Lipid Nanoparticle-mRNA Formulations for Therapeutic Applications

Abstract

After decades of extensive fundamental studies and clinical trials, lipid nanoparticles (LNPs) have demonstrated effective mRNA delivery such as the Moderna and Pfizer-BioNTech vaccines fighting against COVID-19. Moreover, researchers and clinicians have been investigating mRNA therapeutics for a variety of therapeutic indications including protein replacement therapy, genome editing, and cancer immunotherapy. To realize these therapeutics in the clinic, there are many formidable challenges. First, novel delivery systems such as LNPs with high delivery efficiency and low toxicity need to be developed for different cell types. Second, mRNA molecules need to be engineered for improved pharmaceutical properties. Lastly, the LNP-mRNA nanoparticle formulations need to match their therapeutic applications.In this Account, we summarize our recent advances in the design and development of various classes of lipids and lipid derivatives, which can be formulated with multiple types of mRNA molecules to treat diverse diseases. For example, we conceived a series of ionizable lipid-like molecules based on the structures of a benzene core, an amide linker, and hydrophobic tails. We identified N¹,N³,N⁵-tris(3-(didodecylamino)propyl)benzene-1,3,5-tricarboxamide (TT3) as a lead compound for mRNA delivery both in vitro and in vivo. Moreover, we tuned the biodegradability of these lipid-like molecules by introducing branched ester or linear ester chains. Meanwhile, inspired by biomimetic compounds, we synthesized vitamin-derived lipids, chemotherapeutic conjugated lipids, phospholipids, and glycolipids. These scaffolds greatly broaden the chemical space of ionizable lipids for mRNA delivery. In another section, we highlight our efforts on the research direction of mRNA engineering. We previously optimized mRNA chemistry using chemically-modified nucleotides to increase the protein expression, such as pseudouridine (ψ), 5-methoxyuridine (5moU), and N¹-methylpseudouridine (me¹ψ). Also, we engineered the sequences of mRNA 5' untranslated regions (5'-UTRs) and 3' untranslated regions (3'-UTRs), which dramatically enhanced protein expression. With the progress of LNP development and mRNA engineering, we consolidate these technologies and apply them to treat diseases such as genetic disorders, infectious diseases, and cancers. For instance, TT3 and its analog-derived lipid-like nanoparticles can effectively deliver factor IX or VIII mRNA and recover the clotting activity in hemophilia mouse models. Engineered mRNAs encoding SARS-CoV-2 antigens serve well as vaccine candidates against COVID-19. Vitamin-derived lipid nanoparticles loaded with antimicrobial peptide-cathepsin B mRNA enable adoptive macrophage transfer to treat multidrug resistant bacterial sepsis. Biomimetic lipids such as phospholipids formulated with mRNAs encoding costimulatory receptors lead to enhanced cancer immunotherapy.Overall, lipid-mRNA nanoparticle formulations have considerably benefited public health in the COVID-19 pandemic. To expand their applications in clinical use, research work from many disciplines such as chemistry, engineering, materials, pharmaceutical sciences, and medicine need to be integrated. With these collaborative efforts, we believe that more and more lipid-mRNA nanoparticle formulations will enter the clinic in the near future and benefit human health.

Figure 1.

Biodegradability of (A) FTT5 and (B) FTT9 in vivo; (C) the level and (D) activity of hFIX in hemophilia A mice after i.v. injection of FTT5-hFIX mRNA. Reproduced with permission from ref . Copyright 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. Distributed under a Creative Commons Attribution License 4.0 (CC BY); https://creativecommons.org/licenses/by/4.0/.

Figure 2.

(A) Illustration of mRNA UTR engineering and its application to COVID-19 mRNA vaccines. (B) Serum IgG level in mouse sera post-intramuscular injection. (C) Dosage dependency effects of TT3-NASAR mRNA formulation as a COVID-19 mRNA vaccine. Reproduced with permission from ref . Copyright 2020 Wiley-VCH.

Figure 3.

(A) Illustration of macrophages containing antimicrobial peptides linked to cathepsin B in the lysosomes (MACs) for the treatment of multidrug resistant bacterial sepsis: (i) adoptive macrophage transfer; (ii) recovery from multidrug resistant bacteria induced sepsis. (B) Survival rate and (C) body weights of septic mice after treatments. Adapted with permission from ref . Copyright 2020 Springer Nature.

Figure 4.

Therapeutic effects in an A20 mouse tumor model: (A) tumor size; (B) percent survival; (C) percent survival of rechallenged mice.

Scheme 1.

Synthesis of 1,3,5-Benzenetricarbonyl Trichloride (TT)-Derived Lipid-Like Molecules (A) and Functionalized TT (FTT) Molecules (B)^,

Scheme 2.

Chemical Structures of Vitamin-Derived Ionizable Lipids (A) and Chemotherapy Drug-Derived Ionizable Lipids (B)^,

Scheme 3.

Synthesis of Ionizable Phospholipids (A) and Glycolipids (B)

Scheme 4.

Examples of Chemically-Modified Nucleobases