scholarly journals High-Efficiency Transduction of Primary Human Hematopoietic Stem/Progenitor Cells by AAV6 Vectors: Strategies for Overcoming Donor-Variation and Implications in Genome Editing

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Chen Ling ◽  
Kanit Bhukhai ◽  
Zifei Yin ◽  
Mengqun Tan ◽  
Mervin C. Yoder ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Genome Editing ◽  
High Efficiency ◽  
Hematopoietic Stem ◽  
Donor Variation

scholarly journals High-Efficiency Transduction of Primary Human Hematopoietic Stem/Progenitor Cells By AAV6 Vectors: strategies for Overcoming Donor-Variation and Implications in Genome Editing

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 5889-5889
Author(s):  
Chen Ling ◽  
Kanit Bhukhai ◽  
Zifei Yin ◽  
Mengqun Tan ◽  
Mervin Yoder ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Genome Editing ◽  
High Efficiency ◽  
Ex Vivo ◽  
Transduction Efficiency ◽  
Triple Mutant ◽  
Hematopoietic Stem ◽  
Combined Use ◽  
Donor Variation ◽  
Viral Capsid Proteins

Abstract We have reported that of the 10 commonly used AAV serotype vectors, AAV6 is the most efficient in transducing primary human hematopoietic stem/progenitor cells (HSPCs). However, the transduction efficiency of the wild-type (WT) AAV6 vector varies greatly in HSPCs from different donors. Here we report two distinct strategies to further increase the transduction efficiency in HSPCs from donors that are transduced poorly with the WT AAV6 vectors. The first strategy involved modification of the viral capsid proteins where specific surface-exposed tyrosine (Y) and threonine (T) residues were mutagenized to generate a triple-mutant (Y705+Y731F+T492V) AAV6 vector. The second strategy involved the use of ex vivo transduction at high cell density, which revealed a novel mechanism, which we have termed, 'cross-transduction'. The combined use of these strategies resulted in transduction efficiency exceeding ~90% in HSPCs. Our studies have significant implications in the optimal use of capsid-optimized AAV6 vectors in genome editing in HSPCs. Disclosures Leboulch: bluebird bio: Patents & Royalties. Payen:bluebird bio: Patents & Royalties. Srivastava:AGTC; Genzyme: Patents & Royalties.

scholarly journals 10. High-Efficiency Transduction of Primary Human CD34+ Hematopoietic Stem/Progenitor Cells by AAV6 Serotype Vectors: Strategies for Overcoming Donor Variation and Implications in Genome Editing

2016 ◽  
Vol 24 ◽  
pp. S5-S6
Author(s):  
Zifei Yin ◽  
Kanit Bukhai ◽  
George Aslanidi ◽  
Chen Ling ◽  
Mengqun Tan ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Genome Editing ◽  
High Efficiency ◽  
Hematopoietic Stem ◽  
Human Cd34 ◽  
Donor Variation

scholarly journals Homologous recombination-based genome editing by clade F AAVs is inefficient in the absence of a targeted DNA break

2019 ◽  
Author(s):  
Geoffrey L. Rogers ◽  
Hsu-Yu Chen ◽  
Heidy Morales ◽  
Paula M. Cannon
Keyword(s):  
Homologous Recombination ◽  
Progenitor Cells ◽  
Genome Editing ◽  
High Efficiency ◽  
Zinc Finger Nucleases ◽  
Dna Breaks ◽  
Adeno Associated Virus ◽  
Hematopoietic Stem ◽  
Double Stranded Dna ◽  
Dna Break

AbstractAdeno-associated virus (AAV) vectors are frequently used as donor templates for genome editing by homologous recombination. Although modification rates are typically under 1%, they are greatly enhanced by targeted double-stranded DNA breaks (DSBs). A recent report described clade F AAVs mediating high-efficiency homologous recombination-based editing in the absence of DSBs. The clade F vectors included AAV9 and a series isolated from human hematopoietic stem/progenitor cells (HSPCs). We evaluated these vectors by packaging homology donors into AAV9 and an AAVHSC capsid and examining their ability to insert GFP at the CCR5 or AAVS1 loci in human HSPCs and cell lines. As a control we used AAV6, which effectively edits HSPCs, but only when combined with a targeted DSB. Each AAV vector promoted GFP insertion in the presence of matched CCR5 or AAVS1 zinc finger nucleases (ZFNs), but none supported detectable editing in the absence of the nucleases. Rates of editing with ZFNs correlated with transduction efficiencies for each vector, implying no differences in the ability of donor sequences delivered by the different vectors to direct genome editing. Our results therefore do not support that clade F AAVs can perform high efficiency genome editing in the absence of a DSB.

