Summary
The effect of the three purine antimetabolites 6-mercaptopurine riboside (6-MPr), 6-methylmercaptopurine riboside (6-MeMPr), and azathioprine on the differentiation of the forelimbs of mouse embryos was evaluated by comparing the action of these drugs in vivo with that observed in an organ culture system. The following results have been obtained:
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1.
In vivo a single dose of 6-MPr triggered an abnormal development of the extremities, especially on days 10 and 11 of gestation, with a maximal sensitivity on day 10. The scapula, “long bones” and forepaw are dose-dependently affected on day 10 of gestation whereas the abnormalities are mostly confined to the forepaw skeleton when the drug is given on day 11.
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2.
In the NMRI strain, azathioprine had about 1/3 to 1/4 of the teratogenic capacity of 6-MPr when given subcutaneously. Azathioprine is considerably less embryotoxic when given as a single, oral dose. Malformations of the extremities, produced by azathioprine, closely resembly those seen after the application of 6-MPr.
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3.
6-MeMPr, which is believed to be a metabolite of 6-MPr and of azathioprine, up to 60 mg/kg s.c., is not able to induced any limb abnormalities in the NMRI strain used.
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4.
When tested in the organ culture system, initiated with limb buds from 11 day mouse embryos, 6-MPr, 6-MeMPr, and azathioprine, in concentrations exceeding 1 to 5 μg/ml, are capable of inducing a disturbance of limb development.
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5.
A closer analysis of the dose-response relationship by using a technique for a “semiquantitative” evaluation, revealed that the three purine antimetabolites do not act via the same mechanism. Surprisingly, in vitro, 6-MeMPr was found to be a stronger teratogen than the other two antimetabolites.
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6.
The phenomenon of “phase specificity” can also be demonstrated in the organ culture system: the malformation pattern depended on the stage at which the limb bud was put into culture and on the time interval during which the drug acted on the explant in vitro.
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7.
The considerable difference in the teratogenic capacity of the three purine antimetabolites observed in vivo apparently is not due to a different susceptibility of the embryonic tissue to the drugs but can only be explained by pharmacokinetic variances in different species.
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Abbreviations
- 6-MP:
-
6-Mercaptopurine
- 6-MPr:
-
6-mercaptopurine riboside
- 6-MeMPr:
-
6-methylmercaptopurine riboside
References
Aydelotte, M. B., Kochhar, D. M.: Development of mouse limb buds in organ culture: chondrogenesis in the presence of a proline analogue, l-azetidine-2-carboxylic acid. Develop. Biol. 28, 191–201 (1972)
Caplan, R. M., Dossetor, J. B., Mauhan, G. B.: Pregnancy following cadaver kidney homotransplantation. Amer. J. Obstet. Gynec. 106, 644–648 (1970)
Holden, H. E., Ray, V. A., Wahrenburg, M. G., Zelenski, J. D.: Mutagenicity studies with 6-mercaptopurine. 1. Cytogenetic activity in vivo. Mut. Res. 20, 257–263 (1973)
Horáková, K., Navarová, J., Paterson, A. R. P.: The delayed cytotoxic effect of 6-mercaptopurine, characterization of the unbalanced growth in HeLa cells produced by 6-mercaptopurine. Biochim. biophys. Acta (Amst.) 366, 333–340 (1974)
Kaufman, J. J., Dignam, W., Goodwin, W. E., Martin, D. C., Goldman, R., Maxwell, M. H.: Successful, normal childbirth after kidney homotransplantation. J. Amer. med. Ass. 200, 338–341 (1967)
Kury, G., Chaube, S., Murphy, M. L.: Teratogenic effects of some purine analogues on fetal rats. Arch. Path. 86, 395–402 (1968)
LePage, G. A.: Biochemical effects and mechanism of action of 6-thioguanine. Cancer Res. 23, 1202–1206 (1963)
Lessmöllmann, U., Hinz, N., Neubert, D.: In vitro system for toxicological studies on the development of mammalian limb buds in a chemically defined medium. Arch. Toxicol. 36, 169–176 (1976)
