姜黄素衍生物C817在体外可抑制具有野生型或突变型Bcr-Abl的伊马替尼耐药慢性髓性白血病细胞的增殖。
Curcumin derivative C817 inhibits proliferation of imatinib-resistant chronic myeloid leukemia cells with wild-type or mutant Bcr-Abl in vitro.
作者信息
Wu Li-xian, Wu Ying, Chen Rui-jia, Liu Yang, Huang Li-sen, Lou Li-guang, Zheng Zhi-hong, Chen Yuan-zhong, Xu Jian-hua
机构信息
1] Department of Pharmacology, School of pharmacy, Fujian Medical University, Fuzhou 350004, China [2] Institute of Materia Medica, School of pharmacy, Fujian Medical University, Fuzhou 350004, China [3] Fuijan Key Laboratory of Natural Medicine Pharmacology, School of pharmacy, Fujian Medical University, Fuzhou 350004, China.
1] Institute of Materia Medica, School of pharmacy, Fujian Medical University, Fuzhou 350004, China [2] Fuijan Key Laboratory of Natural Medicine Pharmacology, School of pharmacy, Fujian Medical University, Fuzhou 350004, China.
出版信息
Acta Pharmacol Sin. 2014 Mar;35(3):401-9. doi: 10.1038/aps.2013.180. Epub 2014 Feb 3.
AIM
To find new kinase inhibitors that overcome the imatinib resistance in treatment of chronic myeloid leukemia (CML), we synthesized C817, a novel derivative of curcumin, and tested its activities against wild-type (WT) and imatinib-resistant mutant Abl kinases, as well as in imatinib-sensitive and resistant CML cells in vitro.
METHODS
32D cells harboring WT or mutant Abl kinases (nucleotide binding P-loop mutants Q252H, Y253F, and imatinib contact residue mutant T315I), as well as K562/G01 cells (with whole Bcr-Abl gene amplication) were tested. Kinase activity was measured using Kinase-Glo Luminescent Kinase Assay Platform in recombinant WT and mutant (Q252H, Y253F, and T315I) Abl kinases. Cell proliferation and apoptosis were examined using MTT assay and flow cytometry, respectively. The phosphorylation levels of Bcr-Abl initiated signaling proteins were analyzed using Western blotting. Colony forming units (CFU) growth and long term culture-initiating cells (LTC-ICs) were used to test the effects of C817 on human leukemia progenitor/stem cells.
RESULTS
C817 potently inhibited both WT and mutant (Q252H, Y253F, and T315I) Abl kinase activities in a non-ATP competitive manner with the values of IC₅₀ at low nanomole levels. In consistent with above results, C817 suppressed the growth of both imatinib-sensitive and resistant CML cells, including wild-type K562, K562/G01, 32D-T315I, 32D-Q252H, and 32D-Y253F cells with the values of IC₅₀ at low micromole levels. C817 (0.5 or 1 μmol/L) dose-dependently inhibited the phosphorylation of Bcr-Abl and downstream proteins STAT-5 and CrkL in imatinib-resistant K562/G01 cells. Furthermore, C817 significantly suppressed CFU growth and LTC-ICs, implicating that C817 could eradiate human leukemia progenitor/stem cells.
CONCLUSION
C817 is a promising compound for treatment of CML patients with Bcr-Abl kinase domain mutations that confer imatinib resistance.
目的
为了寻找能够克服慢性髓性白血病（CML）治疗中伊马替尼耐药性的新型激酶抑制剂，我们合成了姜黄素的新型衍生物C817，并在体外测试了其对野生型（WT）和伊马替尼耐药突变型Abl激酶的活性，以及对伊马替尼敏感和耐药的CML细胞的活性。
方法
对携带WT或突变型Abl激酶（核苷酸结合P环突变体Q252H、Y253F以及伊马替尼接触残基突变体T315I）的32D细胞，以及K562/G01细胞（具有完整的Bcr-Abl基因扩增）进行了测试。使用Kinase-Glo发光激酶检测平台测量重组WT和突变型（Q252H、Y253F和T315I）Abl激酶的激酶活性。分别使用MTT法和流式细胞术检测细胞增殖和凋亡。使用蛋白质印迹法分析Bcr-Abl起始信号蛋白的磷酸化水平。使用集落形成单位（CFU）生长和长期培养起始细胞（LTC-ICs）来测试C817对人白血病祖细胞/干细胞的影响。
结果
C817以非ATP竞争性方式有效抑制WT和突变型（Q252H、Y253F和T315I）Abl激酶活性，IC₅₀值处于低纳摩尔水平。与上述结果一致，C817抑制了伊马替尼敏感和耐药的CML细胞的生长，包括野生型K562、K562/G01、32D-T315I、32D-Q252H和32D-Y253F细胞，IC₅₀值处于低微摩尔水平。C817（0.5或1μmol/L）剂量依赖性地抑制伊马替尼耐药的K562/G01细胞中Bcr-Abl以及下游蛋白STAT-5和CrkL的磷酸化。此外，C817显著抑制CFU生长和LTC-ICs，这表明C817可以根除人白血病祖细胞/干细胞。
结论
C817是一种有前景的化合物，可用于治疗具有导致伊马替尼耐药的Bcr-Abl激酶结构域突变的CML患者。
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