Aurora A Kinase As a Promising Therapeutic Target in Philadelphia Chromosome Positive Leukemia Cells

Introduction: ABL tyrosine kinase inhibitor (TKI), imatinib and second-generation ABL TKIs, such as nilotinib and dasatinib have demonstrated the potency against chronic myeloid leukemia (CML) and Philadelphia chromosome (Ph) positive acute lymphoblastic leukemia (Ph+ALL) patients. However, resistance to ABL TKI can develop in CML patients due to BCR-ABL point mutations. Moreover, ABL TKIs do not eliminate the leukemia stem cells (LSCs). Residual CML cells are present in bone marrow microenvironment. These cells are contained within a niche in the bone marrow and are often impervious to current treatments. Aurora kinases are novel family of serine/threonine kinases that play a critical role in regulating many of the processes that are pivotal to mitosis. Aurora kinases comprises three members, designated aurora A, B, and C. Amplification of aurora kinase gene is oncogenic and has been observed in several cancers including hematologic malignancies. Therefore, new approach to the treatment for Ph-positive cells with aurora kinase inhibitors may improve the outcome of leukemia patients.

Materials and methods: In this study, we investigated whether aurora kinase inhibitor could suppress Ph-positive leukemia cells including T315I mutation and primary samples.

Results: One of aurora kinase A inhibitor, alisertib inhibits proliferation of leukemia cells. We found alisertib treatment inhibited the growth of Ph-positive leukemia cells in a dose dependent manner and phosphorylation of aurora kinase A was reduced. Alisertib inhibits proliferation in imatinib resistant cells including T315I mutation. In the cell cycle analysis, alisertib induces a G2/M phase arrest in Ph-positive cells. Intracellular ROS levels were also increased in a dose dependent manner. Combined treatment of Ph-positive cells with ABL tyrosine kinase inhibitors, imatinib or ponatinib and alisertib caused significantly more cytotoxicity than each drug alone. Apoptotic and Caspase 3/7 positive cells were also increased. Ph-positive cells were treated with alisertib and cellular senescence was determined by senescence- associated β-galactosidase (SA-β-gal) staining. Senescent cells were increased in a time and dose dependent manner. SA-β-gal positive cells were reduced by N-Acetyl-L-cysteine treatment. Small interfering RNA mediated knockdown of the Aurora A gene causes a proliferation defect and enhances ABL TKI activity in Ph-positive cells. We next examined tumor formation in mice model. We injected subcutaneously 1×10⁷ T315I mutant cells in nude mice. A dose of 20 mg/kg/day p.o of ponatinib and 30 mg/kg/day p.o of alisertib inhibited tumor growth and reduced tumor volume compared with control mice. We also found that co-treatment with ponatinib and alisertib increased mouse survival. Co-treatment with ponatinib and alisertib showed reduced mouse spleen weight and size. The treatments were well tolerated with no animal health concerns observed.

Conclusion: The results of our study indicate that the aurora kinase inhibitor may be a powerful strategy against ABL TKI resistant cells including T315I mutation and enhance cytotoxic effects of ABL TKI against those Ph-positive leukemia cells.