Study on the Relevance of Inducing Apoptosis and Reversal Effect of Nilotinib In Combination with Tet on Multidrug Reststahoe of K562/A02 Cell Line

To investigate the relevance of Inducing apoptosis and reversal effect of nilotinib in combination with Tet on multidrug reststahoe of K562/A02 cell line and its mechanism.

Methyl-thiazol tetrazolium(MTT) assay is employed to examine the pharmacological effect of Nilotinib or Tet alone on K562/A02 cell line, to calculate the inhibitor concerntration 50(IC₅₀) of daunorubicin(DNR) in K562/A02 cell line and the resistance multiple of DNR to K562/A02 cell line. Flow cytometry(FCM) was employed to comple the effect on the inducing apoptosis of K562/A02. The expression of bax/survivin mRNA was determined by RT-PCR, and the expression of bax/survivin protein was assessed by Western blot.

After 48 h 5 nmol/L nilotinib or 1.0μmoL/L Tet treatment, IC₅₀ of DNR to K562/A02 was 5.71mg/L or 6.52 mg/L respectively;While on combinative treatment, its IC₅₀ decreased to 3.12 mg/L. Nilotinib or Tet alone was able to increase the DNR induced apoptosis rate of K562/A02 cell (P<0.05), while on combination treatment the apoptosis rate increased remarkably. After 48 h 5 nmoL/L nilotinib or 1.0μmoL/L Tet treatment alone, gray—scale vaule of bax mRNA Was 2.90±0.31 or 3.6±0.46, respectively Gwhile on combinative treatment the value increased to 5.9±0.98. The bax protein expression level in K562/A02 cells was 2.1±0.07 or 2.90±0.09 when treated with 5 nmoL/L nilotinib or 1.0μmoL/L Tet alone for 48 h, but on combination treatment, the level increased to 4.8±0.31. After 48 h 5 nmoL/L nilotinib or 1.0μmoL/L Tet treatment alone, gray—scale vaule of survivin mRNA was 0.70±0.01 or 0.55±0.02, respectively;while on combinative treatment the value decreased to 0.35±0.08. The survivin protein expression level in K562/A02 cells was 0.74±0.03or 0.61±0.04 when treated with 5 nmoL/L nilotinib or 1.0μmoL/L Tet alone for 48 h, but on combination treatment, the level decreased to 0.42±0.06.

Nilotinib or Tet alone can pattially reverse drug resistance of K562/A02 cells. There is resistance phenomena of DNR to K562/A02 cells, and Nilotinib or Tet alone alone is able to reserve the resistance of DNR and enhance the inducing apoptasis effect of DNR. The mechanism may be associated with the increase of bax mRNA and protein expression and decrease of survivin mRNA and protein expression and increase of the apoptosis rate. And there is a synergistic action with these two agants in combination.

No relevant conflicts of interest to declare.