Delivery of Genome Editing Reagents to Hematopoietic Stem/Progenitor Cells

2016 ◽  
Vol 36 (1) ◽  
Author(s):  
Megan D. Hoban ◽  
Zulema Romero ◽  
Gregory J. Cost ◽  
Matthew Mendel ◽  
Michael Holmes ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Genome Editing ◽  
Hematopoietic Stem

Adipose tissue is an extramedullary reservoir for functional hematopoietic stem and progenitor cells

Blood ◽  
2010 ◽  
Vol 115 (5) ◽  
pp. 957-964 ◽  
Author(s):  
Jinah Han ◽  
Young Jun Koh ◽  
Hye Rin Moon ◽  
Hyun Gee Ryoo ◽  
Chung-Hyun Cho ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Progenitor Cells ◽  
High Efficiency ◽  
Stromal Vascular Fraction ◽  
Hematopoietic Stem ◽  
Blood Transplantation ◽  
Variable Extent ◽  
Stem And Progenitor Cells

Abstract The stromal vascular fraction (SVF) in adipose tissue contains a pool of various stem and progenitor cells, but the existence of hematopoietic stem and progenitor cells (HSPCs) in the SVF has not been seriously considered. We detected the presence of HSPCs in the SVF by phenotypically probing with Lin−Sca-1+c-kit+ (LSK) and functionally confirming the presence using colony-forming cell assay and assessing the long-term multilineage reconstitution ability after SVF transplantation. The LSK population in the SVF was 0.004% plus or minus 0.001%, and 5 × 105 freshly isolated SVF cells gave rise to 13 plus or minus 4 multilineage colonies. In addition, 0.15% plus or minus 0.03% of SVF cells was home to bone marrow (BM), especially near vascular and endosteal regions, 24 hours after blood transplantation. SVF transplantation was capable of generating a long-term (> 16 weeks), but variable extent (2.1%-32.1%) multilineage reconstitution in primary recipients, which was subsequently transferred to the secondary recipients by BM transplantation. All HSPCs within the SVF originated from the BM. Furthermore, the granulocyte–colony-stimulating factor (G-CSF) mobilization of HSPCs from BM markedly elevated the number of phenotypic and functional HSPCs in the SVF, which induced a high efficiency long-term reconstitution in multilineage hematopoiesis in vivo. Our results provide compelling evidence that adipose tissue is a novel extramedullary tissue possessing phenotypic and functional HSPCs.

Understanding and overcoming adverse consequences of genome editing on hematopoietic stem and progenitor cells

2021 ◽  
Author(s):  
Byung-Chul Lee ◽  
Richard J. Lozano ◽  
Cynthia E. Dunbar
Keyword(s):  
Progenitor Cells ◽  
Genome Editing ◽  
Hematopoietic Stem ◽  
Stem And Progenitor Cells

scholarly journals 124. Cas9-Mediated Genome Editing in Hematopoietic Stem/Progenitor Cells

2015 ◽  
Vol 23 ◽  
pp. S51
Author(s):  
Jennifer L. Gori ◽  
G. Grant Welstead ◽  
McKensie A. Collins ◽  
Justin W. Fang ◽  
Ari E. Friedland ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Genome Editing ◽  
Hematopoietic Stem

scholarly journals Global Transcriptional Response to CRISPR/Cas9-AAV6-Based Genome Editing in CD34+ Hematopoietic Stem and Progenitor Cells

2018 ◽  
Vol 26 (10) ◽  
pp. 2431-2442 ◽  
Author(s):  
M. Kyle Cromer ◽  
Sriram Vaidyanathan ◽  
Daniel E. Ryan ◽  
Bo Curry ◽  
Anne Bergstrom Lucas ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Genome Editing ◽  
Transcriptional Response ◽  
Hematopoietic Stem ◽  
Stem And Progenitor Cells
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