title: Targeting urokinase and the transferrin receptor with novel, anti-mitotic N-alkylisatin cytotoxin conjugates causes selective cancer cell death and reduces tumor growth.
abstract: Tumor-specific delivery of ligand-directed prodrugs can increase the therapeutic window of chemotherapeutics by maintaining efficacy whilst decreasing toxic side effects. We have previously described a series of synthetic N-alkylated isatin cytotoxins that destabilize microtubules and induce apoptosis with 10-fold greater potency than conventional anti-mitotics in vitro. Here, we report the characterization, in vitro cytotoxicity and in vivo efficacy of a lead compound, 5,7-dibromo-N-(p-hydroxymethylbenzyl)isatin (N-AI) conjugated via an esterase-labile linker (N-AIE) to two proven targeting ligands, transferrin (Tf) and plasminogen activator inhibitor type 2 (PAI-2/serpinB2). N-AI was released from N-AIE and the targeting ligands Tf/PAI-2 in an esterase-dependent manner at 37 C and both Tf- and PAI-2-N-AIE conjugates were stable at physiological pH. Human cancer cell lines which vary in their expression levels of Tf receptor (TfR/CD71) and PAI-2 target, receptor bound urokinase (uPA) selectively internalized the conjugates. Tf-N-AIE was up to 24 times more active than the free drug and showed clear selectivity patterns based on TfR levels. PAI-2-N-AIE showed equivalent activity compared to the parent drug and strong selectivity patterns for uPA levels. In preliminary in vivo experiments, the PAI-2- and Tf-N-AIE conjugates were efficacious at 1/20(th) and 1/10(th) of the dose of the free N-AI, respectively, in a metastatic, orthotopic human breast tumor xenograft mouse model. Thus, this strategy specifically delivers and concentrates a novel class of isatin-based, tubulin destabilizing agents to tumors in vivo and warrants further detailed preclinical investigation.
authors: School of Biological Sciences, University of Wollongong, NSW, 2522, Australia. kara@uow.edu.au
authors: K L
authors: KL
authors: Vine
authors: V
authors: V
authors: Indira Chandran
authors: J M
authors: JM
authors: Locke
authors: L
authors: L
authors: Matesic
authors: J
authors: J
authors: Lee
authors: D
authors: D
authors: Skropeta
authors: J B
authors: JB
authors: Bremner
authors: M
authors: M
authors: Ranson
mesh_terms: D000818:Animals; D050256:Antimitotic Agents / Q000008:administration & dosage*; D000970:Antineoplastic Agents / Q000008:administration & dosage* / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects / Q000502:physiology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D003603:Cytotoxins / Q000008:administration & dosage*; D016503:Drug Delivery Systems / Q000379:methods*; D005260:Female; D006801:Humans; D007510:Isatin / Q000008:administration & dosage*; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D011990:Receptors, Transferrin / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D020298:U937 Cells; D014568:Urokinase-Type Plasminogen Activator / Q000037:antagonists & inhibitors* / Q000378:metabolism; D023041:Xenograft Model Antitumor Assays / Q000379:methods
chemical_list: D050256:Antimitotic Agents; D000970:Antineoplastic Agents; D003603:Cytotoxins; D011990:Receptors, Transferrin; D007510:Isatin; D014568:Urokinase-Type Plasminogen Activator