Pro‐survival signal inhibition by CDK inhibitor dinaciclib in Chronic Lymphocytic Leukaemia

Abstract

Short title: Effects of Dinaciclib on pro-survival signals in CLL. 2 SUMMARYDinaciclib is a cyclin-dependent kinase inhibitor with clinical potential in different cancers, including Chronic Lymphocytic Leukemia (CLL). In order to better understand its cytotoxic action, we characterized its effects on signalling pathways important for the survival of CLL cells. We found that dinaciclib induced apoptosis through the activation of caspases 8 and 9, which was independent of the presence of cytokines to mimic the environment of proliferation centres or IGVH mutation status. Moreover, treatment with dinaciclib led to the inhibition of oncogenic pathways normally activated in stimulated CLL cells, such as STAT3, NF-κB and p38. Also, PI3K/AKT and RAF/MEK/ERK pathways showed a transient and early activation, followed by a later, permanent inhibition. Dinaciclib was also able to block the expression of anti-apoptotic proteins of the BCL-2 family such as MCL-1 and BCL-xL. Finally, we show that low concentrations of dinaciclib enhanced cell sensitivity to ibrutinib and BCL-2 inhibitor ABT-199, two drugs with known effects on CLL. Taken together, our data show that dinaciclib targets multiple pro-survival signalling pathways in CLL, which provides a mechanistic explanation for its potent induction of apoptosis. They also support a therapeutic application of CDK inhibitors in CLL in combination with other relevant targeted therapies.

CUDC‐907 blocks multiple pro‐survival signals and abrogates microenvironment protection in CLL

“…Chronic lymphocytic leukaemia cells were cultured in RPMI‐1640 medium containing soluble 10 ng/mL interleukin (IL)‐4 and CD40 ligand (CD40L or CD154) to mimic the microenvironment of proliferation centres as previously described . Cells were incubated for 24 hours in these conditions before applying any treatments.…”

“…Cells were incubated for 24 hours in these conditions before applying any treatments. Human CLL cell line MEC‐1 was cultured as previous described . Goat F(ab’)2 anti‐human IgM was purchased from Bio‐Rad (Hercules, CA, USA), recombinant human BAFF (soluble) was purchased from Enzo (Farmingdale, NY, USA).…”

“…Cell viability was assessed by the CellTiter 96 Aqueous One Solution Cell Proliferation MTS Assay (Promega, Madison, WI, USA), following the manufacturer's instructions as previously described . The absorbance at 490 nm was recorded on a TECAN infinite F50 reader (Labtech International, Heathfield, UK).…”

“…Multiple signalling pathways, including PI3K/AKT, RAF/MEK/ERK, STAT3, and NF‐κB, are constitutively activated in CLL cells . Given the pivotal role of PI3K/AKT in cancer initiation, growth, proliferation, and survival, an increasing number of small molecular inhibitors that target this pathway have been developed.…”

Differential activation of pro‐survival pathways in response to CD40LG/IL4 stimulation in chronic lymphocytic leukaemia cells

“…Chronic lymphocytic leukaemia (CLL) is characterized by the progressive accumulation of mature monoclonal B lymphocytes, which are dependent on the microenvironment of proliferation centres for survival (Burger et al , ). The pro‐survival signals activated in these tissues include the ERK, AKT, STAT3 and NF‐κB pathways (Chen et al , ). The secretion of the T cell chemokines CCL3/4/17/22 by CLL cells is also important for proliferation (Burger, ).…”