Synthetic delivery systems that control release of anticancer agents from early endosomes

Abstract

Tubulin-binding agents are an important class of therapeutics for cancer chemotherapy. However, their application can be limited by systemic toxicity, poor tissue specificity, and drug resistance. To address these issues, modification of tubulin-binding agents with drug delivery systems has shown significant promise. Our group has developed a novel technology for delivery of small molecules and macromolecules into mammalian cells. By mimicking small natural cell surface receptors, cargo linked to the membrane anchor N-alkyl-3beta-cholesterylamine can be internalized through a clathrin-mediated endocytic pathway and trapped in early/recycling endosomes. Co-administration with endosome-disruptive peptides enable 3beta-cholesterylamine-conjugated cargo to escape these early/recycling endosomes of living mammalian cells. This approach can be highly efficient as evidenced by fluorescence-based assays. To explore the potential of this system for delivery of tubulin-binding agents, we designed and synthesized a series of colchicine- and colchinol methyl ether-cholesterylamine conjugates. These disulfide-linked compounds were designed to deliver tubulin-binding agents into early/recycling endosomes to minimize their cytotoxic effect. Only upon the activation by an endosome disruptive peptide would these conjugates be released into the cytosol. Cleavage of a disulfide bond in a linker region by glutathione provides a mechanism of release. Cell viability assays demonstrated that carefully-designed conjugates to cholesterylamine can substantially reduce the toxicity of tubulin-binding agents by trapping the warhead in endosomes. After activation by endosome disruptive peptides, conjugates with a glutamic acid residue in the linker region proximal to colchinol methyl ether showed high potency against several cancer cell lines including Jurkat lymphocytes (T-cell leukemia) and PC3 prostate cancer cells.