Oligonucleotide-based therapeutics represent a transformative approach in modern biomedicine, enabling targeted modulation of gene expression, splicing, protein interactions, and immune activation. From antisense oligonucleotides to siRNA, these therapies are redefining treatment strategies for rare diseases, cancer, and metabolic disorders. This article explores the eight mechanistic pathways, examines drug delivery challenges, and discusses future opportunities for clinical application.
Introduction to Oligonucleotide Therapeutics
The field of biomedicine is witnessing a revolutionary shift with the emergence of oligonucleotide-based therapeutics. These precision medicines, capable of modulating gene expression, correcting splicing defects, inhibiting pathogenic proteins, and even activating immune responses, are transforming the treatment landscape for rare genetic disorders, cancers, and metabolic diseases. However, their mechanisms of action extend far beyond simple “gene silencing.”
A pivotal review ‘Delivery of oligonucleotide‐based therapeutics: challenges and opportunities’ published in EMBO Molecular Medicine (DOI: 10.15252/emmm.202013243) systematically categorizes oligonucleotide drugs into eight distinct mechanistic pathways, each with unique subcellular targeting strategies. Below, we delve into these mechanisms, illustrated in Figure 2, to provide a comprehensive “battle map” for oligonucleotide therapeutics.
Figure 1. Mechanisms and sites of action of oligonucleotides1,2
Representative mechanisms of action and intracellular localisation for (1) gapmer and mRNA degradation, (2) aptamer, (3) nuclear steric blockage for splice switching, (4) blockage the assembly of RNA‐binding factors, (5) TLR activation of innate immunity, (6) miRNA and antagomir, steric block, translational upregulation, (7) agomir, translational inhibition, and (8) siRNA, RISC, RNAi silencing ONs.
Seven Strategic Pathways of Oligonucleotide Therapeutics
1. Gapmer-Mediated RNase H-Dependent mRNA Degradation
Type: Antisense oligonucleotide (ASO)
- Mechanism: Gapmers (chimeric DNA-RNA molecules) bind target mRNA, recruiting endogenous RNase H to degrade the RNA strand.
- Action Sites: Cytoplasm (mature mRNA) and nucleus (pre-mRNA).
- Key Application: IONIS-HTT Rx (Huntington’s disease).
2. Aptamers: Protein-Binding Nucleic Acid Therapeutics
Type: Aptamer
- Mechanism: Structured single-stranded oligonucleotides bind proteins (e.g., VEGF) with high specificity, mimicking antibodies.
- Action Sites: Extracellular, membrane, or cytoplasmic proteins.
- Key Drug: Pegaptanib (anti-VEGF for macular degeneration).
3. Splice-Switching Oligonucleotides (SSO)
Type: Non-RNase H ASO
- Mechanism: Modulate pre-mRNA splicing by blocking/enhancing splice sites, restoring functional protein expression.
- Action Site: Nucleus.
- Key Drug: Nusinersen (SMA, correcting SMN2 splicing).
4. RNA Structure-Targeting ASOs
Type: Repeat-expansion interceptors
- Mechanism: Disrupt toxic RNA foci (e.g., CUG repeats in DM1) to release sequestered RNA-binding proteins.
- Action Site: Nuclear RNA foci.
- Potential Use: Myotonic dystrophy, ALS.
5. CpG Oligonucleotides: TLR9 Immune Activation
Type: Immunomodulatory oligonucleotide
- Mechanism: Unmethylated CpG motifs activate TLR9 in dendritic cells, boosting cytokine production.
- Action Site: Endosomes.
- Key Use: CpG-1018 (HBV vaccine adjuvant).
6. miRNA Inhibitors (Antagomirs) & Mimics (Agomirs)
- Antagomirs: Block endogenous miRNAs to derepress target mRNAs.
- Agomirs: Mimic miRNAs to suppress mRNA translation/degradation.
- Action Site: Cytoplasm.
- Applications: miR-122 inhibitors (hepatitis), miR-34a mimics (cancer).
7. siRNA: RNA Interference (RNAi)
Type: Small interfering RNA
- Mechanism: Guide strand directs RISC complex to cleave target mRNA.
- Action Site: Cytoplasm.
- Key Drug: Patisiran (hereditary ATTR amyloidosis).
Challenges in Oligonucleotide Drug Delivery
Despite their promise, oligonucleotide therapeutics face delivery hurdles:
- Cellular Uptake: Naked oligonucleotides struggle to cross membranes; lipid nanoparticles (LNPs) and conjugate technologies (e.g., GalNAc-siRNA) are breakthroughs.
- Endosomal Escape: >90% of internalized oligonucleotides remain trapped in endosomes.
- Off-Target Effects: Mismatched binding can trigger unintended gene modulation.
Recent advances, such as antibody-targeted LNPs (e.g., IL-10 mRNA delivery for cerebral hemorrhage), highlight the potential of precision delivery systems.
Conclusion: A Tactical Roadmap for Oligonucleotide Drugs
The diversity of oligonucleotide mechanisms—from mRNA degradation to immune activation—demands tailored design and delivery strategies. Figure 2 serves as an essential guide for researchers developing next-generation therapies.
Table. Oligonucleotide Therapeutics: Mechanisms and Applications
| Class | Example Drug | Mechanism | Site of Action | Application |
| Gapmer | IONIS-HTT Rx | RNase H-mediated degradation | Nucleus/Cytoplasm | Neurodegenerative diseases |
| SSO | Nusinersen | Splicing modulation | Nucleus | SMA |
| siRNA | Patisiran | RNAi cleavage | Cytoplasm | Hereditary amyloidosis |
| Aptamer | Pegaptanib | Protein binding inhibition | Extracellular/membrane | Macular degeneration |
| CpG ON | CpG-1018 | TLR9 immune activation | Endosome | Vaccine adjuvant |
| Antagomir | miR-122 inhibitor | miRNA inhibition | Cytoplasm | Hepatitis |
| Agomir | miR-34a mimic | miRNA mimicry | Cytoplasm | Cancer |
| Structure-ASO | CUG-targeting ASO | RNA foci disruption | Nuclear foci | Myotonic dystrophy (DM1) |
By decoding these “molecular tactics,” we inch closer to curing previously untreatable diseases. The future of oligonucleotide therapeutics is not just about silencing genes—it’s about rewriting medical possibilities.
Oligonucleotide Development Services
If you are exploring oligonucleotide-based drug development, our end-to-end solutions can accelerate your research and clinical translation:
- Custom Oligonucleotide Synthesis (ASO, siRNA, aptamers)
- Advanced Delivery Systems (LNPs, GalNAc, antibody conjugates)
- Preclinical & Clinical Support (stability, biodistribution, safety studies)
Contact us to discuss how we can help bring your oligonucleotide therapy from concept to clinic.
References
- Image retrieved from Figure 2 “Mechanisms and location of action for oligonucleotides,” Hammond, Suzan M., et al., 2021, used under [CC BY 4.0](https://creativecommons.org/licenses/by/4.0/). The original image was modified by extracting and using only part a, and the title was changed to “Mechanisms and sites of action of oligonucleotides”
- Hammond, Suzan M., et al. “Delivery of oligonucleotide‐based therapeutics: challenges and opportunities.” EMBO molecular medicine 13.4 (2021): e13243.