Targeting the casein kinase II /ikaros axis sensitizes TKI activity by suppression of GLUT1 expression and glycolysis in ph+ ALL

Abstract

Abstract Background The survival rate of Philadelphia (Ph) chromosome-positive acute lymphoblastic leukemia (Ph+ ALL) was significantly increased by tyrosine kinase inhibitors (TKIs); however, many Ph+ ALL patients were resistant to TKIs. IKZF1 deletions are common in Ph+ ALL, and the hyperphosphorylation of the IKZF1-encoding protein IKAROS, caused by casein kinase II (CK2), contributes to its malfunction in high-risk ALL. In ALL, IKAROS function is restored by the CK2 inhibitor CX-4945. Glycolysis aids in the development of chemoresistance and the growth of cancer. The underlying molecular mechanisms that link glycolysis to TKI chemoresistance in Ph+ ALL are still unknown, despite the metabolic process's critical importance in the context of malignant transformation. In this work, we examined the impact of CK2/IKAROS axis targeting on TKI sensitivity in Ph+ ALL and the underlying molecular mechanism by altering the disease's glycolysis. Methods The CCK8 and Annexin V apoptosis assays were carried out in SUP-B15 Ph+ B-ALL and patient samples. NCG mice that received intravenous injections of SUP-B15 cells were used to examine the effect of the drugs in vivo. For glycolysis, the seahorse test, the glucose consumption assay, and the lactic acid generation assay were applied. Results TKI resistance, particularly Imatinib, is observed in SUP-B15 Ph+ ALL with IKZF1 deletion.To test if restoring the IKAROS function by CK2 inhibition sensitizes the TKI activity, we examined the combined effect of TKIs with CX-4945. Results showed that the combination ofvarious doses of TKI Imatinib or Ponatinib with 2.5 μM (1/2 IC50) or 5 μM (IC50) CX-4945 showed significant synergistic effects on the cell proliferation arrest of Sup-B15 cells (CalcuSyn analysis CI < 1, and the Bliss model synergy score of 6.617 or 5.885, respectively). The combination of Imatinib or Ponatinib with IC50 CX-4945 significantly increased the apoptosis compared to single drug control in SUP-B15 (Imatinib: 13% vs. 50%, Ponatinib: 25% vs. 91%, P<0.05). Moreover, the combination of Imatinib or Ponatinib with 1.5 μM (1/2 IC50) or 3 μM (IC50) CX-4945 demonstrated the synergistic effect on Ph+ ALL patient samples (CI < 1, and the Bliss score of 3.173 or 3.757, respectively), and both Imatinib and Ponatinib combined with CX-4945 significantly promoted the apoptosis compared to the single drug control (P<0.001). Utilizing a human leukemia-xenograft model (SUP-B15-xenograft), the effect of CX-4945 on the sensitivity of Imatinib was also evaluated in vivo. The combination of Imatinib and CX-4945 considerably increased survival, reduced spleen weights and sizes, and reduced the % hCD45+hCD19+ Ph+ALL cells in the spleen and bone marrow of the mouse models when compared to single therapy controls. Taken together, these data indicate that CX-4945 sensitizes the TKI activity in Ph+ALL. Transcriptome analysis shows that GLUT1 is the top gene upon the two-drug treatment, and higher levels of GLUT1 expression are observed in the cohort of newly diagnosed Ph+ALL patients compared to the twenty normal BM controls. Additionally, patients who expressed GLUT1highversus GLUT1low had a considerably worse OS and RFS. IKAROS directly regulates GLUT1 transcription, and CX-4945 inhibits GLUT1 expression in an IKAROS-dependent manner. The combination treatment of CX-4945 with Imatinib or Ponatinib dramatically inhibits GLUT1 expression. In comparison to the single-drug controls, the Seahorse assay demonstrated that the combination of imatinib and CX-4945 decreased the extracellular acidification rate (ECAR), which includes glycolysis, glycolytic capacity, and the glycolytic reserve. Conclusions CX-4945 sensitizes the sensitivity of TKI activity in Ph+ALL, particularly with IKZF1 deletion, by affecting GLUT1 expression and glycolysis. Our findings reveal the potential of the new combination in the therapy of ALL patients.