Cepharanthine inhibits autophagic degradation in breast cancer MDA-MB-231 cells

Qin DENG; Ruoqiu FU; Ning GAO

doi:10.16016/j.1000-5404.201709083

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Cepharanthine inhibits autophagic degradation in breast cancer MDA-MB-231 cells

Basic Medicine | 更新时间：2026-01-09

- Cepharanthine inhibits autophagic degradation in breast cancer MDA-MB-231 cells
- Journal of Third Military Medical University Vol. 39, Issue 24, Pages: 2352-2358(2017)
- 作者机构：
  
  400038 重庆，第三军医大学药学院生药学教研室
- 作者简介：
- 基金信息：
  
  the General Program of National Natural Science Foundation of China(31571425)
- DOI：10.16016/j.1000-5404.201709083
  CLC： R282.71; R285.1; R737.9
- Received：12 September 2017，
  
  Revised：2017-10-18，
  
  Published：30 December 2017
- 稿件说明：
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Qin DENG, Ruoqiu FU, Ning GAO. Cepharanthine inhibits autophagic degradation in breast cancer MDA-MB-231 cells[J]. Journal of Third Military Medical University, 2017, 39(24): 2352-2358.
DOI：

Qin DENG, Ruoqiu FU, Ning GAO. Cepharanthine inhibits autophagic degradation in breast cancer MDA-MB-231 cells[J]. Journal of Third Military Medical University, 2017, 39(24): 2352-2358. DOI： 10.16016/j.1000-5404.201709083.

摘要

目的

探讨千金藤素对乳腺癌细胞MDA-MB-231自噬的影响及其机制。

方法

激光共聚焦显微镜观察千金藤素对转染了EGFP-LC3、tfLC3(mRFP-EGFP-LC3)质粒的MDA-MB-231细胞自噬体形成及自噬流变化的影响；Western blot检测自噬相关蛋白LC3B、p62以及Beclin1的表达水平。激光共聚焦显微镜观察千金藤素、Bafilomycin A1、Rapamycin分别对转染了EGFP-LC3、tfLC3(mRFP-EGFP-LC3)质粒的MDA-MB-231细胞自噬体形成及自噬流变化的影响；Western blot检测溶酶体相关膜蛋白LAMP1、LAMP2的表达。

结果

千金藤素处理后细胞中的自噬体(EGFP-LC3荧光团)数量明显多于对照组，自噬相关蛋白LC3B-Ⅱ、p62的表达也呈量效和时效增多。转染tfLC3质粒后，千金藤素组和Bafilomycin A1组红绿荧光重叠呈现较多的黄色荧光团，而Rapamycin组出现较多的红色荧光团。千金藤素联用Bafilomycin A1并未进一步升高p62和LC3B-Ⅱ蛋白的表达水平，但联用Rapamycin明显升高了LC3B-Ⅱ蛋白的表达水平，同时，逆转了Rapamycin导致的p62蛋白表达水平减少。千金藤素既不影响对溶酶体的染色，也不减少LAMP1、LAMP2蛋白的表达水平，但影响了mRFP-LC3和mGFP-LAMP1的共定位。

结论

千金藤素通过阻断自噬体与溶酶体的融合抑制MDA-MB-231细胞自噬体降解。

Abstract

Objective

To investigate the effect of cepharanthine on autophagy in breast cancer MDA-MB-231 cells.

Methods

Confocal laser scanning microscopy was used to observe the formation of autophagosome and autophagic flux in breast cancer MDA-MB-231 cells after the transfection with EGFP-LC3 or tfLC3 plasmids. Western blot analysis was used to detect the expression of autophagy related proteins

including LC3B

p62 and Beclin1. Confocal laser scanning microscopy was used to observe the LysoTracker staining and the co-localization of autophagosome (mRFP-LC3) and lysosome (mGFP-LAMP1). Western blot analysis was also used to detect the expression of lysosomal-associated membrane proteins LAMP1 and LAMP2.

