The Heat Shock Protein 32 (Hsp32)/HO-1-Targeting Drug SMA-ZnPP and the Triterpenoid CDDO-Me Exert Synergistic Growth-Inhibitory Effects on TKI-Resistant Leukemic Cells in Ph+ CML

Abstract 4414

Resistance against one or more tyrosine kinase inhibitors (TKI) prevents eradication of Ph+ chronic myeloid leukemia (CML). In many patients BCR/ABL1 mutations are detectable. We have recently identified two targeted drugs that exert major growth-inhibitory effects on drug-resistant CML cells, the triterpenoid CDDO-Me (Bardoxolone-methyl, REATA Pharma) that blocks several signalling molecules including mTOR, Akt, and STAT3, and upregulates expression of heat shock protein 32 (Hsp32 = heme oxygenase 1, HO-1), and styrene-maleic acid-copolymer micelle-encapsulated ZnPP (SMA-ZnPP), a water-soluble inhibitor of Hsp32/HO-1. In the current project, we asked whether CDDO-Me exerts inhibitory effects on growth of TKI-resistant CML cells and whether the combination of CDDO-Me and SMA-ZnPP would produce synergistic effects in drug-resistant CML cells. As determined by ³H-thymidine incorporation, CDDO-Me was found to inhibit the proliferation of imatinib-responsive and imatinib-resistant K562, imatinib-resistant KCL-22, KU812, and Ba/F3 cells transfected with various TKI-resistant mutants of BCR/ABL1 (T315I, E255K, Y253F, H396P). In each case, IC₅₀ values <1 μM were obtained without major differences between imatinib-responsive and imatinib-resistant cells. Growth-inhibition was accompanied by apoptosis as assessed by combined AnnexinV/PI staining as well as by an increase in expression of HO-1 in KU812 and KCL-22 cells. CDDO-Me was also found to inhibit proliferation of leukemic cells in all patients with TKI-resistant CML (n=4), with IC₅₀ values ranging between <0.1 and 0.5 μM. No differences in IC₅₀ values were observed between treatment-naïve and TKI-resistant cells. Next, we applied the combination CDDO-Me+SMA-ZnPP and found that this combination acts highly synergistically on imatinib-responsive and imatinib-resistant K562 cells as well as primary CML cells isolated from imatinib-naïve CML patients (n=2) or from patients with imatinib-resistant CML (n=2), including one patient in whom BCR/ABL1 T315I was detected. We also examined whether CDDO-Me would exert synergistic effects on CML cells when combined with BCR/ABL1 TKI. In these experiments, we applied the combinations CDDO-Me+dasatinib and CDDO-Me+nilotinib on K562 cells. Both combinations were found to synergistically induce growth inhibition. In conclusion, CDDO-Me inhibits the proliferation of imatinib-resistant BCR/ABL1+ cells, including primary CML cells isolated from untreated patients and cells derived from patients with TKI-resistant CML carrying the BCR/ABL1 mutant T315I. Our data also show that CDDO-Me + SMA-ZnPP and CDDO-Me + BCR/ABL1 TKI synergize in producing growth inhibition in CML cells. Whether these drug combinations also produce synergistic effects in vivo in patients with TKI-resistant CML remains to be evaluated.