Test Id : BALFP

This test includes a charge for the probe application, analysis, and professional interpretation of results for 9 probe sets (19 individual fluorescence in situ hybridization [FISH] probes). Additional charges will be incurred for all reflex or additional probe sets performed. Analysis charges will be incurred based on the number of cells analyzed per probe set. If no cells are available for analysis, no analysis charges will be incurred.

This test is performed as panel testing only using the following analysis algorithm.

The diagnostic pediatric/young adult B-lymphoblastic leukemia (B-ALL) FISH panel includes testing for the following abnormalities using the FISH probes listed:

CRLF2 (Xp22.33) or (Yp11.32) rearrangement, CRLF2 break-apart probe set

t(1;19)(q23;p13) or TCF3::PBX1 fusion, PBX1/TCF3 probe set

Hyperdiploidy or +4,+10,+17, D4Z1/D10Z1/D17Z1 probe set

t(8;14)(q24.21;q32) or IGH::MYC fusion, MYC/IGH probe set

t(8q24.21;var) or MYC rearrangement, MYC break-apart probe set

t(9;22)(q34;q11.2) or BCR::ABL1 fusion, ABL1/BCR probe set

t(11q23;var) or KMT2A rearrangement, KMT2A break-apart probe set

t(12;21)(p13;q22) or ETV6::RUNX1 fusion or iAMP21, ETV6/RUNX1 probe set

t(14q32;var) or IGH rearrangement, IGH break-apart probe set

If results for the initial panel are negative or demonstrate nonclassical abnormalities, the B-lymphoblastic leukemia/lymphoma with BCR::ABL1-like features panel will be performed as a secondary panel. This panel includes testing for the following kinase activating chromosome abnormalities, using the FISH probes listed below, as well as IKZF1 deletion, which often accompanies BCR::ABL1-like ALL.

t(1q25;var) or ABL2 rearrangement, ABL2 break-apart probe set

t(5q32;var) or PDGFRB rearrangement, PDGFRB break-apart probe set

t(9p24.1;var) or JAK2 rearrangement, JAK2 break-apart probe set

t(9q34;var) or ABL1 rearrangement, ABL1 break-apart probe set

7p- or IKZF1 deletion, IKZF1/CEP7 probe set

Appropriate ancillary probes may be performed at consultant discretion to render comprehensive assessment. Any additional probes used will have the results included within the final report and will be performed at an additional charge. In the following situations, additional (reflex) testing may be performed at the laboratory's discretion and may be influenced by available karyotype results or other FISH testing.

When a KMT2A rearrangement is identified, testing with 1 or more dual-fusion FISH (D-FISH) probe sets may be performed in an attempt to identify the translocation partner for the following abnormalities:

t(4;11)(q21;q23) or KMT2A::AFF1 fusion, AFF1/KMT2A probe set

t(6;11)(q27;q23) or KMT2A::AFDN ;fusion, AFDN/KMT2A probe set

t(9;11)(p22;q23) or KMT2A::MLLT3 fusion, MLLT3/KMT2A probe set

t(10;11)(p12;q23) or KMT2A::MLLT10 fusion, MLLT10/KMT2A probe set

t(11;19)(q23;p13.1) or KMT2A::MLLT1 fusion, KMT2A/ELL probe set

t(11;19)(q23;p13.3) or KMT2A::ELL fusion, KMT2A/MLLT1 probe set

When an unbalanced CRLF2 rearrangement is identified concurrently with an IGH rearrangement, testing using the CRLF2/IGH probe set will be considered to identify a potential t(X;14)(p22.33;q32) or t(Y;14)(p11.32;q32) cryptic translocation.

When an extra or atypical ABL1 signal is identified in the absence of the BCR::ABL1 fusion, testing using the ABL1 break-apart probe set may be performed to identify a potential variant translocation involving ABL1, t(9;var)(q34;?).

When an extra ETV6 signal is identified in the absence of ETV6::RUNX1 fusion, testing using the ETV6 break-apart probe set may be performed to evaluate for the presence or absence of a potential rearrangement involving ETV6 t(12;var)(p13;?).

