Inhibition of metabolism-mediated cytotoxicity by 1,1-disubstituted hydrazines in mouse mastocytoma (line p815) cells

Abstract Cyclophosphamide (CPA), a compound requiring metabolic activation to form toxic metabolites, has been used as a model compound to validate the use of a hepatocyte/mouse mastocytoma (line P815) cell culture system capable of measuring metabolism-mediated cytotoxicity in vitro . A number of hydrazines were tested in this system and were found to be relatively non-toxic. However, an attempt has been made to correlate complex formation between hydrazine metabolites and cytochrome P-450 in rat hepatocytes and the inhibition of the metabolism-mediated cytotoxicity of CPA. N -Aminopiperidine (NAP) was the most potent hydrazine tested in this system, since it significantly reduced the metabolism-mediated toxicity of CPA. The concentration dependency of this phenomenon permitted the calculation of the amount of NAP that reduced CPA cytotoxicity by 50%. This value (approximately 1 mM) correlated well with the apparent Michaelis constant (1.2 mM) for the formation of the inhibitory metabolite complex with cytochrome P-450 in microsomal suspensions. There was good correlation between the abilities of a number of 1,1-disubstituted hydrazines to reduce the metabolism-mediated toxicity of CPA and the extent to which they form the inhibitory metabolite complex in microsomes. 1,2-Dimethylhydrazine did not form an observable inhibitory complex in microsomal suspensions and only slightly reduced the metabolism-mediated toxicity of CPA in the mouse mastocytoma cell culture system. However, some other hydrazine derivatives were shown to alter significantly cytochrome P-450 function in isolated hepatocytes. This in vitro system may be of use in the evaluation of the biological effects of xenobiotics, particularly those requiring metabolic activation by cytochrome P-450.