Edward G. Saravolac

Catalytic DNA: A Novel Tool for Gene Suppression

RNA, as an intermediate in the production of every gene encoded protein and the genetic material of many pathogenic viruses, presents an attractive target for both biological and therapeutic manipulation. Despite its extensive involvement in living systems, its chemical diversity based on four units is relatively low compared with protein. This provides the opportunity for a generic approach to targeting with specificity based on primary structure rather than complex higher order structures. This form of recognition occurs naturally in complementary nucleic acids, due to an ability to bind their single stranded target through Watson-Crick interactions. The most established nucleic acid based approach to gene suppression at the RNA level is through antisense oligodeoxynucleotides (ODNs). These compounds form heteroduplex with target RNA which are thought to either block its function or mediate its destruction by activation of RNase H. Alternatively, RNA can be targeted by catalytic RNA such as the hammerhead ribozyme. Ribozymes have the advantage of being equipped with their own RNA cleavage apparatus and are therefore independent of host nuclear protein activity. At present, the utility of ribozyme oligonucleotides is restricted by the relative difficulty synthesising active molecules with sufficient resistance to nuclease degradation. Recently the power of in vitro selection has been used to evolve catalytic DNA sequences with RNA cleavage specificity and activity rivalling the very best ribozymes, while maintaining the more robust chemistry of an ODN. These deoxyribozymes or DNAzymes have tremendous potential as gene suppression agents for both target validation and therapeutic applications. A number of studies evaluating the biological activity of these compounds have shown promising results. However, as with other oligonucleotide based strategies, future exploitation of this approach may depend on accessory technology to assist with the accessibility of a target which is folded by its own secondary structure and hidden within the intracellular compartment.

Immunoprophylactic strategies against respiratory influenza virus infection

The objective of this report is to evaluate the prophylactic efficacy of liposome-mediated immunotherapy for prevention of respiratory influenza virus infection in mice. Antiviral antibody, interferon-gamma and poly (ICLC) were encapsulated in liposomes and they were evaluated for their ability to induce protective immunity against lethal influenza infection. Passive immunization using liposome-encapsulated antiviral antibody was found to offer complete protection against the virus challenge. However, this pretreatment must be administered within 24 h prior to virus challenge to be protective. Pretreatment with liposome-encapsulated interferon-gamma was found to stimulate cellular immune responses, but the protection is partial. Immunoprophylaxis using liposome-encapsulated double-stranded (ds) RNA poly (ICLC) provided complete and longer-lasting protection against influenza infection. These results suggest liposome-mediated immunoprophylactic approaches are effective in the prevention of respiratory influenza virus infection.

Recent Patents on development of nucleic acid-based antiviral drugs against seasonal and pandemic influenza virus infections.

Influenza viruses are etiological agents of deadly flu that continue to pose global health threats, and have caused global pandemics that killed millions of people worldwide. The global crisis involving the avian H5N1 influenza provides compelling reasons to accelerate fast track development of novel antiviral drugs against the potential pandemic virus. The availability of neuraminidase inhibitors such as oseltamivir (tamiflu) improves our ability to defend against influenza viruses, but the incidences of tamiflu-resistance are rising rapidly. Nucleic acid-based antiviral drugs are promising classes of experimental antiviral drugs that have been shown in pre-clinical studies to be effective against seasonal and avian influenza viruses. The potency and versatility of these drugs make them potential candidates to be used in seasonal and pandemic influenza scenarios. The review will assess the recent patents, research and development of antisense oligonucleotides, small interfering RNA, immunomodulating RNA for the prevention and treatment of influenza infection.

Recent patents in antiviral siRNAs

The advent of gene silencing siRNA technology has created opportunities to develop therapeutics based on targeting the genomics of the disease state. Amongst the first applications of siRNA technology, antiviral applications have been quickly and extensively exploited allowing emergence of a range of antiviral therapeutic strategies. Patent activity has encompassed a range of the components required to utilize this technology ranging from the identification of susceptible genomic targets through to the development of vector systems to express the siRNA endogenously or the synthesis of stable RNA oligonucleotides for in vivo therapeutics. Indeed the primary focus of research effort in this area has been to overcome the challenge common to all of gene therapeutics - delivery of the oligonucleotide - to the diseased tissues and organs, sites of infection and/or sites of drug action. Here we survey the development of siRNA therapeutics both in terms of the range of virus species targeted and the strategic approaches employed. Our study illustrates features commonly observed in the field of nucleic acid drug development. While in vitro studies provide a broad range of molecules and molecular targets for potential therapeutics, the field is however severely limited in terms of safe, effective means to deliver the potential siRNA therapeutics in vivo, to the intracellular site of action.