Dacomitinib potentiates the efficacy of conventional chemotherapeutic agents via inhibiting the drug efflux function of ABCG2 in vitro and in vivo.
Dacomitinib potentiates the efficacy of conventional chemotherapeutic agents via inhibiting the drug efflux function of ABCG2 in vitro and in vivo.
复制标题
Dacomitinib 通过在体外和体内抑制 ABCG2 的药物流出功能来增强传统化疗药物的疗效。
DOI：
10.1186/s13046-018-0690-x
复制
发表时间：
2018-02-20
期刊：
影响因子：
--
通讯作者：
Fu L
中科院分区：
文献类型：
--
作者：
Guo X;To KKW;Chen Z;Wang X;Zhang J;Luo M;Wang F;Yan S;Fu L
三磷酸腺苷结合盒G亚家族成员2(ABCG2)是ABC转运蛋白超家族的成员之一，通过将底物抗癌药物转运出癌细胞并减少其在细胞内的积聚来介导多药耐药(MDR)。多药耐药是化疗成功的主要障碍。克服MDR的合乎逻辑的方法是抑制转运蛋白。然而，目前还没有安全有效的MDR抑制剂在临床上被批准。采用四甲基偶氮唑盐比色法检测细胞毒作用和多药耐药逆转效果。用流式细胞仪检测药物外排和细胞内药物蓄积。建立H460/MX20细胞异种移植模型，评价达科米替尼对拓扑替康体内抗癌作用的影响。为了确定达科米尼与ABCG2底物结合部位的相互作用，进行了[125I]-碘芳基氮杂环己酮(IAAP)对ABCG2光标记物的竞争。在不同浓度的Dacomitinib存在下，检测ABCG2对钒敏感的ATPase活性，以评价作为转运体能量来源的Dacomitinib对ATP水解酶的影响。采用流式细胞术和Western blotting方法检测达科米丁布是否能抑制ABCG2的表达。实时荧光定量RT-PCR检测ABCG2mRNA的表达水平。Western blotting检测AKT、ERK蛋白表达水平及其磷酸化。在这里，我们发现Dacomitinib，一种处于III期临床试验的不可逆泛ErbB酪氨酸激酶抑制剂(TKI)，可以增强常规化疗药物对ABCG2高表达MDR癌细胞的疗效，但不能增强亲代敏感细胞的疗效。达科米替尼通过抑制转运体外排功能显著增加ABCG2探针底物[阿霉素(DOX)、罗丹明123(ROH123)和Hoechst 33342]的积聚。此外，达科米替尼还以浓度依赖的方式刺激ABCG2 ATPase活性，并与ABCG2的[125I]-IAAP光标记物竞争。但达考米替尼不改变ABCG2在蛋白和mRNA水平的表达，也不抑制ErbB下游的AKT和ERK信号转导。重要的是，达科米替尼显著增强了拓扑替康在ABCG2高表达H460/MX20细胞裸鼠异种移植瘤中的疗效，而不会产生额外的毒性。这些结果表明，达科米替尼通过抑制ABCG2外排功能和增加抗癌药物在细胞内的积聚来逆转ABCG2介导的多药耐药。我们的发现支持对ABCG2过表达多药耐药肿瘤的癌症患者进行达科米替尼联合常规化疗的进一步临床研究。
ATP-binding cassette subfamily G member 2 (ABCG2), a member of the ABC transporter superfamily proteins, mediates multidrug resistance (MDR) by transporting substrate anticancer drugs out of cancer cells and decreasing their intracellular accumulation. MDR is a major hurdle to successful chemotherapy. A logical approach to overcome MDR is to inhibit the transporter. However, no safe and effective MDR inhibitor has been approved in the clinic. The MTT assay was used to evaluate cell cytotoxicity and MDR reversal effect. Drug efflux and intracellular drug accumulation were measured by flow cytometry. The H460/MX20 cell xenograft model was established to evaluate the enhancement of anticancer efficacy of topotecan by dacomitinib in vivo. To ascertain the interaction of dacomitinib with the substrate binding sites of ABCG2, the competition of dacomitinib for photolabeling of ABCG2 with [125I]- iodoarylazidoprazosin (IAAP) was performed. Vanadate-sensitive ATPase activity of ABCG2 was measured in the presence of a range of different concentrations of dacomitinib to evaluate the effect of dacomitinib on ATP hydrolysis as the energy source of the transporter. A flow cytometry-based assay and western blotting were employed to study whether dacomitininb could inhibit the expression level of ABCG2. The mRNA expression levels of ABCG2 were analyzed by real-time quantitative RT-PCR. The protein expression level of AKT, ERK and their phosphorylations were detected by Western blotting. Here, we found that dacomitinib, an irreversible pan-ErbB tyrosine kinase inhibitor (TKI) in phase III clinical trial, could enhance the efficacy of conventional chemotherapeutic agents specifically in ABCG2-overexpressing MDR cancer cells but not in the parental sensitive cells. Dacomitinib was found to significantly increase the accumulation of ABCG2 probe substrates [doxorubicin (DOX),Rhodamine 123 (Rho 123) and Hoechst 33342] by inhibiting the transporter efflux function. Moreover, dacomitinib stimulated ABCG2 ATPase activity and competed with [125I]-IAAP photolabeling of ABCG2 in a concentration-dependent manner. However, dacomitinib did not alter ABCG2 expression at protein and mRNA levels or inhibit ErbB downstream signaling of AKT and ERK. Importantly, dacomitinib significantly enhanced the efficacy of topotecan in ABCG2-overexpressing H460/MX20 cell xenografts in nude mice without incurring additional toxicity. These results suggest that dacomitinib reverses ABCG2-mediated MDR by inhibiting ABCG2 efflux function and increasing intracellular accumulation of anticancer agents. Our findings advocate further clinical investigation of combinations of dacomitinib and conventional chemotherapy in cancer patients with ABCG2-overexpressing MDR tumors.
登录
查看更多内容
影响因子：
11.2
作者：
Dai CL;Tiwari AK;Wu CP;Su XD;Wang SR;Liu DG;Ashby CR Jr;Huang Y;Robey RW;Liang YJ;Chen LM;Shi CJ;Ambudkar SV;Chen ZS;Fu LW
通讯作者：
Fu LW
影响因子：
4.8
作者：
Hrycyna, CA;Ramachandra, M;Gottesman, MM
通讯作者：
Gottesman, MM
影响因子：
5.8
作者：
Tong, Xiu-zhen;Wang, Fang;Fu, Li-wu
通讯作者：
Fu, Li-wu
影响因子：
3.5
作者：
Jain, Hiteshkumar D.;Zhang, Chunchun;Cook, James M.
通讯作者：
Cook, James M.
影响因子：
2.9
作者：
Shukla, Suneet;Robey, Robert W.;Ambudkar, Suresh V.
通讯作者：
Ambudkar, Suresh V.