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Molecular diagnosis and targeted treatment of advanced follicular cell-derived thyroid cancer in the precision medicine era

  • Jaume Capdevila
    Correspondence
    Corresponding author at: Medical Oncology Department, Gastrointestinal and Endocrine Tumor Unit, Vall d'Hebron University Hospital, Vall d'Hebron Institute of Oncology (VHIO), Pg Vall d'Hebron, 119-129, 08035 Barcelona, Spain.
    Affiliations
    Medical Oncology Department, Vall d'Hebron University Hospital, Vall d'Hebron Institute of Oncology (VHIO), IOB-Teknon, Barcelona, Spain
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  • Ahmad Awada
    Affiliations
    Oncology Medicine Department, Institut Jules Bordet, Université Libre de Bruxelles, Brussels, Belgium
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  • Dagmar Führer-Sakel
    Affiliations
    Department of Endocrinology, Diabetes and Metabolism, Endocrine Tumor Center at West German Cancer Center, University Hospital Essen, University of Duisburg-Essen, Essen, Germany
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  • Sophie Leboulleux
    Affiliations
    Department of Nuclear Medicine and Endocrine Oncology, Gustave Roussy and University Paris Saclay, Villejuif, France

    Department of Endocrinology, Diabetes, Nutrition and Therapeutic Patient Education, Geneva University Hospitals, Geneva, Switzerland
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  • Patrick Pauwels
    Affiliations
    Department of Pathology, Center for Oncological Research, University Hospital of Antwerp, Edegem, Belgium
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Open AccessPublished:March 16, 2022DOI:https://doi.org/10.1016/j.ctrv.2022.102380

      Highlights

      • Actionable alterations in thyroid cancer include those in BRAF, RET, NTRK, ALK, and MTOR.
      • Targeted therapies for these alterations have shown promising efficacy.
      • Routine testing can identify patients who could benefit from targeted therapies.

      Abstract

      Most malignant thyroid tumours are initially treated with surgery or a combination of surgery and radioactive iodine (RAI) therapy. However, in patients with metastatic disease, many tumours become refractory to RAI, and these patients require alternative treatments, such as locoregional therapies and/or systemic treatment with multikinase inhibitors. Improvements in our understanding of the genetic alterations that occur in thyroid cancer have led to the discovery of several targeted therapies with clinical efficacy. These alterations include NTRK (neurotrophic tyrosine receptor kinase) gene fusions, with the tropomyosin receptor kinase inhibitors larotrectinib and entrectinib both approved by the European Medicines Agency and in other markets worldwide. Inhibitors of aberrant proteins resulting from alterations in RET (rearranged during transfection) and BRAF (B-Raf proto-oncogene) have also shown promising efficacy, and so far have received approval by the US Food and Drug Administration. Selpercatinib, a RET kinase inhibitor, was approved for use in Europe in early 2021. With the discovery of multiple actionable targets, it is imperative that effective testing strategies for these genetic alterations are integrated into the diagnostic armamentarium to ensure that patients who could potentially benefit from targeted treatments are identified. In this review, we offer our recommendations on the optimal testing strategies for detecting genetic alterations in thyroid cancer that have the potential to be targeted by molecular therapy. We also discuss the future of treatments for thyroid cancers, including the use of immune checkpoint inhibitors, and new generations of targeted treatments that are being developed to counter acquired tumour resistance.

