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Introduction of the T315I gatekeeper mutation of BCR/ABL1 into a Philadelphia chromosome-positive lymphoid leukemia cell line using the CRISPR/Cas9 system.
- Source :
-
International journal of hematology [Int J Hematol] 2022 Oct; Vol. 116 (4), pp. 534-543. Date of Electronic Publication: 2022 May 06. - Publication Year :
- 2022
-
Abstract
- Imatinib and second-generation tyrosine kinase inhibitors (TKIs) have dramatically improved the prognosis of Philadelphia chromosome-positive (Ph+) acute lymphoblastic leukemia (ALL). However, overcoming TKI resistance due to the T315I gatekeeper mutation of BCR/ABL1 is crucial for further improving the prognosis. The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system is appropriate for establishing a human model of Ph+ ALL with the T315I mutation, because it can induce specific mutations via homologous recombination (HR) repair in cells with intact endogenous HR pathway. Here we used CRISPR/Cas9 to introduce the T315I mutation into the Ph+ lymphoid leukemia cell line KOPN55bi, which appeared to have an active HR pathway based on its resistance to a poly (ADP-Ribose) polymerase-1 inhibitor. Single-guide RNA targeting at codon 315 and single-strand oligodeoxynucleotide containing ACT to ATT nucleotide transition at codon 315 were electroporated with recombinant Cas9 protein. Dasatinib-resistant sublines were obtained after one-month selection with the therapeutic concentration of dasatinib, leading to T315I mutation acquisition through HR. T315I-acquired sublines were highly resistant to imatinib and second-generation TKIs but moderately sensitive to the therapeutic concentration of ponatinib. This authentic human model is helpful for developing new therapeutic strategies overcoming TKI resistance in Ph+ ALL due to T315I mutation.<br /> (© 2022. Japanese Society of Hematology.)
- Subjects :
- CRISPR-Associated Protein 9 genetics
CRISPR-Associated Protein 9 metabolism
CRISPR-Cas Systems
Cell Line
Dasatinib therapeutic use
Drug Resistance, Neoplasm genetics
Fusion Proteins, bcr-abl
Humans
Imatinib Mesylate pharmacology
Imatinib Mesylate therapeutic use
Mutation
Nucleotides therapeutic use
Oligodeoxyribonucleotides therapeutic use
Philadelphia Chromosome
Poly (ADP-Ribose) Polymerase-1 genetics
Poly (ADP-Ribose) Polymerase-1 metabolism
Protein Kinase Inhibitors pharmacology
Protein Kinase Inhibitors therapeutic use
RNA, Guide, CRISPR-Cas Systems therapeutic use
Antineoplastic Agents therapeutic use
Leukemia, Myelogenous, Chronic, BCR-ABL Positive drug therapy
Precursor Cell Lymphoblastic Leukemia-Lymphoma drug therapy
Precursor Cell Lymphoblastic Leukemia-Lymphoma genetics
Subjects
Details
- Language :
- English
- ISSN :
- 1865-3774
- Volume :
- 116
- Issue :
- 4
- Database :
- MEDLINE
- Journal :
- International journal of hematology
- Publication Type :
- Academic Journal
- Accession number :
- 35524023
- Full Text :
- https://doi.org/10.1007/s12185-022-03369-x