Thyroid Cancer Molecular Assessment

Medical Experts

Contributor

Matynia

Anna P. Matynia, MD

Associate Professor of Pathology (Clinical), University of Utah

Subspecialty Director, Solid Tumor Non-NGS Molecular Pathology, ARUP Laboratories

Contributor

Straseski

Joely A. Straseski, PhD, MS, MT(ASCP), DABCC, FAACC

Professor of Pathology (Clinical), University of Utah

Head of Clinical Operations for Clinical Chemistry, Toxicology, and Biochemical Genetics; Medical Director, Endocrinology; Co-Director, Automated Core Laboratory, ARUP Laboratories

Thyroid cancers are usually detected as nodules found during physical examination or as incidental findings during imaging procedures (refer to the ARUP Consult Thyroid Nodules topic for more information). If a malignancy is confirmed or suspected based on cytology, or if cytology is indeterminate, pre- and postoperative histologic examination and testing for molecular markers can be used to confirm the diagnosis, differentiate between thyroid cancer subtypes, inform prognosis, and guide treatment. If a familial syndrome is suspected, germline molecular testing may be warranted. Laboratory tests for tumor markers (including thyroid-stimulating hormone [TSH], thyroglobulin [Tg], and thyroglobulin antibody [TgAb]) are also used in thyroid cancer monitoring and surveillance. Medullary thyroid cancer (MTC) entails additional testing (for more information, refer to the Testing for Medullary Thyroid Cancer section).

For in-depth information regarding testing of fine needle aspiration (FNA) specimens from thyroid nodules, refer to the ARUP Consult Thyroid Nodules topic.

Quick Answers for Clinicians

When should molecular testing be considered in thyroid cancer?

It is recommended that molecular testing be considered in addition to, rather than as a replacement for, cytologic evaluation. Molecular testing is recommended only if results will influence clinical decision-making and is generally not recommended in nodules that are clearly benign or clearly malignant based on cytologic evaluation. If cytology samples obtained via fine needle aspiration (FNA) are indeterminate, molecular testing may be useful to support tumor diagnosis, stratify risk for progression and recurrence, and guide therapeutic decision-making.^,^,

What is the role of molecular marker panel tests in thyroid cancer?

Either multigene assays or single gene molecular marker tests may be useful in thyroid cancer, although specifically designed thyroid cancer gene panels may be particularly helpful to assess nodules with indeterminate cytology results for cancer, given that there is no single gene molecular test that can definitively confirm or rule out malignancy. However, mutation panel tests are not standardized, may use different technologies (e.g., next generation sequencing [NGS], RNA expression signature analysis, microRNA [miRNA] analysis, or a combination of technologies), and are proprietary, each with their own predictive advantages and disadvantages. For more information, refer to the Molecular Markers in Tumor Tissue section.

How does pediatric thyroid cancer differ from adult thyroid cancer, and how does this affect laboratory testing?

The evaluation and management of thyroid nodules are similar in children and adults. However, pediatric thyroid nodules are more likely to be malignant than adult nodules.^, Furthermore, some thyroid cancer treatments may not be appropriate for children. This ARUP Consult topic focuses on laboratory testing for thyroid cancer in adults; for detailed information on thyroid cancer testing in children, refer to the the American Thyroid Association's management guidelines.

Which factors have contributed to the overdiagnosis of thyroid cancer?

The overdiagnosis of thyroid cancer has become a problem worldwide. Although mortality rates have remained stable and generally low, the number of patient diagnoses has significantly increased. This increase is due to the fact that a considerable number of small thyroid cancers are now detected on radiologic tests, such as ultrasound, computed tomography (CT), and magnetic resonance imaging (MRI) scans that are performed to investigate other conditions. These small cancers are generally low risk and may not need treatment. Overdiagnosis may account for 60-90% of thyroid cancer cases globally. As a result of overdiagnosis, healthy individuals may be subject to unnecessary and sometimes harmful diagnostic tests and interventions, including surgery.

Indications for Testing

Molecular testing for thyroid cancer is appropriate in individuals with:

Thyroid nodule(s) that are indeterminate, suspicious for malignancy, or malignant based on cytology
Individuals at risk for a germline genetic variant due to family history of thyroid cancer or identification of a germline variant in a first-degree relative

Screening for thyroid cancer in the general population is not recommended.

