Stimulation of Transgene Expression by RNA Export Elements

Inventors: Thomas Hope, Romain Zufferey, Didier Trono, John Edward Donello
Potential Uses: Gene Expression
Enhance transgene expression by promoting correct processing and transport of RNA from the cell nucleus to the cytoplasm.

High levels of transgene expression are desired for most gene therapy protocols. This invention is directed to cis-acting RNA export elements and the use of such elements to enhance transgene expression. Specifically, the RNA export elements of the invention enhance transgene expression by promoting correct processing and transport of RNA from the cell nucleus to the cytoplasm. Nuclear export of RNAs containing invention cis acting RNA export elements is mediated by cellular factors which interact with structural elements contained within the cis acting elements. Accordingly, such export requires no exogenous factors. Introduction of a cis-acting RNA export element of the invention into a retroviral vector substantially increases the expression of transgene contained in the vector. For example, introduction of an export element derived from the woodchuck hepatitis virus (WHV) downstream of GFP and luciferase cDNAs, within the context of an HIV-based vector, increased expression of these reporters from 5- to 10-fold compared to homologous vectors which did not contain the export element. Stimulation of transgene expression by the export element is vector and promoter independent. Similar stimulatory effects are seen when the transgene is delivered in the widely utilized MLV based vector system. Moreover, the invention export element derived from the WHV stimulates transgene expression to similar levels when stable cell populations are generated by transfection. Transgene expression utilizing the export element derived from WHV is also stimulated when either the strong CMV promoter or the weaker thymidine kinase promoter are utilized to drive transgene expression. The posttranscriptional regulatory element isolated from WHV, namely WPRE, is notable because it is an extremely efficient RNA export element with the ability to activate a reporter for RNA export up to levels achieved with HIV-1 Rev and the Rev-responsive element (RRE).