B-cell chronic lymphocytic leukemia (CLL) is the most common adult human leukemia. The pathogenetic mechanisms involved in development of CLL are unknown. Several studies demonstrated the central role of deletion of the region 13q14.3 involving microRNA cluster mir15a and mir16-1 that downregulated in more than 60% of the patients. In this study, we investigated the effect of the replacement of microRNA 15 and 16 in CLL cells with del13q14 by transfection of miRNA mimics into CLL cells in vitro and in a NOD/Shi-scid,γcnull(NGS) mice in vivo. Administration of miRNA mimics determined a decrease of CLL cells viability in vitro, with exception of certain CLL bearing TP53 mutated. Transfection of miRNA mimics in NGS mice, previously injected with CLL cells, resulted in substantial tumor regression. In contrast, transfection of miRNA inhibitors in CLL cells without del13q14 and variable expression of miR15 and miR16, lead to an increase of cells viability in vitro and an enhanced capacity of CLL cells to growth in vivo. These data demonstrate the capacity of the mir15-16 cluster to control clonal expansion providing indications for a target therapeutic strategy based on microRNAs.