Correction of mouse models of sickle cell disease and beta-thalassemia using stem cell and genetic-based therapy

Inherited disorders of hemoglobin occur with a greater frequency than any other monogenic disease, with over 1,000 mutations now known to cause thalassemia. The recent development of mouse models for sickle cell disease (SCD) and beta thalassemia created an important reagent to test stem cell and gene-based therapies for correction of these diseases. Gene therapy for the correction of blood disorders has been considered since the first molecular characterization of a monogenic disorder, an A to T transversion in the sixth codon of beta-globin resulting in SCD. Early gene therapy vectors were derived from retroviruses because of their ability to integrate into the host genome. However, retroviruses require breakdown of the nuclear envelope for integration into chromatin. As a consequence, retroviral vectors can only deliver transgenes to actively dividing cells. Lentiviruses have redundant nuclear-localizing proteins that enable stable integration into nondividing cells. We anticipated that lentiviral vectors could be used to transduce the highly quiescent hematopoietic stem cell population. Work shown here demonstrates for the first time correction of a mouse model of SCD using unmobilized, highly purified bone marrow hematopoietic stem cells transduced with a self-inactivating (SIN) lentiviral vector. Future treatment of hemoglobinopathies in the clinic may proceed by the correction of embryonic stem (ES) cells derived from skin fibroblasts of the patient using nuclear transfer and reimplanted following their differentiation into hematopoietic stem cells. In anticipation of this strategy, we have corrected our mouse model of beta-thalassemia by rescuing the beta-globin defect in ES cells with a lentiviral vector and cloned the animals by tetraploid embryo complementation. This is the first step toward curing a hemoglobinopathy by "therapeutic cloning."...