YAP蛋白在HER2阳性乳腺癌拉帕替尼耐药中的作用

陈鹏; 徐婷; 郭丹; 黄佳祎

doi:10.16016/j.1000-5404.201911189

您当前的位置：

首页 >

文章列表页 >

YAP蛋白在HER2阳性乳腺癌拉帕替尼耐药中的作用

基础医学 | 更新时间：2026-01-09

- YAP蛋白在HER2阳性乳腺癌拉帕替尼耐药中的作用
- Role of yes-associated protein in resistance of HER2⁺ breast cancer to lapatinib
- 第三军医大学学报 2020年42卷第7期页码：692-698
- 作者机构：
  
  1.400016 重庆，重庆医科大学基础医学院病理生理学教研室
  2.400016 重庆，重庆医科大学附属第二医院三腺外科
- 作者简介：
  
  [黄佳祎，E-mail: 102561@cqmu.edu.cn HUANG Jiayi, E-mail: 102561@cqmu.edu.cn
- 基金信息：
- DOI：10.16016/j.1000-5404.201911189
  中图分类号： R737.9;R915;R979.1
- 收稿：2019-11-22，
  
  修回：2019-12-17，
  
  纸质出版：2020-04-15
- 稿件说明：
移动端阅览
陈鹏, 徐婷, 郭丹, 等. YAP蛋白在HER2阳性乳腺癌拉帕替尼耐药中的作用[J]. 第三军医大学学报, 2020,42(7):692-698.

Peng CHEN, Ting XU, Dan GUO, et al. Role of yes-associated protein in resistance of HER2⁺ breast cancer to lapatinib[J]. Journal of Third Military Medical University, 2020, 42(7): 692-698.
陈鹏, 徐婷, 郭丹, 等. YAP蛋白在HER2阳性乳腺癌拉帕替尼耐药中的作用[J]. 第三军医大学学报, 2020,42(7):692-698. DOI： 10.16016/j.1000-5404.201911189.

Peng CHEN, Ting XU, Dan GUO, et al. Role of yes-associated protein in resistance of HER2⁺ breast cancer to lapatinib[J]. Journal of Third Military Medical University, 2020, 42(7): 692-698. DOI： 10.16016/j.1000-5404.201911189.

摘要

目的

建立乳腺癌拉帕替尼耐药细胞株，探讨Hippo通路中YAP蛋白在HER2阳性乳腺癌细胞拉帕替尼耐药中的作用及机制。

方法

采用逐步增加剂量与高浓度反复间歇诱导乳腺癌细胞株Skbr3对拉帕替尼耐药，耐药株命名为Skbr3-LR。用拉帕替尼同时处理亲本细胞和耐药细胞株，CCK-8法检测其耐药指数；结晶紫染色和平板克隆形成实验检测细胞的存活状况；定量PCR检测上皮-间质化改变。定量PCR及Western blot检测YAP、TAZ的表达。以维替泊芬阻断耐药细胞株Skbr3-LR中YAP下游通路的激活，CCK-8法检测其细胞活性的改变并计算IC

值的变化；结晶紫染色检测细胞存活数量的变化。定量PCR及Western blot检测YAP表达的改变。

结果

与亲本细胞比较，结晶紫染色和平板克隆形成实验结果均显示，耐药株存活细胞数量明显增多。亲本细胞耐药指数为3.61 μmol/L，耐药株耐药指数上升约8倍，为29.26 μmol/L。PCR结果显示，与亲本细胞比较，耐药细胞株的上皮标志物E-cadherin表达降低，而间质标志物Twist、Vinemtin和N-cadherin表达上升，YAP/TAZ表达上升，差异均有统计学意义(

0.05)。Western blot结果显示，耐药细胞株中总YAP和磷酸化YAP表达均较亲本细胞显著上升(

0.05)。以维替泊芬阻断耐药细胞株YAP下游通路的激活后，其耐药指数为2.98 μmol/L，较未处理的Skbr3-LR耐药指数下降约4倍；其存活细胞数量明显降低，YAP表达也明显降低(

0.05)。

结论

成功建立了拉帕替尼获得性耐药的乳腺癌细胞模型，其耐药机制可能与Hippo信号通路下游的关键效应因子YAP表达异常有关。

Abstract

Objective

To construct a lapatinib-resistant breast cancer cell line

and to study the role and underlying mechanism of yes-associated protein (YAP) in Hippo signaling pathway in the process.

