TRIAMF: A New Method for Delivery of Cas9 Ribonucleoprotein Complex to Human Hematopoietic Stem Cells

Abstract CRISPR/Cas9 mediated gene editing of patient-derived hematopoietic stem and progenitor cells (HSPCs) ex vivo followed by autologous transplantation of the edited HSPCs back to the patient can provide a potential cure for monogenic blood disorders such as β-hemoglobinopathies. One challenge...

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Autores principales:	Jonathan Yen, Michael Fiorino, Yi Liu, Steve Paula, Scott Clarkson, Lisa Quinn, William R. Tschantz, Heath Klock, Ning Guo, Carsten Russ, Vionnie W. C. Yu, Craig Mickanin, Susan C. Stevenson, Cameron Lee, Yi Yang
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2018
Materias:	Hematopoietic Stem And Progenitor Cells (HSPCs) Purified Cas9 Protein HSPC Expansion Neon Electroporation Syringe Connector Medicine R Science Q
Acceso en línea:	https://doaj.org/article/7126e75f7f804ba7a87cf87864786123
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spelling	oai:doaj.org-article:7126e75f7f804ba7a87cf878647861232021-12-02T15:08:00ZTRIAMF: A New Method for Delivery of Cas9 Ribonucleoprotein Complex to Human Hematopoietic Stem Cells10.1038/s41598-018-34601-62045-2322https://doaj.org/article/7126e75f7f804ba7a87cf878647861232018-11-01T00:00:00Zhttps://doi.org/10.1038/s41598-018-34601-6https://doaj.org/toc/2045-2322Abstract CRISPR/Cas9 mediated gene editing of patient-derived hematopoietic stem and progenitor cells (HSPCs) ex vivo followed by autologous transplantation of the edited HSPCs back to the patient can provide a potential cure for monogenic blood disorders such as β-hemoglobinopathies. One challenge for this strategy is efficient delivery of the ribonucleoprotein (RNP) complex, consisting of purified Cas9 protein and guide RNA, into HSPCs. Because β-hemoglobinopathies are most prevalent in developing countries, it is desirable to have a reliable, efficient, easy-to-use and cost effective delivery method. With this goal in mind, we developed TRansmembrane Internalization Assisted by Membrane Filtration (TRIAMF), a new method to quickly and effectively deliver RNPs into HSPCs by passing a RNP and cell mixture through a filter membrane. We achieved robust gene editing in HSPCs using TRIAMF and demonstrated that the multilineage colony forming capacities and the competence for engraftment in immunocompromised mice of HSPCs were preserved post TRIAMF treatment. TRIAMF is a custom designed system using inexpensive components and has the capacity to process HSPCs at clinical scale.Jonathan YenMichael FiorinoYi LiuSteve PaulaScott ClarksonLisa QuinnWilliam R. TschantzHeath KlockNing GuoCarsten RussVionnie W. C. YuCraig MickaninSusan C. StevensonCameron LeeYi YangNature PortfolioarticleHematopoietic Stem And Progenitor Cells (HSPCs)Purified Cas9 ProteinHSPC ExpansionNeon ElectroporationSyringe ConnectorMedicineRScienceQENScientific Reports, Vol 8, Iss 1, Pp 1-11 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	Hematopoietic Stem And Progenitor Cells (HSPCs) Purified Cas9 Protein HSPC Expansion Neon Electroporation Syringe Connector Medicine R Science Q
spellingShingle	Hematopoietic Stem And Progenitor Cells (HSPCs) Purified Cas9 Protein HSPC Expansion Neon Electroporation Syringe Connector Medicine R Science Q Jonathan Yen Michael Fiorino Yi Liu Steve Paula Scott Clarkson Lisa Quinn William R. Tschantz Heath Klock Ning Guo Carsten Russ Vionnie W. C. Yu Craig Mickanin Susan C. Stevenson Cameron Lee Yi Yang TRIAMF: A New Method for Delivery of Cas9 Ribonucleoprotein Complex to Human Hematopoietic Stem Cells
description	Abstract CRISPR/Cas9 mediated gene editing of patient-derived hematopoietic stem and progenitor cells (HSPCs) ex vivo followed by autologous transplantation of the edited HSPCs back to the patient can provide a potential cure for monogenic blood disorders such as β-hemoglobinopathies. One challenge for this strategy is efficient delivery of the ribonucleoprotein (RNP) complex, consisting of purified Cas9 protein and guide RNA, into HSPCs. Because β-hemoglobinopathies are most prevalent in developing countries, it is desirable to have a reliable, efficient, easy-to-use and cost effective delivery method. With this goal in mind, we developed TRansmembrane Internalization Assisted by Membrane Filtration (TRIAMF), a new method to quickly and effectively deliver RNPs into HSPCs by passing a RNP and cell mixture through a filter membrane. We achieved robust gene editing in HSPCs using TRIAMF and demonstrated that the multilineage colony forming capacities and the competence for engraftment in immunocompromised mice of HSPCs were preserved post TRIAMF treatment. TRIAMF is a custom designed system using inexpensive components and has the capacity to process HSPCs at clinical scale.
format	article
author	Jonathan Yen Michael Fiorino Yi Liu Steve Paula Scott Clarkson Lisa Quinn William R. Tschantz Heath Klock Ning Guo Carsten Russ Vionnie W. C. Yu Craig Mickanin Susan C. Stevenson Cameron Lee Yi Yang
author_facet	Jonathan Yen Michael Fiorino Yi Liu Steve Paula Scott Clarkson Lisa Quinn William R. Tschantz Heath Klock Ning Guo Carsten Russ Vionnie W. C. Yu Craig Mickanin Susan C. Stevenson Cameron Lee Yi Yang
author_sort	Jonathan Yen
title	TRIAMF: A New Method for Delivery of Cas9 Ribonucleoprotein Complex to Human Hematopoietic Stem Cells
title_short	TRIAMF: A New Method for Delivery of Cas9 Ribonucleoprotein Complex to Human Hematopoietic Stem Cells
title_full	TRIAMF: A New Method for Delivery of Cas9 Ribonucleoprotein Complex to Human Hematopoietic Stem Cells
title_fullStr	TRIAMF: A New Method for Delivery of Cas9 Ribonucleoprotein Complex to Human Hematopoietic Stem Cells
title_full_unstemmed	TRIAMF: A New Method for Delivery of Cas9 Ribonucleoprotein Complex to Human Hematopoietic Stem Cells
title_sort	triamf: a new method for delivery of cas9 ribonucleoprotein complex to human hematopoietic stem cells
publisher	Nature Portfolio
publishDate	2018
url	https://doaj.org/article/7126e75f7f804ba7a87cf87864786123
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TRIAMF: A New Method for Delivery of Cas9 Ribonucleoprotein Complex to Human Hematopoietic Stem Cells

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