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FDA RNA-based therapies can regulate gene expression and generate therapeutic proteins or antigens that stimulate immune responses to treat a wide range of disease types, including cancers, infectious diseases, immune disorders, and genetic disorders. However, RNA-based gene therapy requires therapeutic RNAs to function within target cells and not be degraded by RNases nor trigger unrelated immune responses. To overcome these obstacles, scientists have developed the viral and nonviral delivery systems.
Therapeutic RNA Delivery
Small RNA molecules can be chemically modified at specific sites to alter stability and pharmacokinetics and potentially trigger immune responses. Meanwhile, they can be delivered using nanoparticles or conjugate-based delivery systems.
- Large RNA molecules (mRNA)
mRNA can encode proteins with therapeutic activities or antigens that trigger immune responses; it can also express DNA-modifying nucleases transiently, such as Zinc finger protein nucleases, Cas nucleases, etc.
Fig. 1 Therapeutic RNA Delivery (Paunovska K, 2022)
RNA delivery system
- Lipid-based delivery system
Lipid-based delivery systems include micelles, liposomes, and lipid nanoparticles (LNP). Chemical reactions can be applied to synthesize different lipids, and lipid structures are influential to the overall delivery. Scientists have invented a library containing tens of thousands of lipid delivery systems of different chemical structures, which supported the efficiency of RNA therapeutics delivered to the liver dramatically. In addition, changing the lipid fraction of LNP or adding new lipid molecules can also facilitate LNP delivery to tissues other than the liver.
Fig. 2 FDA-approved lipid-based structures contain variations on four basic components: cholesterol, helper lipid, PEG-lipid, and cationic or ionizable lipid (Paunovska K, 2022)
- Polymers and polymer-based nanoparticles
Various nonviral delivery systems use polymers and polymer-based nanoparticles, and the efficiency of RNA delivery can be altered by modulating the molecular weights, charges and degradability of polymers. Commonly used polymers include poly (lactic-co-glycolic acid) (PLGA), poly(beta-amino ester)s (PBAEs) and dendrimers.
Fig. 3 Delivering RNA using nanoparticles formulated polymers or dendrimers (Paunovska K, 2022)
- Active targeting delivery
Active targeting strategies incorporate ligands that bind to specific biomolecules with delivery systems. Clinically, most of the validated active targeting strategies involved GalNAc-conjugated siRNA and ASO, and FDA-approved therapeutic examples include givosiran, lumasiran, and inclisiran. Other evidence has demonstrated that conjugation of small molecules or antibodies/antibody fragments to RNA can sufficiently deliver RNA to tissues other than the liver. For example, siRNA can be delivered to cardiac and skeletal muscle when conjugated with anti-CD71 antibodies or antibody fragments, resulting in long-term gene silencing. In addition, siRNA or mRNA is delivered by lipoproteins carried in the LNP, and this system has been used to target extrahepatic cells to treat inflammatory bowel diseases.
- Passive targeting delivery
Passive targeting delivery of RNA relies on nanoparticles adsorbed on the surface of biomolecules to direct specific organ delivery (endogenous or passive targeting). For example, apolipoprotein E (ApoE) adsorption is critical for ionizable lipid delivery of siRNA into hepatocytes, which leads to LNP uptake by hepatocytes via the low-density lipoprotein receptor (LDLR).
Fig. 4 Two mechanisms of action can be used for drug vehicles in reaching target cells (Paunovska K, 2022)
Characteristics of successful RNA delivery systems
- Scalable and biodegradable chemical synthetic strategies; for example, introducing ester bonds in ionizable lipids can improve the overall safety of LNP.
- Simple and straightforward chemical synthetic routes to support scalable production for human uses
- Acceptable on-target/off-target delivery ratios (On-target and off-target delivery need to be measured using both biodistribution and functionalization metrics because 95% of the RNA is likely to be retained in the endosome while biodistribution does not necessarily predict functional RNA delivery)
- Effective dosage should be significantly lower than toxic dosage in terms of drug safety; toxicology studies should be conducted in non-human primates prior to use
- Bath to batch consistency of drug activity
- No loss of efficacy or safety during repeated administration in clinical settings
Reference
- Paunovska K; et al. Drug delivery systems for RNA therapeutics. Nat Rev Genet. 2022 Jan 4: 1-16.
* Only for research. Not suitable for any diagnostic or therapeutic use.
