Inducible and reversible breaching of the blood brain barrier by RNAi

John J Rossi

doi:10.1002/emmm.201100132

Abstract

Sequence-specific knockdown of gene expression is a goal that has been long sought by both basic and clinical investigators. In this regard, the discovery of RNA interference (RNAi) in Caenorhabditis elegans was immediately recognized as a potential breakthrough for studying gene function (Fire et al, 1998). These findings demonstrated that double-stranded (ds)RNAs are triggers for sequence-specific, post-transcriptional gene silencing via targeted degradation of messenger RNAs harbouring a complementary sequence to one of the two strands. Initially, it was thought that such post-transcriptional regulation of gene expression could not be achieved in mammalian systems due to the strong induction of interferon by dsRNAs. This potential restriction was short lived with the demonstration that endonuclease processed dsRNAs of 21–25 nucleotides in length, designated small interfering RNAs (siRNAs), were able to elicit sequence-specific degradation of mRNAs in mammalian cells without triggering interferon responses (Elbashir et al, 2001). These findings provided a huge impetus to develop RNAi as a therapeutic modality. The dream to selectively block the expression of deleterious proteins and treat formerly non-drugable diseases led to the rapid establishment of new biotech companies and branches of major pharmaceutical companies devoted to RNAi therapeutics.

Highlights

Sequence-specific knockdown of gene expression is a goal that has been long sought by both basic and clinical investigators
An alternative approach is to use promoter expressed short hairpinRNAs in gene therapy settings. These short hairpins are processed by the RNA interference (RNAi) machinery into small interfering RNAs (siRNAs), which trigger sequence-specific degradation of target transcripts
Despite the relative ease designing and expressing shRNAs, the use of promoter expressed shRNAs received somewhat of a setback when it was demonstrated that the high level expression of Pol III expressed shRNAs incorporated into an adeno associated viral (AAV) vector resulted in extreme liver toxicity and death of mice (Grimm et al, 2006)

Summary

Introduction

Sequence-specific knockdown of gene expression is a goal that has been long sought by both basic and clinical investigators. These startling findings put the brakes on the development of all RNAi-based drugs for a time, the major problem was found to be toxicity related to high levels of expression of the shRNAs resulting in competition for cellular RNAi components and consequent off-target effects.

Results

Conclusion