Small-interfering RNA (siRNA)-based therapeutics work by suppressing disease-causing genes, and they hold tremendous promise for targeting a large part of currently undruggable diseases, ranging from viral infections to hereditary disorders and cancers.1 A Nobel prize was awarded to the discovery of RNA interference in 2006. However, the majority of the siRNA-based drug candidates having entered clinical trials target diseases of/originating from the liver, with only one lipid particle-formulated siRNA, patisiran, being approved by the U.S. Food and Drug Administration for the treatment of polyneuropathy caused by hereditary transthyretin amyloidosis.2
A key obstacle of siRNA-based therapeutics is the delivery to the non-liver organs/tissues and across the plasma membrane of cells in vivo. Naked siRNA does not readily enter unperturbed cells even at high concentrations. Additionally, they can be rapidly filtered from the body by the kidneys and are subject to digestion by serum and cellular nucleases, resulting in a half-life too short for clinical activity. Chemically modified siRNA, which addresses the problems of degradation, primarily accumulates in the liver and may induce liver/cardiovascular toxicity. A variety of cationic materials have been developed for siRNA-delivery through electrostatic interaction with negatively charged siRNA. However, these materials often create additional problems, such as intrinsic toxicity and immunogenicity, which limit their clinical translation. Therefore, to realize the full potential of siRNA drugs, a safe, simple, and efficient vector system is still very much sought after.
Here, researchers from Northeastern University and MIT address these challenges by using a non-charged, brush polymer-siRNA conjugate, termed polymer-assisted-compaction of RNA (pacRNA). The brush polymer-RNA conjugate consists of approximately two siRNA strands covalently attached to the backbone of a sterically-congested brush polymer with the biocompatible polyethylene glycol (PEG) side chains via a bio-reductive linker. The spatial congestion of the bottlebrush polymer resulting from the densely spaced PEG side chains provides steric shielding over the siRNA and prevents proteins from accessing, thus circumventing many of the side effects associated with siRNA-protein interactions, such as coagulopathy and inflammation.
PacRNA-enhanced cellular uptake of siRNA and augmented stability against nuclease, which makes it survive the digestive endosomal environment and successfully deliver a fraction of the intact conjugate to the cytosol, where released siRNA can function in RNA interference. The authors have shown successful regulation of target genes in different cancer cell lines. In addition, pacRNA also has substantially increased blood circulation times after systemic administration due to the large size (~30 nm) and diminished interaction with serum proteins. Over 50% of pacRNAs remain in the bloodstream an hour after injection, while only less than 1% of unmodified siRNA remains in the circulation. The increased circulation time results in passive targeting of pacRNA to the tumor through enhanced permeation and retention effect. They have shown that pacRNA accumulate in the tumor ~20 times more than free siRNA. Collectively, these improvements result in outstanding efficacy on knocking down antiapoptotic B cell lymphoma 2 (Bcl-2) gene and suppress tumor growth in a mouse model.
Importantly, all observed enhancements of the pacRNA over naked siRNA are realized through the architecture change of PEG, which should promote the overall safety of the vector. Indeed, this common delivery material shows minimum toxicity and immunogenicity in vivo and has been used in a variety of biopharmaceutical formulations, as well as in food, cosmetics, and industrial products. Being a molecular entity with a well-defined structure as opposed to a supramolecular assembly of various components is another advantage for the pacRNA over heterogeneous vectors in terms of large-scale manufacturing and batch-to-batch consistency.
This study represents a significant advance on non-cationic delivery vehicles for siRNA. In addition, the brush polymer platform created in this study can serve as the basis for delivery of other kinds of oligonucleotides, such as antisense, microRNA, etc., to treat theoretically any genetically targetable diseases. These data also imply a different design strategy for drug delivery platforms by considering the immediate local environment of the therapeutics.
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- X. Shen, D. R. Corey, Chemistry, mechanism and clinical status of antisense oligonucleotides and duplex RNAs. Nucleic Acids Res. 46, 1584–1600 (2018).
- M. S. Shim, Y. J. Kwon, Efficient and targeted delivery of siRNA in vivo. FEBS J. 277, 4814–4827 (2010).