Methods

The drug-resistant cell model of human breast cancer was established by gradually increasing the dose of drug and repeated intermittent high concentration inducing. After lapatinib treatment

the parent cell line Skbr3 and the drug-resistant cell line Skbr3- LR were detected by CCK-8 assay for drug resistance index

crystal violet staining and plate colony for cell survival

and quantitative PCR for the expression of epithelial mesenchymal transformation (EMT) markers. The expression of YAP/TAZ at mRNA and protein levels were measured by quantitative PCR and Western blotting

respectively. After verteporfin was used to block the downstream pathway of YAP in Skbr3-LR cells

CCK-8 assay was employed to measure the cell viability to obtain IC

cell crystal violet staining was performed to count the number of survival cells

and the change of YAP expression was be detected by quantitative PCR and Western blotting.

Results

The results of crystal violet staining and plate colony showed that there were more survival cells in the Skbr3-LR cells when compared with the parent Skbr3 cells. The drug resistance index was 3.61 μmol/L in the parent Skbr3 cells

and was increased by 8 times in the drug-resistant Skbr3-LR cells (29.26 μmol/L). Compared with the parent cells

the expression level of the marker of epithelial cells

E-cadherin was decreased

while those of mesenchymal markers

Twist

Vinemtin

N-cadherin

and of YAP/TAZ were increased in the drug resistant cells (all

0.05). And Western blot assay also indicated the enhanced expression of YAP and p-YAP proteins (

0.05). After the Skbr3-LR cells were treated with verteporfin

the IC

to Lapatinib was decreased from 11.25 to 2.98 μmol/L

by 4 times

and there were less survival cells and the expression of YAP was decreased (

0.05).

Conclusion

The lapatinib-resistant model of breast cancer Skbr3-LR cells is successfully established. And the mechanism of cell resistant to lapatinib may be associated with the abnormal expression of YAP protein.

关键词

Keywords

references

CAMERON D, PICCART-GEBHART M J, GELBER R D, et al . 11 years' follow-up of trastuzumab after adjuvant chemotherapy in HER2-positive early breast cancer: final analysis of the HERceptin Adjuvant (HERA) trial[J ] . Lancet, 2017, 389(10075): 1195-1205. DOI:10.1016/S0140-6736(16)32616-2.

PARONI G, FRATELLI M, GARDINI G, et al . Synergistic antitumor activity of lapatinib and retinoids on a novel subtype of breast cancer with coamplification of ERBB2 and RARA[J ] . Oncogene, 2012, 31(29): 3431-3443. DOI:10.1038/onc.2011.506.

GUJRAL T S, KIRSCHNER M W. Hippo pathway mediates resistance to cytotoxic drugs[J]. Proc Natl Acad Sci USA, 2017, 114(18): E3729-E3738. DOI:10.1073/pnas.1703096114.

SONG Q H, MAO B B, CHENG J B, et al . YAP enhances autophagic flux to promote breast cancer cell survival in response to nutrient deprivation[J ] . PLoS ONE, 2015, 10(3): e0120790. DOI:10.1371/journal.pone.0120790.

JEONG W, KIM S B, SOHN B H, et al . Activation of YAP1 is associated with poor prognosis and response to taxanes in ovarian cancer[J ] . Anticancer Res, 2014, 34(2): 811-817.

SONG S M, HONJO S, JIN J K, et al . The hippo coactivator YAP1 mediates EGFR overexpression and confers chemoresistance in esophageal cancer[J ] . Clin Cancer Res, 2015, 21(11): 2580-2590. DOI:10.1158/1078-0432.CCR-14-2191.

HSU P C, YOU B, YANG Y L, et al . YAP promotes erlotinib resistance in human non-small cell lung cancer cells[J ] . Oncotarget, 2016, 7(32): 51922-51933. DOI:10.18632/oncotarget.10458.

LIU H R, DU S Y, LEI T T, et al . Multifaceted regulation and functions of YAP/TAZ in tumors (Review)[J ] . Oncol Rep, 2018, 40(1): 16-28. DOI:10.3892/or.2018.6423.

