., 15 resp. 35 mg/kg bw/d with lifetime exposure. Data on Mesna alone indicate that both doses could have been chosen much higher without resulting in any effects, as e.g. a NOAEL was determined to be 350 mg/kg bw/d over an exposure period of 39 weeks. Again this indicates that this group of chemicals does not trigger any relevant adverse effects upon repeated dosage. Last but not least, Mesna was not identified to be a developmental and or reproductive toxicant in several available studies on that endpoint, up to and including limit dosages of 2000 mg/kg bw/d.

Qualifier:

according to guideline

Guideline:

other: OECD Guideline 451 (Carcinogenicity Studies)

Deviations:

not specified

Principles of method if other than guideline:

92 male Sprague-Dawley rats, 8 weeks old, were divided into two groups. Group A received 70 mg/kg N-nitroso-N-butyl-N-(4-hydroxybutyl)amine (BBNOH) twice a week by gavage, whereas to Group B, BBNOH was administered at the same dose level together with 350 mg/kg Mesna given 5 times a week in drinking water. Two control groups of 40 animals each were given Mesna alone or were totally untreated.

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Süddeutsche Versuchstierfarm, Tuttlingen, F.R.G
- Age at study initiation: 8 weeks
- Weight at study initiation: no data (the weight was checked regularly during the entire period of the treatment).
- Housing: 2 per cage
- Diet (e.g. ad libitum): Altrominm pellets diet; ad libitum
- Water (e.g. ad libitum): tap water; ad libitum

Route of administration:

oral: drinking water

Vehicle:

water

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: 20 mL Mesna solutions were given to rats 5 days a week.

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

39 weeks

Frequency of treatment:

5 times a week

Remarks:

Doses / Concentrations:
350 mg/kg bw
Basis:
nominal in water

No. of animals per sex per dose:

40

Control animals:

yes, concurrent no treatment

Details on study design:

- Dose selection rationale: The high dose of Mesna was chosen in order to provide an excess amount that could interfere with the metabolites of BBNOH in the urinary bladder.

Positive control:

N-nitroso-N-butyl-N-(4-hydroxybutyl)amine (BBNOH) alone.

Observations and examinations performed and frequency:

The body weight was checked regularly during the entire period of the treatment.

Sacrifice and pathology:

At the end of the treatment animals were observed for life, with the exception of a few that were killed when moribund. After dissection, urinary bladders and all other organs showing macroscopical alterations were collected for histological examination.

Statistics:

Kaplan-Meier method.

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Water consumption and compound intake (if drinking water study):

no effects observed

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Details on results:

CLINICAL SIGNS AND MORTALITY
No significant differences were observed in the animals of the 4 experimental groups from a general point of view. Control animals did not show any pathological alteration at the end of the experiment. There were no signs of toxicity in the animals that received Mesna alone. A statistically significant increase of the lifespan of the Mesna treated animals was observed (P< 0.05).

BODY WEIGHT AND WEIGHT GAIN
Administration of Mesna did not have a significant effect on the weight; an increase in body weight was observed at the beginning of the treatment, up to the eighteenth week, followed by a slight decrease and a steady phase at the end of the experiment. Values ranged between 300 g and 500 g for all experimental groups.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study)
Mesna solutions were completely consumed within 24 h from the time of administration. The total Mesna consumption was 63.35 g/kg.

GROSS PATHOLOGY
No findings.

HISTOPATHOLOGY: NON-NEOPLASTIC
No findings.

HISTOPATHOLOGY: NEOPLASTIC (if applicable)
No findings.

Dose descriptor:

NOAEL

Effect level:

350 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: see 'Remark'

Critical effects observed:

not specified

Conclusions:

The analogue mesna was not carcinogenic in a two-year carcinogenicity study in rats. 350 mg/kg bw, which rats received daily during the exposure period of 39 weeks, can be considered as NOAEL since no clinical signs of toxicity and no findings of necropsy were noted in treated animals.

Executive summary:

Mesna is known as a cytoprotective agent that helps to prevent haemorrhagic cystitis caused by a widely used class of antineoplastic drugs, comprising cyclophosphamide, ifosfamide, and trofosfamide (Tacchi et al., 1984). A direct chemical interaction between Mesna and the active metabolites of antineoplastic drugs is supposed to be involved in the protective effects on the urinary bladder. In this respect, the influence of mesna on urinary bladder cancer induced by N-nitroso-N-butyl-N-(4 hydroxybutyl)amine (BBNOH) was studied in male Sprague-Dawley rats.
The treatment consisted of the administration of BBNOH alone per gavage, BBNOH with Mesna, Mesna alone and untreated control. The health effects in treated animals receiving Mesna alone is relevant for this endpoint. The group of 40 male animals (8-week old) received 350 mg/kg bw (total Mesna consumption was 63 g/kg bw (mean for all animals)) in drinking water over a period of 39 weeks. The rats received daily the dose in 20 mL water solution 5 days a week. Health condition of the animals and their body weights were checked regularly during the entire period of the treatment. At the end of the treatment animals were killed and dissected and urinary bladders and all other organs showing macroscopical alterations were collected for histological examination.
No significant differences were observed in the animals of the 4 experimental groups from a general point of view. Mesna solutions were completely consumed within 24 h from the time of administration. Administration of mesna did not have a significant effect on the weight; an increase in body weight was observed at the beginning of the treatment, up to the eighteenth week, followed by a slight decrease and a steady phase at the end of the experiment. Values ranged between 300 g and 500 g for all experimental groups. Control animals (untreated and receiving mesna alone) did not show any pathological alteration at the end of the experiment and there were no signs of toxicity. A statistically significant increase of the lifespan of the mesna treated animals was observed (P< 0.05).There were no findings at necropsy.
As no toxicological findings were observed in animals treated with mesna alone, 350 mg/kg bw can be considered as chronic NOAEL. Mesna is a suitable Read-Across substance for SPS as Dimesna and Mesna could be considered a metabolite of each other, which allows the conclusion that the same also applies for SPS and MPS, and their respective alkyl side chains differ only in one –CH₂- group. This is in detail outlined in the read-across justification. Hence, it can be concluded that the derived NOAEL of 350 mg/kg is applicable for SPS, too.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

