Therapeutics


or near-perfect base pairing with the mRNA target (Figure 1).


miRNA miRNAs are small non-coding RNAs that play key roles in cell differentiation, proliferation and sur- vival. The dysregulation of endogenous miRNAs occurs in multiple diseases including hepatitis, car- diovascular diseases and cancer (where miRNAs act as tumour suppressors or oncogenes). miRNAs are loaded on to the RNA-induced silencing com- plex (RISC) and interact with partially comple- mentary targets on mRNA to suppress protein expression (Figure 1). Antisense RNAs comple- mentary to miRNA can block activity, whereas double- or single-stranded RNAs that mimic miRNA can enhance activity. Both miRNA inhibitors and mimics are currently being devel- oped and have shown encouraging results10. For example, RG-012 (Regulus Therapeutics Inc) is a miRNA drug currently being evaluated in Phase I trials for the treatment of Alport syndrome11.


siRNA In contrast to miRNAs, which attenuate protein production, when an siRNA recognises mRNA it causes cleavage and degradation of the mRNA and completely silences the gene, shutting down pro- tein production (Figure 1). siRNAs arose as a nat- ural defence mechanism against RNA viruses and are double-stranded RNAs acting as prodrugs: the antisense strand is pharmacologically active where- as the sense strand facilitates drug delivery, trans- porting the antisense strand to the intracellular Argonaute (Ago) loading complex. There are four Ago proteins that can be loaded with miRNAs or siRNAs and alter translation and/or RNA stability: siRNAs preferentially bind to Ago2. siRNAs can also compete with miRNAs loaded on to Ago2, thereby altering the half-lives of other cellular RNAs. Exogenous siRNAs operate via a sequence- specific mechanism with perfect complementarity to the target mRNA but can also have miRNA-like effects on some partially complementary mRNA sequences, leading to a lack of specificity. Therefore, a single siRNA sequence can potentially modulate expression of hundreds of off-target genes, which can impact on the efficacy of the RNA drug. Following systemic injection, siRNAs encapsu-


lated in LNPs often tend to accumulate in the liver and spleen12. For systematic delivery, synthetic car- riers are usually decorated with cell-specific ligands or aptamers that facilitate receptor-mediated uptake (Figure 1). Furthermore, biodegradable


Drug Discovery World Fall 2018


nanoparticle carriers allow for slow drug release within the cell to regulate dose. Patisiran (brand name Onpattro; Alnylam Pharmaceuticals Inc) represents the first FDA approval of an RNAi ther- apeutic in an LNP formulation for hereditary transthyretin-mediated amyloidosis (hATTR) in adults (FDA approved in August 2018; Table 1).


RNA aptamers RNA aptamers are short, single-stranded RNAs that are usually selected in vivo to bind to specific molecular targets using SELEX (systematic evolu- tion of ligands by exponential enrichment). RNA aptamers have a propensity to form complementary base pairs, which drives the formation of aptamer- target complexes. Aptamers feature the high affini- ty of antibodies but also offer several distinct advantages: their relatively small size and flexibility allow engagement with binding sites inaccessible to larger antibodies; improved transport and tissue penetration; quick synthesis and comparatively lower manufacturing costs; and high stability and minimal immunogenicity. Many aptamers are inter- nalised upon binding to cell-specific receptors, making them useful drug carriers to deliver small- molecule chemotherapeutics, siRNAs, miRNAs or antisense RNAs into targeted tissues (Figure 1). However, the inherent physiochemical characteris- tics of aptamers, which affect metabolic stability and limit in vivo potency, combined with a lack of available safety data, have hindered their develop- ment. As with other classes of RNA-based thera- peutics, unmodified aptamers are susceptible to nuclease-mediated degradation leading to very short in vivo half-lives (typically less than 10 min- utes). Therefore, most aptamers in clinical develop- ment feature chemical modifications to improve nuclease resistance and pharmacokinetic proper- ties13. For example, Macugen is PEGylated and conjugated to polyethylene glycol (PEG) to extend its half-life in vivo. Aptamers can act as antagonists to block pro-


tein-protein or receptor-ligand interactions; as ago- nists to activate receptors; or as cell-specific deliv- ery systems. All aptamers currently in clinical development are inhibitors that disrupt the func- tion of a target protein. In addition, aptamers can be designed to act as RNA decoys that compete with a natural RNA sequence that represents the target of an RNA-binding protein, sequestering its interaction.


References 1 Crooke, ST, Witztum, JL, Baker, BF. RNA-targeted therapeutics. Cell Metabolism. 2018; 27: 714-739. 2 Burnett, JC and Rossi, JJ. RNA-based Therapeutics- Current Progress and Future Prospects. Chem Biol. 2012;19(1): 60-71. 3 Dowdy, SF. Overcoming cellular barriers for RNA therapeutics. Nature Biotechnology. 2017; 35(3): 222-229. 4 Kulkarni, JA, Cullis, PR and van der Meel R. Lipid Nanoparticles Enabling Gene Therapies: From Concepts to Clinical Utility. Nucleic Acid Ther. 2018;28(3):146-157. 5 Song, J and Yi, C. Chemical Modifications to RNA: A New Layer of Gene Expression Regulation. ACS Chem. Biol. 2017; 12 (2): 316-325. 6 Mauer, J, Luo, X, Blanjoie, A, Jiao, X, Grozhik, AV, Patil, DP, Linder, B, Pickering, BF, Vasseur, JJ, Chen, Q, Gross, SS, Elemento, O, Debart, F, Kiledjian, M and Jaffrey, SR. Reversible methylation of


In December 2004, Macugen


(Pfizer/Valeant Pharmaceuticals International Inc), a VEGF-specific modified RNA aptamer, gained FDA approval for the treatment of age-related macular degeneration (AMD) and several other


m6Am in the 5’ cap controls mRNA stability. Nature. 2017; 541(7637): 371-375. 7 Johannes, L and Lucchino, M. Current Challenges in Delivery and Cytosolic Translocation of Therapeutic RNAs. Nucleic Acid Ther. 2018;28(3):178-193. 8 Juliano, RL. The delivery of therapeutic oligonucleotides. Nucleic Acids Res. 2016; 44(14): 6518-6548. 9 Sahin, U, Karikó, K and Türeci, Ö. mRNA-based therapeutics – developing a new class of drugs. Nat Rev Drug Discov. 2014;13(10): 759-780. 10 Rupaimoole, R and Slack, FJ. MicroRNA therapeutics: towards a new era for the management of cancer and other diseases. Nat Rev Drug Discov. 2017; 16(3):203-222. 11 A Study of RG-012 in Subjects With Alport Syndrome. Available from: https://clinicaltrials.gov/ct2/sho w/NCT03373786.


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