is particularly interesting among the cyclin family because it can activate two different CDKs and functions in both S phase and mitosis. In S phase, phosphorylated cyclin A2
-CDK2 complexes are suggested to play an important role in the initiation of DNA replication. In mitosis, cyclin A2
may contribute to the control of cyclin B stability. Consistent with its role as a key cell cycle regulator, overexpression of cyclin A2
is associated with transformed cells 
. However, it is difficult to determine whether elevation of cyclin A2
is a contributing factor or a mere consequence of the increased cell proliferation. In haematological malignancies, cyclin A2
is associated with proliferation rate of these disorders and can be used for molecular diagnostics as a proliferation marker 
Single-walled carbon nanotubes (SWNTs) have been considered as the leading candidate for nanodevice applications ranging from gene therapy and novel drug delivery to membrane separations. We have previously showed that siRNA transfection efficiency of lipofectamine 2000 in K562 cells was low (28%) and some cells underwent apoptosis and necrosis during the process 
. However, SWNTs could efficiently facilitate the coupling of siRNA to form siRNA
SWNTs complexes and carry siRNA into K562 cells, significantly knocking down the expression of target gene. No apparent cell toxicology was observed. Hence, in order to directly probe whether cyclin A2
participates in cell apoptosis and differentiation, we employed SWNTs as transfection vector to deliver cyclin A2
siRNA into K562 cells to specifically knock down the expression of cyclin A2
and found that carbon nanotubes vector showed no additive or synergetic effect on cell toxicology of DOX, which was consistent with our previous report 
DOX, a prominent member of anthracycline antibiotics, has been extensively used for treatment of solid tumors and leukemia. It exerts its cytotoxic activity against cancer cells mainly by intercalation into DNA, inhibition of topoisomerase II and helicase activity, leading to cell-cycle arrest at the G2/M phase and apoptosis 
. In clinical applications, doses of DOX are strictly limited by its cardiotoxicity 
. It should be noted that the dose of DOX administrated here (0.4 µM) is pharmacological relevant compared to the initial or steady-state plasma concentrations observed in patients after standard bolus infusions (5 µM and 25–250 nM, respectively).
It has been reported that there is a link between cyclin A2
and apoptosis 
. Hoang et al.
showed that in c-myc overexpressing serum deprived rat 1A fibroblasts undergoing apoptosis, cyclin A2
mRNA expression was increased, in contrast to the invariant expression for cyclin B, C, D1 and E 
. Moreover, serum-deprived rat 1A fibroblast stably transfected with cyclin A2
exhibited increased apoptosis following stimulation of cyclin A2
expression. Meikrantz et al.
reported that induction of apoptosis was uniformly associated with activation of cyclin A2
-dependent kinases but not associated with cyclins E or B, and overexpression of the cyclin A2
could circumvent the anti-apoptosis activity of the oncogene BCL-2 in human Hela cells 
. Furthermore, Hiromura et al.
indicated that apoptosis was associated with an increase in cytoplasmic cyclin A2
-CDK2 activity following UV irradiation, under these conditions, nuclear cyclin A2
-Cdk2 activity decreased significantly 
. Our results showed knocking down the expression of cyclin A2
in K562 cells significantly suppressed the apoptosis induced by DOX and a positive correlation between the levels of cyclin A2
and apoptosis was observed. The findings also indicate that the cytoplasmic subcellular distribution of cyclin A2
correlates with its pro-apoptotic role. We speculate that cyclin A2
associated kinases are involved in DOX-induced apoptotic cell death pathways in K562 cells, although the exact downstream mechanisms are not known.
