Introduction of the T315I gatekeeper mutation of BCR/ABL1 into a Philadelphia chromosome-positive lymphoid leukemia cell line using the CRISPR/Cas9 system.
Int J Hematol
; 116(4): 534-543, 2022 Oct.
Article
em En
| MEDLINE
| ID: mdl-35524023
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.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Leucemia Mielogênica Crônica BCR-ABL Positiva
/
Leucemia-Linfoma Linfoblástico de Células Precursoras
/
Antineoplásicos
Limite:
Humans
Idioma:
En
Revista:
Int J Hematol
Assunto da revista:
HEMATOLOGIA
Ano de publicação:
2022
Tipo de documento:
Article
País de afiliação:
Japão