Abstract 1792: A novel lipid nanoparticle formulation transfects acute myeloid leukemia cells and primary AML samples with high efficiency and low toxicity

Abstract RNA interference is a powerful research tool for studying gene function in cancer cells. However, functional genomic studies in Acute Myeloid Leukemia (AML) remain challenging. Classic lipid-based transfection reagents are ineffective in delivering siRNA to AML cells and lentiviral delivery of shRNA is complex and time-consuming. Lipid Nanoparticles (LNPs) are a novel approach to deliver genetic material to cells that are difficult to transfect. However, a formulation of LNP capable of transfecting AML cells and primary AML samples with high efficiency and low toxicity is lacking. Here, we developed an LNP formulation that delivers siRNA to AML cell lines and primary samples without impacting their growth and viability. To identify LNPs able to transfect AML cell lines, we generated a library of LNPs that differed in lipid compositions, aqueous phase pH, and RNA-to-ionizable lipid weight ratio through the Design of Experiment (DOE). We synthesized 54 unique LNPs formulations encapsulating siRNA targeting luciferase. We then screened this library of LNPs against OCI-AML2 leukemia cells over-expressing luciferase. From this screen and subsequent validation studies, we identified a novel formulation, LNP 1-28, as the top hit based on transfection efficiency and safety. LNP1-28 also suppressed luciferase expression in 3 AML cell lines (OCI-AML2, OCI-AML3, MOLM13) with > 80% target knockdown with 100nM of siRNA 3 days post-transfection. Next, we asked if LNP 1-28 could deliver siRNA to knockdown an endogenous protein, CD45. AML cell lines (OCI-AML2, OCI-AML3, MOLM13) were treated with LNP 1-28 carrying siRNA targeting CD45 or control sequences. Knockdown of CD45 protein was observed by flow cytometry in all tested AML cells with knockdown being time and concentration dependent. CD45 knockdown was observed as early as 1-day post-transfection and persisted for 7 days before gradually returning towards baseline. Greater than 82.5% knockdown was observed 4 days after 100nM siRNA-LNP treatment in all tested cell lines. LNP 1-28 had no effect on cell growth and viability. Using the low passage primary AML culture model 8227 cells, we demonstrated that LNP 1-28 could deliver siRNA to both bulk and CD34+/CD38- stem populations of AML. Finally, LNP 1-28 efficiently transfected primary AML patient samples and delivered siRNA to knock down CD45. In summary, we developed a novel LNP formulation capable of transfecting AML cells and primary AML samples with siRNA at high efficiency and without toxicity. Thus, we have developed a nanocarrier system for in vitro transfection that will aid in functional genomics studies of AML. Citation Format: Lan Xin Zhang, Yongran Yan, Jingan Chen, Rose Hurren, Jiachuan Bu, Juan Chen, Dakai Ling, Nathan Duong, Andrea Arruda, Hansen He, Mark D. Minden, Rod Bremner, Gang Zheng, Bowen Li, Aaron D. Schimmer. A novel lipid nanoparticle formulation transfects acute myeloid leukemia cells and primary AML samples with high efficiency and low toxicity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 1792.