In vitro and in vivo growth inhibition of human cervical cancer cells via human papillomavirus E6/E7 mRNAs’ cleavage by CRISPR/Cas13a system

Sustained infection of high-risk human papillomavirus (HR-HPVs), especially HPV16 and HPV18, is a major cause of cervical cancer. E6 and E7 oncoproteins, encoded by the HPV genome, are critical for transformation and maintenance of malignant phenotypes of cervical cancer. Here, we used an emerging p...

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Published inAntiviral research Vol. 178; p. 104794
Main Authors Chen, Yili, Jiang, Hongye, Wang, Ting, He, Dan, Tian, Rui, Cui, Zifeng, Tian, Xun, Gao, Qinglei, Ma, Xin, Yang, Jianrong, Wu, Jun, Tan, Songwei, Xu, Hongyan, Tang, Xiongzhi, Wang, Yan, Yu, Zhiying, Han, Hui, Das, Bhudev C., Severinov, Konstantin, Hitzeroth, Inga Isabel, Debata, Priya Ranjan, Xu, Wei, Fan, Weiwen, Jin, Zhuang, Cao, Chen, Yu, Miao, Xie, Weiling, Huang, Zhaoyue, Hu, Zheng, You, Zeshan
Format Journal Article
LanguageEnglish
Published Netherlands Elsevier B.V 01.06.2020
Subjects
Animals
Apoptosis
Cell Line, Tumor
Cell Proliferation
Cervical cancer
CRISPR-Cas Systems
CRISPR/Cas13a system
DNA Breaks, Double-Stranded
DNA-Binding Proteins - genetics
Down-Regulation
E6/E7
Female
Genetic Therapy
Growth inhibition
HeLa Cells
HPV 16/18
Human papillomavirus 16 - genetics
Human papillomavirus 18 - genetics
Humans
Mice
Mice, Inbred BALB C
Oncogene Proteins, Viral - genetics
Papillomavirus E7 Proteins - genetics
Papillomavirus Infections - virology
Repressor Proteins - genetics
Retinoblastoma Binding Proteins - genetics
RNA, Messenger - genetics
RNA, Messenger - metabolism
RNA, Viral - genetics
RNA, Viral - metabolism
Tumor suppression
Tumor Suppressor Protein p53 - genetics
Ubiquitin-Protein Ligases - genetics
Up-Regulation
Uterine Cervical Neoplasms - pathology
Uterine Cervical Neoplasms - virology
Xenograft Model Antitumor Assays
HPV 16/18
Tumor suppression
CRISPR/Cas13a system
Growth inhibition
E6/E7
Cervical cancer
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Abstract Sustained infection of high-risk human papillomavirus (HR-HPVs), especially HPV16 and HPV18, is a major cause of cervical cancer. E6 and E7 oncoproteins, encoded by the HPV genome, are critical for transformation and maintenance of malignant phenotypes of cervical cancer. Here, we used an emerging programmable clustered regularly interspaced short palindromic repeat (CRISPR)/Cas13a system to cleave HPV 16/18 E6/E7 messenger RNAs (mRNAs). The results showed that customized CRISPR/Cas13a system effectively and specifically knocked down HPV 16/18 E6/E7 mRNAs, inducing growth inhibition and apoptosis in HPV16-positive SiHa and HPV18-positive HeLa Cell lines, but not in HPV-negative C33A cell line. Simultaneously, we detected downregulation of E6/E7 oncoproteins and upregulation of tumor suppressor P53 and RB proteins. In addition, we used subcutaneous xenograft tumor growth assays to find that the weight and volume of tumors in the SiHa-16E6CR1 group knocked down by the CRISPR/Cas13a system were significantly lower than those in the SiHa-VECTOR group lacking crRNA. Our study demonstrated that targeting HPV E6/E7 mRNAs by the CRISPR/Cas13a system may be a candidate therapeutic strategy for HPV-related cervical cancer. [Display omitted] •CRISPR/Cas13a system was designed and constructed to cleave HPV 16/18 E6/E7 mRNAs.•CRISPR/Cas13a system effectively and specifically knocked down E6/E7 mRNAs and restored P53 and RB proteins.•CRISPR/Cas13a system induced growth inhibition and apoptosis in human cervical cancer cells.•CRISPR/Cas13a system suppressed tumor growth in vivo effectively and specifically.
