The College of American Pathologists (CAP) offers these templates to assist pathologists in providing clinically useful and relevant information when reporting results of biomarker testing. The CAP regards the reporting elements in the templates as important elements of the biomarker test report, but the manner in which these elements are reported is at the discretion of each specific pathologist, taking into account clinician preferences, institutional policies, and individual practice.

The CAP developed these templates as educational tools to assist pathologists in the useful reporting of relevant information. It did not issue them for use in litigation, reimbursement, or other contexts. Nevertheless, the CAP recognizes that the templates might be used by hospitals, attorneys, payers, and others. The CAP cautions that use of the templates other than for their intended educational purpose may involve additional considerations that are beyond the scope of this document.

Completion of the template is the responsibility of the laboratory performing the biomarker testing and/or providing the interpretation. When both testing and interpretation are performed elsewhere (eg, a reference laboratory), synoptic reporting of the results by the laboratory submitting the tissue for testing is also encouraged to ensure that all information is included in the patient's medical record and thus readily available to the treating clinical team.

Thyroid

Select a single response unless otherwise indicated.

Note: Use of this template is optional.

Adequacy Assessment of Thyroid Fine-Needle Aspirates (note A)

___ Adequate

___ Inadequate

___ Suboptimal (explain): _____________________________

Adequacy of Resected Specimens or Cell Blocks for Testing (note A)

___ Adequate

 Estimated tumor cellularity (area used for testing): ______%

___ Suboptimal (explain): _____________________________

Note: If “Adequate” not selected, please refer to original laboratory report for explanation.

BRAF Mutational Analysis (note B)

___ No mutation detected

___ Mutation identified

  ___ p.V600E, c.1799T>A

  ___ p.K601E, c.1801A>G

  ___ Other BRAF mutation (specify): ________________

    Indicate mutant allele frequency: ______%

___ Cannot be determined (explain): ___________________

TERT Mutational Analysis (note B)

___ No mutation detected

___ Mutation identified

  ___ c.1-124 (C228T)

  ___ c.1-146 (C250T)

  ___ Other TERT mutation (specify): ________________

___ Cannot be determined (explain): ___________________

NRAS Mutational Analysis (note C)

___ No mutation detected

___ Mutation identified

  ___ p.Q61R, c.182A>G

  ___ p.Q61K, c.181C>A

  ___ Other NRAS mutation (specify): ________________

___ Cannot be determined (explain): ___________________

HRAS Mutational Analysis (note C)

___ No mutation detected

___ Mutation identified

  ___ p.Q61R, c.182A>G

  ___ p.G12V, c.35G>T

  ___ Other HRAS mutation (specify): ________________

___ Cannot be determined (explain): ___________________

KRAS Mutational Analysis (note C)

___ No mutation detected

___ Mutation identified

  ___ p.G12D, c.35G>A

  ___ Other KRAS mutation (specify): ________________

___ Cannot be determined (explain): ___________________

AKT1 Mutational Analysis (note D)

___ No mutation detected

___ Mutation identified

  ___ p.E17K, c.49G>A

  ___ Other AKT1 mutation (specify): ________________

___ Cannot be determined (explain): ___________________

TP53 Mutational Analysis (note D)

___ No mutation detected

___ Mutation identified (specify): ______________________

___ Cannot be determined (explain): ___________________

PIK3CA Mutational Analysis (note D)

___ No mutation detected

___ Mutation identified

  ___ p.H1047R, c.3140A>G

  ___ Other PIK3CA mutation (specify): ______________

___ Cannot be determined (explain): ___________________

CTNNB1 (β-catenin) Mutational Analysis (note E)

___ No mutation detected

___ Mutation identified

  ___ p.S33A, c.97T>G

  ___ Other CTNNB1 mutation (specify): _____________

___ Cannot be determined

RET Mutational Analysis (note F)

___ No mutation detected

___ Mutation identified

  ___ p.M918T, c.2753T>C

  ___ Other RET mutation (specify): _________________

Mutation type

    ___ Germline (inherited)

    ___ Somatic (sporadic)

    ___ Unknown

___ Cannot be determined (explain): ___________________

ALK Rearrangement (note G)

___ No rearrangement detected

___ Rearrangement identified

  ___ STRN/ALK

  ___ EML4/ALK

  ___ Other ALK rearrangement (specify): ____________

___ Cannot be determined (explain): ___________________

NTRK1 Rearrangement (note H)

___ No rearrangement detected

___ Rearrangement identified

  ___ NTRK1/TPM3

  ___ NTRK1/TFG

  ___ Other NTRK1 rearrangement (specify): __________

___ Cannot be determined (explain): ___________________

NTRK3 Rearrangement (note H)

___ No rearrangement detected

___ Rearrangement identified

  ___ NTRK3/ETV6

  ___ Other NTRK3 rearrangement (specify): __________

___ Cannot be determined (explain): ___________________

RET Rearrangement (note F)

