The Pan-PIM Kinase Inhibitor LGB321 Strongly Increases Ibrutinib Effects in CLL, By Blocking Anti-Apoptotic Pathways and Microenvironmental Interactions

Despite remarkable anti-tumor activity of ibrutinib in CLL, some patients develop resistance due to acquired mutations. By adding drugs which target survival pathways and the CLL protective microenvironmental interactions, we aim to increase ibrutinib efficacy and to prevent ibrutinib resistance. The three PIM kinases are involved in important disease mechanisms in CLL, with PIM1 regulating CXCR4 surface expression impacting the interaction with the protective microenvironment, and PIM2/3 affecting the apoptotic machinery by regulating BAD.

Here, we investigated the effect of the Pan-PIM kinase inhibitor LGB321 in CLL in vitro and in vivo and its cooperative effect with ibrutinib. LGB321 was highly effective in inducing apoptosis in primary CLL cells, independent of prognostic factors or microenvironmental interactions, and reduced pBAD and total BAD protein levels in CLL. In vitro combination of ibrutinib and LGB321 demonstrated strong additive effects with doubled apoptosis rates compared to single treatment. Further, LGB321 treatment blocked the CXCR4/CXCL12 axis by dephosphorylating the CXCR4 receptor on Ser339, by reducing total CXCR4 protein levels, and by blocking the externalization of the CXCR4 receptor. Concordantly, LGB321 blocked CXCR4 functions regarding migration towards a CXCL12 gradient and homing of LGB321-pretreated primary CLL cells towards the bone marrow of NOG mice.

In vivo experiments confirmed the efficacy of LGB321 in 5 different CLL xenograft studies. Transplantation of primary CLL cells into NOG mice and treatment with LGB321 for 2 weeks strongly reduced WBC counts, spleen size and spleen infiltration with human CLL cells in all 5 CLL cases, and blocked BAD as well as CXCR4 phosphorylation also in vivo.

Xenograft studies combining ibrutinib with LGB321 demonstrated much faster reduction of WBC counts than ibrutinib single treatment and strongly reduced disease burden in the double treated mice, confirming the cooperative effects of those two inhibitors also in vivo.

Our results demonstrate that LGB321 might be an effective treatment option for CLL patients by impairing PIM2/3 mediated CLL-cell survival, and by blocking the PIM1/CXCR4-mediated interaction of CLL cells with the protective microenvironment. Furthermore, LGB321 enhances ibrutinib treatment effects and might help to increase its efficacy and to avoid the development of ibrutinib resistance.