An Optimized Lentiviral Vector Efficiently Corrects the Human Sickle Cell Disease Phenotype

Molecular Therapy. Methods & Clinical Development
Leslie WeberAnnarita Miccio

Abstract

Autologous transplantation of hematopoietic stem cells transduced with a lentiviral vector (LV) expressing an anti-sickling HBB variant is a potential treatment for sickle cell disease (SCD). With a clinical trial as our ultimate goal, we generated LV constructs containing an anti-sickling HBB transgene (HBBAS3), a minimal HBB promoter, and different combinations of DNase I hypersensitive sites (HSs) from the locus control region (LCR). Hematopoietic stem progenitor cells (HSPCs) from SCD patients were transduced with LVs containing either HS2 and HS3 (β-AS3) or HS2, HS3, and HS4 (β-AS3 HS4). The inclusion of the HS4 element drastically reduced vector titer and infectivity in HSPCs, with negligible improvement of transgene expression. Conversely, the LV containing only HS2 and HS3 was able to efficiently transduce SCD bone marrow and Plerixafor-mobilized HSPCs, with anti-sickling HBB representing up to ∼60% of the total HBB-like chains. The expression of the anti-sickling HBB and the reduced incorporation of the βS-chain in hemoglobin tetramers allowed up to 50% reduction in the frequency of RBC sickling under hypoxic conditions. Together, these results demonstrate the ability of a high-titer LV to express elevated levels of a ...Continue Reading

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Citations

Feb 19, 2019·Molecular Therapy. Methods & Clinical Development·Simone Merlin, Antonia Follenzi
Aug 29, 2021·Viruses·Valentina Poletti, Fulvio Mavilio
May 20, 2020·Molecular Therapy. Methods & Clinical Development·Richard A MorganDonald B Kohn
Aug 22, 2021·Molecular Therapy : the Journal of the American Society of Gene Therapy·Sophie RamadierAnnarita Miccio

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Methods Mentioned

BETA
transfection
flow cytometry
PCR

Clinical Trials Mentioned

NCT02212535

Software Mentioned

Plerixafor
Prism4
GraphPad
ImageJ
Kaluza
LC Solution

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