AbstractList Sustained infection of high-risk human papillomavirus (HR-HPVs), especially HPV16 and HPV18, is a major cause of cervical cancer. E6 and E7 oncoproteins, encoded by the HPV genome, are critical for transformation and maintenance of malignant phenotypes of cervical cancer. Here, we used an emerging programmable clustered regularly interspaced short palindromic repeat (CRISPR)/Cas13a system to cleave HPV 16/18 E6/E7 messenger RNAs (mRNAs). The results showed that customized CRISPR/Cas13a system effectively and specifically knocked down HPV 16/18 E6/E7 mRNAs, inducing growth inhibition and apoptosis in HPV16-positive SiHa and HPV18-positive HeLa Cell lines, but not in HPV-negative C33A cell line. Simultaneously, we detected downregulation of E6/E7 oncoproteins and upregulation of tumor suppressor P53 and RB proteins. In addition, we used subcutaneous xenograft tumor growth assays to find that the weight and volume of tumors in the SiHa-16E6CR1 group knocked down by the CRISPR/Cas13a system were significantly lower than those in the SiHa-VECTOR group lacking crRNA. Our study demonstrated that targeting HPV E6/E7 mRNAs by the CRISPR/Cas13a system may be a candidate therapeutic strategy for HPV-related cervical cancer.Sustained infection of high-risk human papillomavirus (HR-HPVs), especially HPV16 and HPV18, is a major cause of cervical cancer. E6 and E7 oncoproteins, encoded by the HPV genome, are critical for transformation and maintenance of malignant phenotypes of cervical cancer. Here, we used an emerging programmable clustered regularly interspaced short palindromic repeat (CRISPR)/Cas13a system to cleave HPV 16/18 E6/E7 messenger RNAs (mRNAs). The results showed that customized CRISPR/Cas13a system effectively and specifically knocked down HPV 16/18 E6/E7 mRNAs, inducing growth inhibition and apoptosis in HPV16-positive SiHa and HPV18-positive HeLa Cell lines, but not in HPV-negative C33A cell line. Simultaneously, we detected downregulation of E6/E7 oncoproteins and upregulation of tumor suppressor P53 and RB proteins. In addition, we used subcutaneous xenograft tumor growth assays to find that the weight and volume of tumors in the SiHa-16E6CR1 group knocked down by the CRISPR/Cas13a system were significantly lower than those in the SiHa-VECTOR group lacking crRNA. Our study demonstrated that targeting HPV E6/E7 mRNAs by the CRISPR/Cas13a system may be a candidate therapeutic strategy for HPV-related cervical cancer.
Sustained infection of high-risk human papillomavirus (HR-HPVs), especially HPV16 and HPV18, is a major cause of cervical cancer. E6 and E7 oncoproteins, encoded by the HPV genome, are critical for transformation and maintenance of malignant phenotypes of cervical cancer. Here, we used an emerging programmable clustered regularly interspaced short palindromic repeat (CRISPR)/Cas13a system to cleave HPV 16/18 E6/E7 messenger RNAs (mRNAs). The results showed that customized CRISPR/Cas13a system effectively and specifically knocked down HPV 16/18 E6/E7 mRNAs, inducing growth inhibition and apoptosis in HPV16-positive SiHa and HPV18-positive HeLa Cell lines, but not in HPV-negative C33A cell line. Simultaneously, we detected downregulation of E6/E7 oncoproteins and upregulation of tumor suppressor P53 and RB proteins. In addition, we used subcutaneous xenograft tumor growth assays to find that the weight and volume of tumors in the SiHa-16E6CR1 group knocked down by the CRISPR/Cas13a system were significantly lower than those in the SiHa-VECTOR group lacking crRNA. Our study demonstrated that targeting HPV E6/E7 mRNAs by the CRISPR/Cas13a system may be a candidate therapeutic strategy for HPV-related cervical cancer.
Sustained infection of high-risk human papillomavirus (HR-HPVs), especially HPV16 and HPV18, is a major cause of cervical cancer. E6 and E7 oncoproteins, encoded by the HPV genome, are critical for transformation and maintenance of malignant phenotypes of cervical cancer. Here, we used an emerging programmable clustered regularly interspaced short palindromic repeat (CRISPR)/Cas13a system to cleave HPV 16/18 E6/E7 messenger RNAs (mRNAs). The results showed that customized CRISPR/Cas13a system effectively and specifically knocked down HPV 16/18 E6/E7 mRNAs, inducing growth inhibition and apoptosis in HPV16-positive SiHa and HPV18-positive HeLa Cell lines, but not in HPV-negative C33A cell line. Simultaneously, we detected downregulation of E6/E7 oncoproteins and upregulation of tumor suppressor P53 and RB proteins. In addition, we used subcutaneous xenograft tumor growth assays to find that the weight and volume of tumors in the SiHa-16E6CR1 group knocked down by the CRISPR/Cas13a system were significantly lower than those in the SiHa-VECTOR group lacking crRNA. Our study demonstrated that targeting HPV E6/E7 mRNAs by the CRISPR/Cas13a system may be a candidate therapeutic strategy for HPV-related cervical cancer. [Display omitted] •CRISPR/Cas13a system was designed and constructed to cleave HPV 16/18 E6/E7 mRNAs.•CRISPR/Cas13a system effectively and specifically knocked down E6/E7 mRNAs and restored P53 and RB proteins.•CRISPR/Cas13a system induced growth inhibition and apoptosis in human cervical cancer cells.•CRISPR/Cas13a system suppressed tumor growth in vivo effectively and specifically.
