| Ph-like ALL(Philadelphia chromosome-like ALL)is a subtype of ALL that displays a gene expression profile similar to BCR-ABL1 positive ALL(Ph+ ALL).It has adverse clinical features and poor prognosis.Ph-like ALL account for 20%of ALL.It has a diverse range of genetic alterations that activate cytokine receptor genes and kinase signaling pathways,frequently accompanied by abnormal transcription factors related to lymphatic development.Tyrosine kinase inhibitors can significantly improve the prognosis of Ph-like ALL,According to the molecular cytogenetic abnormalities,Therefore,the detection of Ph-like ALL related molecular genetic abnormalities is of great clinical value.Using real-time PCR technology platform,combined with mediator probe technology and HAND system(Homo-Tag Assisted Non-Dimer),a four-color multiplex real-time PCR method was developed for screening of 32 Ph-like ALL related fusion genes comprising 42 fusion types and an independent prognostic factor.Firstly,the breakpoints and the sequences of each fusion gene were thoroughly investigated from relevant literatures.Secondly,each primer and probe set was confirmed by simplex PCR using 42 artificial plasmids as templates.The individual assays were then combined and optimized,and finally a multiplex real-time PCR system with 4 parallel reactions was developed.This system was clinically evaluated by 86 leukemic samples.As a result,9 positive samples were identified,8 of them were concordant with sequencing results,1 sample was failed to sequence.This novel system was sensitive,specific,reliable and cost-effective,comprehensive,and thus a robust clinical tool for screening of Ph-like ALL related fusion genes.In addition,we used the primers and probes of multiplex real-time PCR to establish a duplex qPCR be used for subsequent quantification of minimal residual disease(MRD),monitor patient outcomes.Further extending the clinical application value of the multiplex real-time PCR. |
PDF Full Text Request