Utilisation of RT-PCR assay for the detection of t(9;22)BCR-ABL fusion gene mRNA in diagnosis of chronic myeloid leukaemia at the University Teaching Hospital,Lusaka, Zambia

Abstract

Chronic MyeloidLeukaemia (CML) is consistently associated with a characteristic chromosome translocation called the Philadelphia (Ph) chromosome. This translocation fuses sequences of theBreakpoint Cluster Region(BCR) gene from band q11 on chromosome 22 with regions of theAbelson(ABL) gene from band q34 on chromosome 9, resulting in the production of a chimeric molecule, the BCR-ABL fusion mRNA product. The Philadelphia chromosome, t(9;22) (q34; q11), is the hallmark of CML, and the discovery of this molecular pathway has resulted in more accurate diagnosis, more targeted drug therapy and the application of molecular methodologies for post-therapy follow-up for the presence of minimal residual disease (MRD). Demonstration of the t(9;22) chromosomal aberration confirms the diagnosis. PCR is the most sensitive technique available for detecting BCR-ABL gene fusion in the diagnosis of CML as well as monitoring of CML patients on therapy and as such molecular diagnosis and monitoring of CML patients has become clinically very important. The aim of this study was to evaluate the utilisation of Xpert BCR-ABL Ultra, a commercially available real time quantitativeReverse Transcriptase Polymerase Chain Reaction(RT-PCR) assay for detection and measurement of Major BCR-ABL(p210 BCR-ABL mRNA), to routine molecular diagnosis of CML, demonstrating that the same assay could be used both for diagnosis and monitoring in the routine management of CML patients. A cross sectional descriptive laboratory study was undertaken at the University Teaching Hospital in Lusaka, Zambia. The GeneXpert BCR-ABL Assay was used with the GeneXpertDx System designed to detect the BCR-ABL p210 (b2a2 and b3a2) major break point translocation transcript in peripheral blood specimens. A total of 15 samples were analysed, of which 8 were known positive confirmed to have BCR-ABL by conventional PCR assay collected from known CML patients, and 7 negative taken from healthy blood donors using Xpert BCR-ABL Ultra Assay. Our results showed100% sensitivity, 85.7% specificity, 99.9%positive predictive value, 100% negative predictive value and 93.3% test efficiency. This study showed that the Xpert BCR-ABL Ultra, quantitative RT-PCR assay could also be reliably used as a diagnostic tool in the diagnosis and management of CML. This study showed that the Xpert BCR-ABL Ultra, a commercially available real time quantitative RT-PCR assay, designed for quantitative monitoring of CML patients undergoing Tyrosine Kinase Inhibitors therapy, can also be reliably used as a diagnostic tool, for detection of Major BCR-ABL (p210 BCR-ABL mRNA), in the diagnosis and management of CML. That means the same assay could be adopted for both diagnosis and monitoring in the routine management of CML patients in Zambia.