ArticleNumber 104794
Author Das, Bhudev C.
Severinov, Konstantin
Yang, Jianrong
Chen, Yili
Tian, Xun
Tan, Songwei
Gao, Qinglei
Tian, Rui
Xu, Wei
Wang, Yan
Yu, Zhiying
Cui, Zifeng
Wu, Jun
Han, Hui
Ma, Xin
Jin, Zhuang
He, Dan
Xu, Hongyan
Cao, Chen
Jiang, Hongye
Fan, Weiwen
Xie, Weiling
Hitzeroth, Inga Isabel
Debata, Priya Ranjan
Wang, Ting
Yu, Miao
Huang, Zhaoyue
Tang, Xiongzhi
Hu, Zheng
You, Zeshan
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  organization: Department of Obstetrics and Gynecology, The First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, China
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  givenname: Qinglei
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– sequence: 9
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  fullname: Ma, Xin
  organization: Department of Urology, The General Hospital of the People's Liberation Army, Beijing, China
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  fullname: Yang, Jianrong
  organization: Department of Biology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China
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  surname: Wu
  fullname: Wu, Jun
  organization: School of Biomedical Engineering, Sun Yat-sen University, Guangzhou, China
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  organization: Tongji School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
– sequence: 13
  givenname: Hongyan
  surname: Xu
  fullname: Xu, Hongyan
  organization: Department of Obstetrics and Gynecology, Yuebei People's Hospital, Medical College of Shantou University, Shaoguan, Guangdong, China
– sequence: 14
  givenname: Xiongzhi
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  organization: Key Laboratory of Molecular Biophysics of the Ministry of Education, School of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, Hubei, China
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  surname: Han
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  organization: State Key Laboratory of Oncology in Southern China, Collaborative Innovation Center for Cancer Medicine & Department of Urology, Sun Yat-sen University Cancer Center, Guangzhou, Guangdong, China
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  givenname: Chen
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  organization: Department of Obstetrics and Gynecology, Academician Expert Workstation, The Central Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China
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  givenname: Miao
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Keywords HPV 16/18
Tumor suppression
CRISPR/Cas13a system
Growth inhibition
E6/E7
Cervical cancer
Language English
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PublicationDecade 2020
PublicationPlace Netherlands
PublicationPlace_xml – name: Netherlands
PublicationTitle Antiviral research
PublicationTitleAlternate Antiviral Res
PublicationYear 2020
Publisher Elsevier B.V
Publisher_xml – name: Elsevier B.V
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Snippet Sustained infection of high-risk human papillomavirus (HR-HPVs), especially HPV16 and HPV18, is a major cause of cervical cancer. E6 and E7 oncoproteins,...
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SubjectTerms Animals
Apoptosis
Cell Line, Tumor
Cell Proliferation
Cervical cancer
CRISPR-Cas Systems
CRISPR/Cas13a system
DNA Breaks, Double-Stranded
DNA-Binding Proteins - genetics
Down-Regulation
E6/E7
Female
Genetic Therapy
Growth inhibition
HeLa Cells
HPV 16/18
Human papillomavirus 16 - genetics
Human papillomavirus 18 - genetics
Humans
Mice
Mice, Inbred BALB C
Oncogene Proteins, Viral - genetics
Papillomavirus E7 Proteins - genetics
Papillomavirus Infections - virology
Repressor Proteins - genetics
Retinoblastoma Binding Proteins - genetics
RNA, Messenger - genetics
RNA, Messenger - metabolism
RNA, Viral - genetics
RNA, Viral - metabolism
Tumor suppression
Tumor Suppressor Protein p53 - genetics
Ubiquitin-Protein Ligases - genetics
Up-Regulation
Uterine Cervical Neoplasms - pathology
Uterine Cervical Neoplasms - virology
Xenograft Model Antitumor Assays
Title In vitro and in vivo growth inhibition of human cervical cancer cells via human papillomavirus E6/E7 mRNAs’ cleavage by CRISPR/Cas13a system
URI https://dx.doi.org/10.1016/j.antiviral.2020.104794
https://www.ncbi.nlm.nih.gov/pubmed/32298665
https://www.proquest.com/docview/2391977804
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