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Mayo Test ID MPNCM Myeloproliferative Neoplasm, CALR with Reflex to MPL, Varies


Shipping Instructions


Specimen must arrive within 7 days of collection.



Necessary Information


The following information is required:

1. Pertinent clinical history

2. Clinical or morphologic suspicion

3. Date of collection

4. Specimen source



Specimen Required


Submit only 1 of the following specimens:

 

Specimen Type: Whole Blood

Container/Tube: Lavender top (EDTA) or yellow top (ACD-B)

Specimen Volume: 3 mL

Collection Instructions:

1. Invert several times to mix blood.

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

3. Label specimen as blood.

Specimen Stability Information: Ambient (preferred)/Refrigerate 7 days

 

Specimen Type: Bone marrow

Container/Tube: Lavender top (EDTA) or yellow top (ACD-B)

Specimen Volume: 2 mL

Collection Instructions:

1. Invert several times to mix specimen.

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

3. Label specimen as bone marrow.

Specimen Stability Information: Ambient (preferred)/Refrigerate 7 days

 

Specimen Type: Extracted DNA from blood or bone marrow

Container/Tube: 1.5 to 2 mL tube

Specimen Volume: Entire specimen

Collection Instructions:

1. Indicate volume and concentration of DNA

2. Label specimen as extracted DNA from blood or bone marrow.

Specimen Stability Information: Frozen (preferred)/Refrigerate/Ambient


Useful For

Aiding in the distinction between a reactive cytosis and a myeloproliferative neoplasm when JAK2V617F testing result is negative

Reflex Tests

Test ID Reporting Name Available Separately Always Performed
MPNML MPL Exon 10 Sequencing, Reflex No, (Bill Only) No

Testing Algorithm

This test reflexively evaluates for variants in the CALR and MPL genes commonly associated with BCR::ABL1-negative myeloproliferative neoplasms. The testing sequence is based on the reported frequency of gene variants in this disease group. It is usually ordered when a JAK2 V617F result is known to be negative. Initial testing evaluates for the presence of the CALR insertions and deletions. If out-of-frame CALR insertions or deletions are detected, the testing algorithm ends. If the CALR result is negative or an in-frame CALR insertion or deletion is identified, then testing proceeds, at an additional charge, to evaluate for variants in exon 10 of the MPL gene by Sanger sequencing. An integrated report is issued with the summary of test results.

 

For more information the following algorithms are available:

-Myeloproliferative Neoplasm: A Diagnostic Approach to Bone Marrow Evaluation

-Myeloproliferative Neoplasm: A Diagnostic Approach to Peripheral Blood Evaluation

Method Name

Polymerase Chain Reaction (PCR) and Fragment Analysis

Reporting Name

MPN (CALR, MPL) Reflex

Specimen Type

Varies

Specimen Minimum Volume

Blood, bone marrow: 0.5 mL; Extracted DNA: 50 mcL at 20 ng/mcL concentration

Specimen Stability Information

Specimen Type Temperature Time Special Container
Varies Varies 7 days

Reject Due To

Gross hemolysis Reject
Paraffin-embedded bone marrow aspirate clot or biopsy blocks
Slides
Paraffin shavings
Moderately to severely clotted		 Reject

Clinical Information

The JAK2 V617F variant is present in 95% to 98% of patients with polycythemia vera , 50% to 60% of patients with primary myelofibrosis (PMF), and 50% to 60% of patients with essential thrombocythemia (ET) patients. Detection of the JAK2 V617F variant helps establish the diagnosis of a myeloproliferative neoplasm (MPN). However, a negative JAK2 V617F result does not indicate the absence of MPN. Other important molecular markers in BCR::ABL1-negative MPN include CALR exon 9 variants (20%-30% of PMF and ET) and MPL exon 10 variants (5%-10% of PMF and 3%-5% of ET). Variants in JAK2, CALR, and MPL are essentially mutually exclusive. A CALR variant is associated with decreased risk of thrombosis in both ET and PMF and confers a favorable clinical outcome in patients with PMF. A triple negative (JAK2 V617F, CALR, and MPL-negative) genotype is considered a high-risk molecular signature in PMF.

