Suppression of mTOR complex 2-dependent AKT phosphorylation in melanoma cells by combined treatment with rapamycin and LY294002
Suppression of mTOR complex 2-dependent AKT phosphorylation in melanoma cells by combined treatment with rapamycin and LY294002
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DOI：
10.1111/j.1365-2133.2008.08991.x
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发表时间：
2009-05-01
影响因子：
10.3
通讯作者：
Pratscher, B.
中科院分区：
文献类型：
--
作者：
Werzowa, J.;Cejka, D.;Pratscher, B.
在一些癌细胞中，雷帕霉素抑制mTOR复合物1 （mTORC1）可导致AKT磷酸化，其生物学后果未知。这种磷酸化在黑色素瘤中的作用尚不清楚，尽管初步的临床数据表明在黑色素瘤中rapalogues活性较差。我们旨在阐明mTORC1抑制后AKT磷酸化在黑色素瘤细胞中的作用。采用Western blotting、细胞凋亡、细胞周期分析和细胞活力分析，分析雷帕霉素和LY294002对黑色素瘤细胞的影响。为了抑制mTOR复合物2 (mTORC2)，使用了靶向载体的siRNA。雷帕霉素对细胞活力的影响有限，但在黑色素瘤细胞中导致强烈而持久的AKT磷酸化。LY294002联合抑制PI3K/mTOR对细胞活力有显著影响，但在长期治疗后也导致AKT磷酸化增加。相比之下，雷帕霉素与LY294002联合抑制AKT磷酸化。抑制AKT磷酸化与细胞活力降低无关。抑制mTORC2导致磷酸化AKT水平降低。雷帕霉素和LY294002抑制mTORC1可通过mTORC2导致黑色素瘤细胞中AKT磷酸化。雷帕霉素联合LY294002可抑制AKT磷酸化，但对治疗效果无显著影响。
Inhibition of mTOR complex 1 (mTORC1) with rapamycin leads to phosphorylation of AKT in some cancer cells, with unknown biological consequences. The role of this phosphorylation in melanoma is unknown, although preliminary clinical data indicate poor activity of rapalogues in melanoma.We aimed at elucidating the role of AKT phosphorylation after mTORC1 inhibition in melanoma cells.Western blotting, apoptosis assays, cell cycle analyses and viability assays were performed to analyse the effects of rapamycin and LY294002 treatment on melanoma cells. For suppression of mTOR complex 2 (mTORC2) an siRNA directed against rictor was used.Rapamycin showed limited effects on cell viability but resulted in strong and lasting AKT phosphorylation in melanoma cells. Combined PI3K/mTOR inhibition with LY294002 had pronounced effects on viability but also led to increased AKT phosphorylation after prolonged treatment. In contrast, combination of rapamycin plus LY294002 suppressed AKT phosphorylation. Suppression of AKT phosphorylation did not correlate with decreases in cell viability. Inhibition of mTORC2 led to reduced levels of phosphorylated AKT.mTORC1 inhibition with rapamycin and with LY294002 can lead to AKT phosphorylation in melanoma cells via mTORC2. Combination of rapamycin and LY294002 suppresses AKT phosphorylation but without significant effect on treatment efficacy.