**4. Antisense oligonucleotides**

Antisense oligonucleotide, first recognized in 1978 by Zamecnik and Stevenson, is a small synthetic piece of DNA (usually 15–18 mer in length) that can bind complementary RNA by Watson-Crick base pairing. ASOs can target most RNA transcripts and have emerged as the ideal therapeutic agents for a broad number of diseases [52, 53]. Upon binding to their target, ASOs can modulate the intermediary metabolism of RNA by the recruitment of endogenous RNase H1 to interfere with RNA function [54]. Human RNase H1 is a ubiquitous enzyme that hydrolyzes the duplex formed between a DNA containing ssASO and target RNA through its N-terminus RNA-binding domain. In order to cleave the RNA in the duplex, the RNase H1 catalytic domain needs at least 5 consecutive DNA/RNA base pairs, and cleavage usually occurs within 7–10 nucleotides from the 5'-end of the RNA. After cleavage, the exposed phosphate on the 5'-end and hydroxyl on the 3'-end are recognized, and the RNA is subse‐ quently degraded by cellular nucleases. At some point after RNase H1 cleaves the RNA, the ssASO is released and is available to reengage another transcript.

Even though much progress has been made in the ASO field so far, there are still many questions that might result in nonspecific effects. One of the principle challenges for success is efficacious delivery to target organs. Because initial ASO molecules are either of low affinity or low membrane permeability, they suffered from poor solubility and rapid degradation by nucleases. In the field, many studies to improve the therapeutic potential of ASOs have focused on chemical modifications to either improve nuclease resistance, such as 2'-O-methoxyethyl (2'-MOE), or to facilitate cellular uptake, like phosphorothioate backbone that improves membrane penetration [55, 56]. Moreover, too many heparin-binding cell surface proteins have been identified to bind the phosphorothioate oligo with nanomolar affinity. The delivery of ASO drug, encapsulating with materials ranging from cationic lipids to dendrimers to alginate/ chitosan nanoparticles, has reached new heights of clinical acceptance [52].

Over the past several years, antisense oligonucleotide-based targeted therapy has emerged rapidly. Interest in the field has ramped-up dramatically, as numerous ongoing clinical trials are evaluating the treatment effect on diseases with ASOs. Antisense oligonucleotide sodium LY2181308 (LY2181308), hybridizing to the human survivin mRNA, is well tolerated in patients with acute myeloid leukemia (AML). In combination with chemotherapy, LY2181308 does not cause additional toxicity, though 1/16 patients had incomplete responses, and 4/16 patients had cytoreduction [57]. Thus, future clinical trials are needed to further confirm its clinical benefit. In another open-label, parallel-group study, reducing factor XI levels by a second-generation antisense oligonucleotide FXI-ASO (ISIS 416858) is an effective method for prevention of postoperative venous thromboembolism. With respect to the risk of bleeding, FXI-ASO received once daily appeared to be safe [58]. In another phase II trial, compared with those who received placebo, the participants with Crohn's disease who received SMAD7 ASO Mongersen (formerly GED0301) had significantly higher rates of remission and clinical response [59]. Even more important, mipomersen, an antisense agent targeted to apolipopro‐ tein B, has recently received FDA (United States Food and Drug Administration) approval for the treatment of familial hypercholesterolemia (http://www.fda.gov/newsevents/newsroom/ pressannouncements/ucm337195.htm). This compelling therapeutic potential powerfully supports further clinical investigations of ASOs in subjects in the near future.
