Sunday, February 28, 2016

BCR/ABL1 Testing: Introduction, indications, methods and interpretation

Fig. BCR-ABL1 fusion gene (Philadelphia chromosome)
BCR-ABL1 refers to a gene sequence found in an abnormal chromosome 22 of some people with certain forms of leukemia. Unlike most cancers, the cause of chronic myelogenous leukemia (CML) and some other leukemias can be traced to a single, specific genetic abnormality in one chromosome. The presence of the gene sequence known as BCR-ABL1 confirms the diagnosis of CML and a form of acute lymphoblastic lymphoma (ALL). Chronic myelogenous leukemia (CML) is part of a group of diseases called the myeloproliferative disorders, with an estimated 4600 newly diagnosed cases and 850 deaths in 2005. More than 95% of patients with CML have the distinctive Philadelphia chromosome (Ph1) that results from a reciprocal translocation between the long arms of
chromosomes 9 and 22. The translocation involves the transfer of the Abelson (ABL) gene on chromosome 9 to the breakpoint cluster region (BCR) of chromosome 22, resulting in a fused BCR-ABL gene. The fusion gene produces a protein, p210 (b2a2 and b3a2), a tyrosine kinase with deregulated activity that plays a key role in the development of CML.

Humans have 23 pairs of chromosomes containing inherited genetic information. Those genes contain the blueprints, in the form of DNA, for producing the proteins that our bodies rely on to function properly. While some genetic abnormalities are inherited, they can also come from changes that occur to genes or chromosomes after a person is born. This can happen through exposure to various environmental factors (e.g., radiation, certain chemicals) but more often for unknown reasons.

The BCR-ABL1 gene sequence is one such acquired change that is formed when pieces of chromosome 9 and chromosome 22 break off and switch places. When this occurs, the ABL1 region in chromosome 9 fuses with the BCR gene region in chromosome 22. This type of change is called a reciprocal translocation and is often abbreviated as t(9;22). The resulting chromosome 22 that has the BCR-ABL1 gene sequence is known as the Philadelphia (Ph) chromosome because that is where it was first discovered.

The resulting Philadelphia chromosome or BCR-ABL1 gene encode an abnormal protein that is responsible for the development of CML and a type of ALL. At diagnosis, 90-95% of cases of CML show a characteristic t(9;22) BCR-ABL1 reciprocal chromosomal translocation. About one in four adults with ALL have the translocation.

The protein formed by BCR-ABL1 is a type of enzyme called a tyrosine kinase. That enzyme is responsible for the uncontrolled growth of leukemic cells. When large numbers of abnormal leukemic cells start to crowd out the normal blood cell precursors in the bone marrow, signs and symptoms of leukemia start to emerge. Treatment of these leukemias typically involves a tyrosine kinase inhibitor (TKI).

Testing for BCR-ABL1 detects the Philadelphia chromosome and BCR-ABL1 fusion gene or its transcripts, which are the RNA copies made by the cell from the abnormal stretches of DNA. The presence of the BCR-ABL1 abnormality confirms the clinical diagnosis of CML, a type of ALL, and rarely AML


This test is done for diagnostic workup and monitor the treatment for patients with high probability bcr/abl1- positive hematopoietic neoplasms, predominantly chronic myelogenous leukemia (CML) and acute lymphoblastic leukemia (ALL). When you have results of a complete blood count (CBC) and/or signs and symptoms that suggest that you may have leukemia.

A blood sample drawn from a vein in your arm or a bone marrow sample collected using a bone marrow aspiration and/or biopsy procedure.

There are several different types of BCR-ABL1 tests available, including:
·         Cytogenetics (chromosome analysis or karyotyping)
This test looks at chromosomes under a microscope to detect structural and/or numerical abnormalities. For example, the Philadelphia chromosome looks shorter than normal. Cells in a sample of blood or bone marrow are grown in the laboratory and then examined to determine if the Philadelphia chromosome is present. Other chromosomal abnormalities can also be detected.
·         Fluorescence in situ hybridization (FISH)
This test method uses fluorescent dye-labeled probes to "light up" the BCR-ABL1 gene sequence when it is present. It can also determine the percentage of blood or bone marrow cells that contain the abnormal, fused BCR-ABL1 gene.
·         Genetic molecular testing (qualitative or quantitative)
Polymerase chain reaction (PCR)
-based qualitative and quantitative tests detect and measure the BCR-ABL1 gene in leukemia cells taken from blood or bone marrow samples.
·         Secondary mutations within the BCR-ABL1 are known to cause resistance to therapy. These can be detected by DNA sequencing methods.

