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PML-RARA Mutation Analysis

Promyelocytic leukemia/retinoic acid receptor alpha or PML-RARA refers to an abnormal fusion gene sequence. It is a specific rearrangement of genetic material from two separate chromosomes (chromosomal translocation) and is associated with a specific type of leukemia. This test detects and measures PML-RARA in the blood or bone marrow to determine if an individual has acute promyelocytic leukemia (APL), a subtype of acute myeloid leukemia (AML).

Humans normally have 23 pairs of chromosomes, including 22 pairs of non-sex-determining chromosomes (also known as autosomes) and 1 pair of sex chromosomes (XX for females, XY for males). The genetic makeup of a person is contained on her/his chromosomes. The genes that reside on chromosomes form the blueprints for the production of thousands of proteins. Sometimes changes can occur to a person’s chromosomes and/or genes during their lifetime because of exposures to radiation, toxins, or for unknown reasons. These exposures could lead to gene mutations or to chromosome translocations.

The PML-RARA fusion gene sequence is one such acquired change (mutation) that is formed when pieces of chromosome 15 and chromosome 17 break off and switch places (translocate). The PML gene region in chromosome 15 then fuses with the RARA gene region in chromosome 17. This is referred to as reciprocal translocation, and this particular one is commonly expressed as t(15;17).

Normally, the PML gene codes for a protein that helps prevent uncontrolled cell growth and acts as a tumor suppressor. The RARA gene codes for a protein that is crucial for white blood cell (WBC) maturation, as these cells typically develop through several stages in the bone marrow before release into circulation. The mutated PML-RARA fusion gene codes for an abnormal fusion protein that does neither of these functions but instead leads to the uncontrolled production and accumulation of leukemic WBCs that do not mature or differentiate beyond the promyelocyte stage. As a large number of these abnormal cells start to crowd out the normal blood cell precursors in the bone marrow, signs and symptoms of leukemia start to emerge.

Up to 98% of cases of acute promyelocytic leukemia have a characteristic t(15;17) PML-RARA reciprocal chromosomal translocation. In about 2% of APL cases, other translocations involving the RARA gene and genes other than PML have been reported.

Testing detects the PML-RARA fusion gene or its transcripts, the RNA copies made by the cell from the abnormal gene sequence of DNA. The presence of the PML-RARA abnormality helps confirm the diagnosis of APL.

Testing can also direct APL therapy and monitor minimal residual disease, which could be fatal. Treatment of APL typically involves all-trans retinoic acid (ATRA), a drug that binds to retinoic acid receptors in cells. The drug can overcome the effect of the abnormal PML-RARA protein and induce downstream signaling and maturation of WBCs. This therapy works well in conjunction with chemotherapy but only in those cases where the PML-RARA fusion gene is present. The treatment results in remission in about 80-90% of these patients, according to the American Cancer Society. A small percentage of people with APL have a fusion between the RARA gene and a different gene, and they may or may not benefit from ATRA therapy based upon the specific gene involved.

The Test

This testing is used to detect the abnormal promyelocytic leukemia/retinoic acid receptor alpha or PML-RARA gene sequence. It is used to help diagnose acute promyelocytic leukemia (APL) in which the PML-RARA gene sequence is present, to guide treatment, to monitor response to treatment, and to monitor for disease recurrence.

A few different test methods are available to evaluate for PML-RARA, and they may be used for various purposes:

  • Fluorescence in situ hybridization (FISH) may be used to help diagnose APL and/or help to determine the percentage of a person’s blood or bone marrow cells that contain the abnormal, fused PML-RARA gene. This test method uses fluorescent dye-labeled probes to “light up” the PML-RARA gene sequence when it is present. FISH can also be used to detect the variant translocations involving RARA and genes other than PML. This may help identify drug-resistant (ATRA-resistant) rearrangements.

  • A molecular test (real-time quantitative polymerase chain reaction, RQ-PCR) may be ordered to help establish the initial diagnosis of APL. The PML-RARA PCR test is quantitative, that is, it provides a general estimate of the number of PML-RARA gene sequences present in a person’s blood and/or bone marrow samples. A PML-RARA PCR test is typically ordered at the time of the initial diagnosis to establish a baseline value and then periodically to monitor a person’s response to treatment and, if the person achieves remission, to monitor for recurrence. PCR will only detect the PML-RARA fusion, not the more rare combination of RARA with another gene.

  • Chromosome analysis (standard cytogenetic analysis) to detect chromosomal abnormalities (e.g., translocations or deletions, or gain or loss of chromosome) may help diagnose APL. This test method involves the evaluation of a person’s chromosomes under a microscope to detect structural and/or numerical abnormalities. Cells in a sample of blood or bone marrow are examined to determine if the PML-RARA t(15;17) chromosome translocation is present. This method can detect the more rare combination of RARA with a gene other than PML, so it may be used if one of the other test methods is negative but APL is still strongly suspected.

PML-RARA testing is often performed along with other blood and/or bone marrow tests if a healthcare practitioner suspects that a person has leukemia and is trying to diagnose or rule out APL. Some of these other tests may include:

  • Complete blood count (CBC)—evaluates the number of each type of blood cell

  • Differential—identifies and counts different types of white blood cells

  • Blood smear—blood cells are examined under a microscope

  • Immunophenotyping—classifies cells according to the markers (antigens) on the surfaces of cells

What Does the Test Result mean?

If a person has abnormal promyelocytes in the blood and bone marrow and has the PML-RARA gene sequence, then the person is diagnosed as having APL.

The presence of PML-RARA means the individual will likely benefit from treatment with all-trans retinoic acid (ATRA). This is a drug that can help promyelocytic leukemia cells to continue to differentiate and become more mature. ATRA is typically effective in those cases where the PML-RARA fusion gene is present. A small percentage of people with APL have a fusion between RARA and a different gene, and they may or may not benefit from ATRA therapy depending upon the partner gene involved.

When monitoring treatment, a decrease in the amount of PML-RARA in the blood or bone marrow over time means the person is responding to treatment. If the number of cells that have PML-RARA drops below the test’s detection limit and the person’s blood cell counts are normal, then the person is considered to be in remission. An increase in PML-RARA levels over time indicates disease progression or relapse.

If a person with APL is not positive for the PML-RARA gene sequence, then that person may not be given ATRA therapy and PML-RARA molecular testing cannot be used to monitor the person.

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