Improving Gene Editing in Blood Stem Cells with DNA Barcoding
Updated
Abstract
A barcoded AAV6 donor template (BC-AAV) was developed to monitor edited hematopoietic stem and progenitor cells (HSPCs) following transplantation.
- Edited HSPCs transplanted into immunodeficient mice showed hematopoiesis driven by a limited number of dominant clones despite initial barcode diversity.
- The engraftment pattern of gene-edited cells suggested that editing does not alter clonal dynamics in this model.
- Key improvements, including GMP-grade StemSpan AOF medium and StemRegenin-1, increased clonal diversity while preserving hematopoietic potential.
- The NHEJ inhibitor AZD-7648 significantly enhanced editing efficiency.
- A shorter transduction period improved engraftment and clonal balance without compromising editing outcomes.
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Funding
Competing interests
J.-C.S. is a consultant in Rocket Pharmaceuticals and is co-founder and holds equity in DanausGT Biotechnology. M.P. is on the Scientific Advisory Board of Allogene Therapeutics, holds equity in CRISPR Therapeutics, and is a founder and holds equity in Kamau Therapeutics—none of these entities were involved in these studies nor have any rights, options, nor rights to the work described. L-J.S., J.B. are employees at DanausGT Biotechnology. C.T. and V.L. were employed by Viralgen SL when the present research was conducted. The IT code used in this work is under intellectual property.
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