156. Cationic Lipopolymers for BCR-ABL siRNA Delivery and Growth Arrest in Chronic Myeloid Leukemia Tumors

Abstract

Chronic Myeloid Leukemia (CML) is a disease initiated by hematopoietic stem cells after chromosomal translocation, which results in a BCR-ABL fusion oncogene. This fusion causes permanent activation of ABL tyrosine kinase that leads to myeloid cell expansion and accumulation of immature blast in bone marrow and bloodstream[1]. The shortcomings of current leukemia treatments i.e., acquired drug resistance and drug insensitivity, call for development of new treatments [2]. To this end, we have designed a carrier system using cationic lipo-polymers and evaluated the potential of siRNA therapy in vitro and in vivo using this polymer system. Cationic lipo-polymers were prepared by grafting aliphatic lipids (palmitic acid, PA, α-linoleic acid, αLA and carboxyl end-capped αLA) onto 1.2 kDa polyethyleneimine (PEI) via N-acylation [3]. The carboxyl-end capped αLA (tαLA) was prepared by coupling αLA with mercaptopropionic acid through thio-ester linkage. Structuralfunctionalities of tαLA and modified PEIs were analyzed through 1H-NMR spectroscopy and TNBS assay. GFP-expressing CML cells (GFP-K562 cells) were used as cell model and siRNA specific to BCR-ABL as therapeutic model. To create polymer/siRNA complexes that are more conducive for dissociation, polyanionic additive (hyaluronic acid, HA) was incorporated along with siRNA to PEI- tαLA complexes.GFP silencing efficacy of PEI in GFP-K562 cells was significantly increased (~ 60%) by hydrophobic modification and it was higher/or comparable to commercial agents, e.g. Lipofectamine™ 2000, Turbofect™ and PEI25. Addition of HA into complexes further increased the GFP silencing efficacy (~ 80%). A significant cell growth arrest was observed in day-6 and day-9 in GFP-K562 cells compared to control groups after the treatment with PEI1.2-αLA/BCR-ABL complexes. To evaluate the in vivo therapeutic efficacy, five-week old female nu-nu mice (Taconic Farms) were employed as animal models. Subcutaneous treatment (vicinity of the tumor) with PEI1.2-αLA/BCR-ABL was substantially effective in reducing tumor volumes starting from day 3 (Figure 1). ddPCR supports this silencing effect by revealing a reduction in BCR-ABL mRNA expression. Similar results were found when tumors were injected with siRNA intra-peritoneally. These preliminary results demonstrate the use of hydrophobically modified small molecular weight PEIs as effective carriers for siRNA delivery and therefore their potential for therapeutic use in the treatment of CML. View Large Image | Download PowerPoint Slide