Arsenic Trioxide Sensitizes Glucocorticoid-Resistant T-Cell Acute Lymphoblastic Leukemia to Dexamethasone through Inactivation of AKT and Downregulation of XIAP.

Arsenic trioxide (ATO) is a very effective agent for the treatment of acute promyelocytic leukemia (APL). Because of its pro-apoptotic activity already at low dosage, ATO may be beneficial for combination treatment of drug-resistant disease to augment the effect of other chemotherapeutic agents. In childhood acute lymphoblastic leukemia (ALL), de novo resistance to glucocorticoids (GC) is markedly associated with poor outcome. As a mechanism of action, GCs induce apoptosis in steroid-sensitive leukemic cells. Recent evidence suggests that GC-resistance is associated with altered levels of pro- and anti-apoptotic regulatory proteins. We therefore evaluated the potential of ATO as a chemo-sensitizer in a model of GC-resistant ALL. As expected, ATO effectively induced cell death in ALL cell lines and in primary ALL patient samples at clinically relevant concentrations (IC50 values between 0.4–1.2 uM). Interestingly, ATO re-sensitized the GC-resistant ALL lines CEM C1-15, MOLT-4 and Jurkat to dexamethasone at doses that do not affect cell survival as single agent. The same effect was observed using primary cells from two patients with GC-resistant T-cell ALL. The sensitizing effect of ATO could not be observed in combination with the standard ALL chemotherapeutic agents daunorubicin, asparaginase and vincristine, suggesting that a mechanism specifically relevant for GC-resistance is involved. ATO, alone or at lower dose in combination with dexamethasone, induced cell death by activating caspase-3 and −8, which resulted in PARP cleavage. We then investigated the MAPK and PI3K/AKT pathways as candidate mechanisms of action with key apoptosis regulators such as Bad and Bcl-2 as targets. The MEK1/2 inhibitor PD98059 only marginally augmented the response to dexamethasone, suggesting that this pathway would not play a major role for GC sensitization. In contrast, we found that low dose ATO treatment resulted in a rapid decrease of AKT phosphorylation at the activating serine 473 phosphorylation site, while AKT levels remained unaffected. Concomitantly, the AKT targets Bad and XIAP appeared to be modulated. The pro-apoptotic regulator Bad was induced, and its phosphorylation was decreased. Downregulation of Bad mediated by siRNA partly rescued cells from ATO-induced cell death indicating that Bad might have a sensitizing role in ATO-mediated apoptosis. Moreover, ATO induced downregulation of the NF-kB target XIAP. The protein levels of other NF-kB targets such as Bcl-2 and Bcl-xL were unchanged. Downregulation of XIAP could be due to decreased protein stability resulting from reduced XIAP phosphorylation by AKT. The effects of low dose ATO on AKT phosphorylation as well as Bad and XIAP levels were more pronounced when cells were treated in combination with dexamethasone. Ongoing experiments will further define the role of XIAP and NF-kB as specific therapeutic targets in this model. These results indicate that resistance to GCs is - at least in part - mediated through apoptosis regulators in ALL. Our data provide evidence that ATO may be beneficial as sensitizing agent in defined subgroups of ALL patients, next to its role in APL and multiple myeloma treatment.