___ No rearrangement detected

___ Rearrangement identified

  ___ RET/PTC1

  ___ RET/PTC3

  ___ Other RET rearrangement (specify): ____________

___ Cannot be determined (explain): ___________________

PPARG Rearrangement (note I)

___ No rearrangement detected

___ Rearrangement identified

  ___ PAX8/PPARG

  ___ CREB3L2/PPARG

  ___ Other PPARG rearrangement (specify): __________________________________________________

___ Cannot be determined (explain): ___________________

Other Markers Tested (if applicable)

Specify marker: _____________________________________

Specify results: ______________________________________

Dissection Method(s) (select all that apply)

___ Laser capture microdissection

  Specify test name*: ______________________________

___ Manual under microscopic observation

  Specify test name*: ______________________________

___ Manual without microscopic observation

  Specify test name*: ______________________________

___ Cored from block

  Specify test name*: ______________________________

___ Whole tissue section (no tumor enrichment procedure used)

  Specify test name*: ______________________________

* If more than 1 dissection method used, please specify.

BRAF Mutational Analysis Testing Method(s) (select all that apply)

___ Direct (Sanger) sequencing

___ High-resolution melting analysis

___ Next-generation (high-throughput) sequencing

___ Immunohistochemistry

  ___ VE1 clone

  ___ Other (specify): ______________________________

___ Other (specify): __________________________________

TERT Mutational Analysis Testing Method(s)

___ Direct (Sanger) sequencing

___ Next-generation (high-throughput) sequencing

___ Other (specify): __________________________________

NRAS, HRAS, KRAS, AKT1, TP53, and PIK3CA Mutational Analysis Testing Method(s) (select all that apply)

___ Direct (Sanger) sequencing

___ High-resolution melting analysis

___ Next-generation (high-throughput) sequencing

___ Immunohistochemistry

  ___ Clone (specify): ______________________________

___ Other (specify): __________________________________

NRAS Codons Assessed (select all that apply)

___ Codon 12

___ Codon 13

___ Codon 61

___ Other (specify): __________________________________

HRAS Codons Assessed (select all that apply)

___ Codon 12

___ Codon 13

___ Codon 61

___ Other (specify): __________________________________

KRAS Codons Assessed (select all that apply)

___ Codon 12

___ Codon 13

___ Codon 61

___ Other (specify): __________________________________

ALK Rearrangement Testing Method(s)

___ In situ hybridization

___ Reverse transcription–polymerase chain reaction (RT-PCR)

___ Immunohistochemistry

  ___ ALK 5A4 clone

  ___ ALK D5F3 clone

  ___ Other (specify): ______________________________

___ Next-generation (high-throughput) sequencing

PPARG Rearrangement Testing Method(s)

___ In situ hybridization

___ Reverse transcription–polymerase chain reaction (RT-PCR)

___ Immunohistochemistry

  Clone (specify): __________________________________

___ Next-generation (high-throughput) sequencing

RET/PTC1, RET/PTC3, NTRK1, and NTRK3 Rearrangement Testing Method(s)

___ In situ hybridization

___ Reverse transcription–polymerase chain reaction (RT-PCR)

___ Immunohistochemistry

  Clone (specify): __________________________________

___ Next-generation (high-throughput) sequencing

CTNNB1 Mutational Analysis Testing Method(s)

___ Direct (Sanger) sequencing

___ Next-generation (high-throughput) sequencing

___ Immunohistochemistry

  Clone (specify): _________________________________

Sensitivity/Limit of Mutation Detection (note A)

___ ≥20%

___ ≥10%

___ ≥5%

___ Other (specify): ________%

Other Methods Used (if applicable)

Specify method: ____________

___________________________________________________

___________________________________________________

Note: Fixative type, time to fixation (cold ischemia time), and time of fixation should be reported if applicable in this template or in the original pathology report.

Gene names should follow recommendations of the Human Genome Organisation (HUGO) Nomenclature Committee (www.genenames.org; accessed May 25, 2016).

All reported gene sequence variations should be identified following the recommendations of the Human Genome Variation Society (http://varnomen.hgvs.org; accessed May 25, 2016).

A. Specimen Adequacy

The collection of material for molecular studies should not affect the morphologic cytologic assessment. For fine-needle aspirates (FNAs), at the time of the FNA procedure, a small portion of the (residual) aspirated material may be collected into nucleic acids preservative. The material may represent a part of the first needle pass or a separate pass dedicated for the molecular analysis.1  The storage and transportation conditions (time, temperature) have to be specified by laboratories.

The quantity of isolated nucleic acids is the total amount of extracted nucleic acids. The minimal acceptable amount of nucleic acids will depend on the methodology and should be determined by laboratories. The quality of DNA and RNA can be assessed by amplification of housekeeping genes (eg, GAPDH, PGK1). The troubleshooting procedure for suboptimal specimens should be specified (eg, increasing and decreasing the amount of nucleic acid template).2 

The proportion of follicular thyroid epithelial cells in an FNA sample can be assessed by comparing the expression of the housekeeping gene and a gene known to be expressed predominantly in thyroid follicular cells (eg, keratin 7, thyroid transcription factor 1 [NK2 homeobox 1]), genes expressed in mimics of thyroid nodule (eg, parathyroid hormone), or genes expressed in medullary thyroid carcinoma (ie, calcitonin).35 

The sensitivity of mutation detection and the method used to establish sensitivity should be established by the laboratory for each methodology (eg, serial dilutions of the positive controls in normal blood/lymphocytes or normal formalin-fixed, paraffin-embedded tissue).

