Molecular screening of pointmutation [T3151] using allele specific oligonucleotides-polymerase chain reaction [ASO-PCR]

ABSTRACT

The development of the BCR-ABL targeted Imatinib Mesylate [IM] represent a paradigm shift in the treatment of chronic myeloid leukemia [CML]. However, a considerable number of CML patients have been reported to show resistance to IM, leading to relapses. The purpose of this paper is to screening for T3151 mutation in patients who are showing criteria of resistance or relapse, using Allele Specific Oligonucleotides Polymerase Chain Reaction [ASO-PCR]. A total of 24 DNA samples related to 18 CML patients of non-responder to IM were analyzed for T3151. They received daily dose of IM [300-400] mg for a mean duration 35 months. They were followed up hematologically, cytogenetically and molecularly every 3-6 months. They had not achieved complete hematological response [CHR], major cytogenetic response [MCyR] or major molecular response [MMR] along the follow up duration and until the end of this study. DNA was extracted from 100 micro l of venous blood [VB] using DNA isolation kit. Screening for the presence of T3151 mutation was done using [ASO-PCR]. PCR products were electrophoresed using 2.5% agarose gel electrophoresis. Agarose gel electrophoresis of ASO-PCR amplified products showed just one band of about [158 bp] in the lanes of wild-type alleles, indicating that non of those patients were harboring such mutation. There was no evidence that the initial lack of CHR, CyR or MMR in CML patients treated with IM due to the presence of T3151 mutation but those patients may have carried other types of mutations result in shift in sensitivity that may allow sufficient kinase activity to initiate disease progression in the presence of IM