This proposal is based on the novel approach of designing an engineering novel zinc finger protein transcription factors (ZFPs) to regulate the expression of endogenous genes for the treatment of cardiovascular disease. We have previously validated this approach in vivo and in vitro with engineered ZFPs that activate expression of the VEGF-A gene. We now propose further studies to more vigorously investigate the mechanism of action of our ZFP constructs and to delineate the differences in the biological effects of endogenous gene activation vs. cDNA-based transgene expression. In the context of these experiments we study the determinants of ZFP half-life and the effects of differential ZFP positional DMA- binding on the stoichiometry of VEGF splice variant expression. We will also investigate ZFP-induced gene repression using a ZFP construct engineered to target and repress the phospholamban (PLB) gene. PLB is a critical determinant of cardiac contractility, and the ability to downregulate the expression of PLB has significant clinical implications. In the context of these experiments we will also study the relationship between the physical half-life of the PLB represser ZFP and the functional half-life of PLB repression. These experiments represent the first time an engineered transcription represser will have been used in a clinically relevant in vivo context. Finally, we will further build on work we have recently completed demonstrating the feasibility of building cell- permeable ZFPs that are capable of crossing cell membranes, entering the nucleus and regulating the expression of specifically targeted genes. This represents a novel new therapeutic approach that also has significant clinical implications.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL075616-04
Application #
7683761
Study Section
Vascular Cell and Molecular Biology Study Section (VCMB)
Program Officer
Evans, Frank
Project Start
2006-08-01
Project End
2011-05-31
Budget Start
2009-06-01
Budget End
2010-05-31
Support Year
4
Fiscal Year
2009
Total Cost
$401,751
Indirect Cost
Name
Yale University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
043207562
City
New Haven
State
CT
Country
United States
Zip Code
06520
Li, Qi; Michaud, Michael; Park, Chan et al. (2017) The role of endothelial HIF-1 ?in the response to sublethal hypoxia in C57BL/6 mouse pups. Lab Invest 97:356-369
Liu, Dinggang; Lei, Li; Desir, Matthew et al. (2016) Smooth Muscle Hypoxia-Inducible Factor 1? Links Intravascular Pressure and Atherosclerosis--Brief Report. Arterioscler Thromb Vasc Biol 36:442-5
Huang, Yan; Di Lorenzo, Annarita; Jiang, Weidong et al. (2013) Hypoxia-inducible factor-1? in vascular smooth muscle regulates blood pressure homeostasis through a peroxisome proliferator-activated receptor-?-angiotensin II receptor type 1 axis. Hypertension 62:634-40
Zhang, Pei; Huang, Angela; Morales-Ruiz, Manuel et al. (2012) Engineered zinc-finger proteins can compensate genetic haploinsufficiency by transcriptional activation of the wild-type allele: application to Willams-Beuren syndrome and supravalvular aortic stenosis. Hum Gene Ther 23:1186-99
Tirziu, Daniela; Giordano, Frank J; Simons, Michael (2010) Cell communications in the heart. Circulation 122:928-37
Giordano, Frank J (2007) Therapeutic gene regulation: targeting transcription. Circulation 115:1180-3