DRUG EXPERIENCE

Drug Safety 6 (5): 390-392, 1991 0114-5916/91/0009-0390/$01.50/0 © Adis International Limited. All rights reserved.

DRS1034

Sister Chromatid Exchange in Patients Treated with Nonsteroidal Anti-Inflammatory Drugs Sernra ~arda~, Ali Esat Karakaya, Necla Ugur, Mustafa <;engel and Ahrnet Melin Gazi University. Faculty of Pharmacy, Toxicology Department, Ankara and Turkish State Railways Ankara Hospital, Turkey

Many chemicals with an anti-inflammatory ac­tion have been shown to influence DNA metab­olism and can give rise to later damage in the gen­etic material (Hoffer & Thumb 1984; Klein & Wottawa 1975). Sister chromatid exchange (SeE) is considered to be a more sensitive, rapid and sim­ple cytogenetic end-point test for evaluating the genotoxic potential of a variety of mutagenic and carcinogenic agents. It also appears to be useful for assessing the cytogenetic impact of clastogenic agents administered to patients. SeE represents the interchange of DNA . replication products at ap­parently homologous chromosomal loci; the mo­lecular exchange can easily be visualised under a light microscope at the metaphase stage. An in­creased number of such exchanges in lymphocytes reflects the influence of mutagens (Das 1988). We therefore attempted to determine if there is an in­crease in SeE baseline in rheumatic patients after therapy with nonsteroidal anti-inflammatory drugs (NSAIDs) [indomethacin, ibuprofen, diclofenac).

Patients and Methods

45 patients (30 female and 15 male) with con­firmed diagnoses of degenerative rheumatic dis­eases and intervertebral disc disorders were en­rolled in this study. Each subject was interviewed concerning smoking habits, use of other drugs, de-

tails of work (exposures to physical or chemical agents), past illnesses, food habits, hereditary dis­eases, consumption of alcohol and coffee, etc., in order to eliminate the biological factors which are already known to interfere with SeE results (Das 1988). 40 nonsmoking patients were selected; the other 5 smoked 1 pack of cigarettes a day and were eliminated from the results. These patients had not received previous therapy before, were newly di­agnosed and gave informed consent. The therapy for each patient consisted of 1 of the following NSAIDs: diclofenac ('Voltaren', 200 mg/day); ibu­profen ('Brufen', 1200 mg/day); or indomethacin ('Indocid', 75 mg/day). The first blood sample was taken before commencing therapy, and the second after a 2-week treatment with 1 of the antirheu­matic agents. The ages of the patients were be­tween 26 and 50 years.

Analysis

Peripheral heparinised blood samples were drawn aseptically from each patient and incubated (in duplicate) in 199Technetium medium (Gibeo) supplemented with 10% fetal calf serum (Gibco), reconstituted phytohaemaglutinin 0.2ml, penicillin SOU, streptomycin 10 mg/L, and 5-bromodeoxy­uridine (in dark) at a final concentration of 10-5

mol/L. The lymphocytes were allowed to divide

Sister Chromatid Exchange in NSAID Therapy

for 2 cell cycles at 37C. Colchicine 0.5 mg/L was added to the cultures for the last 3 hours of incu­bation. The cells were centrifuged and resuspended in hypotonic solution (0.075 mOl/L) fixed, and washed in a 3 : 1 mix of methanol and acetic acid.

Chromosomal preparations made by the con­ventional method were stained by fluorescence plus the Giemsa technique of Perry and Wolff (1974), using fluorescent dye (Hoechst 33258, 10-4 moll L) and Giemsa stain. An average of 30 metaphase plates with 46 intact chromosomes and well dif­ferentiated SCE were scored from each subject to calculate the mean SCE/cell, on coded slides, by a single observer.

Results and Discussion

SCE assay has been used quite extensively in genetic toxicology studies. It has been demon­strated that the frequency of exchange is dramat­ically increased when cells are exposed to known mutagens and carcinogens (Latt et al. 1981; Perry & Evans 1975).

