Combination Therapy of Small Molecule Inhibitor PHA-739358 and Tyrosine Kinase Inhibitor Imatinib Yields Synergistic Antiproliferative Effects and Suppresses Emergence of Resistance of Chronic Myeloid Leukemia in Vitro

The success story of Imatinib (IM) in chronic myeloid leukemia (CML) is comprised by primary or acquired resistance to IM, insufficient therapeutic options for advanced stages of disease and limited effects on immature hematopoietic stem cells emphasizing the need for novel therapeutic strategies. We recently reported on PHA-739358, which shares activity of a pan-Aurora and an ABL kinase inhibitor. In this study, the individual contribution of both pathways to the effect of PHA-739358 was further investigated and a dose dependent reduction of BCR-ABL activity starting at very low concentrations was observed. In contrast, inhibition of the phosphorylation of Aurora kinase down stream target phospho-histone H3-Ser10 required higher amount of the inhibitor suggesting a predominant BCR-ABL mediated effect of PHA-739358 at low concentration. Simultaneous short-term treatment with PHA-739358 and IM resulted in pronounced apoptosis of wild type or low-grade IM-resistant BCR-ABL expressing cells while no such effects were observed in BCR-ABL negative or highly IM-resistant T315I mutants. In primary CD34+ cells of CML patients including non-dividing quiescent leukemic stem cells combination therapy with IM and PHA-739358 revealed a synergistic antiproliferativ activity which also affected immature CD34+38-cells. However, neither mono- nor combination therapy led to a significant induction of apoptosis. In addition, PHA-739358 did not influence quiescent stem cells analysed in the CSFE assays. Another focus of interest was the emergence of resistance secondary to treatment with PHA-739358, where resistant clones were found considerably less frequent than in IM-treated cells. Interestingly, PHA-739358 resistant cells did not reveal mutations in Aurora kinases A and B or in BCR-ABL kinase domain and remained IM-sensitive. Furthermore, simultaneous administration of PHA-739358 and IM considerably reduced the number of resistant clones and virtually abolished emergence of resistance at 2-times IC50 concentration of PHA- 739358 (plus IM at IC50). In conclusion, PHA-739358 potently inhibits proliferation of CD34+ stem und progenitor cells including immature CD38- subpopulation. However, in line with other tyrosine kinase inhibitors no induction of apoptosis in quiescent stem cells can be achieved. Remarkably, combination of PHA-739358 and IM prevents emergence of resistance rising the hope for a durable control of CML.