Reference Values

An interpretive report will be provided.

Interpretation

The results will be reported as 1 of the 3 following states:

-Positive for CALR variant

-Positive for MPL variant

-Negative for CALR and MPL variants

 

Positive variants status is highly suggestive of a myeloid neoplasm and clinicopathologic correlation is necessary in all cases.

 

Negative variant status does not exclude the presence of a myeloproliferative neoplasm or other neoplasms.

Cautions

A positive result is not specific for a particular subtype of myeloproliferative neoplasm and clinicopathologic correlation is necessary in all cases.

 

A negative result does not exclude the presence of a myeloproliferative neoplasm or other neoplastic process.

Clinical Reference

1. Klampfl T, Gisslinger H, Harutyunyan AS, et al. Somatic mutation of calreticulin in myeloproliferative neoplasms. N Engl J Med. 2013;369(25):2379-2390

2. Nangalia J, Massie CE, Baxter EJ, et al. Somatic CALR mutation in myeloproliferative neoplasms with nonmutated JAK2. N Engl J Med. 2013;369(25):2391-2405

3. Rotunno G, Mannarelli C, Guglielmelli P, et al. Impact of calreticulin mutations on clinical and hematological phenotype and outcome in essential thrombocythemia. Blood. 2014;123(10):1552-1555

4. Tefferi A, Lasho TL, Finke CM, et al. CALR vs JAK2 vs MPL-mutated or triple-negative myelofibrosis: clinical, cytogenetic and molecular comparisons. Leukemia. 2014;28(7):1472-1477

5. Pikman Y, Lee BH, Mercher T, et al. MPLW515L is a novel somatic activating mutation in myelofibrosis with myeloid metaplasia. PLoS Med. 2006;3(7):e270

6. Pardanani AD, Levine RL, Lasho T, et al. MPL515 mutations in myeloproliferative and other myeloid disorders: a study of 1182 patients. Blood. 2006;108(10):3472-3476

7. Defour JP, Chachoua I, Pecquet C, Constantinescu SN. Oncogenic activation of MPL/thrombopoietin receptor by 17 mutations at W515: implications for myeloproliferative neoplasms. Leukemia. 2016;30(5):1214-1216. doi:10.1038/leu.2015.271

Method Description

Polymerase chain reaction (PCR) amplification of CALR exon 9 is performed on DNA isolated from the patient sample. The PCR product is then run on an ABI Genetic Analyzer for fragment analysis to detect insertions and deletions. An unaltered CALR will show an amplicon at 266 base pairs (bp), an altered CALR with insertion will show an amplicon greater than 266 bp, and an altered CALR with deletion will show an amplicon smaller than 266 bp. This assay has an analytical sensitivity of approximately 6% (ie, 6 variant-containing cells in 100 total cells) in most variant types, except for the rare type of 1-bp deletion, which has a sensitivity of approximately 20%.(Unpublished Mayo method)

 

Genomic DNA is extracted, and Sanger sequencing is used to evaluate for variants in MPL, exon 10. The sensitivity of this assay is approximately 20%, such that samples containing lower percentages of altered DNA will appear negative.(Unpublished Mayo method)

Day(s) Performed

Monday through Friday

Report Available

7 to 10 days

Specimen Retention Time

Whole blood, bone marrow: 2 weeks; Extracted DNA: 3 months

Performing Laboratory

Mayo Clinic Laboratories in Rochester

Test Classification

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.

CPT Code Information

81219-CALR (calreticulin) (eg, myeloproliferative disorders), gene analysis, common variants in exon 9

81339 -MPL (MPL proto-oncogene, thrombopoietin receptor) (eg, myeloproliferative disorder) gene analysis; sequence analysis, exon 10 (if appropriate)

LOINC Code Information

Test ID Test Order Name Order LOINC Value
MPNCM MPN (CALR, MPL) Reflex In Process

 

Result ID Test Result Name Result LOINC Value
42393 MPNCM Reflex Result 82939-0
MP036 Specimen Type 31208-2
42392 Final Diagnosis 50398-7