Sensitivity to Tyrosine Kinase Inhibitors in a Human Philadelphia Chromosome-Positive (Ph+) Leukemia Model With the T315I-Inclusive Compound Mutation

The T315I-inclusive compound mutation, the multiple mutations including the T315I mutation on the same BCR::ABL1 gene, confers resistance to diverse tyrosine kinase inhibitors (TKIs). Development of the F311I/T315I compound mutation has been reported in chronic myeloid leukemia patients who sequentially showed clinical resistance to imatinib and dasatinib. The establishment of a human leukemia model with the T315I-inclusive compound mutation remains an experimental challenge. Here, we introduced the F311I/T315I compound mutation into the intrinsic BCR::ABL1 gene of a human TKI-sensitive Philadelphia chromosome-positive leukemia cell line via homologous recombination using the CRISPR/Cas9 system and obtained three types of sublines: the F311I mutation alone, the T315I mutation alone, and the F311I/T315I compound mutation. The F311I subline was sensitive to dasatinib but moderately resistant to imatinib and nilotinib, while the T315I subline and the F311I/T315I subline were highly resistant to these TKIs. Notably, the T315I subline and the F311I/T315I subline were sensitive to therapeutic concentrations of ponatinib, although more resistant than the F311I subline. Moreover, the T315 subline and the F311I/T315 subline were sensitive to asciminib at therapeutic concentration, as was the F311I subline. This is the first human leukemia model in which the impact of the T315I-inclusive compound mutation on TKI sensitivity was directly confirmed.