<i>BCR-ABL1</i>expression, RT-qPCR and treatment decisions in chronic myeloid leukaemia

Latham, Susan; Bartley, Paul C.; Budgen, Bradley; Ross, David M.; Hughes, Elizabeth; Branford, Susan; White, Deborah L.; Hughes, Timothy P.; Morley, Alexander A.

doi:10.1136/jclinpath-2015-203538

Cited by 9 publications

(7 citation statements)

References 15 publications

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“…In addition, we and others have compared the quantification of BCR-ABL in genomic DNA and RNA in serial samples and have shown that individual patients may have consistently higher or lower expression of BCR-ABL mRNA for a given number of CML cells. 47,48 This interindividual difference results in a measurement bias whereby RQ-PCR may underestimate the level of residual disease in some patients. Although genomic Q-PCR for BCR-ABL is not widely available, it is a promising research tool to elucidate the relationship between MRD and TFR outcome.…”

Section: Areas Of Future Investigationmentioning

confidence: 99%

Moving treatment-free remission into mainstream clinical practice in CML

Hughes

Ross

2016

Blood

290

260

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The dramatic success of tyrosine kinase inhibitors (TKIs) has led to the widespread perception that chronic myeloid leukemia (CML) has become another chronic disease, where lifelong commitment to pharmacologic control is the paradigm. Recent trials demonstrate that some CML patients who have achieved stable deep molecular response can safely cease their therapy without relapsing (treatment free remission [TFR]). Furthermore, those who are unsuccessful in their cessation attempt can safely re-establish remission after restarting their TKI therapy. Based on the accumulated data on TFR, we propose that it is now time to change our approach for the many CML patients who have achieved a stable deep molecular response on long-term TKI therapy. Perhaps half of these patients could successfully achieve TFR if offered the opportunity. For many of these patients ongoing therapy is impairing quality of life and imposing a heavy financial burden while arguably achieving nothing. This recommendation is based on the evident safety of cessation attempts and TFR in the clinical trial setting. We acknowledge that there are potential risks associated with cessation attempts in wider clinical practice, but this should not deter us. Instead we need to establish criteria for safe and appropriate TKI cessation. Clinical trials will enable us to define the best strategies to achieve TFR, but clinicians need guidance today about how to approach this issue with their patients. We outline circumstances in which it would be in the patient's best interest to continue TKI, as well as criteria for a safe TFR attempt.

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Section: Areas Of Future Investigationmentioning

confidence: 99%

Moving treatment-free remission into mainstream clinical practice in CML

Hughes

Ross

2016

Blood

290

260

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“…MR 5.0 at all (Figure S2C). In any cases, the consideration of the detectability of the transcript was not statistically significant.…”

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confidence: 95%

Digital PCR improves the quantitation of DMR and the selection of CML candidates to TKIs discontinuation

et al. 2019

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Treatment‐free remission (TFR) by tyrosine kinase inhibitors (TKI) discontinuation in patients with deep molecular response (DMR) is a paramount goal in the current chronic myeloid leukemia (CML) therapeutic strategy. The best DMR level by real‐time quantitative PCR (RT‐qPCR) for TKI discontinuation is still a matter of debate. To compare the accuracy of digital PCR (dPCR) and RT‐qPCR for BCR‐ABL1 transcript levels detection, 142 CML patients were monitored for a median time of 24 months. Digital PCR detected BCR‐ABL1 transcripts in the RT‐qPCR undetectable cases. The dPCR analysis of the samples, grouped by the MR classes, revealed a significant difference between MR 4.0 and MR 4.5 ( P = 0.0104) or MR 5.0 ( P = 0.0032). The clinical and hematological characteristics of the patients grouped according to DMR classes (MR 4.0 vs MR 4.5‐5.0 ) were superimposable. Conversely, patients with dPCR values <0.468 BCR‐ABL1 copies/µL (as we previously described) showed a longer DMR duration ( P = 0.0220) and mainly belonged to MR 4.5‐5.0 ( P = 0.0442) classes compared to patients with higher dPCR values. Among the 142 patients, 111 (78%) discontinued the TKI treatment; among the 111 patients, 24 (22%) lost the MR 3.0 or MR 4.0 . RT‐qPCR was not able to discriminate patients with higher risk of MR loss after discontinuation ( P = 0.8100). On the contrary, according to dPCR, 12/25 (48%) patients with BCR‐ABL1 values ≥0.468 and 12/86 (14%) patients with BCR‐ABL1 values <0.468 lost DMR in this cohort, respectively ( P = 0.0003). Treatment‐free remission of patients who discontinued TKI with a dPCR <0.468 was significantly higher compared to patients with dPCR ≥ 0.468 (TFR at 2 years 83% vs 52% P = 0.0017, respectively). In conclusion, dPCR resulted in an improved recognition of stable DMR and of candidates to TKI discontinuation.

