Abstract C215: EZN-2208, a novel pegylated SN-38 drug conjugate, markedly inhibits tumor growth and metastatic spreading in preclinical models of human neuroblastoma

Abstract

Abstract Treatment of neuroblastoma (NB) is successful in less than half of patients with high-risk disease. Camptothecin and its analog irinotecan (CPT-11), hold great promise but several limitations suggest that chemical modifications may improve the therapeutic index. EZN-2208 is a water soluble pegylated SN38 drug conjugate, composed of a four-arm 40 KDa polyethylene glycol (PEG) linked via glycine residue to SN38. In various preclinical models of solid tumors EZN-2208 was more efficacious than CPT-11. In phase I trials in adult tumors, EZN-2208 was well-tolerated with neutropenia as the dose limiting toxicity. Here, the anti-tumor activity of EZN-2208 was first evaluated in preclinical, pseudometastatic and an orthotopic, models of human NB. Mice were treated every other day for 5 total doses with 10 mg/kg of CPT-11 or with the SN38 equivalents of EZN-2208. In the first model, mice treated with EZN-2208 displayed significant increased life span compared to control mice or those treated with CPT-11. After 150d post cell implantation, all EZN-2208-treated mice were still disease-free, while control and CPT-11-treated animals died with metastatic disease. In the orthotopic model, mice treated with EZN-2208 showed a dramatic arrest and regression in primary tumor growth compared to control mice. While CPT-11-treated mice died with widespread tumor masses within 80 days, long term survival was seen in 100% of EZN-2208-treated animals. 21 days after the end of treatment tumors had almost disappeared, as assessed by staining histological sections of the tumors with antibodies recognizing NB cells and the cell proliferation marker, Ki-67. In a second set of in vivo experiments, MTD doses of both CPT-11 and EZN-2208 were compared in immunodeficient (GI-LI-N cells) and immunocompetent (NXS2 cells) orthotopic NB animal models. While CPT-11 at MTD dose led to a partial increased in long term survival, EZN-2208-treated, GI-LI-N-bearing mice were 100% cured after 180 days post cells implantation. In the very aggressive syngeneic NB animal model (NXS2 cells), while CPT-11 did not exert any anti-tumor effect, EZN-2208 led to 100% and 40% of long term survivors, in mice challenged with 5x104 and 5x105 tumor cells, respectively. The differences in the anti-angiogenic activity between CPT-11 and EZN-2208 was evaluated by chorioallantoic membrane (CAM) assay. Incubation of the CAMs with EZN-2208 significantly reduced the number of radiating vessels that invaded the implant compared to either specimens alone or CAMs incubated CPT-11. In the last set of in vivo experiments, the effects of EZN-2208 and CPT-11 were compared in a luciferase-transfected human NB cells inoculated in the right flank of mice. EZN-2208 led to a significant tumor regression compared to CPT-11. Finally, mechanistic experiments showed enhanced TUNEL and Histone H2ax staining in tumors removed from mice treated with EZN-2208, indicating its effect on tumor cell apoptosis. In conclusion, EZN-2208 could be considered as a new, promising anti-NB agent, to be administered alone and/or in combination with traditional chemotherapeutics. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):C215.