Treatment of β654‐thalassaemia by TALENs in a mouse model.

Objectives: This study explored whether TALENs‐mediated non‐homologous end joining (NHEJ) targeting the mutation site can correct the aberrant β‐globin RNA splicing, and ameliorate the β‐thalassaemia phenotype in β654 mice. Material and methods: TALENs vectors targeted to the human β‐globin gene (HBB) IVS2‐654C >T mutation in a mouse model were constructed and selected to generate double heterozygous TALENs+/β654 mice. The gene editing and off‐target effects were analysed by sequencing analysis. β‐globin expression was identified by RT‐PCR and Western blot analysis. Various clinical indices including haematologic parameters and tissue pathology were examined to determine the therapeutic effect in these TALENs+/β654 mice. Results: Sequencing analysis revealed that the HBBIVS2‐654C >T point mutation was deleted in over 50% of the TALENs+/β654 mice tested, and off‐target effects were not detected. RT‐PCR and Western blot analysis confirmed the expression of normal β‐globin in TALENs+/β654 mice. The haematologic parameters were significantly improved as compared with their affected littermates. The proportion of nucleated cells in bone marrow was considerably decreased, splenomegaly with extramedullary haematopoiesis was reduced, and significant decreases in iron deposition were seen in spleen and liver of the TALENs+/β654 mice. Conclusion: These results suggest effective treatment of the anaemia phenotype in TALENs+/β654 mice following deletion of the mutation site by TALENs, demonstrating a simple and straightforward strategy for gene therapy of β654‐thalassaemia in the future. [ABSTRACT FROM AUTHOR]