HONG W J, GUAN K L. The YAP and TAZ transcription co-activators: key downstream effectors of the mammalian Hippo pathway[J]. Semin Cell Dev Biol, 2012, 23(7): 785-793. DOI:10.1016/j.semcdb.2012.05.004.

HANSEN C G, NG Y L, LAM W L, et al . The Hippo pathway effectors YAP and TAZpromote cell growth by modulating amino acid signaling to mTORC1[J ] . Cell Res, 2015, 25(12): 1299-1313. DOI:10.1038/cr.2015.140.

XIE M, ZHANG L, HE C S, et al . Prognostic significance of TAZ expression in resected non-small cel l lung cancer[J ] . J Thorac Oncol, 2012, 7(5): 799-807. DOI:10.1097/JTO.0b013e318248240b.

WANG Y, DONG Q Z, ZHANG Q F, et al . Overexpression of yes-associated protein contributes to progression and poor prognosis of non-small-cell lung cancer[J ] . Cancer Sci, 2010, 101(5): 1279-1285. DOI:10.1111/j.1349-7006.2010.01511.x.

ZHOU J Y, ZHANG S Z, LI Z J, et al . Yap-Hippo promotes A549 lung cancer cell death via modulating MIEF1-related mitochondrial stress and activating JNK pathway[J ] . Biomedecine Pharmacother, 2019, 113: 108754. DOI:10.1016/j.biopha.2019.108754.

ZHOU H L, LI G Q, HUANG S, et al . SOX9 promotes epithelial-mesenchymal transition via the Hippo-YAP signaling pathway in gastric carcinoma cells[J ] . Oncol Lett, 2019, 18(1): 599-608. DOI:10.3892/ol.2019.10387..

KARPOWICZ P, PEREZ J, PERRIMON N. The Hippo tumor suppressor pathway regulates intestinal stem cell regeneration[J]. Development, 2010, 137(24): 4135-4145. DOI:10.1242/dev.060483.

VISSER S, YANG X L. LATS tumor suppressor: A new governor of cellular homeostasis[J]. Cell Cycle, 2010, 9(19): 3892-3903. DOI:10.4161/cc.9.19.13386.

COOPER J, GIANCOTTI F G. Cancer: a new role for non-canonical Hippo signaling[J]. Cell Res, 2017, 27(4): 459-460. DOI:10.1038/cr.2017.27.

WANG X D, SU L L, OU Q S. Yes-associated protein promotes tumour development in luminal epithelial derived breast cancer[J]. Eur J Cancer, 2012, 48(8): 1227-1234. DOI:10.1016/j.ejca.2011.10.001.

YUN M R, CHOI H M, LEE Y W, et al . Targeting YAP to overcome acquired resistance to ALK inhibitors in ALK-rearranged lung cancer[J ] . EMBO Mol Med, 2019, 11(12): e10581. DOI:10.15252/emmm.201910581.

LEE B S, PARK D I, LEE D H, et al . Hippo effector YAP directly regulates the expression of PD-L1 transcripts in EGFR-TKI-resistant lung adenocarcinoma[J ] . Biochem Biophys Res Commun, 2017, 491(2): 493-499. DOI:10.1016/j.bbrc.2017.07.007.

KIM M H, KIM C G, KIM S K, et al . YAP-induced PD-L1 expression drives immune evasion in BRAFi-resistant melanoma[J ] . Cancer Immunol Res, 2018, 6(3): 255-266. DOI:10.1158/2326-6066.CIR-17-0320.

MOORING M, FOWL B H, LUM S Z C, et al . Hepatocyte stress increases expression of YAP and TAZ in hepatocytes to promote parenchymal inflammation and fibrosis[J ] . Hepatology, 2019.[Epub ahead of print ] . DOI:10.1002/hep.30928.

浏览量

下载量

CSCD

文章被引用时，请邮件提醒。

提交

工具集

关联资源

基于中国人群的雌激素受体表达新分层（低/中/高）可更精准地预测HR⁺/HER2^-早期乳腺癌患者的预后

二氢杨梅素通过抑制mTOR信号通路抑制乳腺癌细胞迁移和侵袭

BIRC5基因rs9904341多态性与乳腺癌易感性的关联研究

长链非编码ATB通过抑制miR-200c的表达促进乳腺癌细胞的侵袭转移

多柔比星对乳腺癌细胞核内丝切蛋白相互作用蛋白影响的比较蛋白质组学研究