      Keywords

      Introduction

      Thyroid cancer is the most common endocrine malignancy, accounting for 3% of all cancers diagnosed annually worldwide [
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      ]. Thyroid cancers can originate from two types of endocrine cells: parafollicular C cells and follicular thyroid cells. Cancers derived from parafollicular C cells are termed medullary thyroid cancers (MTCs) and make up a small proportion of thyroid cancers (2–3%) [
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      ]. Follicular cell-derived thyroid malignancies account for most thyroid tumours and may be subdivided into: follicular thyroid cancer (FTC; 6–10% of all thyroid cancers); papillary thyroid cancer (PTC; accounting for 65–93% of all thyroid cancers); poorly differentiated thyroid cancer (PDTC; 0.3–6.7% of all cases); Hürthle cell cancers and anaplastic thyroid cancer (ATC; the rarest, but most aggressive type of the follicular cell-derived malignancies) [
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      ]. PTC and FTC are grouped as differentiated thyroid cancer (DTC) [
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      ]. For a recent review of Hürthle cell cancer, please see literature by Ganley et al. [
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      ] and Tiedje & Fagin [
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      ]. PDTC was recognized as a distinct thyroid cancer subtype in the 2004 WHO Classification of Tumours [
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      ]. It has an intermediate prognosis between DTC and ATC, and usually presents in older patients [
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      ]. Guidelines for treating PDTC have not been standardized, with most treatment options being derived from those for DTC. Whilst surgery may be effective for patients with PDTC, the advantages of adjuvant treatment are inconclusive [
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      ].
      The prognosis for ATC is the poorest of the follicular cell-derived malignancies because of the often advanced, metastatic stage at presentation, with large and fast-growing primary tumours rendering the possibility of total resection unlikely [
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      ]. Furthermore, treatment options for ATC are currently more limited than for DTC, and typically involve a multimodal approach comprising surgery followed by high-dose external beam radiation therapy (EBRT), with or without concomitant chemotherapy [
      • Filetti S.
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      ]. There is some evidence that multimodal treatment can provide some survival benefit in patients with local ATC; however, this approach can also negatively impact on quality of life [
      • Filetti S.
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      ]. A recent retrospective cohort study of patients with ATC over a near 20-year period found that survival has increased significantly over time, with changes in the management of patients cited as the primary reason for this improvement [
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      ].
      Primary treatment options for thyroid cancers are determined by preoperative risk assessment. Surgery is indicated for most patients, the extent of which (lobectomy, total thyroidectomy with or without central neck dissection, and extended resection) depends on patient and disease characteristics. Radioactive iodine (RAI) is indicated to prevent/treat recurrent disease for patients with FTC and PTC who have undergone total thyroidectomy [
      • Filetti S.
      • Durante C.
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      • Leboulleux S.
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      ]. However, approximately 60–70% of patients with PDTC and metastatic DTC eventually have thyroid cancer that displays resistance to RAI (RAI-refractory [RAI-R]). Life expectancy for patients with RAI-R thyroid cancer is 3–5 years [
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      ]. Locoregional treatments, including surgery or EBRT, may be options for patients with targetable, locally oligometastatic disease that is not curable with RAI [
      • Filetti S.
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      • et al.
      Thyroid cancer: ESMO clinical practice guidelines for diagnosis, treatment and follow-up.
      ]. For patients who have locally advanced and/or symptomatic metastatic RAI-R DTC, the multikinase inhibitors (MKIs) lenvatinib and sorafenib are standard first-line systemic therapy in Europe [
      • Filetti S.
      • Durante C.
      • Hartl D.
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      • Locati L.D.
      • Newbold K.
      • et al.
      Thyroid cancer: ESMO clinical practice guidelines for diagnosis, treatment and follow-up.
      ,
      • Fugazzola L.
      • Elisei R.
      • Fuhrer D.
      • Jarzab B.
      • Leboulleux S.
      • Newbold K.
      • et al.
      2019 European Thyroid Association guidelines for the treatment and follow-up of advanced radioiodine-refractory thyroid cancer.
      ]. The efficacy of lenvatinib and sorafenib has been demonstrated in the SELECT and DECISION phase III trials, respectively [
      • Schlumberger M.
      • Tahara M.
      • Wirth L.J.
      • Robinson B.
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      Lenvatinib versus placebo in radioiodine-refractory thyroid cancer.
      ,
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      • Elisei R.
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      ], and confirmed in real-world studies [
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      ,
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      ,
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      Tertiary care experience of sorafenib in the treatment of progressive radioiodine-refractory differentiated thyroid carcinoma: a Korean multicenter study.
      ,
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      • Garcia-Aleman J.
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      • Trigo-Perez J.M.
      • Tinahones-Madueno F.
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      ,
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      • Felicetti F.
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      • et al.
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      ]. There are currently no biomarkers to identify patients who may respond favourably to MKI treatment.
      Recent advances in our understanding of the molecular pathways involved in tumorigenesis have been made by identification of a wide spectrum of targetable genetic alterations in the pathogenesis of thyroid cancer. These oncogenic drivers include, in particular, neurotrophic tyrosine receptor kinase (NTRK) gene fusions, B-Raf proto-oncogene (BRAF) fusions and mutations, and, in cases of MTC, rearranged during transfection (RET) receptor tyrosine kinase fusion and mutations. The identification of these oncogenic drivers has led to the development of new and specific targeted therapies to add to the treatment armamentarium for advanced thyroid cancer. To identify patients for whom specific targeted treatments may be beneficial, it is important that genomic testing becomes a routine step in the clinical assessment of patients with advanced thyroid cancer. Here we offer our opinion on how genetic testing for targetable oncogenic drivers can be integrated into the testing and treatment algorithms for DTC, PDTC, and ATC, with a focus on European practice.

