Ribonucleoprotein transfection agents
Provided herein are compositions and methods useful, inter alia, for the delivery of ribonucleoprotein complexes (e.g., Cas9/guide RNA complexes) into cells. The compositions and methods provided herein are particularly useful for the delivery of ribonucleoprotein complexes into pluripotent cells and lymphatic cells.
1. A composition comprising a ribonucleoprotein complex, a lipid aggregate-forming cationic lipid and an enhancer element;
wherein said lipid aggregate-forming cationic lipid has the structure of formula:
wherein
R 1 and R 2 are independently hydrogen, substituted or unsubstituted heteroalkyl, or substituted or unsubstituted alkyl,
R 5 , R 6 , R 7 , R 8 and R 9 are independently hydrogen or unsubstituted alkyl;
X is an anion; and
m is an integer from 1-10;
n is an integer from 1-6; and
o and p are independently integers from 8-30,
and wherein said enhancer element comprises a polyamine moiety covalently attached to a nuclear localization signal sequence.
2. The composition of claim 1 , wherein said ribonucleoprotein complex comprises an endonuclease and a ribonucleic acid.
3. The composition of claim 2 , wherein said endonuclease comprises CRISPR associated protein 9 (Cas9) and said ribonucleic acid comprises a guide RNA.
4. The composition of claim 1 , wherein R 1 is substituted or unsubstituted heteroalkyl.
5. The composition of claim 1 , wherein said polyamine moiety is a spermine moiety.
6. The composition of claim 1 , wherein said polyamine moiety is a plurality of spermine moieties.
7. The composition of claim 1 , wherein said polyamine moiety is covalently attached to the N-terminus of said nuclear localization signal sequence.
8. The composition of claim 1 , wherein said enhancer element comprises a peptide comprising the sequence of SEQ ID NO:1-SEQ ID NO:46.
9. The composition of claim 1 , wherein said enhancer element comprises a peptide comprising at least two nuclear localization signal (NLS) sequences.
10. The composition of claim 1 , further comprising a donor nucleic acid.
11. The composition of claim 1 , further comprising a eukaryotic cell.
12. The composition of claim 11 , wherein said eukaryotic cell is a pluripotent cell, a lymphatic cell, a T cell, or a hepatocyte.
13. The composition of claim 12 , wherein said pluripotent cell is an induced pluripotent cell or an embryonic stem cell.
14. The composition of claim 1 , wherein R 5 , R 6 , R 7 , R 8 and R 9 are hydrogen.
15. The composition of claim 1 , wherein R 2 is hydrogen or substituted or unsubstituted alkyl.