A Novel LNP That Delivers mRNA to Bone Marrow Hematopoietic Stem Cells

Researchers from Georgia Institute of Technology and Emory University School of Medicine published a research paper titled "Lipid nanoparticle-mediated mRNA delivery to CD34+ cells in rhesus monkeys" in Nature Biotechnology, a subsidiary of Nature. The study developed a lipid nanoparticle (LNP) called LNP⁶⁷, which does not require bone marrow mobilization or chemotherapy pretreatment and is not modified with targeting ligands. It can deliver mRNA to hematopoietic stem/progenitor cells (HSPC) in rhesus monkeys at a dose as low as 0.25 mg/kg.

Previous studies have shown that LNPs modified with CD117 antibodies can deliver mRNA to mouse bone marrow. But a potential limitation of these studies is that, according to the human protein atlas, human CD117 protein is highly expressed in the respiratory system and lowly expressed in the bone marrow. In another approach, an adenovirus-based delivery system actively targets CD46 (CD46 is expressed in mouse and human HSCs), achieving efficient delivery in non-human primates. However, this approach also requires cell mobilization and chemotherapy. Therefore, clinically relevant doses of LNPs have proven difficult to deliver in non-human primates without active targeting ligands or any other interventions.

Currently, the three FDA-approved LNP-RNA drugs all use four-component LNPs, which are ionizable lipids, auxiliary lipids (phospholipids), cholesterol, and PEGylated lipids. All four components can change the behavior of LNPs in vivo, but it is difficult to predict the tropism of LNPs in vivo using cell culture.

In this new study, the research team used 4 different phospholipids and 4 PEGylated lipid combinations (16 combinations in total), combined with 8 different molar ratios, to generate 128 chemically different LNPs. These LNPs were used to deliver DNA barcodes and mRNA encoding glycosylated phosphatidylinositol (GPI)-anchored camelid VHH antibodies (aVHHs), respectively. Then, the hydrodynamic diameter and polydispersity of all 128 LNPs were analyzed, and monodisperse LNPs with diameters less than 200 nanometers (a total of 105 types) were mixed together.

Considering the scarcity of hematopoietic stem cells (HSCs) and the difficulty in isolating enough cells for sequencing using flow cytometry, the research team used their previously developed single-cell nanoparticle targeted sequencing (SENT-seq) technology to evaluate the effectiveness of the above 105 LNPs in delivering RNA to mouse bone marrow in vivo.

The evaluation results showed that LNP⁶⁷ was able to deliver mRNA to mouse bone marrow and human primary hematopoietic stem cells cultured in vitro. It was also able to deliver mRNA to CD34+ cells in rhesus monkeys at doses of 0.25 mg/kg and 0.4 mg/kg (CD34 is a biomarker for hematopoietic stem/progenitor cells).

Figure 1. LNP⁶⁷ delivers mRNA to CD34+ cells in rhesus monkeys.

In the absence of bone marrow mobilization and pretreatment, LNP⁶⁷ delivered mRNA to hematopoietic stem/progenitor cells (HSPCs) and liver of rhesus monkeys without serum cytokine activation.

These data indicate that it is feasible to use lipid nanoparticles (LNPs) to target hematopoietic stem cells (HSCs) and hematopoietic stem/progenitor cells (HSPCs) in non-human primates for in vivo mRNA delivery without the use of targeting ligand modification.

Reference

Kim H, et al. Lipid nanoparticle-mediated mRNA delivery to CD34+ cells in rhesus monkeys. Nature Biotechnology, 2024: 1-8.