Laboratory Testing

Histology and molecular testing can be used to further investigate thyroid nodules that have been categorized as malignant, suspicious for malignancy, or indeterminate by FNA and cytology.

Histology

Histologic examination of thyroid tissue samples is used to establish or confirm a diagnosis of thyroid cancer, classify the cancer, stratify risk, and guide management after thyroid surgery.^, Histologic features may also suggest the need for germline testing for a variant associated with a familial cancer syndrome; for example, the cribriform-morular form of papillary thyroid carcinoma (PTC), characterized by distinctive histologic features, is associated with familial adenomatous polyposis (FAP).

Molecular Markers in Tumor Tissue

Molecular markers are useful to establish malignancy if cytology samples obtained via FNA are indeterminate, and these markers are also used in risk stratification.^,^, The National Comprehensive Cancer Network (NCCN) recommends molecular diagnostic testing for nodules suspicious for follicular thyroid cancer (FTC) or atypia of undetermined significance (AUS) on FNA. Molecular markers may also guide targeted therapeutic decision-making. The appropriate specimen type (e.g., fine needle aspirate or formalin-fixed, paraffin-embedded tissue) depends on the test used.

Specifically designed thyroid cancer multigene assays may be especially helpful to establish the diagnosis of thyroid cancer because there is no single gene molecular test that can definitively confirm or rule out malignancy. Several different methodologies (e.g., next generation sequencing [NGS], multiplex polymerase chain reaction [PCR], and messenger RNA [mRNA] gene expression) may be used for these panel tests. In many cases, these molecular panel tests are based on proprietary algorithms, involve in-house interpretation, and are not standardized, factors that make it difficult to compare one assay with another.

Molecular Markers in Thyroid Cancer
Tumor Type (WHO Classification)	Common Molecular Markers
FTC	Variants in RAS family genes (NRAS, HRAS, KRAS) PAX8::PPARG rearrangements PTEN variants PIK3CA variants (activating mutations and copy number gain) DICER1 and DGCR8 variants TERT promoter variants
IEFVPTC	Variants in RAS family genes (NRAS, HRAS, KRAS) PAX8::PPARG rearrangements
PTC	BRAF V600E missense variant TERT promoter variant RET, NTRK1/3, BRAF, MET, ALK fusions PLEKHS1 aberrations TP53 variants
Oncocytic carcinoma of the thyroid (previously referred to as Hürthle cell carcinoma)	Deleterious mtDNA variants Loss of chromosomes 2, 8, and 22 Gain of chromosomes 7, 12, and 17 Genome haploidization
Follicular cell-derived carcinomas, high grade (DHGTC and PDTC)	BRAF V600E variant or RAS family variants (depending on whether associated with PTC or FTC) TP53 variants TERT promoter variants DICER1 variants
ATC	BRAF variants (associated with PTC or DHGTCs) RAS family variants (NRAS, HRAS, KRAS; when arising from FTC or EFVPTC) TERT promoter variants TP53 variants CDKN2A/B deletions
MTC	RET variants RAS variants
ATC, anaplastic thyroid carcinoma; DHGTC, differentiated high-grade thyroid carcinoma; EFVPTC, encapsulated follicular variant papillary thyroid carcinoma; IEFVPTC, invasive EFVPTC; mtDNA, mitochondrial DNA; PDTC, poorly differentiated thyroid carcinoma; WHO, World Health Organization Source: Filetti, 2019; Juhlin, 2022

Germline Genetic Testing

The cribriform-morular variant of PTC may occur in individuals with FAP and pathogenic variants in the APC gene; therefore, genetic counseling should be considered.

FTC may occur in individuals with PTEN hamartoma tumor syndrome and pathogenic variants in the PTEN gene; if the thyroid has the characteristic appearance associated with the syndrome, genetic counseling is recommended.

For more information on germline genetic testing in MTC (e.g., RET mutations), refer to the Testing for Medullary Thyroid Cancer section.

Monitoring and Surveillance

Testing is recommended after treatment with thyroidectomy, both before and after radioactive iodine (RAI) remnant ablation (if indicated).