      Actionable molecular alterations in advanced thyroid cancer

      DTC and ATC are characterized by molecular alterations that lead to expression of aberrant proteins involved in different cellular pathways. PDTC shares certain molecular features with DTC and ATC, and shows no specific genetic pattern [
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      ,
      • Cabanillas M.E.
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      ]. Therefore, the transition from PTC or FTC to PDTC and ATC is broadly attributed to additional molecular alterations [
      • Tirrò E.
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      • Romano C.
      • Vitale S.R.
      • Motta G.
      • Di Gregorio S.
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      ] (although there is conflicting evidence regarding the evolution of ATC [
      • Cabanillas M.E.
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      Targeted therapy for advanced thyroid cancer: kinase inhibitors and beyond.
      ,
      • Capdevila J.
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      • Iglesias C.
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      ]). The identification of several molecular alterations in advanced thyroid cancer has provided the opportunity to target these aberrant cellular pathways and offer treatment options for patients with otherwise poor prognoses (Fig. 1 and Table 1).
      Figure thumbnail gr1
      Fig. 1Potential therapeutic strategies targeting cellular aberrations in thyroid cancer [
      • Schlumberger M.
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      Lenvatinib versus placebo in radioiodine-refractory thyroid cancer.
      ,
      • Cabanillas M.E.
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      Targeted therapy for advanced thyroid cancer: kinase inhibitors and beyond.
      ,

      Novartis Pharmaceuticals Corporation. Dabrafenib Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2018/202806s010lbl.pdf; 2018 [accessed 25 November 2021].

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      ,

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      ,
      • Kheder E.S.
      • Hong D.S.
      Emerging targeted therapy for tumors with NTRK fusion proteins.
      ]. AKT, protein kinase B; ALK, anaplastic lymphoma kinase; CTLA-4, cytotoxic T-lymphocyte-associated protein 4; ERK, extracellular signal-regulated kinase; FGFR, fibroblast growth factor receptor; KIT, proto-oncogene c-Kit; MEK, mitogen-activated protein/extracellular signal-regulated kinase kinase; mTOR, mechanistic target of rapamycin; PD-1, programmed cell death-1; PD-L1, programmed death-ligand 1; PI3K, phosphatidylinositol 3-kinase; PKC, protein kinase C; PLCɣ, phospholipase C-gamma; RAF, rapidly accelerated fibrosarcoma; RAS, rat sarcoma; RET, rearranged during transfection; ROS1, c-ros oncogene 1; TRK, tropomyosin receptor kinase; VEGFR, vascular endothelial growth factor receptor.
      Table 1Molecular alterations in follicular cell-derived thyroid cancer with selected targeted treatments.
      Oncogenic driverTargeted treatmentNumber of patients with thyroid cancer included in efficacy analysisEfficacy in patients with thyroid cancer
      Response rate*Median duration of response (months)Median OS (months)Median PFS (months)
      NTRK gene fusionLarotrectinib

      Bayer AG. VITRAKVI Summary of Product Characteristics, https://www.ema.europa.eu/en/documents/product-information/vitrakvi-epar-product-information_en.pdf; 2022 [accessed 1 March 2022].

