ALBUMIN CONJUGATES OF THE ANTICANCER DRUG CHLORAMBUCIL - SYNTHESIS, CHARACTERIZATION, AND IN-VITRO EFFICACY

In our efforts to improve the selectivity and toxicity profile of anti tumor agents, four maleimide derivatives of chlorambucil (1-4) were bo und to thiolated human serum albumin which differ in the stability of the chemical link between drug and spacer. 1 is an aliphatic maleimide ester derivative of chlorambucil, whereas 2-4 are acetaldehyde, aceto phenone, and benzaldehyde carboxylic hydrazone derivatives. HPLC stabi lity studies at pH 5.0 with the related model compounds 5, 7, 8, and 9 , in which chlorambucil was substituted by 4-phenylbutyric acid, demon strated that the carboxylic hydrazone derivatives have acid-sensitive properties; the acid lability of 7 was particular prominent with a hal f-life of only a few hours. The alkylating activity of albumin-bound c hlorambucil was determined with the aid of 4-(4-nitrobenzyl)-pyridine (NBP), demonstrating that on average three equivalents were protein-bo und. Evaluation of the cytotoxicity of free chlorambucil and the respe ctive albumin conjugates in the MCF7 mamma carcinoma and MOLT4 leukemi a cell Line employing a propidium iodide fluorescence assay demonstrat ed that the conjugate in which chlorambucil was bound to albumin throu gh an ester bond was not as active as chlorambucil. In contrast, the c onjugates in which chlorambucil was bound to albumin through carboxyli c hydrazone bonds were as or more active than chlorambucil in both cel l lines. In particular, the conjugate in which chlorambucil was bound to albumin through an acetaldehyde carboxylic hydrazone bond exhibited IC50 values which were approximately 4-fold (MCF7) to 13-fold (MOLT4) lower than those of chlorambucil. Preliminary toxicity studies in mic e showed that this conjugate can be administered at higher doses in co mparison to unbound chlorambucil.