Drosophila Aurora A regulates mitotic timing in cancer stem cells: Possible therapeutic implications

ABSTRACT

Loss of Aurora A in Drosophila neuroblasts promotes loss of cell fate, leading to brain tumors. We showed that these tumor stem cells are delayed during mitosis and efficiently segregate their chromosomes even without the spindle assembly checkpoint. Here, we discuss the possible relevance of our results to human cancers.

To prevent chromosome segregation defects, all dividing cells, including tumor cells, possess a key mechanism called the spindle assembly checkpoint (SAC), which is at play during cell division. The effector of the SAC is the mitotic checkpoint complex (MCC). The MCC is present before mitosis but is also produced by unattached kinetochores. The MCC inhibits the anaphase promoting complex/cyclosome (APC/C) to prevent ubiquitylation and subsequent degradation of securin (required for cohesion of sister chromatids) and cyclin B (required for mitotic exit). When all kinetochores pairs are correctly attached to the mitotic spindle, the MCC is no longer produced. The SAC is therefore satisfied and securin and cyclin B are degraded, triggering loss of chromatid cohesion and mitotic exit. Under normal conditions the SAC is essential in mammals, but not in flies or yeasts. However, the SAC becomes necessary when spindle assembly is compromised.²,¹⁰

In cultured human cancer cells, inhibition of the SAC triggers premature mitotic exit and segregation of sister chromatids leading to severe aneuploidy and cell death. This outcome can be counterintuitive because aneuploidy can trigger genome instability, which is one of the causes of cancer. However, this strategy seems to be efficient because the severe gain and loss of chromosomes induced by SAC inhibition is not compatible with survival of the daughter cells.⁴ We recently challenged this strategy in vivo by suppression of the SAC in fly tumor neural stem cells. Interestingly, the use of Drosophila melanogaster neural stem cells (or neuroblasts [NBs]) has recently been proven to be a traceable system in cancer stem cell biology. Drosophila neural stem cells can produce tumors and, similar to mammalian models, these tumors can be transplanted into adult host flies leading to rapid death of the host. As in other cancer stem cell models, the triggering event that leads to tumor formation is a loss of cell fate.⁸

We used 2 mutant lines carrying null mutations in sas-4 and aurora A respectively. In both mutant flies, the neuroblasts display loss of cell fate (triggering tumor formation). These cells also show spindle assembly defects that are corrected after SAC-mediated mitotic delay.^1,6,9

Ablation of the SAC severely impairs normal chromosome segregation in sas-4 mutant tissues (sas-4 SACΔ) and 70% of the mitotic cells were aneuploid. In addition, these tissues were poorly developed and their ability to induce tumor formation was lost, likely because premature differentiation (but not apoptosis) occurs in this system.⁷

Surprisingly, we found that disruption of the SAC did not cause the expected aneuploidy in the aurora A null mutant (aurA SACΔ); instead, tumors are still formed and, importantly, the cells retain normal ploidy.³

Careful examination of aurora A mutant neural stem cell divisions by live microscopy revealed that the mitotic delay is both SAC dependent (due to spindle assembly defects) and SAC independent (due to defective cyclin B degradation). The SAC-independent mitotic delay is long enough to allow correction of the defective chromosome attachments in aurA SACΔ. For this reason, aurA SACΔ tumor cells do not harbor the severe aneuploidy observed in sas-4 SACΔ mutants (Fig. 1) and tumor growth is not affected.³

Figure 1.

Spindle assembly checkpoint ablation triggers aneuploidy and impairs tumor formation in sas-4 mutant neuroblasts (NBs) but not in aurora A (aurA) mutant NBs that show defective cyclin B degradation. Top panel, sas-4 mutant NBs do not harbor centrosomes but assemble a bipolar spindle after a spindle assembly checkpoint (SAC)-dependent mitotic delay. These NBs continue to proliferate. Ablation of the SAC (second panel from the top) triggers premature anaphase onset, severe chromosome segregation, and severe aneuploidy. The sas-4 SACΔ double mutant is not able to trigger tumor formation. AurA (third panel from top) mutant NBs show defective spindle assembly and SAC activation but the defective cyclin B degradation also contributes to an increase in the mitotic duration. In the aurA SACΔ  μυταντ (bottom panel), defective cyclin B degradation prolongs mitosis to promote correct spindle assembly and correct chromosome segregation. In this background, aneuploidy is weak and tumors still form. NEBD: nuclear envelope breakdown.

Are these results in fly tumor stem cells important for anticancer therapies?

Aurora A loss in Drosophila promotes the proliferation of tumor stem cells that are resistant to spindle defects. Although it is always difficult to make shortcuts between different systems, many of the current anticancer therapies aim to activate or inhibit the SAC by the use of compounds that produce spindle assembly defects and mitotic delay. These inhibitors include microtubule interfering agents and inhibitors against kinases required for spindle assembly and the SAC.⁴ We believe that the contribution of Aurora A, and probably other kinases (such as one of its downstream effectors polo-like kinase 1, best known as PLK1), to mitotic timing deserves to be analyzed in human cells because the duration of mitosis seems to have a relevant impact on the response of tumor cells to chemotherapy.⁵ Moreover, some tumor tissues display high levels of mitotic proteins that are normally degraded during mitotic exit, which is thought to be the consequence of the large number of cells undergoing mitosis. It is possible to speculate that the high levels of these proteins may in some cases reflect the ability of certain tumor types to remain delayed in mitosis and therefore resist SAC targeting agents. However, it should be noted that the mammalian genome harbors many more chromosomes than the fly genome (46 chromosomes versus 4 in Drosophila). This more complex genome is therefore intrinsically more sensitive to spindle damage leading to aneuploidy and subsequent cell death. Only extensive analyses will reveal whether mitotic timing really impacts tumor resistance to SAC-targeted chemotherapies.

Disclosure of potential conflicts of interest

No potential conflicts of interest were disclosed.