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“…The reverse transcription quantitative polymerase chain reaction (RT-qPCR) is considered as the most sensitive, reliable and accurate technique to analyze differential gene expression at the messenger RNA (mRNA) level [1, 2]. Nevertheless, performing the experimental procedures will introduce a variety of potential errors, amongst which pipetting errors, starting material quality variations, differences in mRNA extractions, errors in sample quantifications, altered reverse transcription efficiencies and cDNA sample loading differences [3].…”

Section: Introductionmentioning

confidence: 99%

The validation of Short Interspersed Nuclear Elements (SINEs) as a RT-qPCR normalization strategy in a rodent model for temporal lobe epilepsy

et al. 2019

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BackgroundIn gene expression studies via RT-qPCR many conclusions are inferred by using reference genes. However, it is generally known that also reference genes could be differentially expressed between various tissue types, experimental conditions and animal models. An increasing amount of studies have been performed to validate the stability of reference genes. In this study, two rodent-specific Short Interspersed Nuclear Elements (SINEs), which are located throughout the transcriptome, were validated and assessed against nine reference genes in a model of Temporal Lobe Epilepsy (TLE). Two different brain regions (i.e. hippocampus and cortex) and two different disease stages (i.e. acute phase and chronic phase) of the systemic kainic acid rat model for TLE were analyzed by performing expression analyses with the geNorm and NormFinder algorithms. Finally, we performed a rank aggregation analysis and validated the reference genes and the rodent-specific SINEs (i.e. B elements) individually via Gfap gene expression.ResultsGeNorm ranked Hprt1, Pgk1 and Ywhaz as the most stable genes in the acute phase, while Gusb and B2m were ranked as the most unstable, being significantly upregulated. The two B elements were ranked as most stable for both brain regions in the chronic phase by geNorm. In contrast, NormFinder ranked the B1 element only once as second best in cortical tissue for the chronic phase. Interestingly, using only one of the two algorithms would have led to skewed conclusions. Finally, the rank aggregation method indicated the use of the B1 element as the best option to normalize target genes, independent of the disease progression and brain region. This result was supported by the expression profile of Gfap.ConclusionIn this study, we demonstrate the potential of implementing SINEs -notably the B1 element- as a stable normalization factor in a rodent model of TLE, independent of brain region or disease progression.

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BCR-ABL1expression, RT-qPCR and treatment decisions in chronic myeloid leukaemia

Abstract: Variation between individuals in expression of BCR-ABL1 can materially affect interpretation of the RT-qPCR when this test is used to make decisions on treatment.

Cited by 9 publications

References 15 publications

Moving treatment-free remission into mainstream clinical practice in CML

Moving treatment-free remission into mainstream clinical practice in CML

Digital PCR improves the quantitation of DMR and the selection of CML candidates to TKIs discontinuation

The validation of Short Interspersed Nuclear Elements (SINEs) as a RT-qPCR normalization strategy in a rodent model for temporal lobe epilepsy

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