title: Engineering Periodic shRNA for Enhanced Silencing Efficacy.
abstract: RNA interference (RNAi) provides a versatile therapeutic approach via silencing of specific genes, particularly undruggable targets in cancer and other diseases. However, challenges in the delivery of small interfering RNA (siRNA) have hampered clinical translation. Polymeric or periodic short hairpin RNAs (p-shRNAs)-synthesized by enzymatic amplification of circular DNA-are a recent development that can potentially address these delivery barriers by showing improved stability and complexation to enable nanoparticle packaging. Here, we modify these biomacromolecules via structural and sequence engineering coupled with selective enzymatic digestion to generate an open-ended p-shRNA (op-shRNA) that is cleaved over ten times more efficiently to yield siRNA. The op-shRNA induces considerably greater gene silencing than p-shRNA in multiple cancer cell lines up to 9 days. Furthermore, its high valency and flexibility dramatically improve complexation with a low molecular weight polycation compared to monomeric siRNA. Thus, op-shRNA provides an RNAi platform that can potentially be packaged and efficiently delivered to disease sites with higher therapeutic efficacy.
authors.affiliation: Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA; Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
authors.forename: Connie
authors.initials: C
authors.lastname: Wu
authors.affiliation: Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA; Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
authors.forename: Kevin E
authors.initials: KE
authors.lastname: Shopsowitz
authors.affiliation: Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA; Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA; Institute for Soldier Nanotechnologies, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA. Electronic address: hammond@mit.edu.
authors.forename: Paula T
authors.initials: PT
authors.lastname: Hammond
mesh_terms: D045744:Cell Line, Tumor; D004270:DNA, Circular / Q000378:metabolism*; D020868:Gene Silencing; D005818:Genetic Engineering / Q000379:methods; D006801:Humans; D009690:Nucleic Acid Conformation; D011073:Polyamines / Q000737:chemistry; D000071228:Polyelectrolytes; D034741:RNA, Small Interfering / Q000096:biosynthesis* / Q000737:chemistry; D043244:Ribonuclease III / Q000235:genetics*; D014161:Transduction, Genetic
chemical_list: D004270:DNA, Circular; D011073:Polyamines; D000071228:Polyelectrolytes; D034741:RNA, Small Interfering; D043244:Ribonuclease III