Correction of beta654-thalassaemia mice using direct intravenous injection of siRNA and antisense RNA vectors.

摘要

Although the therapeutic efficacy of beta(654)-thalassaemia treatment using a combination of RNAi and antisense RNA to balance the synthesis of alpha- and beta-globin chains has been demonstrated previously, and the safety of lentiviral delivery remains unclear. Herein, we used the same beta(654)-thalassaemia mouse model to develop a therapy involving direct delivery of siRNA and antisense RNA plasmids via intravenous injection to simultaneously knock down alpha-globin transcript levels and restore correct beta-globin splicing. The amount of alpha-globin mRNAs in siRNA-treated MEL cells decreased significantly, and the properly spliced beta-globin mRNA was restored in HeLabeta(654) cells transfected with pcDNA-antisense plasmid. Furthermore, treatment of beta(654)-thalassaemic mice with siRNA and antisense RNA plasmids resulted in significant reduction of poikilocytosis and reticulocyte counts in blood samples, decreased nucleated cell populations in bone marrow, and reduced intrasinusoidal extramedullary haematopoiesis loci and iron accumulation in liver. RT-PCR analysis revealed that treatment resulted in down-regulation of alpha-globin mRNA synthesis by ~50% along with an increase in the presence of normally spliced beta-globin transcripts, indicating that the phenotypic changes observed in beta(654)-thalassaemic mice following treatment resulted from restoration of the balance of alpha/beta-globin biosynthesis.