Evidence of off-target effects of bosutinib that promote retinoic acid-induced differentiation of non-APL AML cells.

In the present study, we determined the effects of the Src family kinase (SFK) inhibitor, Bosutinib, and the engineered loss of the Lyn SFK on all-trans retinoic acid-induced leukemic cell differentiation. Retinoic acid (RA) is an embryonic morphogen and dietary factor that demonstrates chemotherapeutic efficacy in inducing differentiation of a non-APL AML cell model, the HL-60 human myeloblastic (FAB-M2) leukemia cell line, via activation of a novel signalsome containing an ensemble of signaling molecules that drive differentiation. Bosutinib is an inhibitor of SFKs used to treat myeloid leukemias where prominent high expression of SFKs, in particular Lyn, has been observed. Using either Bosutinib or loss of Lyn expression due to shRNA promoted RA-induced phenotypic differentiation, G0 arrest, and respiratory burst (functional differentiation) of HL-60 cells. Signaling events putatively seminal to RA-induced differentiation, the expression of Fgr, Cbl, Slp-76 and Vav, and the phosphorylation of c-Raf (pS259), Vav (p-tyr), and Slp76 (p-tyr) were not inhibited by Bosutinib or loss of Lyn. Nor was RA-induced upregulation of p-tyr phosphorylation of p47phox, a member of the NADPH complex that produces ROS, a putative phosphorylation dependent signaling regulator. Surprisingly, Bosutinib still works in the absence of Lyn to enhance RA-induced differentiation and neither compromised RA-induced expression, nor phosphorylation of signaling molecules that drive differentiation. These findings suggested there is a novel, off-target, Lyn-independent effect of Bosutinib that is of therapeutic significance to differentiation therapy.