Molecular Testing Enhances Decision-Making Process in Thyroid Cancer of Indeterminate Malignancy

OncologyLive, Vol. 20/No. 17, Volume 20, Issue 17

In thyroid cancer, molecular diagnostic tests enable oncologists to evaluate thyroid nodules with atypical, suspicious, or indeterminate fine-needle aspiration cytology to determine the best course of treatment based on cytopathological result.

Andrew Turk, MD

In thyroid cancer, molecular diagnostic tests enable oncologists to evaluate thyroid nodules with atypical, suspicious, or indeterminate fine-needle aspiration (FNA) cytology to determine the best course of treatment based on cytopathological result.1,2 The Bethesda System for Reporting Thyroid Cytopathology ranks thyroid FNA across 6 categories, assigns a risk of malignancy to each category, and suggests a path for clinical management based on tier.3

“We don’t need a lot of guidance about how to deal with patients who have category II cytology [benign] because the risk of malignancy is low, and we don’t need a tremendous amount of guidance on how to deal with categories V and VI [suspicious for malignancy and malignant, respectively] because risk of malignancy is relatively high,” said Andrew Turk, MD, assistant professor of pathology and cell biology, Department of Pathology and Cell Biology, Columbia University, New York, New York, during a presentation at the 3rd Annual International Congress on Oncology and Pathology™, hosted by Physicians’ Education Resource, LLC (PER®). “What we need molecular pathology and molecular diagnostics to provide is information about how to triage and manage patients with indeterminate nodules.”

Only 2 of the Bethesda system’s 6 diagnostic categories indicate cytological indeterminacy: category III, atypia of undetermined significance or follicular lesion of undetermined significance (AUS/FLUS), and category IV, follicular neoplasm or suspicious for a follicular neoplasm (SFN).1 Bethesda scores inform treatment of patients with nodules, specifically whether surgical intervention is necessary and, if so, to what extent. For example, patients whose cytology result qualifies as category V typically undergo a near-total thyroidectomy or surgical lobectomy.3 Those with a category VI—classified sample receive a near-total thyroidectomy.

Measuring Malignancy

Table. NCCN Treatment Recommendations Based on FNA Result5

Turk highlighted mutation/fusion panels (ThyroSeq v2, ThyGenX), microRNA (miRNA) classifiers (ThyraMIR, RosettaGX Reveal), and messenger RNA gene expression classifiers (Afirma) as some of the FNA tests used to interrogate single nucleotide variants and rearrangements in thyroid nodules.1 ThyroSeq v2 and ThyGenX use next-generation sequencing (NGS) to perform FNA testing, ThyraMIR and RosettaGX Reveal facilitate quantitative reverse transcriptase polymerase chain reactions (RT-qPCR), and Afirma conducts an expression array. “The patients in whom we’re really concerned about how these tests perform, the scenarios in which these assays are really clinically relevant, are patients whose cytology results are indeterminate,” Turk said.

Table 2. NCCN Treatment Recommendations Based on Molecular Diagnostics Results5

The reporting method varies by test: ThyraMIR displays either a positive or a negative result and RosettaGX Reveal and Afirma deem the nodule to be benign or suspicious. Turk noted that sequencing is associated with high positive predictive values (PPVs). A single assay or combination of assays that includes a sequencing or mutation panel, such as ThyroSeq v2 and ThyGenX, leads to more elevated PPVs than miRNA classifiers or gene expression classifiers such as ThyraMIR, RosettaGX Reveal, and Afirma. In molecular diagnostic testing for thyroid FNAs, PPV is the cancer risk based on an abnormal (positive) test result, whereas the negative predictive value (NPV) signifies the probability of benignity based on a negative test result.4 Past study results have shown that NPVs are relatively high and comparably negative across NGS, RT-qPCR, and expression array methods; however, false-negative results can occur with the FNA test, given its high sensitivity. Therefore, a favorable FNA result should not supplant clinical or radiographic findings that suggest malignancy.5 Molecular diagnostic tests can also detect molecular aberrations associated with the pathogenesis of thyroid cancer to help identify alterations in BRAF, RAS, RET/PTC, and PAX8/PPARy.2 Knowledge of the actionable mutations present in a nodule provides valuable insight that can further sculpt the treatment decision and guide administration of systemic and targeted therapies.