When to order:
BCR-ABL1 testing is ordered when a health practitioner suspects that a person has CML or Philadelphia chromosome (Ph)-positive ALL. Initial testing may be indicated when a person has nonspecific signs or symptoms such as:
·         Fatigue
·         Weight loss
·         Joint or bone pain
·         Enlarged spleen
·         As follow-up to abnormal findings on a complete blood count (CBC)

Early in the disease, a person may have few or no symptoms. As time passes and normal blood cells are crowded out of the bone marrow and the number of abnormal leukemic cells increases, a person may experience anemia, prolonged bleeding, and recurrent infections.
Once CML or Ph chromosome-positive ALL has been diagnosed, BCR-ABL1 quantitative genetic testing is ordered periodically (typically every 3 months) to monitor the response to treatment and monitor for recurrence.
When a person is not responding and treatment resistance is suspected or disease recurrence occurs after remission, the BCR-ABL1 kinase domain mutation analysis may be performed.


Interpretation of BCR-ABL1 test:

·    If a person has abnormal white blood cells in the bone marrow and has the Philadelphia (Ph) chromosome and BCR-ABL1 gene sequence, then the individual is diagnosed with CML or Ph-positive ALL.
·     Of those who have CML, 90-95% have the Ph chromosome if tested by cytogenetics and 100% have the BCR-ABL1 gene sequence by FISH and/or qualitative BCR-ABL1 molecular testing. About 25% of adults with ALL and 2-4% of children with ALL are positive for the Ph chromosome and/or the BCR-ABL1 gene sequence.
·    A small percentage of people with CML will have the BCR-ABL1 gene sequence but not the Ph chromosome. These cases either have variant translocations that involve a third or even a fourth chromosome in addition to 9 and 22 or have a hidden translocation involving 9 and 22 that can not be identified by routine chromosomal analysis. Since the treatment for BCR-ABL1-related leukemias specifically targets the tyrosine kinase protein produced, these people can still be monitored with quantitative BCR-ABL1 molecular testing.
·     In general, if the amount of BCR-ABL1 in the blood or bone marrow decreases over time, then the person is responding to treatment. If the quantity of BCR-ABL1 drops below the test's detection level and the person's blood cell counts are normal, then the person is considered to be in remission.
·       If the BCR-ABL1 level rises, then it indicates disease progression or recurrence. It may also indicate that the person has become resistant to imatinib, the first-generation tyrosine kinase inhibitor. Additional genetic testing is often performed to detect the development of BCR-ABL1 kinase domain mutations associated with imatinib resistance.
·       If an individual's leukemia is resistant to imatinib, a second generation tyrosine kinase inhibitor may be given. Those tyrosine kinase inhibitors, developed after imatinib, are also given in case the side effects from imatinib are too severe or in case of a rare BCR-ABL mutation. They include ponatib, bosutinib, nilotinib, and dasatinib.
·       If a person with ALL is not positive for the Ph chromosome and the BCR-ABL1 gene sequence, then that person will not be given a tyrosine kinase inhibitor drug and BCR-ABL1 molecular testing cannot be used to monitor the person.

Different phases of CML:
Recognition of disease progression and transformation is important for prognosis and treatment. CML goes through three phases:
·    Chronic phase—most people with CML are diagnosed in the chronic phase, which usually has an insidious onset, meaning that leukemia symptoms may be absent or subtle. The chronic phase may last for a month to several years. This is the phase when there are few or no symptoms and also the time period when treatment is most successful.
·   Accelerated phase—changes include but are not limited to increasing white blood cell (WBC) counts and additional changes to cells like an increase in blasts in blood and/or bone marrow (but less than 20%) and lack of therapeutic response to standard treatment
·        Blast phase—when blasts are 20% or more of the cells in the blood or bone marrow or when there is blast proliferation outside the bone marrow
Both blood and bone marrow are often evaluated as part of the initial diagnosis, but the majority of follow-up monitoring is performed on blood samples. There is significant test variability among laboratories using different test platforms. Therefore, for a given patient, the quantitative BCR-ABL1 molecular testing should be performed by the same laboratory or referred to a laboratory that follows universal reporting criteria. Rising and falling levels of BCR-ABL1 are usually more important than a single test result.

(References: See more at: and )
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