UBC Theses and Dissertations
Targeting tyrosine kinase inhibitor-insensitive chronic myeloid leukemia stem/progenitor cells by effective inhibition of a novel PP2A-AHI-1-BCR-ABL-JAK2-complex Lai, Damian
Imatinib Mesylate (IM) and other tyrosine kinase inhibitors (TKIs) have had a major impact on treatment of early phase Chronic Myeloid Leukemia (CML) patients. However, TKI monotherapies are not curative and initial and acquired resistance remain challenges. Particularly, CML stem cells are less responsive to TKIs and are a critical target population for TKI resistance. Thus, improved treatments targeting key elements active in CML stem cells are needed. One candidate is Abelson helper integration site-1 (AHI-1), an oncogene that is highly upregulated in CML stem cells and interacts with multiple kinases, including BCR-ABL and JAK2. AHI-1-mediated complexes regulate TKI response/resistance of CML stem/progenitor cells, indicating that AHI-1 is a new therapeutic target in CML. By screening the Prestwick Chemical Library, a specific growth inhibitory compound that potentially targets AHI-1 was identified: Cantharidin (CAN), an inhibitor of protein phosphatase 2A (PP2A). CAN is toxic however, so two new PP2A inhibitors, LB100 and LB102, were identified for this study. These new inhibitors specifically inhibit PP2A activity and suppress growth of CML cell lines. Importantly, these new PP2A inhibitors selectively target CML stem/progenitor cells while sparing healthy stem/progenitor cells. When combined with TKIs there is significant further suppression of growth in cell lines and in CD34+ treatment-naïve IM-nonresponder cells. Furthermore, this combination effect was determined to be synergistic. Cell cycle analysis showed that treatment with PP2A inhibitors alone induced a shift from G1 to G2/M phase. Confocal microscopy confirmed that the G2/M arrest led to mitotic catastrophe. However a similar shift in cell population was observed after combination with IM, suggesting that the G2/M phase arrest is solely due to PP2A inhibition. Mechanistically, the PP2A-PR55α subunit was identified as a new AHI-1 interacting protein. Western blot analysis showed that, compared to single agents, the combination treatment greatly suppresses protein expression of AHI-1, BCR-ABL, JAK2, STAT5, AKT, β-catenin, P-38 and JNK. The combination treatment also affected PP2A and BCR-ABL-mediated β-catenin dephosphorylation/phosphorylation. These results indicate that simultaneously targeting both BCR-ABL and PP2A activities in CML stem/progenitor cells may provide a novel treatment option for CML patients, through destabilization of the protein-protein interactions mediated by AHI-1.
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