The proposed POI is a multi-institutional grant that will develop a stem cell based therapy for the treatment of sickle cell disease (SCD) and ?-thalassemia (?-thal) as well as other hemoglobinopathies, using patient derived somatic cells and reprogramming them into induced pluripotent stem (IPS) cells that will have their mutations corrected and ultimately differentiated into hematopoietic stem cells (HSCs) to reconstitute the patient's hematopoietic system. Development of an effective cellular therapy for the treatment of hemoglobinopathies, the most common inherited diseases worldwide, would significantly improve the quality of life of individuals afflicted with SCD and B-thalassemia that are common among the peoples of Africa, the Mediterranean, the Middle East, and Asia as well as their descendents in the U.S. This proposal will test the hypothesis that an effective cellular and genetic therapy for these diseases can be achieved in the context of this PPG through the generation, modification, and the hematopoietic differentiation of patient derived iPS cells. This will be accomplished through the following Projects: Project 1 will involve the conversion of a patient's somatic cells into IPS cells using phiC31 Integrase-mediated, sequence-specific integration of a plasmid carrying 2A peptide linked Oct4, Sox2, Klf4, and cMyc reprogramming cDNAs or by using small activating double stranded RNA (saRNA) to transiently enhance the expression of these reprogramming genes. Project 2 will involve correction of the disease causing mutations in the somatic cells and the iPS cells by sequence specific modification using either classical homologous recombination (HR) or by oligo/polynucleotide-based small fragment homologous replacement (SFHR) in the presence or absence of targeted zinc finger nucleases (ZFNs) or other meganucleases. Project 3 will involve exposure of uncorrected and corrected iPS cells to conditions to direct hematopoietic differentiation to generate HSCs which have the capacity to engraft and reconstitute the hematopoietic system. In the course of this PPG, all Projects will develop xeno-free systems to optimize safety. The science in the Projects will be augmented by an administrative (Core A) and 2 scientific Cores: Core B: Cell and Molecular Biology, and Core C: Cell Transplantation and Analysis.

Public Health Relevance

This Program Project will focus on the development of a therapy for sickle cell anemia and B-thalassemia the most common genetic diseases worldwide. It aims to devise new methods of treating these diseases by genetically correcting patient cells to generate stems cells for transplantation, thus avoiding rejection due to histo-incompatibility. Correcting these diseases would significantly improve the quality of life among afflicted individuals and decrease the social and economic burden that they impose on the healthcare system.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Program Projects (P01)
Project #
5P01DK088760-03
Application #
8532884
Study Section
Special Emphasis Panel (ZDK1-GRB-6 (M3))
Program Officer
Bishop, Terry Rogers
Project Start
2011-09-30
Project End
2016-07-31
Budget Start
2013-08-01
Budget End
2014-07-31
Support Year
3
Fiscal Year
2013
Total Cost
$1,255,622
Indirect Cost
$291,338
Name
University of California San Francisco
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
094878337
City
San Francisco
State
CA
Country
United States
Zip Code
94143
Muench, Marcus O; Beyer, Ashley I; Fomin, Marina E et al. (2014) The adult livers of immunodeficient mice support human hematopoiesis: evidence for a hepatic mast cell population that develops early in human ontogeny. PLoS One 9:e97312
Ye, Lin; Wang, Jiaming; Beyer, Ashley I et al. (2014) Seamless modification of wild-type induced pluripotent stem cells to the natural CCR5?32 mutation confers resistance to HIV infection. Proc Natl Acad Sci U S A 111:9591-6
Fomin, M E; Togarrati, P P; Muench, M O (2014) Progress and challenges in the development of a cell-based therapy for hemophilia A. J Thromb Haemost 12:1954-65
Xie, Fei; Ye, Lin; Chang, Judy C et al. (2014) Seamless gene correction of ?-thalassemia mutations in patient-specific iPSCs using CRISPR/Cas9 and piggyBac. Genome Res 24:1526-33
Sargent, R Geoffrey; Suzuki, Shingo; Gruenert, Dieter C (2014) Nuclease-mediated double-strand break (DSB) enhancement of small fragment homologous recombination (SFHR) gene modification in human-induced pluripotent stem cells (hiPSCs). Methods Mol Biol 1114:279-90
Fomin, Marina E; Zhou, Yanchen; Beyer, Ashley I et al. (2013) Production of factor VIII by human liver sinusoidal endothelial cells transplanted in immunodeficient uPA mice. PLoS One 8:e77255
Ng, Terry Fei Fan; Kondov, Nikola O; Hayashimoto, Nobuhito et al. (2013) Identification of an astrovirus commonly infecting laboratory mice in the US and Japan. PLoS One 8:e66937
Ye, Lin; Muench, Marcus O; Fusaki, Noemi et al. (2013) Blood cell-derived induced pluripotent stem cells free of reprogramming factors generated by Sendai viral vectors. Stem Cells Transl Med 2:558-66