Large-scale generation of functional mRNA-encapsulating exosomes via cellular nanoporation
Authors: Xiuting Zhao
Junfeng Shi
Yifan Ma
Yuanxin Chen
Yarong Zhao
Veysi Malkoc
Jing Xie
Andrew S Lee
Weiye Deng
Jingyao Sun
Paul Bertani
Betty Y S Kim
Yuan Fu
Jose Otero
June Rhee
Zhaogang Yang
L James Lee
Chen Kang
Chiling Chiang
Yamin Fan
Kwang J Kwak
Changcheng Yin
Kyuson Yun
Lesheng Teng
Tongzheng Liu
Yifan Wang
Xinmei Wang
Wu Lu
Wen Jiang
Journal: Nature Biomedical Engineering (2020)
Discipline: BIOLOGY & BIOCHEMISTRY
Keywords: Exosomes; nucleic-acid carriers; pharmacokinetic; immunological properties; physiological barriers
Affiliations: Department of Chemical and Biomolecular Engineering, The Ohio State University, Columbus, OH, USA., Department of Radiation Oncology, University of Texas Southwestern Medical Center, Dallas, TX, USA., School of Life Sciences, Jilin University, Changchun, China., College of Pharmacy, Jinan University, Guangzhou, China., College of Medicine, The Ohio State University, Columbus, OH, USA., Department of Neurological Surgery, Mayo Clinic, Jacksonville, FL, USA., Carver College of Medicine, University of Iowa, Iowa City, IA, USA., Department of Biophysics, Health Science Center, Peking University, Beijing, China., Institute for Stem Cell Biology, Stanford University, Palo Alto, CA, USA., Department of Electrical and Computer Engineering, The Ohio State University, Columbus, OH, USA., Department of Neuroscience, The Ohio State University, Columbus, OH, USA., Peak Center for Pituitary and Brain Tumor, Department of Neurosurgery, Houston Methodist Research institute, Weill-Cornell Medical College, Houston, TX, USA., Institute of Cancer Research, Shenzhen Bay Laboratory, Shenzhen, China., School of Life Sciences, Jilin University, Changchun, China. ., Department of Neurological Surgery, Mayo Clinic, Jacksonville, FL, USA. ., Department of Neurosurgery, The University of Texas MD Anderson Cancer Center, Houston, TX, USA. ., Department of Chemical and Biomolecular Engineering, The Ohio State University, Columbus, OH, USA. .

Abstract:  Exosomes are attractive as nucleic-acid carriers because of their favourable pharmacokinetic and immunological properties and their ability to penetrate physiological barriers that are impermeable to synthetic drug-delivery vehicles. However, inserting exogenous nucleic acids, especially large messenger RNAs, into cell-secreted exosomes leads to low yields. Here we report a cellular-nanoporation method for the production of large quantities of exosomes containing therapeutic mRNAs and targeting peptides. We transfected various source cells with plasmid DNAs and stimulated the cells with a focal and transient electrical stimulus that promotes the release of exosomes carrying transcribed mRNAs and targeting peptides. Compared with bulk electroporation and other exosome-production strategies, cellular nanoporation produced up to 50-fold more exosomes and a more than 103-fold increase in exosomal mRNA transcripts, even from cells with low basal levels of exosome secretion. In orthotopic phosphatase and tensin homologue (PTEN)-deficient glioma mouse models, mRNA-containing exosomes restored tumour-suppressor function, enhanced inhibition of tumour growth and increased survival. Cellular nanoporation may enable the use of exosomes as a universal nucleic-acid carrier for applications requiring transcriptional manipulation. 

Retracted: No

PubPeer comments:
 One animal and figure 8a appears to be featured twice, with different dimensions and fluorescence. If correct, this may not affect the conclusions, but would nonetheless be a serious ethical concern. ***  , 
 Same issue in 8g. If this concern is valid, this might affect the conclusions. I do not see how these fluorescence differences could occur by accident. ***  , 
 #1 Thanks for raising these concerns. First, in order to save the animal processing time, we took both the luciferase and fluorescence at 1 hour after iv injection. We should have stated this clearly in the method part. For the second question, we are terribly sorry for the mistake we made in plotting this fig. The luciferase images were taken at different days with different luminescence scales. In order to bring them to the same scale into one fig, we adjusted the scales after we obtained all the images. I noticed that image (PBS, day 12) is the original image we saved. However, during the data processing, image (Exo-T, day 3) was wrongly saved and it replaced the original (Exo-T, day 3) image. We contacted with the person who helped us process these images, and he confirmed that he made the mistake during this process. Meanwhile, we are double-checking all the original images to see if we made any other mistakes during the image processing. Thanks again for bring up these issues. , 
 #2 After carefully checking all the original images, we noticed that we wrongly processed all the images in Day 0 and Day 3 in Fig 8g. We are contacting the editors for this. Thanks again for letting us know. ,