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Series GSE229212 Query DataSets for GSE229212
Status Public on Apr 01, 2024
Title Development of a ß-Globin Gene Replacement Strategy as a Therapeutic Approach for ß-Hemoglobinopathies
Organisms Homo sapiens; Mus musculus
Experiment type Expression profiling by high throughput sequencing
Genome variation profiling by high throughput sequencing
Summary β-hemoglobinopathies are the most common genetic disorders worldwide. In sickle cell disease (SCD) a single mutation (E6V) in the b-globin (HBB) gene results in dysfunctional hemoglobin protein, while in β-thalassemia, over 300 mutations distributed across the HBB gene are known to reduce the production of β-globin and cause severe anemia. A genetic engineering approach that replaces the whole HBB gene is an ideal strategy to rescue HBB expression for most genotypes but is technically challenging as the insert cannot be homologous to the endogenous gene and codon-optimized, intron-less sequences may not reconstitute adequate HBB levels. Here, we developed a novel approach for a “one-size-fits-all" HBB gene repair strategy using CRISPR-Cas9 which successfully addresses these problems. First, our DNA donor design avoids sequence homology through a diverged HBB coding sequence and second, incorporates heterologous introns from the fetal g-globin gene, further reducing homology to endogenous HBB while mimicking its intron composition. Screening DNA donors with various heterologous globin introns, polyadenylation signals and truncated sequences, identified a heterologous intron DNA donor that highly expresses β-globin and rescued β-globin expression in two in vitro hemoglobinopathy models in hematopoietic stem and progenitor cells (HSPCs). Furthermore, healthy donor HSPCs modified with this HBB gene replacement approach showed successful engraftment in immunodeficient mice at 16 weeks. In summary, we developed a universal HBB gene replacement strategy that results in physiological b-globin production, offering a potential differentiated approach for treating patients with β-thalassemia, SCD or compound heterozygous individuals.
Overall design HBB gene repair strategy using CRISPR-Cas9 which includes: RNAseq on donors 1125 and 3404; mouse engraftment study, and long read sequencing data.
Contributor(s) Wallace KA, Wienert B
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Submission date Apr 07, 2023
Last update date Apr 01, 2024
Contact name Kristen Seim
Organization name GraphiteBio
Street address 201 Haskins Way
City San Francisco
State/province CA
ZIP/Postal code 94080
Country USA
Platforms (3)
GPL16417 Illumina MiSeq (Mus musculus)
GPL24676 Illumina NovaSeq 6000 (Homo sapiens)
GPL28352 Sequel II (Homo sapiens)
Samples (69)
GSM7156933 gb102 Engraftment study; Donor 1125 HBB only
GSM7156934 gb102 Engraftment study; Donor 1125 RNP only
GSM7156935 gb102 Engraftment study; Donor 1125 unedited sample
BioProject PRJNA953414

Download family Format
SOFT formatted family file(s) SOFTHelp
MINiML formatted family file(s) MINiMLHelp
Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE229212_Expression_Norm_gene7april2023.txt.gz 316 b (ftp)(http) TXT
GSE229212_KW-100-13.vcf.gz 28.5 Mb (ftp)(http) VCF
GSE229212_KW-100-21.vcf.gz 22.3 Mb (ftp)(http) VCF
GSE229212_KW-100-29.vcf.gz 32.0 Mb (ftp)(http) VCF
GSE229212_KW-100-5.vcf.gz 22.8 Mb (ftp)(http) VCF
GSE229212_KW-106-13.vcf.gz 16.5 Mb (ftp)(http) VCF
GSE229212_KW-106-5.vcf.gz 26.8 Mb (ftp)(http) VCF
GSE229212_Mouse_OnTarget_Summary_Table_1csv.csv.gz 3.8 Kb (ftp)(http) CSV
GSE229212_Mouse_OnTarget_Summary_Table_2.csv.gz 1.2 Kb (ftp)(http) CSV
GSE229212_PacBio_Results.csv.gz 659 b (ftp)(http) CSV
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