We have demonstrated that SWNTs could effectively deliver cyclin A2
siRNA into K562 cells, significantly suppressing the expression of cyclin A2
with specificity and cell proliferation, and cells with reduced cyclin A2
showed a decrease in the percentage of cells in S phase 
. Several studies have indicated that cancer cells with a high S-phase fraction/high proliferative activity are more sensitive to apoptosis induced by chemotherapy 
. As for DOX, it is active throughout the cell cycle, but the effect is most pronounced for cells in S phase-G2 phase, especially in S phase, where it interferes with the DNA replication and transcription 
. Therefore, it was not surprising that K562 cells with reduced cyclin A2
showed a marked decrease in DOX susceptibility.
Most of chemotherapeutic agents show significant side effects and not all patients benefit from aggressive chemotherapy. Therefore, searching for tumor biological factors which can predict patient prognosis and chemotherapy response would be of most importance. Several studies have indicated that a high level of cyclin A2
expression may be a marker of poor prognosis in cancers 
. Besides, previous studies have shown that cancer patients with high level of cyclin A2
had better chemotherapy response and survival than those with reduced cyclin A2
and low expression of cyclin A2
, indicating that the patients with high expression of cyclin A2
are more suitable for chemotherapy 
. Our results demonstrate the pro-apoptotic role of cyclin A2
in human myeloid leukemia K562 cells, and indicate that cells with low level of cyclin A2
were more resistant to chemotherapeutic agent DOX. Poon et al.
have suggested that a decrease of cyclin A2
, rather than increase, promotes tumorigenesis, and once the tumor has developed, high levels of cyclin A2
simply reflect a high proliferation rate, which can explain this inconsistency 
. Hence, despite its association with transformed cells, evaluating cyclin A2
level in patients will be an important prognostic marker for use of chemotherapy. Patients with high level of cyclin A2
may be more responsive to anticancer drugs through the induction of apoptotic cell death. Moreover, it should be cautious to combine doxorubicin chemotherapy with any small molecule drug targeting cyclin A2
associated kinases since it can enhance potential drug resistance.
In several systems, it has been reported that down-regulation of cyclin A2
and its associated CDK 2 activity are important for successful differentiation 
. Ito et al
. have suggested cyclin A2
overexpression is directly related with poor differentiation 
. Kiyokawa et al
. have indicated that differentiation of murine erythroleukemia cells induced by hexamethylene is accompanied by a decrease in the level of cyclin A2
and CDK2 proteins and the persistent suppression of cyclin A2
expression may play a role in HMBA-induced commitment to terminal differentiation 
. Yoshizumi et al
. showed down-regulation of cyclin A2
gene expression in vivo at both the RNA and protein levels appears to be important in the permanent withdrawal of human and rat cardiomyocytes from the cell cycle during development 
. Moreover, Rieber et al
. demonstrated that the interaction of cyclin A2
with E2F is the target for tyrosine induction of B16 melanoma terminal differentiation 
. In this work, we found that knocking down the expression of cyclin A2
in K562 cells significantly suppressed DOX-induced erythroid differentiation and a small fraction of cells with reduced cyclin A2
were differentiated along megakaryocytic and monocyte-macrophage pathways upon treatment with DOX. To the best of our knowledge, this is the first report that knocking down expression of one gene can switch K562 cells differentiation pathways. The results suggested that cyclin A2
is directly involved in the checkpoint of cell differentiation pathways and is a key regulator of this process, although the detail downstream mechanisms are not known. For cancer cells with low level of cyclin A2
, which are less responsive to chemotherapeutic agents, induction of differentiation might be an alternative strategy. Combination of cyclin A2
siRNA and DOX may provide a novel option of such therapeutic strategy.
In conclusion, knocking down the expression of cyclin A2 by siRNA delivered by SWNTs suppresses apoptosis and erythroid differentiation, and promotes megakaryocytic and monocyte-macrophage differentiation in human chronic myelogenous leukaemia K562 cells upon administration with DOX. The results demonstrate the pro-apoptotic role of cyclin A2 and suggest that cyclin A2 is a key regulator of cell differentiation, supporting the notion that cyclin A2 is an important regulator for cell cycle as well as for cell apoptosis and differentiation.