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
Suzuki, Shingo; Sargent, R Geoffrey; Illek, Beate et al. (2016) TALENs Facilitate Single-step Seamless SDF Correction of F508del CFTR in Airway Epithelial Submucosal Gland Cell-derived CF-iPSCs. Mol Ther Nucleic Acids 5:e273
Beyer, Ashley I; Muench, Marcus O (2016) Comparison of human hematopoietic reconstitution in different strains of immunodeficient mice. Stem Cells Dev :
Ye, Lin; Wang, Jiaming; Tan, Yuting et al. (2016) Genome editing using CRISPR-Cas9 to create the HPFH genotype in HSPCs: An approach for treating sickle cell disease and β-thalassemia. Proc Natl Acad Sci U S A 113:10661-5
Baimukanova, Gyulnar; Miyazawa, Byron; Potter, Daniel R et al. (2016) Platelets regulate vascular endothelial stability: assessing the storage lesion and donor variability of apheresis platelets. Transfusion 56 Suppl 1:S65-75
Wiemels, J L; de Smith, A J; Xiao, J et al. (2016) A functional polymorphism in the CEBPE gene promoter influences acute lymphoblastic leukemia risk through interaction with the hematopoietic transcription factor Ikaros. Leukemia 30:1194-7
Mahajan, Maya M; Cheng, Betty; Beyer, Ashley I et al. (2015) A quantitative assessment of the content of hematopoietic stem cells in mouse and human endosteal-bone marrow: a simple and rapid method for the isolation of mouse central bone marrow. BMC Hematol 15:9
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
Xie, Fei; Ye, Lin; Chang, Judy C et al. (2014) Seamless gene correction of β-thalassemia mutations in patient-specific iPSCs using CRISPR/Cas9 and piggyBac. Genome Res 24:1526-33
Ye, Lin; Wang, Jiaming; Beyer, Ashley I et al. (2014) Seamless modification of wild-type induced pluripotent stem cells to the natural CCR5Δ32 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

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