Jie Zhu1,2, Ya Zheng1,2, Haiyan Zhang1,2, Jing Zhu1,2, Hong Sun1. 1. Department of Gynecology, Obstetrics and Gynecology Hospital of Fudan UniversityShanghai 200011, China. 2. Shanghai Key Laboratory of Female Reproductive Endocrine Related DiseasesShanghai 200011, China.
Abstract
BACKGROUND: Cisplatin is a common used anti-tumor drug in ovarian cancer therapy with potent effect. Studies have reported that autophagy works as a cell-survival process in cancer, chloroquine has been added to various chemotherapeutic drugs. In the current study, we aim to evaluate whether chloroquine can enhance the effects of cisplatin in treating ovarian cancer. METHODS: CCK-8 assay was used to detect cell viability. Transwell assay was used to examine cell migration and invasion. Flow cytometry assay was applied to evaluate cell apoptosis. Western-blot assay was used to detect proteins related to apoptosis, autophagy and the AKT/mTOR pathway. RESULTS: In the current study, we showed that low concentration of chloroquine alone did not affect cell viability, migration or invasion, but it could enhance the efficacy of cisplatin in inhibiting cell viability, migration and invasion in both SKOV3 and hey cells. Afterwards, we observed that cisplatin triggered apoptosis and autophagy in both SKOV3 and hey cells in a dose-dependent manner. After treatment of cisplatin, SKOV3 and hey cells showed increased apoptotic rate in flow cytometry assay, increased protein levels of cleaved caspase 3, cleaved PARP and Bax, and decreased protein levels of Bcl-2 and Bcl-XL. Cisplatin also induced the formation of autophagosomes and increased autophagy-related proteins ATG 5, ATG 7, Beclin 1 and LC3B II/LC3B I. Meanwhile, cisplatin activated the AKT-mTOR pathway in both SKOV3 and hey cells. Next, chloroquine was added to ovarian cancer cells, flow cytometry assay revealed that chloroquine alone did not affect cell apoptosis and expressions of apoptosis-related proteins, while chloroquine plus cisplatin induced more apoptotic rate than cisplatin alone (p < 0.05). Meanwhile, apoptosis-related proteins had the same change trend. In vivo experiment demonstrated that chloroquine plus cisplatin was more effective than cisplatin alone in suppressing the growth of xenograft tumors, with lower ki-67 expression and higher cleaved caspase 3 expression. CONCLUSION: Based on our study, we propose that cisplatin activates the AKT/mTOR signaling pathway, which subsequently induces cytoprotective autophagy in ovarian cancer cells. Meanwhile, inhibition of autophagy via chloroquine enhances the anti-tumor effect of cisplatin.
BACKGROUND:Cisplatin is a common used anti-tumor drug in ovarian cancer therapy with potent effect. Studies have reported that autophagy works as a cell-survival process in cancer, chloroquine has been added to various chemotherapeutic drugs. In the current study, we aim to evaluate whether chloroquine can enhance the effects of cisplatin in treating ovarian cancer. METHODS: CCK-8 assay was used to detect cell viability. Transwell assay was used to examine cell migration and invasion. Flow cytometry assay was applied to evaluate cell apoptosis. Western-blot assay was used to detect proteins related to apoptosis, autophagy and the AKT/mTOR pathway. RESULTS: In the current study, we showed that low concentration of chloroquine alone did not affect cell viability, migration or invasion, but it could enhance the efficacy of cisplatin in inhibiting cell viability, migration and invasion in both SKOV3 and hey cells. Afterwards, we observed that cisplatin triggered apoptosis and autophagy in both SKOV3 and hey cells in a dose-dependent manner. After treatment of cisplatin, SKOV3 and hey cells showed increased apoptotic rate in flow cytometry assay, increased protein levels of cleaved caspase 3, cleaved PARP and Bax, and decreased protein levels of Bcl-2 and Bcl-XL. Cisplatin also induced the formation of autophagosomes and increased autophagy-related proteins ATG 5, ATG 7, Beclin 1 and LC3B II/LC3B I. Meanwhile, cisplatin activated the AKT-mTOR pathway in both SKOV3 and hey cells. Next, chloroquine was added to ovarian cancer cells, flow cytometry assay revealed that chloroquine alone did not affect cell apoptosis and expressions of apoptosis-related proteins, while chloroquine plus cisplatin induced more apoptotic rate than cisplatin alone (p < 0.05). Meanwhile, apoptosis-related proteins had the same change trend. In vivo experiment demonstrated that chloroquine plus cisplatin was more effective than cisplatin alone in suppressing the growth of xenograft tumors, with lower ki-67 expression and higher cleaved caspase 3 expression. CONCLUSION: Based on our study, we propose that cisplatin activates the AKT/mTOR signaling pathway, which subsequently induces cytoprotective autophagy in ovarian cancer cells. Meanwhile, inhibition of autophagy via chloroquine enhances the anti-tumor effect of cisplatin.
Entities:
Keywords:
AKT/mTOR signaling pathway; Chloroquine; autophagy; cisplatin; ovarian cancer
Authors: J Engel; R Eckel; G Schubert-Fritschle; J Kerr; W Kuhn; J Diebold; R Kimmig; J Rehbock; D Hölzel Journal: Eur J Cancer Date: 2002-12 Impact factor: 9.162
Authors: Brian A Boone; Nathan Bahary; Amer H Zureikat; A James Moser; Daniel P Normolle; Wen-Chi Wu; Aatur D Singhi; Phillip Bao; David L Bartlett; Lance A Liotta; Virginia Espina; Patricia Loughran; Michael T Lotze; Herbert J Zeh Journal: Ann Surg Oncol Date: 2015-04-24 Impact factor: 5.344
Authors: Molly L Bristol; Sean M Emery; Paola Maycotte; Andrew Thorburn; Shweta Chakradeo; David A Gewirtz Journal: J Pharmacol Exp Ther Date: 2013-01-04 Impact factor: 4.030
Authors: Jae Ryoung Hwang; Woo Young Kim; Young-Jae Cho; Ji-Yoon Ryu; Jung-Joo Choi; Soo Young Jeong; Myeong-Sun Kim; Ji Hye Kim; E Sun Paik; Yoo-Young Lee; Hee-Dong Han; Jeong-Won Lee Journal: Cell Death Dis Date: 2020-12-04 Impact factor: 8.469