Seminar: Development of single stranded RNaseH recruiting antisense oligonucleotides for analysis of lncRNA function in cell culture and animals
2014-10-03: by Niels Frandsen, Exiqon. The seminar will take place October 3rd, 12.30-1.15 at University of Copenhagen, SUND/SCIENCE, Dyrlægevej 16, Orangeriet, Frederiksberg C.
Registration is necessary, as refreshments and sandwiches will be served to all registered participants.
Registration deadline: 1st Oct at 12.00 by emailing: email@example.com with subject line "NF- seminar" and your name in the text.
One of the most interesting results from the ENCODE NGS project is the observation that a major part of the genome is transcribed and that the vast majority of the transcriptome is non-coding. Estimates suggest that there are >10.000 lncRNAs, very few of which have been characterized in any detail. The few lncRNAs that have been studied suggest that they have important and highly diverse functions involved in epigenetics and regulation of both transcription and translation. There is therefore an urgent need for the development of tools for functional analysis of lncRNAs. For the past decade we have been accustomed to exploiting RNAi for loss of function studies of protein. However many lncRNA are challenging siRNA targets. Some derive from overlapping antisense transcription which therefore requires absolutely strand specific activity of the double stranded siRNA. And some are either retained or have long residence time in the nucleus which make them poor siRNA targets especially in slowly or non-dividing cells.
To overcome these challenges Exiqon has developed single strand LNA enhanced antisense technology that catalyzes RNaseH dependent degradation of target RNA in the nucleus. This enables efficient and specific KD of lncRNA not only in cell cultures but also in animals. Single strand RNaseH recruiting antisense oligonucleotides (ASO) were first reported 26 years ago, but has apart from pharmaceutical applications, largely been ignored by the life science community, since the discovery of siRNA. This was in part due to the fact that screening of many ASOs was nescessary for identification of one that gave potent KD. We have developed an empirically derived design algorithm that provides ASOs that achieve potent target KD with a high hit-rate. The algorithm factors >30 design parameters of which target accessibility (secondary structure) and specificity are among the most important. The ASOs are typically 16mer in length and potent KD can be achieved in cell cultures by lipid based transfection as well as by simply adding the oligonucleotide to the culture medium (unassisted delivery). By systemic delivery of an ASO targeting ubiquitously expressed lncRNA Malat1 we show that we can achieve effective and long lasting KD in a broad range of tissues in mice. In conclusion LNA enhanced gapmers are important and powerful tools for the analysis of lncRNA function both in both cell cultures as well as animal model.