METHODS AND COMPOSITIONS FOR INCREASING TRANSGENE ACTIVITY

• US 20160326548A1
• Filed: 04/28/2016
• Published: 11/10/2016
• Est. Priority Date: 05/07/2015
• Status: Active Grant

• Current Assignee: Sangamo Therapeutics, Inc.
• Original Assignee: Sangamo Biosciences, Inc. (Sangamo Therapeutics, Inc.)
• Inventors: Cost, Gregory J.
• Granted Patent: US 10,179,918 B2
• Time in Patent Office: Days
• US Class Current: 1/1
• CPC Class Codes: C12N 15/85 for animal cells
• CPC Class Codes: C12N 15/86 Viral vectors
• CPC Class Codes: C12N 2740/16043 viral genome or elements thereof as genetic vector
• CPC Class Codes: C12N 2740/16045 Special targeting system for viral vectors
• CPC Class Codes: C12N 2830/48 regulating transport or export of RNA, e.g. RRE, PRE, WPRE, CTE

Abstract

Methods and compositions for increasing transgene expression and/or activity, including for increasing nuclease-mediated genomic modifications.

Claims

1. A method of increasing nuclease activity in a cell, the method comprising:

introducing a polynucleotide encoding at least one component of one or more nucleases into the cell, wherein the polynucleotide comprises a woodchuck hepatitis virus posttranslational regulatory element (WPRE).

2. The method of claim 1, wherein the WPRE is a wild-type WPRE.

3. The method of claim 1, wherein the WPRE comprises a sequence as shown in any of SEQ ID NOs:

1 to 3.

4. The method of claim 1, wherein the polynucleotide is a plasmid.

5. The method of claim 1, wherein the polynucleotide is a viral vector.

6. The method of claim 1, wherein the polynucleotide is an mRNA.

7. The method of claim 1, wherein the nuclease is a zinc finger nuclease, a TALEN and/or a CRISPR/Cas and/or a Cfp1 CRISPR/Cas nuclease system.

8. The method of claim 1, further comprising subject the cells to cold-shock conditions.

9. An mRNA comprising a WRPE and a sequence encoding at least one component of a nuclease or a transcription factor (TF).

10. The mRNA of claim 9, wherein the sequence encodes a nuclease.

11. The mRNA of claim 9, wherein the nuclease is a zinc finger nuclease, a TALEN and/or at least one component of a CRISPR/Cas nuclease system.

12. The mRNA of claim 9, wherein the Cas protein is a Cas and/or Cfp1 protein.

13. The mRNA of claim 9, wherein the sequence encodes a transcription factor.

14. The mRNA of claim 13, wherein the transcription factor is a ZF-TF, a TALE-TF or a CRISPR/Cas TF.

15. The mRNA of claim 9, further comprising a polyA sequence.