title: Regulation of endogenous gene expression using small molecule-controlled engineered zinc-finger protein transcription factors.
abstract: Small-molecule-regulated gene expression offers the promise of titrating the dose and duration of action of DNA-based therapies. To this end, we show that engineered zinc-finger protein transcription factors (ZFP TFs) can be coupled with a drug-inducible regulatory domain to permit small-molecule control of endogenous gene transcription. We constructed a drug-responsive ZFP TF via the fusion of a ZFP DNA-binding domain (DBD) targeting the human VEGF-A gene and an effector domain containing a truncated progesterone receptor ligand-binding domain linked to the NFkappaB p65 activation domain. Introduction of this engineered ZFP TF into human or murine cells allowed expression of the chromosomal VEGF-A gene to be induced upon addition of mifepristone, a synthetic steroid analog. Mifepristone-dependent VEGF-A induction was rapid, dose-dependent and reversible. Moreover, stable lines expressing the drug-responsive ZFP TF could be maintained in a state of continuous induction for at least 30 days without loss of viability. Potent VEGF-A induction was demonstrated using different engineered ZFP DBDs, thus this approach may represent a general solution to small-molecule regulation of targeted endogenous genes.
authors.affiliation: Sangamo BioSciences Inc., Point Richmond Tech Center, Richmond, CA 94804, USA.
authors.forename: C L
authors.initials: CL
authors.lastname: Dent
authors.forename: G
authors.initials: G
authors.lastname: Lau
authors.forename: E A
authors.initials: EA
authors.lastname: Drake
authors.forename: A
authors.initials: A
authors.lastname: Yoon
authors.forename: C C
authors.initials: CC
authors.lastname: Case
authors.forename: P D
authors.initials: PD
authors.lastname: Gregory
mesh_terms: D000818:Animals; D002460:Cell Line; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D015870:Gene Expression / Q000187:drug effects; D005786:Gene Expression Regulation / Q000187:drug effects; D005818:Genetic Engineering; D015316:Genetic Therapy / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000235:genetics; D006727:Hormone Antagonists / Q000494:pharmacology; D006801:Humans; D051379:Mice; D015735:Mifepristone / Q000494:pharmacology; D009369:Neoplasms / Q000628:therapy*; D011980:Receptors, Progesterone / Q000235:genetics; D013997:Time Factors; D051996:Transcription Factor RelA / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics*; D042461:Vascular Endothelial Growth Factor A / Q000032:analysis / Q000235:genetics*; D016335:Zinc Fingers / Q000235:genetics*
chemical_list: D006727:Hormone Antagonists; D011980:Receptors, Progesterone; D051996:Transcription Factor RelA; D014157:Transcription Factors; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D049452:Green Fluorescent Proteins; D015735:Mifepristone