350 mg/kg bw/day

Study duration:

chronic

Species:

rat

Quality of whole database:

There are three long-term studies on a suitable read-across substance for SPS available, all assessed with Klimisch 2. They consistently reveal no (adverse) effects up to the highest dose tested, i.e. 10 mg/kg resp. 350 mg/kg bw/day over lifetime and 2000 mg/kg bw/day over 6 months, clearly showing the low intrinsic hazard of Mesna and hence SPS. As for Annex IX of REACH only a 90-day study is required, the tonnage-driven data requirements can be considered as fully met, and in summary the database is of high quality.

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

There are three long-term studies on a suitable read-across substance for SPS available, all assessed with Klimisch 2. They consistently reveal no (adverse) effects up to the highest dose tested, i.e. 10 mg/kg resp. 350 mg/kg bw/day over lifetime and 2000 mg/kg bw/day over 6 months, clearly showing the low intrinsic hazard of Mesna and hence SPS.Mesna is a suitable Read-Across substance for SPS as Dimesna and Mesna could be considered a metabolite of each other, which allows the conclusion that the same also applies for SPS and MPS, and their respective alkyl side chains differ only in one –CH₂- group. This is in detail outlined in the read-across justification. Hence, it can be concluded that the derived results are applicable for SPS, too.

Within study from which the result for further risk assessment was derived (NO(A)EL= 350 mg/kg), rats received the substance over 39 weeks via drinking water.Health condition of the animals and their body weights were checked regularly during the entire period of the treatment. At the end of the treatment animals were killed and dissected and urinary bladders and all other organs showing macroscopical alterations were collected for histological examination. So, although the urinary bladder was the organ of interest in that study, it can be concluded that all possible effects seen at other organs would have been documented in that study, and no possibly relevant effect could be missed. Animals receiving Mesna did not show any pathological alteration at the end of the experiment and there were no signs of toxicity. A statistically significant increase of the lifespan of the mesna treated animals was observed (P< 0.05).There were no findings at necropsy. So, it can be safely concluded that the NO(A)EL= 350 mg/kg is sufficiently reliable for further risk assessment.

Similarly, no effects were seen in another lifespan study and the NOAEL was derived as1000 mg/L in drinking water, on mortality, body weight, gross pathology, histology, non-neoplastic and neoplastic lesions in DA and Lewis rats compared to vehicle control. The lower NOAEL is only based on the respective study design and can assumed to be even higher, supporting the above-mentioned NO(A)EL.

In a 6 month oral gavage study,mesna was administered orally as a 40% solution in doses of 500, 1000 and 2000 mg/kg bw. The animals tolerated mesna up to 2000 mg/kg bw without compound-linked mortality and morbidity. The so derived NOAEL of 2000 mg/kg supports the conclusion that the previously derivedNO(A)EL of ≥ 350 mg/kg bw/day is sufficiently safe for further risk assessment.Mesna is a pharmacologically unremarkable, physiologically largely inert and almost totally non-toxic thio compound.

There is no indication given that the derived results are not applicable for SPS. Similarly, there is no doubt on the relevance of these results derived from animal studies for human risk assessment, as no relevant species-specific differences are known. Data derived from two studies on rats and dogs with iv-application (no relevant exposure route for human risk assessment indicate a slight difference, i.e. there were no effects noted in rats at 1000 mg/kg bw, whereas dogs showed vomiting and diarrhoea at a dose of 316 mg/kg. However, this dose is very high and completely systemically available, so possible uncertainties are already considered to be covered by the lower lifetime-NOAEL (which may even be higher and only limited here due to the study design) compared to the high NOAEL of 2000 mg/kg bw/day over 6 months.

As for Annex IX of REACH only a 90-day study is required, the tonnage-driven data requirements can be considered as fully met with the available chronic studies.

Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Due to precautionary reasons, the study with the longest exposure duration (lifetime) was chosen from three equally reliable studies. There are two lifetime studies available, the chosen study revealing a NO(A)EL of ≥ 350 mg/kg bw/day, the other a lower value of ≥ 10 mg/kg. As it was shown in a 6-month study, a dose of 2000 mg/kg bw was tolerated without compound linked mortality or morbidity in this timeframe. Hence, it can be considered as sufficiently safe to choose a NO(A)EL of ≥ 350 mg/kg bw/day for further risk assessment.

Justification for classification or non-classification

No distinct effects on any organs were reported in three oral long-term studies on a suitable read-across substance,up to 2000 mg/kg bw over 6 months were tolerated without compound-linked mortality and morbidity. There is hence no indication given that Mesna and so SPS needs to be classified as STOT RE.