Novel Self-Inactivating Vectors for Reconstitution of Wiskott-Aldrich Syndrome

Wiskott-Aldrich syndrome (WAS) is a life-threatening disorder characterized by immunodeficiency, thrombocytopenia and eczema. Hematopoietic stem cell gene therapy can cure WAS but remains associated with the risk of leukemogenesis. In an effort to decrease the risk of gene-therapy induced leukemia associated with the use of first generation gamma-retroviral vectors, we have developed a series of codon-optimized WASP-expressing alpha-, gammaretro- and lentiviral selfinactivating vectors and have performed comparative studies with regard to expression, vector dose and genotoxicity in vitro. In comparison to the conventional LTR-driven gammaretroviral vector, the self-inactivating (SIN) Elongation Factor 1 short form (EFS) - driven vectors presented decreased expression potency per vector copy. However, the safety profile of SIN vectors was superior, as evidenced by decreased genotoxicity in a murine hematopoietic progenitor cell - based in vitro immortalization assay. Using an in vitro OP9 stroma cell - based co-culture system, we provide evidence that a codon-optimized lentiviral vector improves the B-cell differentiation block characteristic of WAS. Taken together, our studies represent a further step towards the development of a novel codon-optimized coWASP SIN vector for ex-vivo HSC gene therapy for Wiskott-Aldrich syndrome.