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


Ontology type: schema:ScholarlyArticle      Open Access: True


Article Info

DATE

2018-11-02

AUTHORS

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

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. More... »

PAGES

16304

Identifiers

URI

http://scigraph.springernature.com/pub.10.1038/s41598-018-34601-6

DOI

http://dx.doi.org/10.1038/s41598-018-34601-6

DIMENSIONS

https://app.dimensions.ai/details/publication/pub.1107913010

PUBMED

https://www.ncbi.nlm.nih.gov/pubmed/30389991


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36 schema:description 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.
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44 CRISPR/
45 Cas9 protein
46 Cas9 ribonucleoprotein complexes
47 HSPCs
48 Internalization Assisted
49 RNA
50 RNP
51 TRIAMF
52 TRIAMF treatment
53 TRansmembrane Internalization Assisted
54 autologous transplantation
55 blood disorders
56 capacity
57 cell mixtures
58 cells
59 cells ex vivo
60 challenges
61 clinical scales
62 colonies
63 competence
64 complexes
65 components
66 countries
67 cure
68 custom
69 delivery
70 delivery methods
71 disorders
72 edited HSPCs
73 editing
74 effective delivery method
75 efficient delivery
76 engraftment
77 ex vivo
78 filter membrane
79 filtration
80 gene editing
81 genes
82 goal
83 guide RNA
84 hematopoietic stem
85 hematopoietic stem cells
86 hemoglobinopathies
87 human hematopoietic stem cells
88 immunocompromised mice
89 inexpensive components
90 membrane
91 membrane filtration
92 method
93 mice
94 mind
95 mixture
96 monogenic blood disorders
97 multilineage colonies
98 new method
99 patient-derived hematopoietic stem
100 patients
101 post TRIAMF treatment
102 potential cure
103 progenitor cells ex vivo
104 protein
105 ribonucleoprotein complexes
106 robust gene
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