When a MYC rearrangement is identified, testing using both the BCL2 and BCL6 break-apart probe sets will be performed.

If an unbalanced rearrangement of BCL2 is identified, testing using the IGH/BCL2 probe set will be considered to identify a potential t(14;18)(q32;q21) or IGH::BCL2 fusion.

For more information see B-Lymphoblastic Leukemia/Lymphoma Algorithm.

This test is only performed on specimens from patients with B-cell acute lymphoblastic leukemia/lymphoma (B-ALL/LBL) who are aged 30 years or younger.

This test is intended to be ordered when the entire B-ALL fluorescence in situ hybridization (FISH) panel is needed for a pediatric patient.

This test should NOT be used to screen for residual B-ALL/LBL. At follow-up, or if the patient clinically relapses, conventional cytogenetic studies (CHRBM / Chromosome Analysis, Hematologic Disorders, Bone Marrow) is useful to identify cytogenetic changes in the neoplastic clone or the possible emergence of a new therapy-related myeloid clone. Additionally, targeted B-ALL FISH probes can be evaluated based on the abnormalities identified in the diagnostic study.

If targeted B-cell ALL FISH probes are preferred, order BALMF / B-Cell Acute Lymphoblastic Leukemia/Lymphoma (ALL), Specified FISH, Varies, and request specific probes for targeted abnormalities.

If this test is ordered on a patient aged 31 years or older, this test will be canceled and automatically reordered by the laboratory as BALAF / B-Cell Acute Lymphoblastic Leukemia/Lymphoma (ALL), FISH, Adult, Varies.

If this test is ordered and the laboratory is informed that the patient is on a Children's Oncology Group (COG) protocol, this test will be canceled and automatically reordered by the laboratory as COGBF / B-Cell Acute Lymphoblastic Leukemia/Lymphoma (ALL), Children's Oncology Group Enrollment Testing, FISH, Varies.

If either AMLFP / Pediatric Acute Myeloid Leukemia Panel, FISH, Varies; or TALFP / Pediatric T-Lymphoblastic Leukemia/Lymphoma Panel, FISH, Varies, is ordered concurrently with this test, the laboratory may cancel this test and automatically reorder as BALMF / B-Cell Acute Lymphoblastic Leukemia/Lymphoma (ALL), Specified FISH, Varies with the following FISH probes: ETV6/RUNX1, PBX1/TCF3, 4/10/17, break-apart IGH, break-apart CRLF2, break-apart MYC, break-apart ABL2, break-apart PDGFRB, break-apart JAK2, break-apart ABL1, and IKZF1/cep7. If an abnormality is identified that would result in reflex testing in BALFP, the same reflex testing will be performed in the BALMF. This cancellation is necessary to avoid duplicate testing. Probes for ABL1/BCR and break-apart KMT2A will still be performed as part of the pediatric T-ALL FISH panel.

For patients with B-cell lymphoma, order BLPMF / B-Cell Lymphoma, Specified FISH, Varies.

For testing paraffin-embedded tissue samples from patients with B-LBL, order BLBLF / B-Cell Lymphoblastic Leukemia/Lymphoma, FISH, Tissue. If a paraffin-embedded tissue sample is submitted for this test, it will be canceled and BLBLF will be added and performed as the appropriate test.

At diagnosis, conventional cytogenetic studies (CHRBM / Chromosome Analysis, Hematologic Disorders, Bone Marrow) and this fluorescence in situ hybridization panel should be performed. If there is limited specimen available, only this test will be performed.

Advise Express Mail or equivalent if not on courier service.

1. A reason for testing must be provided. If this information is not provided, an appropriate indication for testing may be entered by Mayo Clinic Laboratories.

2. A flow cytometry and/or a bone marrow pathology report should be submitted with each specimen. The laboratory will not reject testing if this information is not provided, but appropriate testing and interpretation may be compromised or delayed.

