Digital PCR for BCR‐ABL1 Quantification in CML: Current Applications in Clinical Practice

Abstract

Molecular monitoring of the BCR-ABL1 transcript for patients with chronic phase chronic myeloid leukemia (CML) has become increasingly demanding. Real-time quantitative PCR (qPCR) is the routinely used method, but has limitations in quantification accuracy due to its inherent technical variation. Treatment recommendations rely on specific BCR-ABL1 values set at timed response milestones, making precise measurement of BCR-ABL1 a requisite. Furthermore, the sensitivity of qPCR may be insufficient to reliably quantify low levels of residual BCR-ABL1 in patients in deep molecular response (DMR) who could qualify for an attempt to discontinue Tyrosine Kinase Inhibitor (TKI) therapy. We reviewed the current use of digital PCR (dPCR) as a promising alternative for response monitoring in CML. dPCR offers an absolute BCR-ABL1 quantification at various disease levels with remarkable precision and a clinical sensitivity reaching down to at least MR5.0. Moreover, dPCR has been validated in multiple studies as prognostic marker for successful TKI treatment discontinuation, while this could not be achieved using classical qPCR. dPCR may thus prospectively be the preferred method to reliably identify patients achieving treatment milestones after initiation of TKI therapy as well as for the selection and timing for TKI discontinuation.

Making Treatment-Free Remission (TFR) Easier in Chronic Myeloid Leukemia: Fact-Checking and Practical Management Tools

“…The panel agreed that standard RT-qPCR is the technique of choice during TFR. ddPCR is a potential alternative to RT-qPCR [95,96] but not yet standard-Table 7 Frequency of molecular monitoring in case of stable and unstable MR4.5 maintained during the first 6 months after discontinuation. After month 6, patients with unstable MR4.5 must be monitored at closer intervals than those with a stable MR4.5.…”

Assessment of droplet digital polymerase chain reaction for measuring BCR‐ABL1 in chronic myeloid leukaemia in an international interlaboratory study

“…The cost of Bio‐Rad’s QXDX BCR‐ABL %IS kit is similar to the automated Cepheid Xpert ® cartridge‐based systems, but is more expensive than conventional laboratory developed RTqPCR tests, especially when the extra wells for deep‐response detection are considered 23 . This is off set by the lack of requirement for a standard curve and the reduced validation required when using a Conformité Européene (CE) marked kit given the imminent changes to Europe’s in vitro diagnostic (IVD) regulations and their impact on laboratory developed tests.…”

Quantification of atypical BCR‐ABL1 fusion transcripts in patients with chronic myeloid leukemia: Which approach for harmonization?

“…Cumulative evidence is building on the similar, if not superior assay characteristics of ddPCR in CML patients expressing the common e13a2 and e14a2 BCR-ABL1 transcript types, particularly in measuring deep molecular response and subsequent prediction of those patients most likely to succeed in a TFR attempt. 8 Proof of principle for quantification of atypical BCR-ABL1 transcripts has been demonstrated with this observation successfully expanded to encompass BCR-ABL1 monitoring by ddPCR during TFR attempts. 9,10 A current disadvantage of ddPCR is the cost compared with conventional laboratory-developed RT-qPCR tests, 11 though throughput might not necessarily be an impediment given the relatively fewer number of CML patients with atypical transcripts than those expressing e13a2 and e14a2 BCR-ABL1.…”