An effective vaginal gel to deliver CRISPR/Cas9 system encapsulated in poly (β-amino ester) nanoparticles for vaginal gene therapy
Gang Niu,
Zhuang Jin,
Chong Zhang,
Dan He,
Xueqin Gao,
Chenming Zou,
Wei Zhang,
Jiahui Ding,
Bhudev C. Das,
Konstantin Severinov,
Inga Isabel Hitzeroth,
Priya Ranjan Debata,
Xin Ma,
Xun Tian,
Qinglei Gao,
Jun Wu,
Zeshan You,
Rui Tian,
Zifeng Cui,
Weiwen Fan,
Weiling Xie,
Zhaoyue Huang,
Chen Cao,
Wei Xu,
Hongxian Xie,
Hongyan Xu,
Xiongzhi Tang,
Yan Wang,
Zhiying Yu,
Hui Han,
Songwei Tan,
Shuqin Chen,
Zheng Hu
Affiliations
Gang Niu
Department of Obstetrics and Gynecology, the First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, China
Zhuang Jin
Department of Obstetrics and Gynecology, the First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, China
Chong Zhang
School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Dan He
Department of Neurology, the First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, Guangdong, China
Xueqin Gao
Department of Pharmacy, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China
Chenming Zou
School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Wei Zhang
School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Jiahui Ding
School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Bhudev C. Das
Amity Institute of Molecular Medicine & Stem Cell Research, Amity University, Uttar Pradesh, Noida 201313, India
Konstantin Severinov
Skolkovo Institute of Science and Technology, Skolkovo, Moscow Region 143025, Russian Federation
Inga Isabel Hitzeroth
Biopharming Research Unit, Department of Molecular and Cell Biology, University of Cape Town, Cape Town 7701, South Africa
Priya Ranjan Debata
Department of Zoology, North Orissa University, Takatpur, Baripada, Odisha 757003, India
Xin Ma
Department of Urology, General Hospital of People's Liberation Army, Beijing 100039, China
Xun Tian
Department of Obstetrics and Gynecology, Academician expert workstation, The Central Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430014, Hubei, China
Qinglei Gao
Department of Obstetrics and Gynecology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, Hubei, China
Jun Wu
School of Biomedical Engineering, Sun Yat-sen University, Guangzhou 510006, Guangdong, China
Zeshan You
Department of Obstetrics and Gynecology, the First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, China
Rui Tian
Department of Obstetrics and Gynecology, the First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, China
Zifeng Cui
Department of Obstetrics and Gynecology, the First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, China
Weiwen Fan
Department of Obstetrics and Gynecology, the First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, China
Weiling Xie
Department of Obstetrics and Gynecology, the First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, China
Zhaoyue Huang
Department of Obstetrics and Gynecology, the First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, China
Chen Cao
Department of Obstetrics and Gynecology, Academician expert workstation, The Central Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430014, Hubei, China
Wei Xu
Department of Obstetrics and Gynecology, the First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, China
Hongxian Xie
Generulor Company Bio-X Lab, Guangzhou 510006, Guangdong, China
Hongyan Xu
Department of Obstetrics and Gynecology, Yuebei People's Hospital, Medical College of Shantou University, Shaoguan 512026, Guangdong, China
Xiongzhi Tang
Department of Obstetrics and Gynecology, Guilin People's Hospital, Guilin, The Guangxi Zhuang Autonomous Region, 541002, China
Yan Wang
Key Laboratory of Molecular Biophysics of the Ministry of Education, School of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, Hubei 430074, China
Zhiying Yu
Department of Obstetrics & Gynecology, First Affiliated Hospital of Shenzhen University, Shenzhen 518000, Guangdong, China
Hui Han
State Key Laboratory of Oncology in Southern China, Collaborative Innovation Center for Cancer Medicine & Department of Urology, Yat-sen University Cancer Center, Guangzhou 510080, Guangdong Province, China
Songwei Tan
School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China; Corresponding authors.
Shuqin Chen
Department of Obstetrics and Gynecology, the First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, China; Corresponding authors.
Zheng Hu
Department of Obstetrics and Gynecology, the First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, China; Department of Obstetrics and Gynecology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, Hubei, China; Precision Medicine Institute, the First Affiliated Hospital, Sun Yat-sen University, Guangzhou 510080, China; Corresponding authors.
Background: Gene therapy has held promises for treating specific genetic diseases. However, the key to clinical application depends on effective gene delivery. Methods: Using a large animal model, we developed two pharmaceutical formulations for gene delivery in the pigs’ vagina, which were made up of poly (β-amino ester) (PBAE)-plasmid polyplex nanoparticles (NPs) based two gel materials, modified montmorillonite (mMMT) and hectorite (HTT). Findings: By conducting flow cytometry of the cervical cells, we found that PBAE-GFP-NPs-mMMT gel was more efficient than PBAE-GFP-NPs-HTT gel in delivering exogenous DNA intravaginally. Next, we designed specific CRISPR/SpCas9 sgRNAs targeting porcine endogenous retroviruses (PERVs) and evaluated the genome editing efficacy in vivo. We discovered that PERV copy number in vaginal epithelium could be significantly reduced by the local delivery of the PBAE-SpCas9/sgRNA NPs-mMMT gel. Comparable genome editing results were also obtained by high-fidelity version of SpCas9, SpCas9-HF1 and eSpCas9, in the mMMT gel. Further, we confirmed that the expression of topically delivered SpCas9 was limited to the vagina/cervix and did not diffuse to nearby organs, which was relatively safe with low toxicity. Interpretation: Our data suggested that the PBAE-NPs mMMT vaginal gel is an effective preparation for local gene therapy, yielding insights into novel therapeutic approaches to sexually transmitted disease in the genital tract. Funding: This work was supported by the National Science and Technology Major Project of the Ministry of science and technology of China (No. 2018ZX10301402); the National Natural Science Foundation of China (81761148025, 81871473 and 81402158); Guangzhou Science and Technology Programme (No. 201704020093); National Ten Thousand Plan-Young Top Talents of China, Fundamental Research Funds for the Central Universities (17ykzd15 and 19ykyjs07); Three Big Constructions—Supercomputing Application Cultivation Projects sponsored by National Supercomputer Center In Guangzhou; the National Research FFoundation (NRF) South Africa under BRICS Multilateral Joint Call for Proposals; grant 17–54–80078 from the Russian Foundation for Basic Research.
