Wednesday, August 8, 2007

Back to Basics: Does RNAi Exist in Humans?

Whenever I try to explain the promise of RNAi for human biology and medicine to friends and family, I feel torn by a conflict of staying scientifically accurate or running the risk of losing my audience by burdening them with too many details. You may ask “why” now- isn’t RNAi this elegant straightforward mechanism in human cells whereby double-stranded siRNAs silence complementary mRNAs? How would you react when I said that a number, if not the majority of notable molecular biologists hold that RNAi does not exist in humans?

RNAi is most easily explained by starting with long double-stranded RNA (dsRNA) that is then chopped up by Dicer into short interfering RNAs which then get incorporated into the RiSC complex. The siRNAs loaded in RiSC consequently seek out and destroy their targets by a slicing mechanism. However, long double-stranded RNAs are not suitable for specific gene suppression in humans due to an innate immune system called the interferon response. The interferon response recognises dsRNA greater than 30bp in length and consequently shuts down almost all protein translation in a cell. This is a mechanism by which the cell protects itself from viruses which often produce dsRNA intermediates as part of their life-cycle.

I therefore vividly remember myself in an elective class (“RNA World”) as an undergrad student at Edinburgh University, when my lecturer, Dr. David Tollervey, predicted that it would only be a matter of time that RNAi would be discovered in humans- 1 year before Tuschl and colleagues published their seminal paper on siRNAs. In retrospect, it is clear that his confidence must have come from the fact that the human genome was littered with RNAi-related genes, such as some coding for Dicer and Argonaute proteins. He also may have been familiar with recent publications on the discovery of RNAi-related small RNAs in plants (Hamilton and Baulcombe) and the finding that double-stranded RNAs were processed into similarly sized small RNAs in an Drosophila in vitro system (Zamore, Tuschl, Sharp, Bartel).

Of course, we all know now that it was Tuschl who had the genius to conclude that siRNAs would be the natural effectors of RNAi in humans without inducing the interferon response. Except that they were not. Of course, synthetic siRNAs would work, but years of searching for bona fide naturally occurring siRNAs in humans derived from long dsRNA failed to convincingly prove their existence. It turns out that the RNAi enzymes are all present in vertebrates for another, albeit related reason, namely microRNA-mediated gene regulation. MicroRNAs are a major class of small RNAs that are not derived from long dsRNA, but hairpin precursors and that have turned out to be ubiquitous mediators of post-transcriptional gene regulation and that require RNAi-related proteins for their biogenesis and function. It is in this pathway that experimentally introduced siRNAs perform their gene silencing function.

It therefore appears that in humans the interferon system may have rendered antiviral roles for RNAi unnecessary. This is unlike in many invertebrates which lack an interferon system and where the ancient antiviral function is still alive and kicking. A fascinating story of evolving overlapping biological pathways with a happy ending for keeping the therapeutic promise of RNAi alive, but when you ask a scientist about RNAi in humans- beware!

Market Watch: Nastech reported “positive non-clinical study results for [their] siRNA program for treatment of influenza [in tissue culture in vitro studies].” Following an announcement this Monday of a webinar on flu RNAi to be hosted by Nastech this Friday, frantic trading led to big gains in Nastech stock in anticipation of major RNAi news. Nastech and Alnylam vie for a potentially lucrative government contract for flu preparedness and progress in this area will be closely watched as flu RNAi may turn out to be the first commercial RNAi Therapeutics product. It is likely that NSTK will give up some of its gain in the wake of today’s news, while ISIS may gather momentum as their RNA modification IP estate is starting to yield a rich harvest. Alnylam is to follow with their report tomorrow.

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By Dirk Haussecker. All rights reserved.

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