The BRAF V600E mutation occurs in about 45% of patients with papillary carcinoma and is the most common mutation.5 Patients with unresectable or metastatic BRAF V600E— positive anaplastic thyroid cancer gained a targeted therapy in May 2018 with the FDA’s approval of the combination of the BRAF inhibitor dabrafenib (Tafinlar) and trametinib (Mekinist).6 Anaplastic thyroid carcinoma is the most aggressive form of thyroid cancer and is known to harbor a mutation in the BRAF oncogene, which renders the disease susceptible to the precision doublet therapy. In an interview with OncologyLive®, Turk said the FDA’s authorization of the 2 agents for use in this setting constitutes “the most significant development in the world of thyroid cancer currently.”

Confronting Indeterminacy

Although the Bethesda system standardizes clinical management based on risk of malignancy, treatment of the nodules that elicit category III and IV scores can be abstruse, and the value of surgery is comparatively less clear than it is for other categories.1 The National Comprehensive Cancer Network (NCCN) updated its Guidelines for Thyroid Carcinoma to recommend molecular diagnostic testing for patients whose cytology samples yield a category III or IV Bethesda score, but did not specify whether this assessment should involve sequence or expression-based testing, or a combination thereof.5

If the cytological result is suggestive of a benign process, the NCCN does not recommend surgical intervention. Instead, it suggests lobectomy or total thyroidectomy for those with AUS/FLUS if there is a high clinical or radiographic suspicion of malignancy; if said suspicion is low, the NCCN advises repetition of the FNA test, along with consideration of a diagnostic lobectomy, molecular diagnostics, and nodule surveillance. Lobectomy or total thyroidectomy is recommended for definitive diagnosis/treatment in those with a follicular neoplasm of supposed malignancy (Table 1 and Table 2).5

Turk expects that the clinical benefit of molecular diagnostic tests will mature in tandem with the method. “The [tests are] pretty new, and it’s an even newer development that testing is standard of care,” he said.

The updated NCCN guidelines for thyroid cancer standardize which patients should ultimately undergo molecular diagnostic testing, and when. This systematized knowledge of test application will be key to the efficacious use of the tests moving forward, Turk said, as will clinicians’ understanding of the different performance characteristics of each molecular diagnostic test.

“I think at the current stage there’s probably variability from academic centers to community practices in terms of how knowledgeable both the ordering physicians and pathologists are about these guidelines and to what extent they’re implemented,” Turk said. “Making the existing guidelines truly universal in terms of physician awareness and implementation is the [next] frontier.


  1. Turk AT. Focus on head and neck cancer: genomic profiling in the contemporary management of thyroid cancer. Presented at: the 3rd Annual Congress on Oncology and Pathology™; June 22, 2019; New York, NY. pdf/ICOP19_1_HNC-Turk.pdf.
  2. Reitz R. Thyroid cancer—the role of molecular testing. Quest Diagnostics website. 30. Published June 4, 2012. Accessed July 22, 2019.
  3. Alshaikh S, Harb Z, Aljufairi E, Almahari SA. Classification of thyroid fine-needle aspiration cytology into Bethesda categories: an institutional experience and review of the literature. Cytojournal. 2018;15:4. doi: 10.4103/cytojournal.cytojournal_32_17.
  4. Nishino M, Nikiforova M. Update on molecular testing for cytologically indeterminate thyroid nodules. Arch Pathol Lab Med. 2018;142(4):446-457. doi: 10.5858/arpa.2017-0174-RA.
  5. NCCN Clinical Practice Guidelines In Oncology. Thyroid Carcinoma, version 1.2019. National Comprehensive Cancer Network website. Published March 28, 2019. Accessed July 22, 2019.
  6. Subbiah V, Kreitman RJ, Wainberg ZA, et al. Dabrafenib and trametinib treatment in patients with locally advanced or metastatic BRAF V600-mutant anaplastic thyroid cancer. J Clin Oncol. 2018;36(1):7-13. doi: 10.1200/JCO.2017.73.6785.