Evaluation of the Cepheid GeneXpert BCR-ABL Assay

Abstract

Patients with chronic myeloid leukemia harbor the chromosomal translocation t(9;22), which corresponds to fusion of the BCR and ABL genes at the DNA level. The translated fusion product is an oncogenic protein with increased ABL tyrosine kinase activity causing cell transformation. To date, reverse transcriptase-polymerase chain reaction is considered the most sensitive method available for detecting low copy numbers of the BCR-ABL gene fusion. Recently, Cepheid introduced its GeneXpert-based assay for the identification of the BCR-ABL gene fusion in cells from blood samples. This system comprises a walkaway self-contained instrument that combines cartridge-based microfluidic sample preparation with reverse transcriptase-polymerase chain reaction-based fluorescent signal detection and BCR-ABL and ABL Ct (threshold cycle) determination. The difference between the BCR-ABL Ct and ABL Ct (⌬Ct) is expected to represent the ratio of the two populations of mRNAs and ultimately the percentage of neoplastic cells present. We tested whether this BCR-ABL fusion detection system could be used as a clinical diagnostic tool for monitoring patients with minimal residual disease of chronic myelogenous leukemia. We report similar performance characteristics, including limit of detection, specificity, sensitivity, and precision, of this automated BCR-ABL fusion detection system to those of a manual TaqMan reverse transcriptase-polymerase chain reaction-based test.

Development of an Integrated Assay for Detection of BCR-ABL RNA

“…This result fits the conventional definition of the lower LOD, in which the sample is measured as positive 95% of the time. Similar results were reported in a separate evaluation by Johns Hopkins University scientists (27 ).…”

Cartridge-based automated BCR-ABL1 mRNA quantification: solving the issues of standardization, at what cost?

“…8 Concordances for CMR were comparable between the two laboratories and positive/negative discordant results were only observed among samples with positivity below 0.1%, in line with previously published data. 9,10 These data suggest that there is no significant bias in achieving CMR and its impact on therapeutic decisions, such as stopping TKI.…”

“…The 5 log range of linearity for automated quantification ( Figure 2) is in line with that previously described. 9,10 Reproducible results at these low levels of minimal residual disease were significantly more frequently obtained when pre-analytical steps were reduced to six hours or less, compared to 24 hours. We emphasize that this situation may only apply to patients undergoing treatment with TKI, in which case the samples remain in contact with the TKI during the pre-analytical phase.…”

“…9,10 We, therefore, chose not to repeat these experiments but to assess reproducibility on patient samples, analyzed in duplicate in two EAC standardized laboratories, with variable pre-analytical times which were systematically less than 24 hours.…”

“…9 Previous studies showed that this automated BCR-ABL1 fusion detection system did indeed have similar analytical performance, including specificity, sensitivity and intra-laboratory reproducibility, to non-automated TaqMan based RT RQ-PCR assays. 9,10 Sensitivity was assessed at 1 K562 cell amongst 100,000, with a linear range of quantitation of 5 logs. Given that neither of these studies fully addressed inter-laboratory reproducibility and alignment on IS, the present study specifically focused on these aspects.…”