A lentiviral vector (CCLc- ?AS3-FB {?AS3LV}) is being investigated for the treatment of severe sickle cell disease, however, it suffers from low titer, sub-optimal gene transfer to CD34+ hematopoietic stem cells (HSCs), and expression likely insufficient to definitively cure ?-thalassemia (although sufficient to prevent sickling in pre-clinical studies). We hypothesize that there are known and unknown human ?-globin genomic sequences within ?AS3LV that are inhibiting vector performance. Studies outlined in this proposal will investigate how removal and/or addition of known or unknown elements within ?AS3LV's human ?-globin genomic sequences affect titer, gene delivery to HSCs, and expression of the anti-sickling ?AS3-globin gene. The outcome of these studies will provide insight into how specific regulatory elements influence the performance of ?AS3LV across multiple categories. Moreover, this research will yield a second generation of improved lentiviral vectors for efficiently transferring and effectively expressing the anti-sickling ?AS3-globin gene for gene therapy of sickle cell disease.
This proposal will provide insight into how combinations of specific regulatory elements influence the performance of ?-globin lentiviral vectors across multiple categories. These studies will produce a ?-globin expression vector with expression levels similar to that of the endogenous ?-globin allele and may also reveal some functional insight into how the human ?-globin Locus Control Region, a regulatory element that influences the expression of globin genes, functions at a level not previously studied.
| Morgan, Richard A; Gray, David; Lomova, Anastasia et al. (2017) Hematopoietic Stem Cell Gene Therapy: Progress and Lessons Learned. Cell Stem Cell 21:574-590 |
