Indometacin

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

other information

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

other: does not meet important criteria of today standard method (insufficent number of animals, lack of positive controls/historical control data)

Data source

Reference

Reference Type:

publication

Title:

Evaluation of the anti-inflammatory drug indomethacin, for its genotoxicity in mice.

Author:

Shoba Devi, P et al.

Year:

1987

Bibliographic source:

Mutation Research 188, 343 - 347

Materials and methods

Principles of method if other than guideline:

Micronucleus test in bone marrow cells, sperm head abnormality (SHA) assay and analysis of meiotic chromosomes in sperm cells was performed in Swiss male mice after oral and intraperitoneal administration of Indomethacin at increasing doses.

GLP compliance:

not specified

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

Indometacin

EC Number:

200-186-5

EC Name:

Indometacin

Cas Number:

53-86-1

Molecular formula:

C19H16ClNO4

IUPAC Name:

2-[1-(4-chlorobenzoyl)-5-methoxy-2-methyl-1H-indol-3-yl]acetic acid

Details on test material:

pure form, supplied by Merck Sharp and Dohme Ltd., Bombay, India
suspended in 4% gum acacia for administration

Test animals

Species:

mouse

Strain:

Swiss

Sex:

male

Administration / exposure

Route of administration:

other: (oral and i.p.)

Duration of treatment / exposure:

Micronucleus test: 1 day
SHA (sperm head analysis): for 5 consecutive days
Meiotic chromosome analysis: for 7, 15 or 30 consecutive days

Frequency of treatment:

Micronucleus test: two times (at 0 and 24 hours) by gavage or i.p.
SHA analysis: once daily by gavage or i.p.
Meiotic chromosome analysis: once daily by gavage

Post exposure period:

Micronucleus test: 6 hours after the 2nd administration
SHA analysis: 30 days after the last administration
Meiotic chromosome analysis: 24 hours after the final treatment

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
12 - 24 - 36 mg/kg bw/day
Basis:
nominal conc.
(for oral adminstration)

Doses / concentrations 2

Remarks:

Doses / Concentrations:
8 - 16 - 24 mg/kg bw/day
Basis:
nominal conc.
(for i.p. administration)

No. of animals per sex per dose:

Micronucleus test: 6
SHA analysis: 6
Meiotic chromosome analysis: data not given (probably 6)

Control animals:

yes, concurrent vehicle

Examinations

Tissues and cell types examined:

Micronucleus test: femoral bone marrow cells (smears)
SHA analysis: sperms from cauda epidymidis
Meiotic chromosome analysis: metaphase chromosomes of spermatocytes of testicular cells

Results and discussion

Test results

Test results 1

Sex:

male

Genotoxicity:

positive

Remarks:

(weak effects in micronucleus test at highest dose)

Toxicity:

not specified

Remarks:

(doses used correspond to 25%, 50% and 75% of LD50 values)

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

not examined

Test results 2

Sex:

male

Genotoxicity:

positive

Remarks:

(SAH analysis: abnormal sperm starting at the oral lowest dose)

Toxicity:

not specified

Remarks:

(doses used correspond to 25%, 50% and 75% of LD50 values)

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

not examined

Test results 3

Sex:

male

Genotoxicity:

positive

Remarks:

(increased percentage of chromosomal aberrations in spermatocytes starting at the lowest dose)

Toxicity:

yes

Remarks:

(lethality/toxic effects at medium and high dose after 15 days of treatment)

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

not examined

Any other information on results incl. tables

Micronucleus assay:

After oral dosing percentages of micronuclei in polyerthrocytes (P) were 0.31%, 0.41% and 0.55% at 12, 24 and 36 mg/kg compared to 0.21% in vehicle controls with statistical significance at the 36 mg/kg dose level. In normochromic erythrocytes (N) micronuclei were 0.07%, 0.09% and 0.08 % compared to 0.06% in vehicle controls. P/N ratio was 0.94, 0.88 and 0.73 compared to 0.94 in controls.

After i.p. dosing percentages of micronuclei in polyerthrocytes (P) were 0.39%, 0.41% and 0.63% at 8, 16 and 24 mg/kg compared to 0.26% in vehicle controls with statistical significance at the 24 mg/kg dose level. In normochromic erythrocytes (N) micronuclei were 0.08 % at all dose levels compared to 0.04% in controls. P/N ratio was 0.94, 0.86 and 0.76 compared to 0.94 in controls.

SHA analysis:

Results are given per group (data summarized for all males in the respective groups) in the following table:

Route of Administration	Dose [mg/kg bw]	Total sperm counted	Abnormal sperm N                           %
Oral	0	10820	302	2.79
	12	12028	433	3.60*
	24	11933	465	3.90*
	36	11652	501	4.30*
i.p.	0	11052	364	3.29
	8	11599	452	3.94
	12^a	13001	559	4.30*
	24	12211	602	4.93*
* significant compared to controls (p < 0.01; Chi- square test) ^a dose given in the table in the publication, according to the method section it should be 16 mg/kg bw There was no animal-to-animal variation

Meiotic chromosome analysis:

Indomethacin was toxic/lethal when given for more than 15 days at doses of 24 mg/kg bw and higher. Animals were sluggish and unhealthy looking.

Incidence of chromosomal aberrations in spermatocytes was increased at all dose levels tested with statistical significance:

Total percentage of anomalies were 14.63%, 15.11% and 17.10% compared to 10.35% in the control after 7 days of oral treatment; 16.92%, 16.31% and 18.61% compared to 12.10% after 15 days of treatment at doses of 12, 24 and 36 mg/kg bw/day and 17.83% at the 12 mg/kg dose compared to 13.62% in the control after 30 days of treatment (only males of the 12 mg/kg group survived until evaluation).

Applicant's summary and conclusion

Executive summary:

Micronucleus test in bone marrow cells, sperm head abnormality (SHA) assay and analysis of meiotic chromosomes in sperm cells were performed in Swiss male mice after oral and intraperitoneal administration of Indomethacin at increasing doses (12, 24, 36 mg/kg bw were given by gavage once daily and in relation to the test employed up to 30 consecutive days and 8, 16 and 24 mg/kg bw were given i.p. once daily up to 5 consecutive days). Micronucleus test was positive at the respective p.o. and i.p. highest doses (36 and 24 mg/kg bw, respectively). Frequency of abnormal sperm and number of total chromosomal aberrations was increased at doses of 12 mg/kg and higher. Doses used in this study induced mortality after multiple administration (none of the animals of the 24 and 36 mg/kg doses survived when adminstered p.o. for > 15 days).

Testing protocol was inadequate (e.g. lack of positive controls/historical control data, insufficent number of animals, only positive at one dose level) so that the study can not be used for hazard and risk assessment.