HIV is an extraordinarily smart virus. Just as researchers track down another potential vehicle for halting its spread, it throws up another roadblock. Research on vaccines has been ongoing for 20 years, with neutralising antibodies remaining the best hope after the recent failure of Merck’s Phase III trial of an adeno-vectored T-cell vaccine, but at the moment combination antiretroviral therapy is our only weapon.
As HIV has a relatively limited number of its own proteins, researchers have obviously zeroed in on these to try to inhibit viral replication. The virus’s ability to mutate has sidestepped many of these attempts, however, so some are looking at inducing the cellular mechanism known as RNA interference to halt the virus in its tracks by silencing gene expression. Even then, however, HIV might have a solution.
Associate Professor Damian Purcell, who heads the Molecular Virology Laboratory at the University of Melbourne, has been experimenting with RNAi since he first heard that it functioned in mammals at the 2001 RNA Society conference addressed by Tom Tuschl, one of the discoverers of mammalian microRNAs (miRNA) who also found that RNAi functions in mammalian species.
Like many, Purcell rushed back to his own laboratory and began designing short interfering RNAs (siRNA) – the small strands of RNA that bind to complementary sequences of messenger RNAs and silence gene expression post-transcriptionally – as a way of targeting HIV. However, as everyone else discovered, HIV is often able to rapidly escape control by siRNAs, its mutability working in its favour yet again.
That doesn’t mean RNAi may not prove a potent weapon against HIV, as the multitude of clinical studies currently underway illustrate. It’s just that many viruses, especially HIV, have evolved with the ability to counter these cellular control mechanisms, so designing the right siRNA is paramount. short hairpin RNAs (shRNAs) are also in development. For example, in October a trial of an shRNA-based RNAi therapy in HIV patients released promising – although very early – results.
Silencing HIV genes using siRNAs and shRNAs does reduce viral replication and as such is a very promising line of enquiry. Like many others, Purcell’s laboratory has designed lentivectors for delivery of siRNA to target HIV, but his team has also pursued other protein targets, in particular cellular proteins that do not mutate and therefore may provide an alternative route.
One target is the HIV trans-activation response RNA binding protein (TRBP), which inhibits the RNA-dependent protein kinase (PKR) response. TRBP binds to PKR, one of the main interferon-response proteins that shut down the expression of viruses in cells, and in HIV infection keeps the PKR pathway from doing its job.
Purcell’s lab has been studying TRBP for many years and has uncovered some surprising activities of the protein in astrocytes. The researchers have also discovered that TRBP turns out to be an essential component of the miRNA biogenesis pathway – it is a binding partner for Dicer, the protein that cleaves double-stranded RNA into siRNAs and sets off the whole RNAi cascade.
“It has been an interesting observation – almost a roadblock – that the TRBP molecule turned out to be an essential component of the RNAi pathway,” Purcell says. “It turns out that it’s a very good target (for inhibition) but that also eliminates the ability to produce siRNAs.
“There’s a lot that we don’t understand in the control of gene expression at a post-transcriptional level and viruses are very busy in this area. Many of them make proteins that are able to defend against these cellular control mechanisms, and curiously the same viral proteins that inhibit PKR also inhibit RNA interference.”