3. If the patient has received an opposite sex bone marrow transplant, note this information on the request.

4. If the patient has Down syndrome, note this information on the request.

Submit only 1 of the following specimens:

Preferred

Specimen Type: Bone marrow

Container/Tube:

Preferred: Yellow top (ACD)

Acceptable: Green top (sodium heparin) or lavender top (EDTA)

Specimen Volume: 2 to 3 mL

Collection Instructions:

1. It is preferable to send the first aspirate from the bone marrow collection.

2. Invert several times to mix bone marrow.

3. Send bone marrow specimen in original tube. Do not aliquot.

Acceptable

Specimen Type: Whole blood

Container/Tube:

Preferred: Yellow top (ACD)

Acceptable: Green top (sodium heparin) or lavender top (EDTA)

Specimen Volume: 6 mL

Collection Instructions:

1. Invert several times to mix blood.

2. Send whole blood specimen in original tube. Do not aliquot.

• B-Lymphoblastic Leukemia/Lymphoma Genetic Testing Guidelines
• Acute Leukemias of Ambiguous Lineage Testing Algorithm

If not ordering electronically, complete, print, and send an Hematopathology/Cytogenetics Test Request (T726) with the specimen.

Bone marrow: 1 mL; Whole blood: 2 mL

Detecting, at diagnosis, recurrent common chromosome abnormalities associated with B-cell acute lymphoblastic leukemia/lymphoma (B-ALL/LBL) and BCR::ABL1-like B-ALL in pediatric/young adult patients using a laboratory-designated probe set algorithm

As an adjunct to conventional chromosome studies in pediatric/young adult patients with B-ALL

Evaluating specimens in which chromosome studies are unsuccessful

This test should not be used to screen for residual B-ALL/LBL.

This test includes a charge for the probe application, analysis, and professional interpretation of results for 9 probe sets (19 individual fluorescence in situ hybridization [FISH] probes). Additional charges will be incurred for all reflex or additional probe sets performed. Analysis charges will be incurred based on the number of cells analyzed per probe set. If no cells are available for analysis, no analysis charges will be incurred.

This test is performed as panel testing only using the following analysis algorithm.

The diagnostic pediatric/young adult B-lymphoblastic leukemia (B-ALL) FISH panel includes testing for the following abnormalities using the FISH probes listed:

CRLF2 (Xp22.33) or (Yp11.32) rearrangement, CRLF2 break-apart probe set

t(1;19)(q23;p13) or TCF3::PBX1 fusion, PBX1/TCF3 probe set

Hyperdiploidy or +4,+10,+17, D4Z1/D10Z1/D17Z1 probe set

t(8;14)(q24.21;q32) or IGH::MYC fusion, MYC/IGH probe set

t(8q24.21;var) or MYC rearrangement, MYC break-apart probe set

t(9;22)(q34;q11.2) or BCR::ABL1 fusion, ABL1/BCR probe set

t(11q23;var) or KMT2A rearrangement, KMT2A break-apart probe set

t(12;21)(p13;q22) or ETV6::RUNX1 fusion or iAMP21, ETV6/RUNX1 probe set

t(14q32;var) or IGH rearrangement, IGH break-apart probe set

If results for the initial panel are negative or demonstrate nonclassical abnormalities, the B-lymphoblastic leukemia/lymphoma with BCR::ABL1-like features panel will be performed as a secondary panel. This panel includes testing for the following kinase activating chromosome abnormalities, using the FISH probes listed below, as well as IKZF1 deletion, which often accompanies BCR::ABL1-like ALL.

t(1q25;var) or ABL2 rearrangement, ABL2 break-apart probe set

t(5q32;var) or PDGFRB rearrangement, PDGFRB break-apart probe set

t(9p24.1;var) or JAK2 rearrangement, JAK2 break-apart probe set

t(9q34;var) or ABL1 rearrangement, ABL1 break-apart probe set

7p- or IKZF1 deletion, IKZF1/CEP7 probe set

Appropriate ancillary probes may be performed at consultant discretion to render comprehensive assessment. Any additional probes used will have the results included within the final report and will be performed at an additional charge. In the following situations, additional (reflex) testing may be performed at the laboratory's discretion and may be influenced by available karyotype results or other FISH testing.

