Potentially therapeutic levels of anti-sickling globin gene expression following lentivirus-mediated gene transfer in sickle cell disease bone marrow CD34+ cells

Abstract

Sickle cell disease (SCD) can be cured by allogeneic hematopoietic stem cell (HSC) transplant. However, this is only possible when a matched donor is available making the development of gene therapy using autologous HSCs a highly desired alternative. We used a culture model of human erythropoiesis to directly compare two insulated, self-inactivating, and erythroid-specific lentiviral vectors, encoding for γ-globin (V5m3-400) or a modified β-globin (βAS3-FB) for production of anti-sickling hemoglobin (Hb) and correction of red cell deformability after deoxygenation. Bone marrow CD34+ cells from three SCD patients were transduced using V5m3-400 or βAS3-FB and compared to mock transduced SCD or healthy donor CD34+ cells. Lentiviral transduction did not impair cell growth or differentiation, as gauged by proliferation and acquisition of erythroid markers. Vector copy number averaged ~1 copy per cell and corrective globin mRNA levels were increased more than 7-fold over mock-transduced controls. Erythroblasts derived from healthy donor and mock-transduced SCD cells produced a low level of HbF that was increased to 23.6 ± 4.1% per vector copy for cells transduced with V5m3-400. Equivalent levels of modified HbA of 17.6 ± 3.8% per vector copy were detected for SCD cells transduced with βAS3-FB. These levels of anti-sickling Hb production were sufficient to reduce sickling of terminal stage RBCs upon deoxygenation. We conclude that the achieved levels of HbF and modified HbA would likely prove therapeutic to SCD patients who lack matched donors.

Gene Insertion Into Genomic Safe Harbors for Human Gene Therapy

“…Globin genes with antisickling properties present such an example. 57 Apart from integration of therapeutic transgenes, GSHs could support addition of exogenous genes that confer new properties to cells. These include drug resistance genes to select gene-modified cells 58 and tumor antigen receptor genes, such as chimeric antigen receptor genes, to mediate antitumor effects.…”

Amelioration of murine sickle cell disease by nonablative conditioning and γ-globin gene-corrected bone marrow cells

“…We demonstrated that transduction of SCD BM CD34 + cells with this vector reduced deoxygenation-induced sickling of in vitro -derived RBC progeny. 31 To further support use of the V5m3-400 vector in SCD clinical trials, we tested whether SCD mice receiving nonablative irradiation would engraft with γ-globin gene-corrected HSCs to a level sufficient for a therapeutic effect. Transplanted animals averaged 35% HSC chimerism and had HbF levels ≥ 20% in circulating RBCs, which improved anemia and reduced SCD-related pathologies.…”

Gene-modified, cell-based therapies—an overview

“…[44]. One of the most successful platforms of chimeric antigen receptor T cells for malignancy uses lentiviral vectors to deliver the transgene [51], and other cell products like dendritic cells [52] and CD34+ cells [53] have also been modified with these vectors.…”