Abstract:
We previously reported on the interaction of 10-chloro-7H-benzo[de]benzo[4,5]imidazo[2,1-a]isoquinolin-7-one (10-Cl-BBQ) with the Aryl hydrocarbon Receptor (AhR) and selective growth inhibition in breast cancer cell lines. We now report on a library of BBQ analogues with substituents on the phenyl and naphthyl rings for biological screening. Herein, we show that absence of the phenyl Cl of 10-Cl-BBQ to produce the simple BBQ molecule substantially enhanced the growth inhibitory effect with GI50 values of 0.001-2.1 μM in select breast cancer cell lines MCF-7, T47D, ZR-75-1, SKBR3, MDA-MB-468, BT20, BT474 cells, while having modest effects of 2.1-7 μM in other cell lines including HT29, U87, SJ-G2, A2780, DU145, BE2-C, MIA, MDA-MB-231 or normal breast cells, MCF10A (3.2 μM). The most potent growth inhibitory effect of BBQ was observed in the triple negative cell line, MDA-MB-468 with a GI50 value of 0.001 μM, presenting a 3,200-fold greater response than in the normal MCF10A breast cells. Additions of Cl, CH3, CN to the phenyl ring and ring expansion from benzoimidazole to dihydroquinazoline hindered the growth inhibitory potency of the BBQ analogues by blocking potential sites of CYP1 oxidative metabolism, while addition of Cl or NO2 to the naphthyl rings restored potency. In a cell-based reporter assay all analogues induced 1.2 to 10-fold AhR transcription activation. Gene expression analysis confirmed the induction of CYP1 oxygenases by BBQ. The CYP1 inhibitor α-naphthoflavone, and the SULT1A1 inhibitor quercetin significantly reduced the growth inhibitory effect of BBQ, confirming the importance of both phase I and II metabolic activation for growth inhibition. Conventional molecular modelling/docking revealed no significant differences between the binding poses of the most and least active analogues. More detailed DFT analysis at the DSD-PBEP86/Def-TZVPP level of theory could not identify significant geometric or electronic changes which would account for this varied AhR activation. Generation of Fukui functions at the same level of theory showed that CYP1 metabolism will primarily occur at the phenyl head group of the analogues, and substituents within this ring lead to lower cytotoxicity.
摘要:
我们先前报道了10-氯-7H-苯并[de]苯并[4,5]咪唑并[2,1-a]异喹啉-7-酮(10-Cl-BBQ)与芳基烃受体(AhR)的相互作用和乳腺癌细胞系中的选择性生长抑制。我们现在报道了在苯基和萘基环上具有取代基的BBQ类似物文库,用于生物筛选。在这里,我们显示,在选择的乳腺癌细胞系MCF-7,T47D中,不存在10-Cl-BBQ的苯基Cl来产生简单的BBQ分子,其GI50值为0.001-2.1μM,ZR-75-1,SKBR3,MDA-MB-468,BT20,BT474细胞,同时在其他细胞系中具有2.1-7μM的适度作用,包括HT29,U87,SJ-G2,A2780,DU145,BE2-C,MIA,MDA-MB-231或正常乳腺细胞,MCF10A(3.2μM)。在三阴性细胞系中观察到BBQ最有效的生长抑制作用,MDA-MB-468,GI50值为0.001μM,呈现比正常MCF10A乳腺细胞大3200倍的反应。Cl的添加,CH3,CN到苯环和从苯并咪唑到二氢喹唑啉的环扩展通过阻断CYP1氧化代谢的潜在位点阻碍了BBQ类似物的生长抑制效力,同时向萘环中添加Cl或NO2恢复了效力。在基于细胞的报道分子测定中,所有类似物诱导1.2至10倍的AhR转录激活。基因表达分析证实了BBQ对CYP1加氧酶的诱导。CYP1抑制剂α-萘黄酮,SULT1A1抑制剂槲皮素显著降低BBQ的生长抑制作用,证实了I期和II期代谢激活对生长抑制的重要性。常规的分子建模/对接显示,活性最高和活性最低的类似物的结合姿势之间没有显着差异。在DSD-PBEP86/Def-TZVPP理论水平上的更详细的DFT分析无法识别出可以解释这种变化的AhR激活的显着几何或电子变化。福井功能的产生在相同的理论水平表明,CYP1代谢将主要发生在类似物的苯基头基,和该环内的取代基导致较低的细胞毒性。