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RNA interference as a potential control of peste des petits ruminants

Final Activity Report Summary - RNA CUREPPR (RNA interference as a potential control of peste des petits ruminants)

Peste des petits ruminants (PPR) and Rinderpest (RP) are highly contagious viral diseases of domestic and wild ruminants inducing high morbidity and mortality. In regions with a large animal production sector, the viruses that cause these diseases have significant repercussions on the local economy and food security. PPR is endemic in Africa, in the Middle East and South-West Asia. Despite large efforts for eradication of RP, this disease still remains endemic in some areas of the Somalian ecosystem. Both PPRV and RPV are members of the Morbillivirus genus in the Paramyxoviridae family, a genus which also includes Measles virus (MMV).

Owing to their animal health and economic relevance, there is a major interest in the efficient control of PPR and RP. Although efficient vaccines are available for both diseases, they can only protect the animals after 14 days post-vaccination. Therefore, there is an interest to develop a curative tool that could help to control the virus replication when animals are newly infected or are at high risk to become infected in the next days. We have developed a new control method against these viruses. The strategy is based on the RNA interference (RNAi), a process which has been the winning research of the Nobel Prize in Physiology or Medicine in 2006. Cells can inhibit the expression of intrinsic or foreign genes at the mRNA level by RNA interference. RNAi is activated when RNA molecules occur as double-stranded pairs in the cell. Double-stranded RNA (dsRNA) activates a biochemical machinery into the cell which degrades those mRNA molecules with a genetic code complementary to that of the dsRNA. When such mRNA molecules disappear, no protein is made and the corresponding gene is silenced. RNA interference occurs in plants, animals, and humans and it is already being used as a method to study the function of genes and as a defence strategy against diseases. RNA interference can be induced by synthetic short double-stranded RNA molecules containing around 21-25 nucleotides (short interfering RNAs - siRNAs). Such silencing RNA molecules can be introduced into the cell and activate the RNA interference machinery to silence the complementary mRNA sequence.

During this Marie Curie action, we have identified siRNA sequences capable of inhibiting more than 90 % of the replication of PPRV and RPV in infected cell cultures. They are targeting the messenger RNA of the nucleoprotein gene of these viruses, then blocking the virus multiplication process. An application for a French patent entitled 'Interfering RNA targeting the nucleoprotein gene of morbilliviruses' (INPI N°05 13029) was deposited in December 2005 and an extension for an international patent was made one year after (PCT / FR2006 / 002819).

Furthermore, to facilitate the transfer of the functional interfering RNA sequences to infected animals, we have constructed a recombinant DNA viral vector commonly used as a vaccine. With this adenovirus vector, an active siRNA sequences (N1-PPRV) was transcribed into the infected cells as a short hairpin RNA precursor (shRNA, with approximately 50 nt). After transcription, the shRNA is processed into active 21-nucleotides RNAs by a cellular enzyme called Dicer, and it recognises and destroys the viral mRNA target. The silencing capacity of this recombinant viral vector, called Ad-shRNAN1, was firstly verified in vitro, i.e. in PPRV infected cell cultures. We have demonstrated the ability of this modified vector to deliver efficiently siRNA and to decrease more than 100 times the amount of PPRV produced into the cells.

Our results suggest the potential use of the Ad-shRNAN1 for the delivery of siRNA in PPRV-infected animals, its use as a therapeutic tool against this disease and to open up vast prospects in terms of animal health.