The present study showed no significant differ­ence in SCE frequency in 40 patients, either treated or untreated (table I) and the incidence ofSCE fre­quency did not differ between drugs. We also ana­lysed our data in relation to sex: no significant dif­ference was observed in the SCE rate between treated and untreated male and female patients. Another variable of concern was the age of the patients, since it is not always possible to obtain accurate age-matching in volunteers. However, an­alysis of the data for age in relation to SCE fre­quency also revealed no change in the rates.

Table I. Mean Sister chromatid exchange (SCE)/cell in human

lymphocytes before and after therapy with nonsteroidal anti-in­

flammatory drugs (not significant at p > 0.05 by paired t-test)

Drug

Oiclofenac

Ibuprofen

Indomethacin

No. of

subjects

15

12

13

Mean SCE/cell (± SO)

before after

7.60 ± 1.59 7.67 ± 1.64

5.98 ± 1.70 6.02 ± 2.05

6.60 ± 1.07 6.90 ± 1.60

391

Cigarette smoking is perhaps the most import­ant confounding factor in the interpretation of hu­man SCE frequencies. Although we attempted to control for cigarette smoking, 5 patients were smokers. Because we were unable to replace these with nonsmokers, we excluded their results from statistical evaluation; their mean SCE/cell values were significantly higher than those observed in nonsmokers, although the number of these subjects was small. This is not surprising, since many po­tent mutagens and carcinogens have been detected in smoke condensates and there are many studies on the cytogenetic effects of cigarette smoking (Lambert et al. 1978, 1982; ~arda~ et al. 1991).

Further studies are needed to verify these data for other antirheumatic agents and for longer pe­riods of therapy. However, our results support the study of Kullich and Klein (1986) who also found no increased risk before and after therapeutic ap­plication of several NSAIDs in human lympho­cytes. Their study included 9 different drugs and mitocycin C, as a mutagen, was also employed as a positive control substance in vitro.

Conclusion

The data generated in this study by using the SCE technique indicate that no genetic damage need be feared in patients receiving 2 weeks' therapy with the NSAIDs used in this study. However, a battery of tests is always recommended in order to estab­lish the mutagenic/carcinogenic risk of chemical and physical agents.

References

Das Be. Factors that influence formation of sister chromatid ex­changes in human blood lymphocytes. CRC Critical Reviews in Toxicology 19: 43-86, 1988

Hoffer L, Thumb N. Anti-inflamatorisch wirksame Substanzen, ihre Wirkung auf DNA Synthese und Reperatur. In Kuem­merle (Ed.) K1inische pharmakologie, Vol. 4, pp. 1-12, Eco­Med Verlag, Landsberg, 1984

Klein G, Wottawa A. Der Einfluss sogennter Basis therapeutica und symptomatisch wirksamer antirheumatica auf Enzyme der DNA-reparatur. Acta Medica Austriaca 2: 153-156, 1975

Kullich W, Klein G. Investigations of the influence of nOnster­oidal antirheumatic drugs on the rates of sister chromatid ex­change. Mutation Research 131-134, 1986

Lambert B, Lindblad A, Nordenskjold M, Werelius B. Increased

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frequency of sister chromatid exchange in cigarette smokers. Hereditas 88: 147-149, 1978

Lambert B, Bemdtsson I, Lindsten J. Smoking and sister chro­matid exchange. Sorsa & Vainio (Eds) In Mutagens in our en­vironment, pp. 401-414, Alan R. Liss Inc, New York, 1982

Latt SA, Allen J, Bloom SE, Carrano A, Falhe E, et al. Sister chromatid exchanges; a report of the Gene-Tox program. Mu­tation Research 87: 17-62, 1981

Perry P, WolffS. New Giemsa method for the differential staining of sister chromatids. Nature 251: 156-158, 1974

Perry P, Evans HJ. Cytological detection of mutagen-carcinogen

Drug Safety 6 (5) 1991

exposure by sister chromatid exchange. Nature 258: 121-125, 1975

~rda, S, G6k S, Karakaya AE. Increased frequency of sister chro­matid exchanges in the peripheral lymphocytes of cigarette smokers. Toxicology In Vitro, in press, 1991

Correspondence and reprints: Dr Semra $ardQfj, Gazi University, Faculty of Pharmacy, Toxicology Department, 06330 Ankara, Turkey.