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Prediction and validation of hematopoietic stem and progenitor cell off-target editing in transplanted rhesus macaques
DC Field | Value | Language |
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dc.contributor.author | AlJanahi, Aisha A. | - |
dc.contributor.author | Lazzarotto, Cicera R. | - |
dc.contributor.author | Chen, Shirley | - |
dc.contributor.author | Shin, Tae-Hoon | - |
dc.contributor.author | Cordes, Stefan | - |
dc.contributor.author | Fan, Xing | - |
dc.contributor.author | Jabara, Isabel | - |
dc.contributor.author | Zhou, Yifan | - |
dc.contributor.author | Young, David J. | - |
dc.contributor.author | Lee, Byung-Chul | - |
dc.contributor.author | Yu, Kyung-Rok | - |
dc.contributor.author | Li, Yuesheng | - |
dc.contributor.author | Toms, Bradley | - |
dc.contributor.author | Tunc, Ilker | - |
dc.contributor.author | Hong, So Gun | - |
dc.contributor.author | Truitt, Lauren L. | - |
dc.contributor.author | Klermund, Julia | - |
dc.contributor.author | Andrieux, Geoffroy | - |
dc.contributor.author | Kim, Miriam Y. | - |
dc.contributor.author | Cathomen, Toni | - |
dc.contributor.author | Gill, Saar | - |
dc.contributor.author | Tsai, Shengdar Q. | - |
dc.contributor.author | Dunbar, Cynthia E. | - |
dc.date.accessioned | 2022-05-16T08:54:23Z | - |
dc.date.available | 2022-05-16T08:54:23Z | - |
dc.date.created | 2022-02-04 | - |
dc.date.created | 2022-02-04 | - |
dc.date.issued | 2022-01-05 | - |
dc.identifier.citation | Molecular Therapy, Vol.30 No.1, pp.209-222 | - |
dc.identifier.issn | 1525-0016 | - |
dc.identifier.uri | https://hdl.handle.net/10371/179811 | - |
dc.description.abstract | The programmable nuclease technology CRISPR-Cas9 has revolutionized gene editing in the last decade. Due to the risk of off-target editing, accurate and sensitive methods for off-target characterization are crucial prior to applying CRISPR-Cas9 therapeutically. Here, we utilized a rhesus macaque model to compare the predictive values of CIRCLE-seq, an in vitro off-target prediction method, with in silico prediction (ISP) based solely on genomic sequence comparisons. We use AmpliSeq HD error-corrected sequencing to validate offtarget sites predicted by CIRCLE-seq and ISP for a CD33 guide RNA (gRNA) with thousands of off-target sites predicted by ISP and CIRCLE-seq. We found poor correlation between the sites predicted by the two methods. When almost 500 sites predicted by each method were analyzed by error-corrected sequencing of hematopoietic cells following transplantation, 19 off-target sites revealed insertion or deletion mutations. Of these sites, 8 were predicted by both methods, 8 by CIRCLE-seq only, and 3 by ISP only. The levels of cells with these off-target edits exhibited no expansion or abnormal behavior in vivo in animals followed for up to 2 years. In addition, we utilized an unbiased method termed CAST-seq to search for translocations between the on-target site and off-target sites present in animals following transplantation, detecting one specific translocation that persisted in blood cells for at least 1 year following transplantation. In conclusion, neither CIRCLE-seq or ISP predicted all sites, and a combination of careful gRNA design, followed by screening for predicted off-target sites in target cells by multiple methods, may be required for optimizing safety of clinical development. | - |
dc.language | 영어 | - |
dc.publisher | Nature Publishing Group | - |
dc.title | Prediction and validation of hematopoietic stem and progenitor cell off-target editing in transplanted rhesus macaques | - |
dc.type | Article | - |
dc.identifier.doi | 10.1016/j.ymthe.2021.06.016 | - |
dc.citation.journaltitle | Molecular Therapy | - |
dc.identifier.wosid | 000744579600003 | - |
dc.identifier.scopusid | 2-s2.0-85114667768 | - |
dc.citation.endpage | 222 | - |
dc.citation.number | 1 | - |
dc.citation.startpage | 209 | - |
dc.citation.volume | 30 | - |
dc.description.isOpenAccess | N | - |
dc.contributor.affiliatedAuthor | Yu, Kyung-Rok | - |
dc.type.docType | Article | - |
dc.description.journalClass | 1 | - |
dc.subject.keywordPlus | CHROMOSOMAL TRANSLOCATIONS | - |
dc.subject.keywordPlus | MOUSE MODEL | - |
dc.subject.keywordPlus | GENOME | - |
dc.subject.keywordPlus | GENE | - |
dc.subject.keywordPlus | CRISPR-CAS9 | - |
dc.subject.keywordPlus | DELIVERY | - |
dc.subject.keywordPlus | SEQ | - |
dc.subject.keywordPlus | DNA | - |
dc.subject.keywordPlus | SPECIFICITY | - |
dc.subject.keywordPlus | EXPRESSION | - |
dc.subject.keywordAuthor | Ca9 | - |
dc.subject.keywordAuthor | CRISPR | - |
dc.subject.keywordAuthor | error-corrected sequencing | - |
dc.subject.keywordAuthor | gene editing | - |
dc.subject.keywordAuthor | gene therapy | - |
dc.subject.keywordAuthor | Macaque | - |
dc.subject.keywordAuthor | off-target | - |
dc.subject.keywordAuthor | translocation | - |
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