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RNAi Broadens its Horizons, Takes Aim at HIV, Cholesterol

Science Editor

Ten years after it was first described as a cellular mechanism to control gene expression, and two years after Andrew Fire and Craig Mello won the Nobel Prize in Physiology or Medicine for its discovery, the therapeutic harnessing of RNA interference continues apace.

Two new papers described using the approach in two very different indications. One group of researchers used short interfering RNAs to combat HIV infection, while another showed that the approach can be used to drastically lower cholesterol levels.

For the latter paper, published in the Aug. 12, 2008, online edition of the Proceedings of the National Academy of Sciences, researchers from the University of Texas Southwestern Medical Center in collaboration with Alnylam Pharmaceuticals and the Massachusetts Institute of Technology, used siRNAs to block the production of proprotein convertase subtilisin/kexin type 9, or PCSK9, a protein that normally raises the level of "bad" low-density lipoprotein cholesterol by influencing the levels of liver LDL receptor.

The scientists made versions of siRNA that blocked the forms of PCSK9 found in rodents, monkeys and humans. Those siRNAs were then injected into normal rodents and monkeys, as well as into mice that had been genetically engineered to produce human PCSK9.

The siRNAs caused the levels of PCSK9 to drop 60 percent to 70 percent in the rodents, and by slightly more than half in monkeys. That in turn led to increased levels of LDL receptors in the liver and decreased levels of plasma LDL cholesterol. "Good" HDL cholesterol and triglycerides were not affected.

The researchers termed the effect on cholesterol levels in rats "unexpected," given that statins, like PCSK9, work by affecting LDL receptor levels, and statins do not lower cholesterol levels in rats.

In the animals, the effect also was comparatively long-lasting. Cholesterol levels fell after a single dose of the siRNA, which was delivered using so-called lipidoid nanoparticles to specifically target the liver, and took several weeks to return back to their baseline values.

The authors concluded that their approach, in animal models anyway, is at least on a par with statins, and that their findings open the way "for the development of PCSK9-lowering agents as a future strategy for treatment of hypercholesterolemia."

Another paper - this one to be published in the Aug. 22, 2008, issue of Cell and now available online - showed that RNAi also may prove useful in the treatment and perhaps even the prevention, of HIV infection.

As often is proving to be the case with RNAi, the key to the success of the researchers hinged on finding a way to deliver the RNA specifically to the cells where it was needed - in that case, T cells.

As the scientists explained in their paper, "Studies have shown the effectiveness of RNAi in suppressing HIV replication in cell lines as well as in primary human T cells and macrophages, the prime targets of HIV." But, they added, "the promise shown in in vitro studies, for RNAi to become clinically useful, many parameters, including delivery to susceptible cells, antiviral efficacy, and toxicity, need to be tested in vivo."

The researchers used a linker peptide to combine three different types of siRNA to part of an antibody that targets a surface molecule on T cells. One of the siRNAs targeted the HIV co-receptor CCR5, while the other two were aimed at viral proteins.

When mice engineered to contain human immune system cells (since mice normally are not prone to HIV infection) were treated with that construct, CD4 cell levels remained essentially stable after infection with HIV; in untreated controls, the levels of CD4 T cells dropped rapidly.

The authors argued that though one school of thought holds that a chronic infection such as HIV is best combated with gene therapy delivery of RNA, delivering exogenous siRNA would allow doctors to vary the sequence if the virus mutates to develop resistance to the initially used sequences.

Published: August 12, 2008