Lower, G. D., Stevens, L. E., Najarian, J. S., Reemtsma, K.: Problems from immunosuppressives during pregnancy. Amer. J. Obstet. Gynec. 111, 1120–1121 (1971)
Luce, J. K., Frenkel, E. P., Vietti, T. J., Isassi, A. A., Hernandez, K. W., Howard, J. P.: Clinical studies of 6-methylmercaptopurine riboside in acute leukemia. Cancer. Chemother. Rep. 51, 535–546 (1967)
Merkatz, I. R., Schwartz, G. H., David, D. S., Stenzel, K. H., Riggio, R. R., Whitsell, J. C.: Resumption of female reproductive function following renal transplantation. J. Amer. med. Ass. 216, 1749–1754 (1971)
Nelson, J. A., Parks, R. E.: Biochemical mechanisms for the synergism between 6-thioguanine and 6-(methylmercapto)purine ribonucleoside in sarcoma 180 cells. Cancer Res. 32, 2034–2041 (1972)
Neubert, D., Merker, H.-J., Tapken, S.: Comparative studies on the prenatal development of mouse extremities in vivo and in organ culture. Naunyn-Schmiedeberg's Arch. Pharmacol. 286, 251–270 (1974a)
Neubert, D., Tapken, S., Merker, H.-J.: Induction of skeletal malformations in organ cultures of mammalian embryonic tissues. Naunyn-Schmiedeberg's Arch. Pharmacol. 286, 271–282 (1974b)
Neubert, D., Merker, H.-J., Barrach, H.-J., Lessmöllmann, U.: Biochemical and teratological aspects of mammalian limb bud development in vitro. In: Tests of teratogenicity in vitro (J. D. Ebert and M. Marois, eds.), pp. 335–365. Amsterdam: North Holland Publishers 1976
Neubert, D., Lessmöllmann, U., Hinz, N., Schmidt, K.: Score system for the quantitative evaluation of drug effects in an organ culture system. Arch. Toxicol. in press (1977)
Penn, I., Makowski, E., Droegemueller, W., Halgrimson, C. G., Starzl, T. E.: Parenthood in renal homograft recipients. J. Amer. med. Ass. 216, 1755–1761 (1971)
Rosenkrantz, J., Githens, J. H., Cox, S. M., Kellum, D. L.: Azathioprine (Imuran) and pregnancy. Amer. J. Obstet. Gynec. 97, 387–394 (1967)
Thiersch, J. B.: Effect of 6-mercaptopurine on the rat fetus and reproduction in the rat. Ann. N.Y. Acad. Sci. 60, 220–227 (1954)
Thiersch, J. B.: Effect of 6-(1′-methyl-4′-nitro-5′-imidazolyl)-mercaptopurine and 2-amino-6-(1′-methyl-4′-nitro-5′-imidazolyl)-mercaptopurine on the rat litter in utero. J. Reprod. Fertil. 4, 297–302 (1962)
Tidd, D. M., Paterson, A. R. P.: Distinction between inhibition of purine nucleotide synthesis and the delayed cytotoxic reaction of 6-mercaptopurine. Cancer Res. 34, 733–737 (1974)
Trowell, O. A.: The culture of mature organs in a synthetic medium. Exp. Cell Res. 16, 118–147 (1959)
Tuchmann-Duplessis, H., Mercier-Parot, L.: Considérations sur les tests tératogènes. Différences de réaction de trois espèces animals à l'égard d'un antitumoral. C. R. Soc. Biol. (Paris) 158 (1964a)
Tuchmann-Duplessis, H., Mercier-Parot, L.: Production de malformations des membres chez le lapin par administration d'un antimétabolite: l'azathioprine. C.R. Acad. Sci. (Paris) 259, 3648 (1964b)
Tuchmann-Duplessis, H., Mercier-Parot, L.: Réactions provoquées chez l'embryon par deux antimétabolites chimiquement voisins. Bull. Acad. Suisse Sci. Med. 22, 153–165 (1966)
Wilcoxon, F.: Individual comparisons by ranking methods. Biometrics 1, 80–83 (1945)
Yahia, C., Hyman, G. A., Phillips, L. L.: Acute leukemia and pregnancy. Obstet. Gynec. Surv. 13, 1–21 (1958)
Zimmerman, T. P., Chu, L.-C., Buggé, C. J. L., Nelson, D. J., Lyon, G. M., Elion, G. B.: Identification of 6-methylmercaptopurine ribonucleoside 5′-diphosphate and 5′-triphosphate as metabolites of 6-mercaptopurine in man. Cancer Res. 34, 221–224 (1974a)
Zimmerman, T. P., Chu, L.-C., Buggé, C. J. L., Nelson, D. J., Miller, R. L., Elion, G. B.: Formation of 5′-nucleotides of 6-methylmercaptopurine ribonucleoside in human tissues in vitro. Biochem. Pharmacol. 23, 2737–2749 (1974b)
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Neubert, D., Lessmöllmann, U., Hinz, N. et al. Interference of 6-mercaptopurine riboside, 6-methylmercaptopurine riboside and azathioprine with the morphogenetic differentiation of mouse extremities in vivo and in organ culture. Naunyn-Schmiedeberg's Arch. Pharmacol. 298, 93–105 (1977). https://doi.org/10.1007/BF00508616
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DOI: https://doi.org/10.1007/BF00508616