Results

The number of EGFP-LC3 puncta in the cells treated with cepharanthine was significantly greater than that in control cells (30.40±6.42

5.00±2.12

0.01)

and the expression levels of LC3B-Ⅱ and p62 were increased in a dose-and time-dependent manner. Exposure to

cepharanthine and bafilomycin A1 caused pronounced formation of LC3 puncta that displayed both green and red fluorescence intensity producing a yellow overlay after transfected with tfLC3. In contrast

the cells exposed to rapamycin led to the production of large amounts of red-only puncta. Cepharanthine combined with bafilomycin A1 did not show any significant increase in the accumulation of LC3B-Ⅱ and p62. In contrast

treatment with rapamycin resulted in a slight increase in the levels of LC3B-Ⅱ that were further enhanced by cepharanthine treatment. Furthermore

treatment with rapamycin resulted in decreased p62 levels that were markedly reversed by cepharanthine treatment. Further studies indicated that cepharanthine did not affect lysosomal staining nor decrease the expression of LAMP1 and LAMP2

but it affected the colocalization of mRFP-LC3 and mEGFP-LAMP1.

Conclusion

Cepharanthine inhibits autophagic degradation in breast cancer MDA-MB-231 cells through blocking autophagosome-lysosome fusion.

关键词

Keywords

references

YEDJOU C, IZEVBIGIE E, TCHOUNWOU P. Preclinical assessment of vernonia amygdalina leaf extracts as DNA damaging Anti-cancer agent in the management of breast cancer[J]. Int J Environ Res Public Health, 2008, 5(5): 337-341.

CHEN W, ZHENG R, BAADE P D, et al. Cancer statistics in China, 2015[J]. CA Cancer J Clin, 2016, 66(2): 115-132. DOI: 10.3322/caac.21338.

周静, 李国兵, 郑怡, 等.莲心碱对乳腺癌细胞MDA-MB-231自噬功能的影[J].第三军医大学学报, 2015, 37(7): 629-633. DOI: 10.16016/j.1000-5404.201411115.

ZHOU J, LI G B, ZHENG Y, et al. Liensinine inhibits autophagy in breast cancer MDA-MB-231 cells[J]. J Third Mil Med Univ, 2015, 37(7): 629-633. DOI:10.16016/j.1000-5404.201411115.

BAUER K R, BROWN M, CRESS R D, et al. Descriptive analysis of estrogen receptor (ER)-negative, progesterone receptor (PR)-negative, and HER2-negative invasive breast cancer, the so-called triple-negative phenotype[J]. Cancer, 2007, 109(9): 1721-1728. DOI: 10.1002/cncr.22618.

DESANTIS C E, LIN C C, MARIOTTO A B, et al. Cancer treatment and survivorship statistics, 2014[J]. CA Cancer J Clin, 2014, 64(4): 252-271. DOI: 10.3322/caac.21235.

JANKU F, MCCONKEY D J, HONG D S, et al. Autophagy as a target for anticancer therapy[J]. Nat Rev Clin Oncol, 2011, 8(9): 528-539. DOI: 10.1038/nrclinonc.2011.71.

LIANG D H, CHOI D S, ENSOR J E, et al. The autophagy inhibitor chloroquine targets cancer stem cells in triple negative breast cancer by inducing mitochondrial damage and impairing DNA break repair[J]. Cancer Lett, 2016, 376(2): 249-258. DOI: 10.1016/j.canlet.2016.04.002.

VALSECCHI M E, KIMMEY G, BIR A, et al. Role of carboplatin in the treatment of triple negative early-stage breast cancer[J]. Rev Recent Clin Trials, 2015, 10(2): 101-110. DOI: 10.2174/1574887110666150624101343.

李兴. 千金藤属植物地不容的生药学及化学成分研究[D]. 重庆: 西南大学, 2009. [http://cdmd.cnki.com.cn/Article/CDMD-10635-2009105399.htm .

LI X. Study on the chemical constituents and pharmacognosy of stephania epigaea[D]. Chongqing: Southwest University, 2009.

陆一安.千金藤素防治抗癌药物引起的白细胞减少症的临床使用经验[J].国外医学参考资料:药学分册, 1976 (5): 296-297. DOI: 10.13220/j.cnki.jipr.1976.05.013.

