A Novel Therapeutic Approach in Acute Promyelocytic Leukemia with All-trans retinoic Acid and Cyclin-dependent Kinase Inhibitors

Background: Acute promyelocytic leukemia (APL) is characterized by chromosomal translocations, which generate fusion oncogenic proteins. In 95% of cases, in frame fusion of the RARα gene to the PML gene, gives rise to the PML-RARα chimeric gene in patients with t(15;17)q22;q21. In addition, in 99% of cases, APL cells also express the germ cell-specific Cyclin A1 protein, which has been shown to contribute to APL-like disease in mouse models. Re-localization and degradation of PML-RARα with pharmacologic levels of all-trans retinoic acid (ATRA) may cause serious side effects such as the development of myelodysplastic syndrome, cutaneous T-cell lymphoma and skin cancers. Here, we exploit the expression of cyclin A1 and of the fusion protein and show that combined, sub-pharmacologic doses of ATRA and cyclin-dependent kinase inhibitors are more efficacious than either agent alone at inducing etc at inducing APL cell differentiation. Methods: Cultured NB4 APL cells were used to compare the degree of differentiation by ATRA and olomoucine or roscovitine, two cyclin-dependent kinase inhibitors alone or in combination. Differentiation was monitored with the NBT reaction, quantitation of proliferation and monitoring the levels of CDK4, PML-RARα and C/EBP proteins using immunoblots. Results: ATRA or roscovitine alone are effective at inducing inhibition of proliferation and differentiation at pharmacologic levels whereas olomoucine alone is not. In contrast, the combined use of sub-pharmacologic doses of ATRA with either olomoucine or roscovitine is more effective at inhibiting APL cell proliferation than ATRA, olomoucine or roscovitine alone with implications for the treatment of APL. Conclusion: We demonstrate that in NB4 cells, the combination of ATRA/ olomoucine or ATRA/roscovitine leads to more potent differentiation accompanied by a reduction in CDK4 protein levels, decreased degradation of the fusion protein and up-regulation of the transcription factor C/EBPε.