Cyclosporin H Improves the Transduction of CD34+ Cells with an Anti-Sickling Globin Vector, a Possible Therapeutic Approach for Sickle Cell Disease.

Sickle cell disease (SCD) is a debilitating monogenic disease originating from mutations in the hemoglobin beta chain gene producing an abnormal hemoglobin HbS. The polymerization of HbS is responsible for the sickling of erythrocytes leading to anemia and vaso-occlusive events. Gene therapy is a promising treatment of SCD, and two different gene therapy drugs, using gene editing or gene transfer, have already reached the marketing stage. There is still a need to improve the efficacy of gene therapy in SCD, particularly when using anti-sickling beta-globin gene transfer strategies, which must outcompete the pathological HbS. One possibility is to increase transduction by inhibiting lentiviral restriction factors such as interferon-induced transmembrane proteins (IFITMs). This can be achieved by the addition of cyclosporin H (CsH) during the transduction process. This strategy was applied here in CD34⁺ hematopoietic progenitor and stem cells obtained from cord blood (CB). A first series of experiments with lentiviral vector coding for a green fluorescent protein (GFP) gene confirmed that the addition of CsH enhanced transgene expression levels and vector copy number per cell (VCN), while CD34⁺ cells remained viable and functional. Notably, the production of colony-forming cells (CFC) remained unaffected unless very high VCN values were reached. In a second step, CD34⁺ cells obtained from the CB of newborns with homozygous (n = 2) or heterozygous (n = 1) SCD mutations were transduced with the GLOBE-AS3 lentiviral vector coding for the HbAS3 anti-sickling beta globin. As with GFP, GLOBE-AS3 lentiviral transduction was clearly enhanced by CsH, leading to VCN > 2 and therapeutic levels of expression of the HbAS3. Moreover, the process did not affect the viability or functions of CFC. The combination of CB progenitors, the GLOBE-AS3 vector, and CsH is thus shown here to be a promising approach for the treatment of SCD.