Tuesday, March 25, 2008
Day 1 of Keystone RNAi Conference: New Small RNAs and Oral RNAi Delivery
Following introductions by the scientific organizers Judy Lieberman (Harvard) and Phil Sharp (MIT), Craig Mello, Nobel laureate and faculty at the University of Massachusetts, gave an account of a variety of relatively recently discovered small RNA populations that are enriched in the worm germline and are characterized by specific structural features, such as 5’ triphosphorylations or 3’ methylations, and the Argonaute proteins (worms have over 20 of those, whereas humans only have 4) with which they are associated.
For many of these small RNAs, the biological function is still somewhat unclear and the field, as was also echoed by the second keynote address by Greg Hannon (Cold Spring Harbor) is busy using high-throughput sequencing techniques and bioinformatics to catalogue them. As an aside, with the speed sequencing technology is progressing, it could very well be that the future of microRNA diagnostics will entail the quantitative high-throughput sequencing of microRNAs rather than interrogating a limited number of defined microRNAs by microarray or PCR.
A particularly interesting small RNA population is one that is marked by 5’ triphosphorylation, unlike siRNAs that are 5’ monophosphorylated. It appears that in their biogenesis, a primary small RNA (siRNA or microRNA) cuts a target mRNA which then becomes the template for RNA-dependent RNA polymerases (RdRPs) that generate the 5’ triphosphorylated RNAs (aka secondary siRNAs). Surprisingly, the secondary small RNAs are the ones that carry out the majority of the gene silencing, while the primary small RNAs are present only in very small amounts and appear to function only in triggering the amplification of gene silencing.
It is, of course, now interesting to speculate whether synthetic small RNAs with various 5’ modifications could likewise function in human cells, possibly with different biological activities from siRNAs and microRNAs. However, in the case of the RdRP-dependent 5’ triphosphorylated small RNAs, one has to keep in mind that a dedicated RdRP appears to be absent in the human genome and that 5’ triphosphorylated RNAs in the cytoplasm may trigger unwanted cytokine responses. But maybe there will be some other type of modification around which one could build a therapeutic platform.
At the start of his presentation, Mello noted that one way to induce gene silencing in worms is by feeding them with bacteria expressing double-stranded RNAs (somewhat reminiscent of Cequent Pharmaceutical’s transkingdom RNAi approach). He finally came full circle at the end of his talk, when he showed some intriguing slides from colleagues at the University of Massachusetts, Michael Czech and Gary Ostroff, on the oral delivery of therapeutic RNAi. Starting with yeast ghosts consisting of essentially a shell of beta-glucans that they filled layer by layer with RNAs, including siRNAs, to create nanoparticles that would be taken up by the Peyer’s patches in the gut. Unfortunately this part of the presentation was quite brief, but it appears that they have succeeded in ameliorating inflammation in a mouse model by targeting TNF-alpha. It will now be important to demonstrate the generality of this phenomenon by targeting a number of other genes unrelated to immune responses. As this approach appears to target immune cells, it may at least initially be applicable for orally delivering RNAi for a number of immune related disorders.
This development could be particularly interesting for RXi given the close relationship between Mello and Czech and this new public company. Also taking into account that UMass RNAi efforts will be a major beneficiary of the Massachusetts life science initiative, it seems that RXi is finally showing some signs of scientific life.
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