Liver-specific inducible promoters and methods of use thereof
The present invention relates to inducible regulatory elements, promoters and vectors, especially gene therapy vectors, and methods of their use.
1 . A vector comprising an expression cassette, the expression cassette comprising a synthetic liver-specific inducible promoter operably linked to a gene, the synthetic liver-specific inducible promoter comprising the at least one cis-regulatory module (CRM) comprising or consisting of a sequence selected from the group consisting of: SEQ ID NO: 42, SEQ ID NO: 43, SEQ ID NO: 44, SEQ ID NO: 45, SEQ ID NO: 46, and SEQ ID NO: 47.
2 . The vector of claim 1 , comprising a plurality of cis-regulatory element (CREs) that are each capable of being bound and activated by a heterodimer of CAR and RXR or a CRE that is capable of being bound and activated by a heterodimer of CAR and RXR operably linked to a minimal promoter or proximal promoter.
3 . The vector of claim 2 , comprising 2 or 3 CREs that are each capable of being bound and activated by a heterodimer of CAR and RXR.
4 . The vector of claim 2 wherein the minimal promoter is HSV thymidine kinase minimal promoter (MinTK), CMV minimal promoter (CMVmp) or SV40 minimal promoter (SV40mp).
5 . The vector of claim 1 , which comprises a sequence according any one of SEQ ID NOs: 10-18 and 59, 67 and 71.
6 . The vector of claim 5 , which comprises a sequence SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 68, SEQ ID NO: 69, SEQ ID NO: 70 or SEQ ID NO: 71.
7 . The vector of claim 1 , wherein the synthetic liver-specific expression cassette comprises sequences providing or coding for one or more of, and preferably all of, a ribosomal binding site, a start codon, a stop codon, a transcription termination sequence, a nucleic acid encoding a posttranscriptional regulatory element and/or a polyA element.
8 . The vector of claim 1 , wherein the gene encodes any one of:
a protein or RNA,
a therapeutic expression product,
a therapeutic protein suitable for use in treating a disease or condition associated with aberrant gene expression in the liver,
a site-specific nuclease, such as a meganuclease, zinc finger nuclease (ZFN), transcription activator-like effector-based nuclease (TALEN), or the clustered regularly interspaced short palindromic repeats system (CRISPR-Cas.
9 . The vector of claim 1 , which is a plasmid or a viral vector.
10 . The vector of claim 1 , wherein the gene encodes a therapeutic expression product.
11 . The vector of claim 1 , formulated as a pharmaceutical composition and comprising a pharmaceutically acceptable carrier.
12 . The vector according to claim 1 present in a cell.
13 . The vector of claim 12 which is a liver cell.