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Monitoring response to therapy in patients with chronic myeloid leukemia who are known to have the e13/a2 or e14/a2 BCR-ABL1 fusion transcript forms
Chronic myelogenous leukemia (CML) is a hematopoietic stem cell neoplasm included in the broader diagnostic category of myeloproliferative neoplasms. CML is consistently associated with fusion of the breakpoint cluster region gene (BCR) at chromosome 22q11 to the Abelson gene (ABL1) at chromosome 9q23. This fusion is designated BCR/ABL1 and may be seen on routine karyotype as the Philadelphia chromosome.
Although various breakpoints within the BCR and ABL1 genes have been described, >95% of CMLs contain a consistent mRNA transcript in which either the BCR exon 13 (e13) or BCR exon 14 (e14) is fused to the ABL1 exon 2 (a2), yielding fusion forms e13/a2 and e14/a2, respectively. The e13/a2 and e14/a2 fusion forms produce a 210-kDa protein (p210). The p210 fusion protein is an abnormal tyrosine kinase known to be critical for the clinical and pathologic features of CML, and agents that block the tyrosine kinase activity (ie, tyrosine kinase inhibitors or TKI, such as imatinib mesylate), have been used successfully for treatment. Monitoring the level of BCR/ABL1 mRNA in CML patients during treatment is helpful for both prognosis and management of therapy.(1-3) Rising BCR-ABL1 mRNA levels following attainment of critical therapeutic milestones (see Clinical References) can be indicative of acquired resistance mutations involving the ABL1 part of the BCR-ABL1 fusion gene.
Quantitative reverse-transcription PCR (qRT-PCR) is the most sensitive method for monitoring BCR-ABL1 levels during treatment. This test detects the BCR-ABL1 mRNA fusion forms found in CML (e13/a2 and e14/a2).
The presence or absence of BCR-ABL1 mRNA fusion form e13/e14-a2 producing the p210 fusion protein is identified. If positive, the quantitative level is reported as the normalized ratio of BCR-ABL1 (p210) to endogenous ABL1 mRNA with conversion to a percentage referenced to the international scale (IS), on which 0.1% BCR-ABL1:ABL1 is designated as a major molecular response (MMR) threshold.
An interpretive report will be provided. When BCR-ABL1 mRNA is present, quantitative results are reported on the international scale (IS), established from data originally reported in the IRIS (International Randomized Study of Interferon versus STI571) trial involving newly diagnosed chronic myeloid leukemia patients. Using the IS, a result <0.1% BCR-ABL1 (p210): ABL1 is equivalent to a major molecular remission. For further discussion of the international scale, see Clinical References.
This test detects only the e13/a2 and e14/a2 fusion forms, which code for the p210 protein. Other fusion forms are not detected, including those containing the BCR e1 exon, which codes for the p190 protein commonly found in acute lymphoblastic leukemia (ALL). If the patient is known to carry an e1/a2 (p190) fusion form, BA190 / BCR/ABL, p190, mRNA Detection, Reverse Transcription-PCR (RT-PCR), Quantitative, Monitoring Assay should be used for monitoring.
This test should not be used to screen for bcr/abl fusions at the time of diagnosis; if a diagnostic screen for BCR-ABL1 is desired, then the BADX / BCR/ABL1, mRNA Detection, Reverse Transcription-PCR (RT-PCR), Qualitative, Diagnostic Assay, which is designed to detect all reported common and rare BCR-ABL1 mRNA fusion variants, should be ordered for this purpose.
The precision of this assay at low BCR-ABL1 levels is more variable, such that inter-run variation can be as high as + or - 0.5 log. Only level changes >0.5 log should be considered clinically significant. For example, if a result is given as 0.1% BCR-ABL1:ABL1, then any result between 0.05% and 0.5% should be considered essentially equivalent. If the results are being used to make major therapeutic decisions, significant changes during monitoring should be verified with a subsequent specimen.
In general, the results of this assay cannot be directly compared with results generated from other PCR assays, including identical assays performed in other laboratories. Monitoring should be performed using the same method and laboratory for each subsequent specimen.
The results of this assay cannot be directly compared with BCR-ABL1 results obtained using FISH technology. FISH measures DNA alleles and PCR-based assays measure mRNA transcripts. Because a single DNA allele can produce many mRNA transcripts, the values are not directly comparable and FISH results are not applicable to the IS or to disease monitoring.
Blood is the specimen of choice for monitoring. While most patients show similar BCR-ABL1 mRNA levels in blood and bone marrow drawn at the same time, some patients may exhibit a difference in the levels between blood and bone marrow such that alternating specimen types during monitoring can lead to interpretive confusion.
Assay precision does not appear to be significantly affected by specimen transport or moderate delays in processing. However, in specimens with low levels of BCR-ABL1 these conditions may cause sufficient RNA degradation to produce false-negative results. Thus, specimens should be shipped as quickly as possible and specimens >3 days old at the time of receipt will be considered unacceptable.
1. Hughes TP, Kaeda J, Branford S, et al: Frequency of major molecular responses to imatinib or interferon alfa plus cytarabine in newly diagnosed chronic myeloid leukemia. N Engl J Med 2003 October 9;349(15):1423-1432
2. Baccarini M, Deininger MW, Rosti G, et al: European LeukemiaNet recommendations for the management of chronic myeloid leukemia: 2013. Blood 2013;122:872-884
3. Press RD, Kamel-Reid S, Ang D: BCR-ABL1 RT-qPCR for monitoring the molecular response to tyrosine kinase inhibitors in chronic myeloid leukemia. J Mol Diagn 2013;15:565-576
4. Cross NC, White HE, Muller MC, et al: Standardized definitions of molecular response in chronic myeloid leukemia. Leukemia 2012;26:2172-2175