Background: Arsenic trioxide (As(2)O(3)) is a well-known and effective treatment that can result in clinical remission for patients diagnosed with acute promyelocytic leukemia (APL). The biologic efficacy of As(2)O(3) in APL and solid tumor cells has been explained through its actions on anti-proliferation, anti-angiogenesis, and apoptotic signaling pathways. We theorize that As(2)O(3) activates a pathway that disrupts microtubule dynamics forming abnormal, nonfunctioning mitotic spindles, thus preventing cellular division. In this study, we investigated how As(2)O(3) induces apoptosis by causing microtubule dysfunction.
Methods: Cultured NB4 cells were treated with As(2)O(3), paclitaxel, and vincristine. Flow cytometric analysis was then performed. An MTT assay was used to determine drug-mediated cytotoxicity. For tubulin polymerization assay, each polymerized or soluble tubulin was measured. Microtubule assembly-disassembly was measured using a tubulin polymerization kit. Cellular microtubules were also observed with fluorescence microscopy.
Results: As(2)O(3) treatment disrupted tubulin assembly resulting in dysfunctional microtubules that cause death in APL cells. As(2)O(3) markedly enhanced the amount of depolymerized microtubules. The number of microtubule posttranslational modifications on an individual tubulin decreased with As(2)O(3) concentration. Immunocytochemistry revealed changes in the cellular microtubule network and formation of polymerized microtubules in As(2)O(3)-treated cells.
Conclusion: The microtubules alterations found with As(2)O(3) treatment suggest that As(2)O(3) increases the depolymerized forms of tubulin in cells and that this is potentially due to arsenite's negative effects on spindle dynamics.
Keywords: Acute promyelocytic leukemia; Antimitotic agents; Apoptosis; Arsenic trioxide; Tubulin.