Targeting strategies for mRNA delivery

If material properties (lipid/polymer chemistry, particle size, charge, formulation components) and routes of administration are systematically analyzed and correlated with organ-specific mRNA expression, then rational design principles can be established to achieve targeted mRNA delivery to desired tissues and cell types, enabling the full therapeutic potential of mRNA medicines.

• Secondary Aims:
• To summarize distinct properties of materials used for organ-targeted mRNA delivery.
• To evaluate various routes of administration of mRNA therapeutics and the applications achievable.
• To serve as a useful guide for the community in developing future materials for mRNA delivery.

Component: Payload; Examples Discussed: mRNA (including self-amplifying mRNA, SAM), sgRNA for CRISPR-Cas9, Cas9 mRNA

Component: Lipid Nanoparticles (LNPs); Examples Discussed: Ionizable lipids (D-Lin-MC3-DMA, cKK-E12, C12-200), helper lipids (DOPE, DSPC, cholesterol), PEG-lipids, cationic lipids (DOTAP, DOTMA), lipidoids, amino-alcohol lipids, alkyne-modified MC3 analogues, oxidized cholesterol variants, zwitterionic amino phosphate lipids

Component: Polymers; Examples Discussed: Poly(beta-amino ester)s (PBAEs), charge-altering releasable transporters (CARTs), poly(ethyleneimine) (PEI), poly(glycoamidoamine) brushes, PEG-PAsp(DET), poly(N-isopropylacrylamide) (PNIPAM), PBAE-co-PCL terpolymers, poly(amine-co-ester) polymers

Component: Hybrid Systems; Examples Discussed: Lipid-polymer hybrid nanoparticles, peptide/poloxamine particles, PEGylated synthetic KL4 peptide, collagen sponge with LNPs, alginate gel

Component: Targeting Ligands; Examples Discussed: Mannose (CD206), folate, anti-CD3, anti-CD8, anti-Ly6c, tri-mannose (DC-SIGN/Langerin), aptamers

Component: Routes of Administration; Examples Discussed: Systemic IV, intramuscular, subcutaneous, intradermal, intratumoral, intrathecal, intracerebral, intracardial, inhalation, intranodal, ocular (subretinal/intravitreal), intraosseous

Component: Organ Targets; Examples Discussed: Liver, spleen, lung, tumor, skin, skeletal muscle, cardiac muscle, CNS, bone, immune cells (T cells, B cells, dendritic cells, macrophages)

• Covers studies in mouse, rat, non-human primate, and human clinical trials.
• Includes applications in infectious disease (COVID-19, Zika, HIV, RSV), cancer immunotherapy, protein replacement (hemophilia B, Fabry disease, HTT-I), regenerative medicine (bone, muscle, skin), and CNS disorders (Friedreich's ataxia, Alzheimer's, spinal cord injury).
• Evaluates material properties (size, zeta potential, lipid tail geometry, cholesterol oxidation, polymer hydrophobicity) and routes of administration.
• Summarizes completed/ongoing clinical trials (Table 3).

• Biodistribution and expression imaging: Luminescence imaging, IVIS, MRI.
• High-throughput screening: FIND (Fast Identification of Nanoparticle Delivery) with Cre recombinase/tdTomato reporter mice, DNA/mRNA barcoding, deep sequencing.
• Flow cytometry: Immune cell transfection analysis (T cells, B cells, DCs, macrophages).
• Reporter gene assays: Luciferase, GFP, tdTomato.
• Clinical trial endpoints: Safety, tolerability, efficacy (e.g., COVID-19 prevention, serum TTR reduction).
• Physicochemical characterization: Size, zeta potential, pKa, cryo-TEM.

• No primary experimental data; conclusions are synthesized from existing literature.
• No systematic search strategy or quantitative meta-analysis; “meta-analysis” is narrative.
• Disparate nature of reports prevented critical global understanding; no central repository of findings.
• Focus primarily on preclinical studies; clinical translation data limited.

Limitations of the field highlighted by the authors:

• In vitro–in vivo disconnect: Weak correlation between in vitro and in vivo nucleic acid delivery efficiency.
• Lack of structure-function understanding: Many individual studies evaluate only a single structure-function relationship unique to the material examined.
• Nucleic acid type specificity: Materials effective for DNA or siRNA delivery are often not effective for mRNA delivery; small structural changes can switch specificity.
• Screening challenges: Large number of formulation variables complicates optimization; DOE and FIND approaches help but are not universally applied.
• Route limitations: Local routes (intratumoral, intranodal, intrathecal) may not be clinically translatable for all indications; repeat dosing challenges for CNS.
• Clinical translation: mRNA therapeutics have not yet achieved technological maturity of small molecules or monoclonal antibodies; regulatory and manufacturing challenges remain.
• Need for better models: 3D spheroids, organ-on-a-chip, and organoids are recommended to improve prediction of in vivo outcomes.