For example, in a series of publications, Huang co-delivery of siRNA and anticancer drugs (194,197,198)

For example, in a series of publications, Huang co-delivery of siRNA and anticancer drugs (194,197,198). Another common approach involves the direct complexation of anionic oligonucleotides with cationic molecules that have some degree of endosome escape capability inherent in their chemistry. advent of antisense and siRNA oligonucleotides sparked high hopes for their eventual use in treatment of disease. However, these early expectations remained largely unfulfilled as first generation oligonucleotides failed to meet therapeutic end points in a number of clinical trials. After a period of disappointment, the field of oligonucleotide therapeutics has now been re-invigorated (1). This is due to the convergence of several developments including improved chemistries, better understanding of the basic biology of oligonucleotides, more sophisticated delivery systems and most importantly, increasing success in the clinic. The 2013 approval of the first major antisense drug, Kynamro? (2), an inhibitor of apolipoprotein B expression, was accompanied by promising clinical trials involving siRNA (3) and splice switching oligonucleotides (SSOs) (4). More recently, a number of clinical trials utilizing various types of oligonucleotides have reported impressive results. Some examples might include a use of a receptor-targeted siRNA conjugate (5), strong effects on liver diseases using antisense with novel chemical modifications (6,7), anti-cancer effects with a miRNA (8) and treatment of a neurodegenerative disease via intrathecal administration of a SSO (9). More detailed summaries of selected current clinical studies are provided in several recent reviews (10C13). Droxidopa Despite these advances at the clinical level, effective delivery of oligonucleotides Ctnnb1 remains a major challenge, especially at extra-hepatic sites (13C15). Various strategies are being pursued including chemical modification of the oligonucleotide itself, use of various lipid or polymeric nanocarriers, linking oligonucleotides to receptor targeting agents such as carbohydrates, Droxidopa peptides or aptamers, and use of small molecules to enhance oligonucleotide effectiveness. The intent of the current article is to provide a broad but analytic review of the oligonucleotide delivery area. The emphasis will be on basic biological aspects rather than recent clinical developments. There are an enormous number of publications in this area, far too many to be cited in their entirety. Thus the focus in this review will be on reports that stand out because of their novelty, or that provide important mechanistic information, or that display significant translational potential. This article will also convey the author’s personal view on the future evolution of the oligonucleotide delivery area. BASIC INFORMATION UNDERLYING OLIGONUCLEOTIDE THERAPEUTICS The scope of the oligonucleotide therapeutics field has expanded substantially over the last few years as additional types of nucleic acids are used and as new targets are addressed. One of the most exciting developments is the realization that thousands of non-coding RNAs play important roles in cellular function (16) and that these entities can be readily manipulated using oligonucleotides (17). A continuing thrust in the field is the pursuit of clinical problems that are Droxidopa not Droxidopa easily addressed with small molecule drugs. Thus there has been emphasis on relatively rare disorders for which no current therapy exists. The various therapeutic approaches currently under investigation involve several types of nucleic acids with different chemistries and mechanisms of action; therefore it seems worthwhile to briefly review some basic aspects of oligonucleotide biology and chemistry. Basic mechanisms of oligonucleotide actions Classic single stranded antisense oligonucleotides (ASOs) primarily act in the nucleus by selectively cleaving pre-mRNAs having complementary sites via an RNase H dependent mechanism (18). Although ASOs can also act by translation arrest, they are currently primarily used as gapmers, having a central region that supports RNase H activity flanked by chemically revised ends that increase Droxidopa affinity and reduce susceptibility to nucleases (19). SSOs are a form of ASO; however they are fully modified so as to ablate RNase H activity and allow connection with nuclear pre-mRNA during the splicing process. SSOs can be designed to bind to 5 or 3 splice junctions or to exonic splicing enhancer or silencer sites. In doing so they can improve splicing in various ways such as promoting alternative use of exons, exon exclusion or exon inclusion (20). SSOs are very flexible tools and are seeing increasing use in therapeutic methods (21). RNA interference (RNAi) is a fundamental endogenous mechanism for control of gene manifestation (22). It can involve selective message degradation, translation arrest or modulation of transcription (23). Both endogenous miRNAs and chemically synthesized externally given siRNAs use Argonaute-containing RISC complexes to regulate gene manifestation (24,25). With siRNA, selective cleavage of mRNA in the cytosol entails Argonaute 2-comprising complexes and requires essentially total complementarity between the siRNA lead strand and the prospective, usually within the coding region of the subject matter. Because of their selectivity, siRNAs.