Ibrutinib synergizes with MDM-2 inhibitors in promoting cytotoxicity in B chronic lymphocytic leukemia

Objective: The aim of this study was to investigate the anti-leukemic activity of the Bruton tyrosine kinase inhibitor Ibrutinib in combination with the small molecule MDM-2 inhibitor Nutlin-3 in preclinical models. Methods: The potential efficacy of the Ibrutinib/Nutlin-3 combination was evaluated in vitro in a panel of B leukemic cell lines (EHEB, JVM-2, JVM-3, MEC-1, MEC-2) and in primary B-chronic lymphocytic leukemia (B-CLL) patient samples, by assessing cell viability, cell cycle profile, apoptosis and intracellular pathway modulations. Validation of the combination therapy was assessed in a B leukemic xenograft mouse model. Results: Ibrutinib exhibited variable anti-leukemic activity in vitro and the combination with Nutlin-3 synergistically enhanced the induction of apoptosis independently from the p53 status. Indeed, the Ibrutinib/Nutlin-3 combination was effective in promoting cytotoxicity also in primary B-CLL samples carrying 17p13 deletion and/or TP53 mutations, already in therapy with Ibrutinib. Molecular analyses performed on both B-leukemic cell lines as well as on primary B-CLL samples, while confirming the switch-offof the MAPK and PI3K pro-survival pathways by Ibrutinib, indicated that the synergism of action with Nutlin-3 was independent by p53 pathway and was accompanied by the activation of the DNA damage cascade signaling through the phosphorylation of the histone protein H2A.X. This observation was confirmed also in the JVM-2 B leukemic xenograft mouse model. Conclusions: Taken together, our data emphasize that the Ibrutinib/Nutlin-3 combination merits to be further evaluated as a therapeutic option for B-CLL.