Extracellular -Glucuronidase for Gene-Directed Enzyme-Prodrug Therapy

Most enzymes used for antibody-directed enzyme-prodrug therapy (ADEPT) and gene-directed enzyme-prodrug therapy (GDEPT) are of bacterial, yeast, or viral origin. These xenogeneic proteins can induce an immune response, which might be a hindrance to their application of these enzymes in humans (1 ). On the other hand, when choosing a human enzyme for prodrug conversion cleavage by the respective endogenuous enzyme obviously needs to be avoided. Lysosomal enzymes seem to be particularly suitable candidates, because under normal circumstances, the endogenous enzymes are restricted to intracellular vesicles and therefore not available for premature prodrug conversion. In addition, lysosomal enzymes leaking out of cells are rapidly internalized via the mannose-6-phosphate (M6P) receptor expressed on the surface of most cells and at particulary high levels on reticuloendothelial cells (2 ,3 ). The human lysosomal enzyme β-glucuronidase has been used for ADEPT (4 –8 ) and GDEPT (9 ,10 ). This exoglycosidase removes terminal β-glucuronic acids from glycosami-noglycans and other glycoconjugates. The enzyme is highly specific for the glucuronyl residue but has little specificity for the conjugated aglycone. Therefore, different drugs could be developed as prodrugs (see Table 1 ). Elevated β-glucuronidase levels in tumor tissue have been reported, released from dying tumor cells and invading macrophages and neutrophils (18 ,19 ,36 ,37 ). β-Glucuronides of different drugs might be exploited for tumor-specific conversion by released endogenous enzymes (“monotherapy”) as well as for ADEPT and GDEPT approaches.

Drug	Prodrug	Application	References
Anthracyclins
Dpirubicin	Epirubicin-glucuronide	ADEPT	11,12
Daunorubicin	Daunorubicin-benzyl carbamate spacer-glucuronide (DNR-GA3)	Monotherapy, ADEPT	13–16
Doxorubicin	Doxorubicin-benzyl carbamate spacer-glucuronide (Dox-GA3)	Monotherapy, ADEPT	7,8,16,17
Doxorubicin	Doxorubicin-nitrophenyl spacer-glucuronide (HMR 1826)	Monotherapy, ADEPT, GDEPT	5,9,10,18–22
Daunorubicin and doxorubicin	Daunorubicin and doxorubicin glucuronides		23
Doxorubicin	Doxorubicin-nitrophenyl spacer-glucuronide		24
Doxorubicin	Doxorubicin-enol ether spacer-glucuronide		25
Mustards
Hydroxyaniline mustard (HAM)	Tetrabutyl HAM glucuronide (BHAMG)	ADEPT	26,27
Mustard	Mustard-carbamate spacer-glucuronide		28
Nitrogen mustard	Nornitrogen mustard-nitrophenyl-glucuronide		29
Other cytotoxic Drugs
Diverse	Acetal-glucopyranosiduronates		30
Rooperol	Hypoxoside	Monotherapy	31
Palitaxel	Palitaxel-carbamate-glucuronide		32
9-Aminocamptothecin	9-Aminocamptothecin-aromatic spacer-glucuronide		33
5-Fluorouracil	5-Fluorouracil-carbamate spacer-glucuronide	MRS a	34
MDR inhibitors
Verapamil, quinine, dipyridamole	MDR inh. glucuronides (for combination with antracyclin prodrugs)		35