TEST:

Xpert® BCR-ABL Ultra

The BCR::ABL1%IS values obtained with this assay showed strong concordance with those generated by local reference methods, and MR classifications were consistent across platforms. These findings confirm the robustness, accuracy, and efficiency of the Xpert® BCR-ABL Ultra assay, supporting its use as a reliable alternative to currently validated systems for the routine clinical monitoring of CML patients.

In addition, we observed that e13a2 and e14a2 BCR::ABL1 transcript types could be discriminated based on the mean fluorescence intensity of BCR::ABL1-positive droplets. BCR::ABL1 digital PCR is feasible for DMR quantification in clinical practice and offers an increased sensitivity over RT-qPCR.

Using an IQCP approach, our lab has established a quality control plan that uses weekly QC with the Xpert BCR-ABL assay. Regular review of this weekly QC data shows that this assay continues to perform well with minimal deviations in QC results during our first 18 months of patient testing.

The validation demonstrated the Xpert P190 BCR-ABL Real-Time RT-PCR assay was very accurate and precise. The assay will be used to confirm p190 BCR::ABL transcripts in whole-blood samples and to quantitate the levels in patients undergoing treatment.

Excellent concordance was seen between the Xpert BCRABL Ultra p210 and p190 assays compared to our institutional RT-qPCR assay in PB/BM samples. The Xpert BCR-ABL Ultra p190 and p210 assays showed distinct benefits, including near full automation, quick setup time without laborious RNA extraction, and fast turnaround time. These advancements allow us to assess PB/BM samples for p190 and p210 transcripts with high sensitivity and specificity for diagnostic and therapeutic monitoring purposes.

Objective: To assess the most mutation in 2 genes: ABL ( T315I) and ASXL1 (c.1934dupG), in Tunisian patients with CML and imatinib resistance... Our findings suggest that both mutations are uncommon in Tunisian patients with CML and TKI resistance. A comprehensive mutational analysis of other genes and epigenetic factors is mandatory to elucidate the molecular mechanisms that lead to kinase inhibitor resistance.

Less than 2-fold difference was observed in %BCR-ABL p190/ABL1 reporting between delta Ct-based and CN-based approaches. Conclusion & Significance: A BCR-ABL p190 copy number estimator for Xpert ® BCR-ABL Ultra p190 test was established, which will provide helpful information for diagnosis and prognosis of diseases relating to BCR-ABL p190.

Hanfstein B, Lauseker M, Hehlmann R, Saussele S, Erben P, Dietz C (2014) Distinct characteristics of e13a2 versus e14a2 BCR-ABL1 driven chronic myeloid leukemia under first-line therapy with imatinib... A %CN e13a2/ABL and e14a2/ABL reporting method with known CN of IVT-RNAs has been successfully established in Xpert BCR-ABL Ultra. Both WHO IS and %CN showed very minor differences between e13a2 and e14a2 for % reporting. %CN method demonstrated comparable %reporting to WHO IS in Xpert BCR-ABL Ultra.

Current results are promising and show high molecular response rates in Tunisian Ph+ALL, though follow-up is still too short to determine their impact on remission duration and long-term survival. A larger study is mandatory to elucidate prognosis impact of TKI in the management of Ph+ALL.

To the best of our knowledge this is the first study to investigate ASXL1 mutation in -TKI-resistant CML patients in Tunisia. Despite the limits of our study, our finding highlights that this truncating ASXL1 mutation may be a potential biomarker for predicting therapeutic efficacy, and a treatment strategy for CML with ASXL1 mutation should be established. Moreover, future studies need to comprehensively identify the landscape and clinical relevance of genetic and epigenetic alterations in CML, which can be used to develop novel therapeutic strategies or to prognosticate treatment response.

A %CN e13a2/ABL and e14a2/ABL reporting method with known CN of IVT-RNAs has been successfully established in Xpert BCR-ABL Ultra. Both WHO IS and %CN showed very minor differences between e13a2 and e14a2 for % reporting. %CN method demonstrated comparable %reporting to WHO IS in Xpert BCR-ABL Ultra.

Statistical analysis of the data obtained with the two systems and on the three different types of material demonstrated a good correlation at all different levels of disease. The CEPHEID cartridge system could therefore be considered as an alternative to the analysis systems currently validated within the Italian network CML LabNet.