Monitoring and surveillance for recurrence in thyroid cancer consist of a combination of imaging and laboratory testing. Follow-up laboratory tests include tests for tumor markers such as TSH, Tg, and TgAb; calcitonin and carcinoembryonic antigen (CEA) are specific tumor markers used in MTC.^,^,

Thyroid-Stimulating Hormone

Serum TSH measurement after discontinuation of levothyroxine is recommended before RAI. Serum TSH measurements are recommended at least every 12 months after thyroid cancer treatment in patients receiving thyroid hormone therapy to guide dosing and ensure TSH is maintained within the appropriate target range.

Serum Thyroglobulin and Antithyroglobulin Antibodies

A serum Tg test is the primary tumor marker test for recurrence after thyroid cancer treatment. An undetectable concentration of Tg has a high negative predictive value for recurrence.^, In addition to its use to detect recurrence, serum Tg measurement can be used to assess the success of treatment, predict the likelihood of metastatic thyroid cancer, predict the likelihood of successful remnant ablation, predict the risk of mortality, and guide clinical management and decision-making.

Because the tumor marker TgAb interferes with Tg, the presence of TgAb makes Tg values uninterpretable.^,^, Thus, simultaneous measurement of TgAb with Tg is required. The same assay from the same laboratory should be used for all measurements of Tg and TgAb during surveillance and monitoring.^,

The European Society for Molecular Oncology (ESMO) and the NCCN recommend that follow-up schedules and techniques be customized according to the type of cancer, type of treatment, response to treatment, and risk level.^, The American Thyroid Association recommends measurement of serum Tg every 6-12 months during initial follow-up, or more frequently in high-risk patients. Serum Tg can be measured every 12-24 months thereafter in patients who have excellent responses, and should be measured at least every 6-12 months in patients at high risk or with incomplete or indeterminate responses to therapy.

Refer to the Thyroid Cancer Monitoring and Surveillance algorithm for additional information.

Testing for Medullary Thyroid Cancer

Molecular markers for MTC are discussed in the Molecular Markers in Thyroid Cancer table. Monitoring and surveillance recommendations for MTC are the same as for other thyroid cancers; refer to the Monitoring and Surveillance section for additional information.

Confirmation of Diagnosis

Calcitonin and CEA are tumor markers that are useful in MTC, and calcitonin expression must be confirmed to diagnose MTC. If the serum calcitonin concentration is elevated, the test should be repeated. If the elevated concentration is confirmed, a calcium stimulation test is recommended. CEA is also expressed in MTC, but not in other primary thyroid cancers, so it may be particularly useful to measure CEA if a calcitonin-negative MTC is suspected. However, CEA expression is not specific to calcitonin-negative MTC but is associated with a number of other malignancies, and results should be interpreted with care. An elevated CEA result warrants ruling out coexisting CEA-expressing tumors and metastases, especially in cases of otherwise undetectable residual disease. As mentioned in the Histology section, calcitonin can also be evaluated by immunostaining in MTC.^,^,

Preoperative Testing

Preoperative testing for pheochromocytoma and hyperparathyroidism is recommended in patients with MTC, unless they are already known to have multiple endocrine neoplasia type 2B (MEN2B).

Prognosis

Because calcitonin and CEA are directly related to the mass of calcitonin-secreting cancer cells, they are also useful tumor markers for prognosis. However, calcitonin is the more specific tumor marker, compared with CEA, in cases of calcitonin-expressing tumors. The measurement of both calcitonin and CEA increases sensitivity for the detection of persistent disease, particularly given that tumors may fail to express or cease to express calcitonin. Measurement of calcitonin and/or CEA is recommended 60-90 days after thyroidectomy to assess for residual disease.

Calcitonin and CEA doubling times are recommended for prognosis in postoperative MTC. When both markers are expressed, calcitonin doubling time is the superior marker of MTC progression. Shorter doubling times are associated with worse prognoses. At least four consecutive measurements over a 2-year period are recommended to enable calculation of doubling time.

Germline Genetic Testing

Genetic counseling and RET germline testing are recommended for all individuals with diagnosed MTC to determine whether MTC is familial. Necessary surgical intervention should not be delayed while awaiting test results.^, RET testing is also recommended for relatives of individuals diagnosed with a familial form of MTC.