When a KMT2A rearrangement is identified, testing with 1 or more dual-fusion FISH (D-FISH) probe sets may be performed in an attempt to identify the translocation partner for the following abnormalities:

t(4;11)(q21;q23) or KMT2A::AFF1 fusion, AFF1/KMT2A probe set

t(6;11)(q27;q23) or KMT2A::AFDN ;fusion, AFDN/KMT2A probe set

t(9;11)(p22;q23) or KMT2A::MLLT3 fusion, MLLT3/KMT2A probe set

t(10;11)(p12;q23) or KMT2A::MLLT10 fusion, MLLT10/KMT2A probe set

t(11;19)(q23;p13.1) or KMT2A::MLLT1 fusion, KMT2A/ELL probe set

t(11;19)(q23;p13.3) or KMT2A::ELL fusion, KMT2A/MLLT1 probe set

When an unbalanced CRLF2 rearrangement is identified concurrently with an IGH rearrangement, testing using the CRLF2/IGH probe set will be considered to identify a potential t(X;14)(p22.33;q32) or t(Y;14)(p11.32;q32) cryptic translocation.

When an extra or atypical ABL1 signal is identified in the absence of the BCR::ABL1 fusion, testing using the ABL1 break-apart probe set may be performed to identify a potential variant translocation involving ABL1, t(9;var)(q34;?).

When an extra ETV6 signal is identified in the absence of ETV6::RUNX1 fusion, testing using the ETV6 break-apart probe set may be performed to evaluate for the presence or absence of a potential rearrangement involving ETV6 t(12;var)(p13;?).

When a MYC rearrangement is identified, testing using both the BCL2 and BCL6 break-apart probe sets will be performed.

If an unbalanced rearrangement of BCL2 is identified, testing using the IGH/BCL2 probe set will be considered to identify a potential t(14;18)(q32;q21) or IGH::BCL2 fusion.

For more information see B-Lymphoblastic Leukemia/Lymphoma Algorithm.

In the United States, the incidence of B-lymphoblastic leukemia/lymphoma (B-ALL/LBL) is roughly 6000 new cases per year or approximately 1 in 50,000. B-ALL/LBL accounts for approximately 70% of all childhood leukemia cases (ages 0 to 19 years), making it the most common type of childhood cancer. It has a peak incidence at 2 to 5 years of age. This incidence decreases with age before increasing again at around age 50. B-ALL/LBL is slightly more common in male patients than female patients. There is also an increased incidence of B-ALL/LBL in individuals with genetic conditions such as Down syndrome, Fanconi anemia, Bloom syndrome, ataxia telangiectasia, Li-Fraumeni syndrome, X-linked agammaglobulinemia, and severe combined immunodeficiency. The overall cure rate for B-ALL/LBL in children is approximately 90%, and about 45% to 60% of adults have long-term disease-free survival. Of note, the IGH::CRLF2 fusion is more commonly observed in patients with Down syndrome or of Hispanic descent.

Specific cytogenetic abnormalities are identified in most cases of B-ALL/LBL by conventional chromosome studies or fluorescence in situ hybridization (FISH) studies. B-ALL genetic subgroups are important to detect and can be critical prognostic markers. For example, a decision for early transplantation may be made if BCR::ABL1 fusion, KMT2A rearrangement, iAMP21, or a hypodiploid clone is identified. In contrast, if the ETV6::RUNX1 fusion or hyperdiploidy is identified, the patient has a more favorable prognosis and transplantation is rarely initially considered.

A newly recognized World Health Organization entity called B-ALL with BCR::ABL1-like features  is increasing in importance due to the poor prognosis seen in pediatric, adolescent, and young adult ALL. Common features of this entity involve rearrangements with tyrosine kinase genes involving the following genes: ABL2, PDGFRB, JAK2, ABL1, CRLF2, and P2RY8, as well as deletions involving IKZF1. Patients who have failed conventional therapies have demonstrated favorable responses to targeted therapies when rearrangements involving these specific gene regions have been identified.

Evaluation of the MYC gene region is included in all diagnostic B-ALL panels to evaluate for Burkitt lymphoma. If a positive result is obtained, additional testing for the BCL2 and BCL6 gene regions will be considered.

