Noncoding RNA therapeutics - challenges and potential solutions

Abstract

Therapeutic targeting of noncoding RNAs (ncRNAs), such as microRNAs (miRNAs) and long noncoding RNAs (lncRNAs), represents an attractive approach for the treatment of cancers, as well as many other diseases. Over the past decade, substantial effort has been made towards the clinical application of RNA-based therapeutics, employing mostly antisense oligonucleotides and small interfering RNAs, with several gaining FDA approval. However, trial results have so far been ambivalent, with some studies reporting potent effects whereas others demonstrated limited efficacy or toxicity. Alternative entities such as antimiRNAs are undergoing clinical testing, and lncRNA-based therapeutics are gaining interest. In this Perspective, we discuss key challenges facing ncRNA therapeutics - including issues associated with specificity, delivery and tolerability - and focus on promising emerging approaches that aim to boost their success.

Fig. 1. MicroRNA biogenesis pathway and ways to interfere therapeutically.

MicroRNA (miRNA) biogenesis is a multistep process (see blue boxes) consisting of transcription of a pri-miRNA by RNA polymerase II or III, its nuclear processing into a pre-miRNA by Drosha and DGCR8, nuclear export of the pre-miRNA by exportin 5, cytoplasmic processing by Dicer and TRBP into a mature miRNA duplex and its helicase-mediated unwinding. The passenger strand is degraded, and the mature miRNA strand is integrated into the RNA-induced silencing complex (RISC) to mediate either translational repression or mRNA degradation depending on the extent of complementarity to the mRNA target. Translational repression is mediated through effects on translation initiation, elongation and termination as well as co-translational degradation. mRNA degradation is mediated through mechanisms resulting first in mRNA deadenylation (step 1), followed by de-capping (step 2) and concluded by exonuclease-mediated 5′ to 3′ degradation (step 3). Ways to interfere with the endogenous miRNA pathway (see red boxes) include inhibition of biogenesis at the nuclear or cytoplasmic level, miRNA replacement therapy and functional inhibition of the mature miRNA or the interaction with its target mRNA. As oligonucleotides are not readily taken up into cells, commonly used delivery methods are shown and include conjugation to various chemical or biological entities as well as delivery within lipid particles, polymers and viral or bacterial vector systems.

Fig. 2. Modes of action of small-molecule inhibitors that target miRNAs and lncRNAs.

a | Small-molecule inhibitors of miRNA (SMIRs) may act at the transcriptional level or may affect the nuclear or cytoplasmic maturation steps of the microRNA (miRNA). The exact mechanism by which azobenzene-2, miR-122 inhibitor 2 and aza-flavanones inhibit the transcription of specific miRNA host genes to primary RNA transcripts (pri-miRNAs) is unknown. Targaprimir-96 binds to the internal loop of pri-miR-96 to prevent its processing by Drosha. Multiple small molecules interfere with Dicer processing, including targapremir-210, which binds to the Dicer cleavage site, and targapremir-18a, which binds to a 1 nt bulge present in three of the six miRNAs of the miR-17–92 cluster. BzDANP similarly binds to a C bulge present in miR-29a and miR-136, causing complex formation and slowing of Dicer processing. Linifanib inhibits the processing of pre-miR-10 via an unknown mechanism. Proximity-enabled Dicer inactivation makes use of two small molecules, a miRNA binder and a weak Dicer inhibitor that is active upon proximation. Use of a photocleavable linker can grant specific Dicer inactivation that can be terminated using light. b | The first small molecules applied to modify long noncoding RNA (lncRNA) expression levels can be classified as interaction element blockers (IEBs) and structural element lockers (SELs). NP-C86 works as an IEB for GAS5, blocking its interaction with UPF1, which normally results in nonsense-mediated decay of GAS5, thus increasing the stability and half-life of GAS5. Multiple SELs are being developed for MALAT1, which carries a stabilizing triple helix structure at its 3′ end. The SELs are aimed at disrupting the stabilizing triple helix, consequently resulting in MALAT1 destabilization and downregulation.