This application is a continuation on our studies of normal and abnormal hemoglobin synthesis in sickle cell anemia and thalassemia. The project in the previous grant period consisted of three aims. 1) DNA diagnosis of sickle cell anemia and thalassemia. 2) Control of globin gene expression. 3) Globin gene transfer. Because the topics in Aim 2 will now be carried on by previous trainees who have become independent investigators, this proposal will continue to pursue only the first and third aims.
hi Aim 1 we have completed our studies on dot blot hybridization for the diagnosis of the mutations in hemoglobinopathies and thalassemia that are commonly found in the American population. We propose to develop and refine methods of prenatal diagnosis from fetal cells isolated using maternal blood. Our preliminary studies show that this is a promising approach, but much more work is needed to make this test practical. We will use phage display to isolate single chain antibodies specific for fetal nucleated red cells and human embryonic hemoglobin. We will investigate the use of image analysis and laser capture microdissections to facilitate the retrieval of fetal nucleated red cells. These developments will greatly enhance the usefulness of this procedure.
The second aim of this proposal is to explore the possibility of in utero gene delivery alpha-thalassemia is a good model for in utero gene therapy because pathological changes in homozygous alpha-thalassemia usually appear before birth. In our laboratory, we have made mouse models of alpha-thalassemia by knocking out the endogenous mouse alpha-globin genes. By crossing various strains, we have mice that have 3,2,1 or no alpha-globin gene and they mimic the clinical manifestation of human alpha-thalassemia. We will use AAV vectors and lentiviral vectors containing the beta-g1obin LCR controlling the human a-globin gene to inject into fetal mice at the 15th week of gestation. We will follow these mice to observe the expression of the. alpha-globin gene and the rescue the disease phenotype. Such an approach could be useful not only for alpha-thalassemia but also for other genetic disorders such as OTC deficiency where the disease begins in utero.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
2R01DK016666-30
Application #
6542371
Study Section
Hematology Subcommittee 2 (HEM)
Program Officer
Badman, David G
Project Start
1976-08-01
Project End
2006-07-31
Budget Start
2002-09-16
Budget End
2003-07-31
Support Year
30
Fiscal Year
2002
Total Cost
$394,771
Indirect Cost
Name
University of California San Francisco
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
073133571
City
San Francisco
State
CA
Country
United States
Zip Code
94143
Ye, Lin; Chang, Judy C; Lu, Ronghua et al. (2008) High oxygen environment during pregnancy rescues sickle cell anemia mice from prenatal death. Blood Cells Mol Dis 41:67-72
Han, Xiao-Dong; Lin, Chin; Chang, Judy et al. (2007) Fetal gene therapy of alpha-thalassemia in a mouse model. Proc Natl Acad Sci U S A 104:9007-11
Kraft, Andrew D; Lee, Jong-Min; Johnson, Delinda A et al. (2006) Neuronal sensitivity to kainic acid is dependent on the Nrf2-mediated actions of the antioxidant response element. J Neurochem 98:1852-65
Calkins, Marcus J; Jakel, Rebekah J; Johnson, Delinda A et al. (2005) Protection from mitochondrial complex II inhibition in vitro and in vivo by Nrf2-mediated transcription. Proc Natl Acad Sci U S A 102:244-9
Lee, Jong-Min; Chan, Kaimin; Kan, Yuet Wai et al. (2004) Targeted disruption of Nrf2 causes regenerative immune-mediated hemolytic anemia. Proc Natl Acad Sci U S A 101:9751-6
Lee, Jong-Min; Calkins, Marcus J; Chan, Kaimin et al. (2003) Identification of the NF-E2-related factor-2-dependent genes conferring protection against oxidative stress in primary cortical astrocytes using oligonucleotide microarray analysis. J Biol Chem 278:12029-38
Huie, M A; Cheung, M C; Muench, M O et al. (2001) Antibodies to human fetal erythroid cells from a nonimmune phage antibody library. Proc Natl Acad Sci U S A 98:2682-7
Chan, K; Han, X D; Kan, Y W (2001) An important function of Nrf2 in combating oxidative stress: detoxification of acetaminophen. Proc Natl Acad Sci U S A 98:4611-6
Chan, K; Kan, Y W (1999) Nrf2 is essential for protection against acute pulmonary injury in mice. Proc Natl Acad Sci U S A 96:12731-6
Chan, J Y; Kwong, M; Lu, R et al. (1998) Targeted disruption of the ubiquitous CNC-bZIP transcription factor, Nrf-1, results in anemia and embryonic lethality in mice. EMBO J 17:1779-87

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