Receptor-mediated delivery of engineered nucleases for genome modification

Zhong Chen, Lahcen Jaafar, Davies G. Agyekum, Haiyan Xiao, Marlene F. Wade, R. Ileng Kumaran, David L. Spector, Gang Bao, Matthew H. Porteus, William S. Dynan, Steffen E Meiler

Research output: Contribution to journalArticle

28 Citations (Scopus)

Abstract

Engineered nucleases, which incise the genome at predetermined sites, have a number of laboratory and clinical applications. There is, however, a need for better methods for controlled intracellular delivery of nucleases. Here, we demonstrate a method for ligand-mediated delivery of zinc finger nucleases (ZFN) proteins using transferrin receptor-mediated endocytosis. Uptake is rapid and efficient in established mammalian cell lines and in primary cells, including mouse and human hematopoietic stem-progenitor cell populations. In contrast to cDNA expression, ZFN protein levels decline rapidly following internalization, affording better temporal control of nuclease activity. We show that transferrin-mediated ZFN uptake leads to site-specific in situ cleavage of the target locus. Additionally, despite the much shorter duration of ZFN activity, the efficiency of gene correction approaches that seen with cDNA-mediated expression. The approach is flexible and general, with the potential for extension to other targeting ligands and nuclease architectures.

Original languageEnglish (US)
JournalNucleic Acids Research
Volume41
Issue number19
DOIs
StatePublished - Oct 1 2013

Fingerprint

Zinc Fingers
Genome
Hematopoietic Stem Cells
Complementary DNA
Ligands
Transferrin Receptors
Transferrin
Endocytosis
Proteins
Cell Line
Population
Genes

ASJC Scopus subject areas

  • Genetics

Cite this

Receptor-mediated delivery of engineered nucleases for genome modification. / Chen, Zhong; Jaafar, Lahcen; Agyekum, Davies G.; Xiao, Haiyan; Wade, Marlene F.; Ileng Kumaran, R.; Spector, David L.; Bao, Gang; Porteus, Matthew H.; Dynan, William S.; Meiler, Steffen E.

In: Nucleic Acids Research, Vol. 41, No. 19, 01.10.2013.

Research output: Contribution to journalArticle

Chen, Z, Jaafar, L, Agyekum, DG, Xiao, H, Wade, MF, Ileng Kumaran, R, Spector, DL, Bao, G, Porteus, MH, Dynan, WS & Meiler, SE 2013, 'Receptor-mediated delivery of engineered nucleases for genome modification', Nucleic Acids Research, vol. 41, no. 19. https://doi.org/10.1093/nar/gkt710
Chen Z, Jaafar L, Agyekum DG, Xiao H, Wade MF, Ileng Kumaran R et al. Receptor-mediated delivery of engineered nucleases for genome modification. Nucleic Acids Research. 2013 Oct 1;41(19). https://doi.org/10.1093/nar/gkt710
Chen, Zhong ; Jaafar, Lahcen ; Agyekum, Davies G. ; Xiao, Haiyan ; Wade, Marlene F. ; Ileng Kumaran, R. ; Spector, David L. ; Bao, Gang ; Porteus, Matthew H. ; Dynan, William S. ; Meiler, Steffen E. / Receptor-mediated delivery of engineered nucleases for genome modification. In: Nucleic Acids Research. 2013 ; Vol. 41, No. 19.
@article{8c6a718fa37f44fca454be197b7c5dd3,
title = "Receptor-mediated delivery of engineered nucleases for genome modification",
abstract = "Engineered nucleases, which incise the genome at predetermined sites, have a number of laboratory and clinical applications. There is, however, a need for better methods for controlled intracellular delivery of nucleases. Here, we demonstrate a method for ligand-mediated delivery of zinc finger nucleases (ZFN) proteins using transferrin receptor-mediated endocytosis. Uptake is rapid and efficient in established mammalian cell lines and in primary cells, including mouse and human hematopoietic stem-progenitor cell populations. In contrast to cDNA expression, ZFN protein levels decline rapidly following internalization, affording better temporal control of nuclease activity. We show that transferrin-mediated ZFN uptake leads to site-specific in situ cleavage of the target locus. Additionally, despite the much shorter duration of ZFN activity, the efficiency of gene correction approaches that seen with cDNA-mediated expression. The approach is flexible and general, with the potential for extension to other targeting ligands and nuclease architectures.",
author = "Zhong Chen and Lahcen Jaafar and Agyekum, {Davies G.} and Haiyan Xiao and Wade, {Marlene F.} and {Ileng Kumaran}, R. and Spector, {David L.} and Gang Bao and Porteus, {Matthew H.} and Dynan, {William S.} and Meiler, {Steffen E}",
year = "2013",
month = "10",
day = "1",
doi = "10.1093/nar/gkt710",
language = "English (US)",
volume = "41",
journal = "Nucleic Acids Research",
issn = "0305-1048",
publisher = "Oxford University Press",
number = "19",

}

TY - JOUR

T1 - Receptor-mediated delivery of engineered nucleases for genome modification

AU - Chen, Zhong

AU - Jaafar, Lahcen

AU - Agyekum, Davies G.

AU - Xiao, Haiyan

AU - Wade, Marlene F.

AU - Ileng Kumaran, R.

AU - Spector, David L.

AU - Bao, Gang

AU - Porteus, Matthew H.

AU - Dynan, William S.

AU - Meiler, Steffen E

PY - 2013/10/1

Y1 - 2013/10/1

N2 - Engineered nucleases, which incise the genome at predetermined sites, have a number of laboratory and clinical applications. There is, however, a need for better methods for controlled intracellular delivery of nucleases. Here, we demonstrate a method for ligand-mediated delivery of zinc finger nucleases (ZFN) proteins using transferrin receptor-mediated endocytosis. Uptake is rapid and efficient in established mammalian cell lines and in primary cells, including mouse and human hematopoietic stem-progenitor cell populations. In contrast to cDNA expression, ZFN protein levels decline rapidly following internalization, affording better temporal control of nuclease activity. We show that transferrin-mediated ZFN uptake leads to site-specific in situ cleavage of the target locus. Additionally, despite the much shorter duration of ZFN activity, the efficiency of gene correction approaches that seen with cDNA-mediated expression. The approach is flexible and general, with the potential for extension to other targeting ligands and nuclease architectures.

AB - Engineered nucleases, which incise the genome at predetermined sites, have a number of laboratory and clinical applications. There is, however, a need for better methods for controlled intracellular delivery of nucleases. Here, we demonstrate a method for ligand-mediated delivery of zinc finger nucleases (ZFN) proteins using transferrin receptor-mediated endocytosis. Uptake is rapid and efficient in established mammalian cell lines and in primary cells, including mouse and human hematopoietic stem-progenitor cell populations. In contrast to cDNA expression, ZFN protein levels decline rapidly following internalization, affording better temporal control of nuclease activity. We show that transferrin-mediated ZFN uptake leads to site-specific in situ cleavage of the target locus. Additionally, despite the much shorter duration of ZFN activity, the efficiency of gene correction approaches that seen with cDNA-mediated expression. The approach is flexible and general, with the potential for extension to other targeting ligands and nuclease architectures.

UR - http://www.scopus.com/inward/record.url?scp=84886043841&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84886043841&partnerID=8YFLogxK

U2 - 10.1093/nar/gkt710

DO - 10.1093/nar/gkt710

M3 - Article

C2 - 23956220

AN - SCOPUS:84886043841

VL - 41

JO - Nucleic Acids Research

JF - Nucleic Acids Research

SN - 0305-1048

IS - 19

ER -