Use of OLIGONUCLEOTIDES as anti viral drugs

Antibiotics are widely used against a wide spectrum of microorganisms but they are not effective on the viruses because viruses are not living and they become active only through the viral transcript. Therefore a different class of drugs known as antiviral drugs is meant for the treatment of viral diseases. Most of the antiviral drugs are the analogues of nucleosides lacking hydroxyl groups, which get incorporated in the viral genome when the viral DNA replicates itself. The double stranded DNA created after replication is inactive due to the presence of nucleoside analogues and the virus is not able to grow. (Mindel 1983)

Conventional antiviral drugs do not destroy the virus but only inhibits their growth by halting the replication. But there is a significant risk associated with these drugs because the nucleoside analogues may also inhibit the replication of cellular DNA.  (Soundararajan 2009)

Therefore there is a requirement of a drug molecule that acts only on the specific target and selectively inhibits the gene expression. There has been an ongoing research on the usage of oligonucleotide as antiviral in antisense and RNA interference against pathogenic RNA viruses. Oligonucleotide as antiviral therapeutics has proved to be effective in this direction and has shown high target specificity so far.  (Lopez-Fraga 2008)

 It utilizes complementary oligonucleotide sequence that inhibits gene transcription either by catalytic degradation of mRNA or binding to mRNA sites for translation. It is different from the optimal approach in being specific because the optimal approach aims at total inhibition of the viral genome. (Spurgersa 2007)

Oligonucleotides in RNA interference (RNAI)

The double stranded small interfering RNA (siRNA) have been used in the mammalian cells for gene silencing. They are about 21 nucleotides in length and generated by the cleavage of long dsRNA by RNase class III riboendonuclease proteins, also known as Dicer. The Dicer generated siRNAs are incorporated into the RNA-induced silencing complex (RISC). RISC is activated by the action of Argonaute 2 (AGO2). After getting activated, the siRNA cleaves the sense strand of targeted RNA and generates a complementary sequence to the mRNA. The endonuclease present in the RISC complex, cleaves mRNA which is then degraded by intracellular proteins and does not gets translated. The RISC complex is recovered at the end of the process. (Lopez-Fraga 2008)

Another method of gene silencing involves micro RNAs (miRNA) which are produced in the cell as a separate species and posses some complementary sequences. They are about 21 nucleotides long non coding RNAs that bind to the mRNA and translation is inhibited leading to degradation of mRNA. (Lopez-Fraga 2008)

The RNAi based antiviral therapy is analogous to this naturally occurring process. Its aim to silence the targeted viral gene without harming the cellular genes and it can be achieved either by transfecting the cell with synthetic siRNA or short hairpin RNA (shRNA) resembling miRNA. There has been a sufficient progress in achieving the stability, drug resistance, toxicity, and cost of siRNA therapy. There are several other advantages associated with siRNA therapy like, they have short pharmacological development time period, very specific in action, can be easily synthesized and a broad spectrum of pathogens could be targeted. These findings support siRNA being used as an antiviral therapy for the treatment of highly pathogenic viruses. (Lopez-Fraga 2008)

Oligonucleotides in Antisense Technology

The antisense technology is not dependent on the host cell mechanism but it is a simple technique where the antisense oligonucleotide combines to the mRNA and inhibits the translation of viral transcript. The analogous antisense oligonucleotides are synthesized artificially and they consist of nuclease activity. Their binding capacity is much higher than the oligonucleotides naturally occurring in the cell. This avoids the binding of the analogue to cellular gene. The analogue is about 20 to 25 nucleotides in length and with 10 to 100% modified bases for effective and specific binding.  (A New Generation of Antiviral Drugs). They are chemically modified and purified to maintain the stability and keep them safe from the exonuclease activity. (Khan 1993)

One of the example of antisense technology is vaccine against Enterovirus and Rhinovirus which are pathogenic viruses and cause several diseases. Antisense antiviral therapy has proved to be very effective in the treatment as no effective treatment was available earlier. Single stranded DNA like antisense compound called, Peptide-conjugated phosphorodiamidate morpholino oligomers   (PPMO) was used. It can easily enter the cell and is specific for internal ribosome entry site (IRES) sequence which is the target sequence against human rhinovirus. The IRES sequence is highly conserved (about 99%) in the Enterovirus and Rhinovirus so it is a good target for a drug. The results showed that PPMO was highly efficient against broad variety of Enterovirus and Rhinovirus.  (Stone 2008)

There are several advantages associated with antisense antiviral therapy like , they do not depend upon the host machinery for functioning, they are very specific in action, can be used against a wide variety of pathogens and several diseases such as, HIV, Hepatitis C, Influenza etc. The main concern in the application of antisense antiviral therapy is to maintain its stability in the cell and prevent from degradation, method of delivery and maintain their bioavailabity. Therefore they are first chemically modified before transfection. The success of PPMO vaccine against Enterovirus and Rhinovirus supports the fact that antisense antiviral therapy can be uses as potential drug again undruggable pathogenic viruses. (A New Generation of Antiviral Drugs)