Test Id : FOXOF

This test does not include a pathology consult. If a pathology consultation is requested, PATHC / Pathology Consultation should be ordered and the appropriate FISH test will be ordered and performed at an additional charge.

This test includes a charge for application of the first probe set (2 FISH probes) and professional interpretation of results. Additional charges will be incurred for all reflex probes 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.

If a pathology consultation is desired, order PATHC / Pathology Consultation.

Advise Express Mail or equivalent if not on courier service.

1. A pathology report is required in order for testing to be performed. Acceptable pathology reports include working drafts, preliminary pathology or surgical pathology reports.

2. 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.

Submit only 1 of the following specimens:

Specimen Type: Tissue

Preferred: Tissue block

Collection Instructions: Submit a formalin-fixed, paraffin-embedded (FFPE) tumor tissue block. Blocks prepared with alternative fixation methods may be acceptable; provide fixation method used.

Acceptable: Slides

Collection Instructions: Four consecutive, unstained, 5-micron thick sections placed on positively charged slides, and 1 hematoxylin and eosin-stained slide.

If not ordering electronically, complete, print, and send an Oncology Test Request (T729) with the specimen.

Two consecutive, unstained, 5-micron thick sections placed on positively charged slides, and 1 hematoxylin and eosin (H and E)-stained slide

Supporting the diagnosis of alveolar rhabdomyosarcomas (ARMS) when used in conjunction with an anatomic pathology consultation

Aiding in the diagnosis of ARMS when reverse transcriptase-PCR results are equivocal or do not support the clinical picture

This test does not include a pathology consult. If a pathology consultation is requested, PATHC / Pathology Consultation should be ordered and the appropriate FISH test will be ordered and performed at an additional charge.

This test includes a charge for application of the first probe set (2 FISH probes) and professional interpretation of results. Additional charges will be incurred for all reflex probes 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.

Rhabdomyosarcomas are a heterogeneous group of malignant tumors showing skeletal muscle differentiation. They can be divided into 3 subtypes: alveolar, embryonal, and pleomorphic. The rarer alveolar rhabdomyosarcomas (ARMS) are seen in older children, are more likely to occur in limbs, and are associated with higher stage disease and an unfavorable prognosis.

The alveolar form consists of 2 variants; classic and solid. The classic form is characterized by small round cells with dark hyperchromatic nuclei containing distinct nucleoli, held together by strands of intercellular collagen, thereby creating a cellular architecture resembling the alveolar spaces of the lungs. The solid form is characterized by a similar cellular morphology but without the formation of alveolar spaces. ARMS are also members of the small round cell tumor group that includes synovial sarcoma, lymphoma, Wilms tumor, Ewing sarcoma, and desmoplastic small round cell tumor.

Most cases of ARMS (75%) are associated with a t(2;13)(q35;q14), where a chimeric gene is formed from the rearrangement of the PAX3 gene on chromosome 2 and the FOXO1(FKHR) gene on chromosome 13. A small subset of ARMS patients (10%) are associated with a variant translocation, t(1;13)(q36;q14), involving the PAX7 gene of chromosome 1 and the FOXO1 gene. Detection of these transcripts by RT-PCR (ARMS / Alveolar Rhabdomyosarcoma by Reverse Transcriptase PCR [RT-PCR]), which allows specific identification of the t(2;13) and t(1;13), has greatly facilitated the diagnosis of ARMS tumors. FISH analysis (using the FOXO1 probe) adds the ability to detect variant FOXO1 rearrangements not detectible by PCR, and will often yield results when the quality of the available RNA is poor or the PCR results are equivocal.

An interpretive report will be provided.

A neoplastic clone is detected when the percent of cells with an abnormality exceeds the normal cutoff for the FOXO1 FISH probe.

A positive result suggests rearrangement of the FOXO1 gene region at 13q14 and is consistent with a subset of alveolar rhabdomyosarcomas (ARMS).

A negative result suggests FOXO1 gene rearrangement is not present, but does not exclude the diagnosis of alveolar rhabdomyosarcomas (ARMS).

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

Fixatives other than formalin (eg, Prefer, Bouin) may not be successful for FISH assays, however, nonformalin-fixed samples will not be rejected.

Paraffin-embedded tissues that have been decalcified are generally unsuccessful for FISH analysis. The pathologist reviewing the hematoxylin and eosin-stained slide may find it necessary to cancel testing.

FISH analysis was performed on 42 formalin-fixed paraffin-embedded tissue samples, including 17 rhabdomyosarcomas and 25 noncancerous soft tissue control specimens (from various anatomic locations). The normal controls were used to generate a normal cutoff for this assay. Rearrangement of FOXO1 was identified in 10 of 17 (59%) of alveolar rhabdomyosarcoma specimens.

1. Fletcher CDM, Unni K, Mertens F: World Health Organization Classification of Tumours. Pathology and Genetics of Tumours of Soft Tissue and Bone. IARC: Lyon 2002, pp 150-152

2. Galili N, Davis RJ, Fredericks WJ, et al: Fusion of a fork head domain gene to PAX3 in the solid tumor alveolar rhabdomyosarcoma. Nat Genet 1993 Nov;5(3):230-235

3. Nishio J, Althof PA, Bailey JM, et al: Use of a novel FISH assay on paraffin-embedded tissues as an adjunct to diagnosis of alveolar rhabdomyosarcoma. Lab Invest 2006 Jun;86(6):547-556

4. Ladanyi M. Bridge JA: Contribution of molecular genetic data to the classification of sarcomas. Hum Pathol 2000 May;31(5):532-538

5. Lae ME, Roche PC, Jin L, et al: Desmoplastic small round cell tumor: a clinicopathologic, immunohistochemical, and molecular study of 32 tumors. Am J Surg Pathol 2002 Jul;26(7):823-835

6. Barr FG: Gene fusions involving PAX and FOX family members in alveolar rhabdomyosarcoma. Oncogene 2001 Sep 10;20(40):5736-5746

The test is performed using a commercially available FOXO1 dual-color break-apart strategy probe (BAP). Formalin-fixed paraffin-embedded tissues are cut at 5 microns and mounted on positively charged glass slides. The selection of selection of tissue and the identification of target areas on the hematoxylin and eosin (H and E)-stained slide is performed by a pathologist. Using the H and E-stained slide as a reference, target areas are etched with a diamond-tipped etcher on the back of the unstained slide to be assayed. The probe set is hybridized to the appropriate target areas and 2 technologists each analyze 50 interphase nuclei each (100 total) with the results expressed as the percent abnormal of nuclei.(Unpublished Mayo method)

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This test was developed using an analyte specific reagent. Its performance characteristics were determined by Mayo Clinic in a manner consistent with CLIA requirements. This test has not been cleared or approved by the US Food and Drug Administration.

88271 x 2, 88291-DNA probe, each (first probe set), Interpretation and report

88271 x 2-DNA probe, each; each additional probe set (if appropriate)

88271-DNA probe, each; coverage for sets containing 3 probes (if appropriate)

88271 x 2-DNA probe, each; coverage for sets containing 4 probes (if appropriate)

88271 x 3-DNA probe, each; coverage for sets containing 5 probes (if appropriate)

88274 w/modifier 52-Interphase in situ hybridization, <25 cells, each probe set (if appropriate)

88274-Interphase in situ hybridization, 25 to 99 cells, each probe set (if appropriate)

88275-Interphase in situ hybridization, 100 to 300 cells, each probe set (if appropriate)