Antibody-directed
enzyme prodrug therapy (ADEPT) targets an
enzyme selectively to a
tumor where it converts a relatively non-toxic
prodrug to a potent cytotoxic
drug. Previous clinical work using antibody-
enzyme chemical conjugates has been limited by the moderate efficiency of
tumor targeting of these molecules. To address this a
recombinant fusion protein composed of MFE-23, an anti-
carcinoembryonic antigen (CEA)
single chain Fv (scFv) antibody, fused to the amino-terminus of the
enzyme carboxypeptidase G2 (CPG2) has been constructed to achieve ADEPT in CEA-producing
tumors. MFE-23::CPG2 fusion
protein was overexpressed in Escherichia coli and purified using CEA affinity chromatography. Efficacy of MFE-23::CPG2 delivery to
tumors in vivo was assessed by measuring catalytic activity after
intravenous injection of purified MFE-23::CPG2 into nude mice bearing CEA-positive LS174T human
colon adenocarcinoma xenografts. Recombinant MFE-23::CPG2 cleared rapidly from circulation and catalytic activity in extracted tissues showed
tumor to plasma ratios of 1.5:1 (6 hr), 10:1 (24 hr), 19:1 (48 hr) and 12:1 (72 hr). (125)I-MFE-23::CPG2 was retained in kidney, liver and spleen but MFE-23::CPG2 catalytic activity was not, resulting in excellent
tumor to normal tissue
enzyme ratios 48 hr after injection. These were 371:1 (
tumor to liver), 450:1 (
tumor to lung), 562:1 (
tumor to kidney), 1,477:1 (
tumor to colon) and 1,618:1 (
tumor to spleen). Favorable
tumor : normal tissue ratios occurred at early time points when there was still 21% (24 hr) and 9.5% (48 hr) of the injected activity present per gram of
tumor tissue. The high
tumor concentrations and selective
tumor retention of active
enzyme delivered by MFE-23::CPG2 establish that this
recombinant fusion protein has potential to give improved clinical efficiency for ADEPT.