Gene therapy introduces healthy genes into cells to replace or inactivate mutated genes. This can cure diseases caused by abnormal gene activity or prevent diseases from arising through genetic predisposition. Lipid nanoparticles are a critical nonviral technology to deliver genetic drugs such as siRNA, mRNA, and plasmid DNA.

Reduced Immune Response

Five years ago, the FDA approved an adeno-associated virus (AAV) based by the FDA to treat inherited forms of blindness, but it has limitations. AAVs are small and cannot carry sizeable gene-editing machinery to perform complex mutations. However, scientists have successfully used a flexible lipid nanoparticle to deliver mRNA-based gene therapy in humans. The results were published in Biomaterials Science. In addition to the mRNA cargo, the lipid nanoparticles were modified to contain a synthetic peptide ligand that binds to the endosomal pH sensor KAT5/KAT2. By selecting peptide ligands with specific binding properties, lipid nanoparticles can be tailored to target different cell types. For example, a lipid nanoparticle that contains an anti-miRNA ligand is targeted to cells expressing the miRNA, while a lipid with an anti-neovascular peptide ligand targets cancer cells. Lipid nanoparticles also have several other properties that make them suitable for gene therapy delivery. For instance, they are relatively non-toxic and can be delivered by intradermal, subcutaneous, or intramuscular injection. This local administration can prime immune responses, as resident and recruited antigen-presenting cells in these tissues can internalize mRNA-encoded antigens and direct them to draining lymph nodes, where they can be processed by T cells and presented to other cells.

Additionally, lipid nanoparticles can be stabilized by adding other lipid components, such as phosphatidylcholine or polyethylene glycol (PEG)-functionalized lipids. These lipids improve particle stability, delivery efficacy, tolerability, and biodistribution.

Ease of Manufacture

Lipid nanoparticles are among the most clinically advanced nonviral gene delivery systems and can overcome many barriers that hamper effective cell transfection. They can encapsulate nucleic acids, efficiently transfect into cells, and deliver mRNA to specific target cells, providing an effective alternative to viral vectors. Lipid-based nanoparticles can be readily formulated to contain hydrophobic molecules such as nucleic acids, drugs, and other biologically active substances. They can be formulated in either liquid or solid form depending on the ratio of the different lipid components and other stabilizing agents. For instance, SLNs can be formulated with water-soluble or liposoluble drugs and can enclose different types of mRNA in their core. The lipid bilayer can also be functionalized with a protein to allow host-guest chemical interactions, such as lipid-anchored antibodies.

Furthermore, LSNs can be formulated for organ-targeting using alterations in the lipid structures. For example, changing the alkyl chain length of cholesterol can result in selective mRNA accumulation in liver endothelial cells, Kupffer cells, and hepatocytes. In addition, the lipid-mRNA conjugates can be labeled with fluorescent or radioactive tags to achieve targeted cell imaging. This allows a precise understanding of the dynamics of the complex interaction between the lipid-mRNA complex and its target cells, allowing for improved design and optimization of future gene therapies.

Multidosing Capabilities

Lipid nanoparticles (LNPs) are among the most clinically advanced nonviral gene delivery systems. They enable efficient delivery of siRNAs to cells and in vivo gene editing in human tissues, addressing a key barrier for developing genetic medicine. They have been used to deliver mRNA to treat various cancers, infectious diseases, and genetic disorders. LNPs can simultaneously encapsulate and release multiple therapeutic agents, increasing a drug compound’s therapeutic index and potential. The size, shape, and lipid composition of a lipid nanoparticle can significantly affect its biological properties. For example, cholesteryl oleate and other cholesterol derivatives have enhanced LNP stability by modifying membrane rigidity multilamellar and lipid partitioning. The molecular geometry of the lipids and their ability to interact with the anionic membranes of blood cells also impact LNP biocompatibility. In addition, LNPs can deliver larger payloads of nucleic acids or proteins into cells without additional carrier molecules.

Larger Payloads

Genetic drugs can treat many diseases by silencing pathological genes or expressing therapeutic proteins. Lipid nanoparticle (LNP) formulations can deliver RNA or DNA and allow multiple dosing. LNPs can carry large payloads — tens to hundreds of genes or oligonucleotides — which significantly expands the therapeutic applications of genetic drugs. Traditional methods for preparing lipid-based LNPs, such as thin-layer hydration or ethanol injection, produce large and heterogeneous particles with low encapsulation efficiency. Alternatively, microfluidic techniques have enabled the preparation of small volumes of lipid-based mRNA particles with high encapsulation efficiency. These can then be rapidly mixed into larger batches in T-mixers for production-scale manufacturing. In addition to cationic lipids, lipid nanoparticle-mRNA formulations often contain non-ionic lipids that improve particle stability, delivery efficacy, and biodistribution. The polarity of the lipids and their molecular geometry can have a significant impact on particle properties. For example, branched tail lipids such as 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) enable lipid-based mRNA particles to fuse with the membrane of endosomes, inducing their osmotic rupturing and release into the cytosol. The structure of lipids can also be modified to achieve organ- or cell-specific delivery of lipid-based mRNAs. For example, changing the length of the hydrophobic tails of cholesterol analogs results in preferential accumulation of lipid-mRNA formulations in liver cells. Barcoding can further enable the high-throughput testing and profiling of lipid-based mRNA formulations at the cell level.

By Sambit