Abstract LB-B23: Prodrug activation by designer enzyme targeted to CD123-expressing tumors

Abstract

Abstract We propose the combination of tumor-targeting antibodies with a drug-activating enzyme and a prodrug, which could be expected to give synergy and enhanced therapeutic efficacy in the treatment of leukemia and other malignancies. We have engineered human glutathione transferase (GST) enzymes for increased catalytic activity with a prodrug with the aim of delivering the enzyme selectively to a tumor. Antibodies, or similar specific binding proteins, will bind to selected epitopes of the cancer cells where the accompanying enzyme will exert its drug-activating function. The approach is generic and could be implemented with alternative binding proteins and GSTs with different functionalities. Myeloid neoplasms represent 20% of leukemias in children and the majority of the pediatric patients have acute myeloid leukemia (AML). There is a need for improved therapies to eradicate tumor cells with particular focus on leukemic stem cells (LSC), which are responsible for recurrent disease. We are designing, by protein engineering and directed in-vitro evolution, glutathione transferase (GST) enzymes that trigger cytotoxicity via chemically activating chemotherapeutic prodrugs. By fusing the enzyme with an antibody with affinity for epitopes selectively expressed by tumor cells, the release of the toxic agent will be targeted to the neoplastic tissue and minimize collateral damage to normal cells. For proof-of-concept a glutathione transferase enzyme activating the prodrug azathioprine is targeted via an anti-CD123 antibody to acute myeloid leukemia (AML) cells ex vivo. The target receptor CD123 is also expressed on LSC, but not on normal hematopoietic stem cells. Following binding to tumor cells GST will catalyze the conversion of the subsequently administered azathioprine to the anti-metabolite 6-mercaptopurine, which will interfere with nucleotide metabolism and DNA synthesis. Alternative prodrugs activated by other GSTs are also available, indicating the versatility of our approach. An engineered GST with 175-fold enhanced activity with azathioprine has been fused with an scFv-anti-CD123 antibody and the designer enzyme produced by heterologous expression. The recombinant fusion protein is catalytically fully competent and maintains the binding activity of the parental scFv antibody. Citation Format: Bengt Mannervik, Birgitta I. Sjödin. Prodrug activation by designer enzyme targeted to CD123-expressing tumors [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-B23.