TAL effector nucleases (TALENs) represent a new class of artificial nucleases capable of cleaving long, specific target DNA sequences in vivo and are powerful tools for genome editing with potential therapeutic applications. Here we report a pair of custom-designed TALENs for targeted genetic correction of the sickle cell disease mutation in human cells, which represents an example of engineered TALENs capable of recognizing and cleaving a human disease-associated gene. By using a yeast reporter system, a systematic study was carried out to optimize TALEN architecture for maximal in vivo cleavage efficiency. In contrast to the previous reports, the engineered TALENs were capable of recognizing and cleaving target binding sites preceded by A, C or G. More importantly, the optimized TALENs efficiently cleaved a target sequence within the human β-globin (HBB) gene associated with sickle cell disease and increased the efficiency of targeted gene repair by >1000-fold in human cells. In addition, these TALENs showed no detectable cytotoxicity. These results demonstrate the potential of optimized TALENs as a powerful genome editing tool for therapeutic applications.