Differential distribution and covalent binding of two labeled forms of methyl-CCNU in the Fischer 344 rat

Abstract

The present study compares the organ distribution and covalent binding of MeCCNU labeled either within the carbamylating [( cyclohexyl-1-14C]MeCCNU; Chx-14C-MeCCNU) or alkylating [( 2-chloroethyl-1,2-14C]MeCCNU; Cle-14C-MeCCNU) region of the compound in an animal model shown to be suitable for studying the nephrotoxicity of the nitrosoureas. Extraction of tissue homogenates with organic solvents of increasing polarity, and subsequent analysis of these extracts by HPLC showed fat to accumulate the highest concentration of parent compound. Kidney accumulated the highest levels of the more polar ether- and methanol-extractable metabolites and/or degradation products of either cyclohexyl-derived or chloroethyl-derived 14C-MeCCNU. Striking differences were apparent in the accumulation, degradation and/or metabolism, and tissue distribution of covalently bound radioactivity for the chloroethyl and cyclohexyl moieties. For example, approximately twice as much cyclohexyl-derived 14C was bound covalently to protein of kidney than to protein of liver or lung. In contrast, approximately twice as much chloroethyl-derived 14C was bound to lung protein than to liver of kidney protein. No radioactivity was bound covalently to tissue DNA following Chx-14C-MeCCNU administration. On the other hand, at 4 h, chloroethyl-derived 14C was irreversibly bound to DNA in the relative amounts of kidney (5.0 nmol/mg), liver (2.7 nmol/mg), and lung (1.25 nmol/mg). These results demonstrate that MeCCNU metabolites and/or degradation products are preferentially accumulated in kidney, a primary target organ for MeCCNU toxicity. Moreover, kidney protein and DNA were subject to extensive carbamylation and alkylation reactions as measured by irreversibly bound cyclohexyl-derived and chloroethyl-derived 14C, respectively. These data suggest that the extent of irreversibly bound drug to tissue macromolecules may be a valid predictor of MeCCNU toxicity. However, the relative toxicological significance of either protein carbamylation or DNA alkylation in mediating MeCCNU-induced nephropathy is not yet understood.