Kinetics of mutations in mutant subclones in 3 representative patients. The kinetics of the BCR-ABL transcripts measured by RQ-PCR are represented by the bold black lines in all 3 panels (expressed as a ratio to the ABL control transcripts × 100%9 ). The proportions of the subclones with the various mutations measured at different time points are represented by the colored lines. (A) Patient no. 10. A 55-year-old female with CML in chronic phase was started on 400 mg daily IM. After 30 months from start of IM the M351T mutant clone had decreased from 80% to 50%, but an additional mutation, H396R, was identified. Subsequently, the 2 mutations evolved discordantly consistent with 2 distinct subclones. After starting dasatinib only H396R mutation was detectable. One month from start of dasatinib therapy, H396R became undetectable, a third mutant subclone, F317L, was detected by Q-SNP. (B) Patient no. 11. A 53-year-old female achieved a partial cytogenetic response to 400 mg daily IM but despite increasing the dose to 600 mg, she entered myeloblastic transformation. Dasatinib was subsequently introduced at 70 mg twice daily, which induced a reduction in BCR-ABL transcripts over the next 6 months, and the F311I mutation became transiently undetectable by Q-SNP. Subsequently, the BCR-ABL levels increased, and the patient was found to have T315I mutation in addition to F311I. The fact that the levels of the 2 mutations coincided closely in the terminal phase of her disease suggests that they were present in the same allele. This was confirmed by RFLP studies and sequencing of cloned PCR products. The patient eventually died in blastic transformation. (C) Patient no. 12. A 23-year-old woman achieved partial cytogenetic response with IM therapy but failed to maintain the response. She then progressed rapidly to lymphoblastic transformation and underwent sibling allogeneic hematopoietic stem cell transplant (HSCT). Four months after HSCT she relapsed to blastic phase, at which time Y253H mutation was detected. She was then started on dasatinib, which induced a 2-log reduction in BCR-ABL transcript numbers, and the Y253H mutant clone became transiently undetectable. Thereafter tumor load increased and T315I was detected in addition to resurgent Y253H mutation. The Q-SNP data implied both mutations were in the same allele; this was confirmed by RFLP studies.
