The Research Of Novel Antisense Drugs

Oligonucleotide therapeutics represents a new paradigm for drug discovery. The paradigm has resulted in substantial enthusiasm because oligonucleotides may display dramatic increases in affinity and selectivity for their nucleic acid targets compared to traditional drugs. The widespread occurrence of diseases caused primarily through retrovirus invasion of a host, such as HIV, has encouraged the development of an ever increasing variety of potential drugs based on both the antisense and antigene strategy. Furthermore, antisense technology may facilitate rational drug design. Oligonucleotides are designed to modulate the information transfer from the gene to protein ?in essence, to alter the intermediary metabolism of RNA. To date, oligonucleotides have been found to inhibit the growth of a large number of viruses in tissur culture, the expression of numerous oncogenes, a variety of normal cellular genes, and a number of transfencted reporter genes controlled by several regulatory elements. Potent antiviral and antitumor activities have been demonstrated with oligonucleotides. There are several obstacles that must be surmounted in order to improve the in vivo efficacy of oligodeoxynucleotide (ODN) analogs. Unmodified ODNs are rapidly degraded by intracellular nucleases and are unable to efficiently passively diffuse through cell membranes. As a first梘eneration antisense oligonucleotides, phosphorthioate modified oligonucleotides are stable to degradation by nucleases, but in general hybridize to target sequences with a lesser affinity than a phosphodiester ODN. The ODNs containing this modification are a mixture of 2~ diastereomers (where n is the number of linkages), and it is possible that an ODN containing all RP or all SP isomers would hybridize with better affinity. Modification of the ODN has been shown to impart stability and may allow for enhanced affinity and increased cellular permeation of ODNs, and for this reason to synthesize more efficient antisense drugs. 4 ABSTRACT 1. The study of oligodeoxynucleotides containing methyleneformacetal The major challenge of ODN analogs is to design backbone modifications which will increase the nuclease stability and cellular permeability while enhancing affinity. ODNs partially substituted phosphodiester backbone with neutral methyleneformacetal are designed to increase the stability against cellular nucleases without disturbing hybridizing affinity of ODNs. Starting from thymidine and 2?deoxycytidine, 5?hydroxyl group of deoxynucleosides is protected selectively by benzoyl chloride, also, 3?terminus is used diphenylphosphinic acid as leaving group. In the presence of trimethylsilyl trifluomethanesulfonate (TMSOTf), condensation of a nucleoside phosphinate, 3?O-CH2-OP(O)Ph2, with 5?unprotected deoxynucleoside acceptor affords in most cases the (3挆+S?methylene acetal linked dimers and trimers. These acetal-linked oligomers, of which 5?hydroxyl group is protected by 4,4?dimethoxytrityl chloride and 3?hydroxyl group is reacted with (2-cyanoethyl N,N-diisopropyl) chlorophosphoramidite, are incorporated into oligonucleotides by using the standed solid梡hase DNA chemistry on controlled pore glass (CPG) support with the phosphoramidite method. The melting temperatures (Tm) of modified oligodeoxynucleotides with their DNA complements are determined.