Per National Comprehensive Cancer Network guidelines, a combination of cytogenetic and FISH testing is currently recommended in all pediatric and adult patients with B-ALL/lymphoblastic lymphoma (LBL). Additional cytogenetic techniques such as chromosomal microarray (CMAH / Chromosomal Microarray, Hematologic Disorders, Varies) may be helpful to resolve questions related to ploidy (hyperdiploid clone vs doubled hypodiploid clone) or to resolve certain clonal structural rearrangements such as the presence or absence of intrachromosomal amplification of chromosome 21 (iAMP21). A summary of the characteristic chromosome abnormalities identified in B-ALL is listed in the following table.

Table. Common Chromosome Abnormalities in B-cell Acute Lymphoblastic Leukemia/Lymphoma 

An interpretive report will be provided.

A neoplastic clone is detected when the percent of cells with an abnormality exceeds the normal reference range for any given probe set.

The absence of an abnormal clone does not rule out the presence of a neoplastic disorder.

This test is not approved by the US Food and Drug Administration, and it is best used as an adjunct to clinical and pathologic information.

Fluorescence in situ hybridization (FISH) is not a substitute for conventional chromosome studies because the latter detects chromosome abnormalities associated with other hematological disorders that would be missed in a targeted B-lymphoblastic leukemia FISH panel test.

Bone marrow is the preferred specimen type for this FISH test. If bone marrow is not available, a blood specimen may be used if there are circulating malignant cells in the blood specimen (as verified by a hematopathologist).

If no FISH signals are observed post-hybridization, the case will be released indicating a lack of FISH results.

1. Moorman AV, Harrison CJ, Buck GA, et al. Karyotype is an independent prognostic factor in adult acute lymphoblastic leukemia (ALL): analysis of cytogenetic data from patients treated on the Medical Research Council (MRC) UKALLXII/Eastern Cooperative Oncology Group (ECOG) 2993 trial. Blood. 2007;109(8):3189-3197

2. Moorman AV. The clinical relevance of chromosomal and genetic abnormalities in B-cell precursor acute lymphoblastic leukemia. Blood Rev. 2012;26:123-135

3. Roberts KG, Li Y, Payne-Turner D, et al. Targetable kinase-activating lesions in Ph-like acute lymphoblastic leukemia. N Engl J Med. 2014;371(11):1005-1015

4. Mullighan CG. The genomic landscape of acute lymphoblastic leukemia in children and young adults. Hematology Am Soc Hematol Educ Program. 2014;2014(1):174-180

5. Swerdlow SH, Campo E, Harris NL, et al, eds: WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues. 4th ed. IARC Press; 2017. WHO Classification of Tumours. Vol 2.

This test is performed using commercially available and laboratory-developed probes. Deletion of IKZF1 on chromosome 7 and gain or losses of chromosomes 4, 10, and 17 are detected using enumeration strategy probes. Rearrangements involving CRLF2, ABL2, BCL3, PDGFRB, MYC, JAK2, ABL1, MLL, ETV6, IGH, and BCL2 are detected using dual-color break-apart (BAP) strategy probes. Dual-color, dual-fusion fluorescence in situ hybridization (D-FISH) strategy probe sets are used to detect t(X/Y;14), t(1;19), t(8;14), t(9;22), t(12;21), t(14;18), and in reflex testing when a rearrangement of the KMT2A gene is detected. Amplification of the RUNX1 gene region is detected using a D-FISH probe to enumerate copies of the RUNX1 probe. For enumeration and BAP strategy probe sets, 100 interphase nuclei are scored; 200 interphase nuclei are scored when D-FISH probes are used. Results are expressed as the percent abnormal nuclei.(Unpublished Mayo method)

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This test was developed and its performance characteristics determined by Mayo Clinic in a manner consistent with CLIA requirements. It has not been cleared or approved by the US Food and Drug Administration.

88271 x19, 88275 x9, 88291 x1-FISH Probe, Analysis, Interpretation; 9 probe sets

88271 x2, 88275 x1-FISH Probe, Analysis; each additional probe set (if appropriate)