Glutathione depletion overcomes resistance to arsenic trioxide in arsenic-resistant cell lines

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Abstract
Arsenic trioxide (As2O3) is an effective treatment for acute promyelocytic leukemia (APL), but is less effective against other leukemias. Although the response of APL cells to As2O3 has been linked to degradation of the PML/RAR fusion oncoprotein, there is evidence that PML/RAR expression is not the only mediator of arsenic sensitivity. Indeed, we found that exogenous expression of PML/RAR did not sensitize a non-APL leukemic line to As2O3. To evaluate possible other determinants of sensitivity of leukemic cells to As2O3, we derived two arsenic-resistant NB4 subclones. Despite being approximately 10-fold more resistant to arsenic than their parental cell line, PML/RAR protein was still degraded by As2O3 in these cells, providing further evidence that loss of expression of the oncoprotein does not confer arsenic sensitivity. Both arsenic-resistant clones contained high glutathione (GSH) levels, however, and we found that GSH depletion coupled with As2O3 treatment dramatically inhibited their growth. Annexin V-staining and TUNEL analysis confirmed a synergistic induction of apoptosis. In addition, these cells failed to accumulate ROS in response to arsenic treatment, in contrast to their arsenic-sensitive parental cells, unless cotreated with buthionine sulfoximine. While other malignant cells did not show a good correlation between arsenic sensitivity and GSH content, GSH depletion nevertheless sensitized all cell lines examined, regardless of their initial response to arsenic alone. These findings suggest that PML/RAR expression is not a determinant of arsenic sensitivity, and further support the coupling of GSH depletion and arsenic treatment as a novel treatment for human malignancies that are unresponsive to arsenic alone.

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