Resection specimens may be inadequate owing to improper fixation, decalcification, low tumor content, or small tumor size.

B. BRAF Mutational Analysis

The presence of BRAF V600E mutation in a FNA is indicative of about a 99% risk of cancer in the sampled thyroid nodule. When identified alone, BRAF V600E mutation may merely reflect the conventional morphology or tall cell variant of papillary thyroid carcinoma. The combination of BRAF V600E mutation with TERT, AKT1, PIK3CA, or TP53 mutations predicts a more aggressive tumor behavior.612  BRAF K601E is an unusual BRAF mutation, which had been reported in follicular variant of papillary thyroid carcinoma and rarely in follicular adenomas.13,14 

C. RAS Mutational Analysis

The finding of RAS mutation in a FNA is associated with about a 80% risk of cancer in a given nodule. The most common types of cancer with RAS mutations are the encapsulated follicular variant of papillary carcinoma and follicular carcinoma. The remaining RAS-positive thyroid nodules are usually diagnosed as follicular adenomas. Sporadic medullary thyroid carcinomas with wild-type RET genes may harbor RAS mutations (HRAS or KRAS).2,4,5,8,15,16 

D. PIK3CA, AKT1, and TP53 Mutational Analysis

PIK3CA, AKT1, and TP53 mutations are usually found in advanced thyroid cancer with propensity for dedifferentiation and distant metastasis.8,17 

E. CTNNB1 Mutational Analysis

The presence of CTNNB1 mutation in a given thyroid nodule is expected to confer a >90% risk of cancer. Point mutations in exon 3 of CTNNB1 stabilize the protein by making it insensitive for adenomatous polyposis coli (APC)–induced degradation, leading to the accumulation of β-catenin in the nucleus. In thyroid tumors, mutations in exon 3 of CTNNB1 were also reported in poorly differentiated and anaplastic carcinomas, but not in well-differentiated carcinomas or benign thyroid nodules.18 

F. RET Mutational Analysis

The presence of RET rearrangements in thyroid FNA is associated with >95% risk of cancer, most frequently classic papillary thyroid carcinoma. Mutations of the RET gene are typically present in sporadic and familial forms of medullary thyroid carcinoma. Among sporadic medullary carcinomas, RET p.M918T mutation accounts for more than 75% of all somatic RET mutations found in medullary carcinomas.19,20 

Laboratories should disclose whether the test was performed on tissue type (tumor versus normal tissue) that allows distinguishing between germline (inherited) and sporadic (acquired) mutation. Nevertheless, the distinction between sporadic and germline mutation can be reliably made only by testing a nontumorous specimen, preferably patient blood. Clinical management of patients, based on the presence of specific RET mutations, has been defined.19,20 

G. ALK Mutational Analysis

The identification of ALK fusions (STRN/ALK or EML4/ALK) in a thyroid FNA is associated with a very high risk of thyroid cancer. ALK fusions were identified in ∼1.5% of papillary thyroid carcinomas and in 4% to 9% of dedifferentiated thyroid cancers.21,22  In advanced papillary thyroid carcinomas and in dedifferentiated thyroid tumors, the presence of an ALK fusion may represent a therapeutic target for crizotinib.21,22 

H. NTRK1 and NTRK3 Mutational Analysis

Rearrangements of the NTRK1 gene occur in <5% of papillary thyroid carcinomas.23  Different fusion partners of NTRK1 have been described including TPM3 and TPR genes. Some studies reported that NTRK1 fusion–positive papillary thyroid carcinomas may have more aggressive biological behavior and higher rate of local recurrence.24  NTRK3 fusions have been reported in papillary thyroid carcinomas.25,26  In vitro studies showed that ETV6/NTRK3 aberrantly activates phosphatidylinositide 3-kinase signaling pathway. A phase 1a/1b clinical trial of the oral tyrosine kinase inhibitor LOXO-101 is available.

I. PPARG Mutational Analysis

The presence of rearrangements involving the PPARG gene, PAX8/PPARG and less frequently CREB3L2/PPARG, correlates with ∼95% risk of cancer, most frequently follicular variant of papillary carcinoma, followed in frequency by follicular carcinoma. Rare cases of follicular adenoma carrying PPARG rearrangements have been reported.27,28  Most of thyroid cancers positive for PPARG rearrangements are low-grade tumors, whereas 5% to 10% of those tumors have aggressive behavior. Of note, PPARG fusions can be exploited as a therapeutic target for advanced thyroid cancer.

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Author notes

The authors have no relevant financial interest in the products or companies described in this article.