LU Y A. Cepharanthine prevention of leukopenia induced by anticancer drugs in clinical experience[J]. Foreign Med Sci: Pharmaceut Sci, 1976(5): 296-297. DOI: 10.13220/j.cnki.jipr.1976.05.013.

PAZYAR N, JAMSHYDIAN N. Cepharanthine: a therapeutic potential candidate for psoriasis[J]. J Altern Complement Med, 2012, 18(7): 639-640. DOI: 10.1089/acm.2012.0314.

彭有梅, 王宁, 王亚峰, 等.盐酸千金藤碱逆转K562/ADR细胞多药耐药性及其机制[J].药学学报, 2012, 47(5): 594-599. DOI: 10.16438/j.0513-4870.2012.05.015.

PENG Y M, WANG N, WANG Y F, et al. Correlation between reversing effect of cepharanthine hydrochloride on multidrug resistance and P-glycoprotein expression and function of K562/ADR cells[J]. Acta Pharm Sin, 2012, 47(5): 594-599. DOI: 10.16438/j.0513-4870.2012.05.015.

GANLEY I G, LAM DU H, WANG J, et al. ULK1.ATG13.FIP200 complex mediates mTOR signaling and is essential for autophagy[J]. J Biol Chem, 2009, 284(18): 12297-12305. DOI: 10.1074/jbc.M900573200.

ROSENFELD M R, YE X, SUPKO J G, et al. A phase Ⅰ/Ⅱ trial of hydroxychloroquine in conjunction with radiation therapy and concurrent and adjuvant temozolomide in patients with newly diagnosed glioblastoma multiforme[J]. Autophagy, 2014, 10(8): 1359-1368. DOI: 10.4161/auto.28984.

CYNTHIA I, CHUDE, RAVI K. Targeting autophagy in cancer: update on clinical trials and novel inhibitors[J]. Int J Mol Sci, 2017, 18(6): 1279. DOI: 10.3390/ijms1806 1279.

KREUZALER P, WATSON C J. Killing a cancer: what are the alternatives[J]. Nat RevCancer, 2012, 12(6): 411-424. DOI: 10.1038/nrc3264.

HUANG H, HU G, WANG C, et al. Cepharanthine, an alkaloid from stephania cepharantha hayata, inhibits the inflammatory response in the RAW264.7 cell and mouse models[J]. Inflammation, 2014, 37(1): 235-246. DOI: 10. 1007/s10753-013-9734-8.

苏彦宏, 李慧兰.左旋千金藤定碱抗心律失常作用的研究[J].昆明医学院学报, 2009, 30(10): 23-25.

SU Y H, LI H L.Anti-Arrhythmic effects of L-stepholidine[J]. J Kunming Med Univ, 2009, 30(10): 23-25.

ROGOSNITZKY M, DANKS R. Therapeutic potential of the biscoclaurine alkaloid, cepharanthine, for a range of clinical conditions[J]. Pharmacol Rep, 2011, 63(2): 337-347.

HUA P, SUN M, ZHANG G, et al. Cepharanthine induces apoptosis through reactive oxygen species and mitochondrial dysfunction in human non-small-cell lung cancer cells[J]. Biochem Biophys Res Commun, 2015, 460(2): 136-142. DOI: 10.1016/j.bbrc.2015.02.131.

SEUBWAI W, VAETEEWOOTTACHARN K, HIYOSHI M, et al. Cepharanthine exerts antitumor activity on cholangiocarcinoma by inhibiting NF-kappaB [J]. Cancer Sci, 2010, 101(7): 1590-1595. DOI: 10.1111/j.1349-7006.2010. 01572.x.

LAW B Y, MOK S W, CHAN W K, et al. Hernandezine, a novel AMPK activator induces autophagic cell death in drug-resistant cancers[J]. Oncotarget, 2016, 7(7): 8090-8104. DOI: 10.18632/oncotarget.6980.

REGGIORI F, UNGERMANN C. Autophagosome maturation and fusion[J]. J Mol Biol, 2017, 429(4): 486-496. DOI: 10.1016/j.jmb.2017.01.002.

NAKAMURA S, YOSHIMORI T. New insights into autophagosome-lysosome fusion[J].J Cell Sci, 2017, 130(7): 1209-1216. DOI: 10.1242/jcs.196352.

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