As shown in Figure 6A, selleck chemicals FTY720 ABT 263 increased the phosphorylation of GSK 3B, but no effect on total GSK 3B. Meanwhile, ABT 263 en hanced the phosphorylation of Akt, an upstream signal molecule of GSK 3B. Suppression of Akt by its inhibitor BEZ 235 dramatically attenuated ABT 263 mediated GSK 3B phosphorylation, Mcl 1 upregulation and apop tosis resistance. Subsequently, we checked whether the phosphorylation of GSK 3B is also affected by ERK, another upstream regulator of GSK 3B. As shown in Figure 6C, inhibition of ERK with U0126 had no effect on ABT 263 triggered GSK 3B phosphoryl ation, indicating that GSK 3B activity was not regulated by ERK in this process. Furthermore, Akt inhibitor also increased the cytoto icity of ABT 263 in HCC cells.

These results indicated that Akt mediated GSK 3B inactivation also involves in ABT 263 induced Mcl 1 stabilization, possibly through regulating the phosphorylation of Mcl 1Ser159. Discussion In the present study, we demonstrated that ABT 263 up regulated Mcl 1 by increasing the stability of Mcl 1 mRNA and protein in HCC cells. As shown in the work ing model, ABT 263 increased Mcl 1 mRNA level by augmenting its stability instead of transcrip tional activation. Meanwhile, ABT 263 enhanced Mcl 1 protein stability by regulating the phosphorylation status of Mcl 1. ERK and JNK mediated Mcl 1Thr163 phos phorylation contributed to ABT 263 induced Mcl 1 pro tein stability. Akt mediated GSK 3B inactivation also played important role in preventing Mcl 1 protein deg radation in the presence of ABT 263.

ABT 263, a newly developed, oral tolerant Bcl 2 L in hibitor, has shown promising anti tumor efficacy in non small cell lung cancer and acute lymphoblastic leukemia as single agent both in vitro and in vivo. Meanwhile, ABT 263 can markedly sensitize several clinical drugs in cancer therapy. However, a recent study has dem onstrated that HCC cells are relatively resistant to ABT 737 compared to leukemia and lung carcinomas. Furthermore, it has been indicated that ABT 737 induced Mcl 1 upregulation Brefeldin_A contributes to this resistance. Consistent with ABT 737, our results showed that both ABT 263 and another Bcl 2 inhibitor AT 101 upregulated Mcl 1 in HCC cells, which at last re sulted in drug resistance. So it is important to clarify the associated mechanisms of ABT 263 induced Mcl 1 upreg ulation in HCC cells. It is known that Mcl 1 is an important anti apoptotic protein, which is now becoming a quite important target for cancer therapy. Characteristically, it has a short half life and is elaborately regulated at different levels. We found that ABT 263 increased Mcl 1 mRNA level in HCC cells.