Combination cancer therapy by hapten-targeted prodrug-activating enzymes and cytokines

Abstract

Combination therapy can help overcome limitations in the treatment of heterogeneous tumors. In the current study, we examined whether multiple therapeutic agents could be targeted to anti-dansyl single-chain antibodies (DNS scFv) that were anchored on the plasma membrane of cancer cells. Functional DNS scFv could be stably expressed on CT-26 colon cancer cells both in vitro and in vivo. Dansyl moieties were covalently attached to recombinant β-glucuronidase (βG) and interleukin 2 (IL-2) via a flexible poly(ethylene glycol) linker to form DNS−PEG−βG and DNS−PEG−IL-2 conjugates. The conjugates displayed enzymatic and splenocyte-stimulatory activities, respectively, that were similar to those of the unmodified proteins. The conjugates selectively bound CT-26 cells that expressed anti-DNS scFv (CT-26/DNS cells) but not CT-26 cells that expressed control scFv (CT-26/phOx cells). DNS−PEG−βG preferentially activated a glucuronide prodrug (BHAMG) of p-hydroxy aniline mustard at CT-26/DNS cells in culture and accumulated in subcutaneous CT-26/DNS tumors after intravenous administration. Systemic administration of DNS−PEG−IL-2 or DNS−PEG−βG and BHAMG significantly delayed the growth of CT-26/DNS but not control CT-26/phOx tumors. Combination treatment with DNS−PEG−βG and BHAMG followed by DNS−PEG−IL-2 therapy significantly suppressed the growth of CT-26/DNS tumors as compared to either single-agent regimen. These results show that at least two DNS-modified therapeutic agents can be selectively delivered to DNS scFv receptors in vitro and in vivo, allowing combination therapy of DNS scFv-modified tumors.