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Stanford: High shRNA Expression Can Be Toxic


A team of researchers from Stanford University has shown that sustained, high-level shRNA expression in murine livers can result in severe toxicity and death — possibly as a result of the shRNAs interfering with endogenous microRNA processing machinery.

Although the findings, which appeared in Nature, have raised questions about the use of shRNA as a therapeutic modality, Mark Kay, a professor at Stanford’s School of Medicine and co-author of the paper, calls them “encouraging.”

“If you take the paper as a whole, it’s actually encouraging because we were able to get therapeutic effects without toxicity if the right sequences were selected,” he says. “In no way do I feel [the paper] should limit the enthusiasm for using shRNA clinically. I think it just points out that, like any drug, you have to monitor how much you give.”

In the paper, the researchers were looking to build on earlier experiments in which plasmid-expressed shRNAs were used to inhibit hepatitis B viral replication in cell culture and in normal and immunodeficient mice transfected with an HBV plasmid.

“What we were working on was a more clinically relevant way to do this, and to also show in a more clinically relevant way that we could get efficacy,” Kay says of the Nature paper. “We wanted to show in a more bona fide preclinical model that we could develop a gene-transfer approach that would be efficacious” for hepatitis B.

“The fact is that we tried to use the highest doses we could generate in the first studies because we didn’t know what to expect,” he says.

Additionally, it remains unclear whether the toxicity observed with high levels of shRNAs would be seen with similar levels of siRNAs. “We’ve never been able to get the same level of siRNA into all the hepatocytes of the mouse — there’s no way to do that,” Kay says.

— Doug Macron


Short Reads

Rosetta Genomics and US Genomics have agreed to collaborate on the development of a microRNA-based, non-invasive diagnostic for lung cancer.

Lentigen has received an exclusive worldwide license from the University of Pennsylvania to use its lentiviral vector technology. According to the company, the technology can be used to deliver genes or RNAi molecules into cells.

Qiagen and CytoPathfinder have teamed up to develop tools for high-throughput RNAi screening. The arrangement will combine Qiagen’s siRNA sets with CytoPathfinder’s transfection microarray technology, which allows for siRNA screening on a microarray chip.

Dharmacon, in its role as originator of the RNAi Global Initiative, aims to have a standards proposal available before the next meeting of the initiative, which will take place in Europe this fall.

Alnylam Pharmaceuticals announced that the USPTO issued one of the key patents from its Tuschl II estate for RNAi therapeutics. The patent covers methods for making siRNAs and is exclusively licensed to Alnylam from the agent for the Max Planck Society.

Frost & Sullivan issued a report on the RNAi market. Their best estimate for the field was a market valuation of $66.2 million in 2005, and they predict that the market will be worth $294.9 million in 2012.


US patent application 20060105360. Diagnosis and treatment of cancers with microRNA located in or near cancer-associated chromosomal features. Inventors: Carlo Croce, Chang-Gong Liu, George Calin, and Cinzia Sevignani. Filed: July 29, 2005.

“MicroRNA genes are highly associated with chromosomal features involved in the etiology of different cancers,” the abstract states. “The perturbations in the genomic structure or chromosomal architecture of a cell caused by these cancer-associated chromosomal features can affect the expression of the miR gene(s) located in close proximity to that chromosomal feature.”

US patent application 20060105377. Screening to optimize RNAi. Inventor: Richard Eglen. Filed: December 7, 2005.

This patent covers “methods and compositions … for screening RNAi molecules for efficiency of modulation, particularly inhibition” of gene expression. The patent provides methods, compositions, kits, and genetic constructs for intracellularly monitoring a b-galactosidase small fragment containing fusion protein gene as a screen for the inhibition of expression of the fusion protein by dsRNA.


$3 million

Amount that NIH has earmarked for funding a set of projects investigating the role of microRNAs and other non-coding RNAs in mental disorders.

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