A chemically-based approach to the design and evaluation of new anticancer drugs for use in conjunction with gene-directed enzyme prodrug therapy (GDEPT) is described. Most of the GDEPT approaches reported so far have used gancyclovir or 5-fluorocytosine as prodrugs with herpes simplex virus thymidine kinase (HSV tk) and cytosine deaminase (CD), respectively, as the drug-activating enzymes. Although encouraging therapeutic findings have been reported with these prodrug/enzyme combinations, HSVtk and CD have some inherent limitations as drug-activating enzymes in the GDEPT approach. First, these enzymes can only accept as substrates compounds that closely resemble the natural substrates. Accordingly, HSVtk and CD cannot be readily used with other classes of potentially cytotoxic prodrugs. Second, the active metabolites derived from gancyclovir and 5-fluorocytosine are only active against cycling cells. It is well established that most cell in common solid tumors are noncycling. The goal of this application is to develop a general GDEPT activation strategy that can be applied to most structural classes of antitumor agents and where the active metabolites are capable of killing cells in all phases of the cell cycle. Three compounds have been selected for initial studies. These are beta-galactoside derivatives of: (a) DNC4 (a potent doxorubicin analog), (b) aldophosphamide (an intermediate in the bioactivation of cyclophosphamide), and (c) SN-38 (a synthetic camptothecin). As independent entities, the galactosides are expected to be biologically inert. However, they should be converted to cytotoxic metabolites in tumor cells that express beta-galactosidase as a consequence of transduction with the Escherichia coli LacZ gene. The cytotoxicities of the new compounds will be determined against mouse colon carcinoma CT26 cells and human breast carcinoma BT474 cells that been genetically modified with LacZ. Wild type tumor cells and HSVtk transduced cells (which should be unable to activate the beta-galactosides) will be used as controls. The antitumor activities of the galactosides will be determined using these same transduced cells in mouse tumor models.
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