MicroRNAs (miRNAs) are master regulators involved in multiple physiological and pathological processes. These naturally occurring, noncoding, single-stranded RNAs (21–25 nucleotides long) base-pair with their target mRNA within the RISC (RNA-induced silencing complex). The latter is the same ribonucleoprotein machinery associated with siRNA-mediated gene silencing.
While siRNAs are double-stranded oligonucleotides that perfectly pair to degrade their target mRNA, miRNAs can pair perfectly or imperfectly. Perfect base pairing leads to the degradation of the mRNA (similar to siRNA), while imperfect complementarity inhibits translation. It’s been suggested that miRNAs regulate up to 50% of all mRNAs in the human genome.
Discoveries linking miRNAs to a number of diseases have helped propel the fast-paced growth of this young field. Release of the first miRNA-based diagnostic test in 2008 helped reduce its failure risk. Keeping pace with new applications and tackling current issues was a focus of Keystone’s “MicroRNAs and Human Disease” conference held last month. Researchers described technological advances as well as novel miRNA therapeutics such as for cardiovascular disease, cancer, and muscular dystrophy.
Some of the same strategies already developed for delivery of siRNA for RNA interference (RNAi) also are being applied to miRNA. “Both miRNAs and siRNAs need to be delivered into the target tissue or cell in order to activate the desired therapeutic effect,” Muthiah Manoharan, Ph.D., senior vp, drug discovery, Alnylam Pharmaceuticals, explained.
“Chemical modifications to provide drug-like properties to RNA molecules are used in the synthesis of both siRNAs and antimicroRNAs (antimiRs). In addition, the lipid nanoparticles (LNP) platform, which has proven to be effective with siRNAs, is also showing promise with miRNAs and miRNA mimics.”
According to Dr. Manoharan, Alnylam is utilizing second-generation LNPs to enhance delivery of siRNAs in several models. “We demonstrated an approximate 100-fold improvement in potency over first-generation formulations, and achieved an effective dose (ED50) at single-digit microgram per kilogram dose levels. Additionally, we’ve made progress in hepatocyte-targeted in vivo gene silencing by modifying the siRNAs.
“We conjugated an N-acetylgalactosamine moiety (GalNac) to siRNAs and achieved improved potency using subcutaneous delivery at low, clinically relevant doses. In the case of a GalNAc-conjugated siRNA targeting transthyretin (TTR), target gene silencing was achieved with an ED50 of approximately 5 mg/kg with a single subcutaneous injection. These results represent a greater than 30-fold improvement in target gene silencing with siRNA conjugates as compared with first-generation siRNA conjugates previously described.”