      PTC: 20

      FTC: 2

      ATC: 7

      Waguespack S, Drilon A, Lin J, Brose M, McDermott R, Almubarak M, et al. Long-term efficacy and safety of larotrectinib in patients with advanced TRK fusion-positive thyroid carcinoma. ATA, 2021, Oral presentation 15.



      ORR: 71%

      (2 CR, 18 PR, 4 SD)

      Waguespack S, Drilon A, Lin J, Brose M, McDermott R, Almubarak M, et al. Long-term efficacy and safety of larotrectinib in patients with advanced TRK fusion-positive thyroid carcinoma. ATA, 2021, Oral presentation 15.

      24-month DoR: 81%

      Waguespack S, Drilon A, Lin J, Brose M, McDermott R, Almubarak M, et al. Long-term efficacy and safety of larotrectinib in patients with advanced TRK fusion-positive thyroid carcinoma. ATA, 2021, Oral presentation 15.

      24-month OS: 76%

      Waguespack S, Drilon A, Lin J, Brose M, McDermott R, Almubarak M, et al. Long-term efficacy and safety of larotrectinib in patients with advanced TRK fusion-positive thyroid carcinoma. ATA, 2021, Oral presentation 15.

      24-month PFS: 69%

      Waguespack S, Drilon A, Lin J, Brose M, McDermott R, Almubarak M, et al. Long-term efficacy and safety of larotrectinib in patients with advanced TRK fusion-positive thyroid carcinoma. ATA, 2021, Oral presentation 15.

      Entrectinib

      Roche Registration GmBH. Rozlytrek Summary of Product Characteristics, https://www.ema.europa.eu/en/documents/product-information/rozlytrek-epar-product-information_en.pdf; 2020 [accessed 25 November 2021].

      TC: 13 (subtype not specified)
      • Bazhenova L.
      • Liu S.
      • Lin J.
      • Lu S.
      • Drilon A.
      • Chawla S.
      • et al.
      Efficacy and safety of entrectinib in patients with locally advanced/metastatic NTRK fusion-positive solid tumours.
      ORR: 53.8%
      • Bazhenova L.
      • Liu S.
      • Lin J.
      • Lu S.
      • Drilon A.
      • Chawla S.
      • et al.
      Efficacy and safety of entrectinib in patients with locally advanced/metastatic NTRK fusion-positive solid tumours.
      13.2
      • Bazhenova L.
      • Liu S.
      • Lin J.
      • Lu S.
      • Drilon A.
      • Chawla S.
      • et al.
      Efficacy and safety of entrectinib in patients with locally advanced/metastatic NTRK fusion-positive solid tumours.
      NRNR
      RET gene fusion or mutationPralsetinib
      FDA approved but not EMA approved.

      Roche. Gavreto Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2020/214701s000lbl.pdf; 2020 [accessed 25 November 2021].

      PTC: 9ORR: 89% (89% PR)NE

      NRNR
      Selpercatinib

      Eli Lilly Nederaland B.V. Retsevmo Summary of Product Characterisitcs, https://www.ema.europa.eu/en/documents/product-information/retsevmo-epar-product-information_en.pdf; 2021 [accessed 25 November 2021].

      RET fusion-positive thyroid cancer, previously treated: 19(PTC: 13; PDTC: 3; ATC: 2; Hürthle cell: 1)

      Eli Lilly Nederaland B.V. Retsevmo Summary of Product Characterisitcs, https://www.ema.europa.eu/en/documents/product-information/retsevmo-epar-product-information_en.pdf; 2021 [accessed 25 November 2021].



      Treatment naïve: 8

      (subtype not specified)

      Eli Lilly and Company. Retevmo Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2020/213246s000lbl.pdf; 2020 [accessed 25 November 2021].



      RET mutation, previously treated ORR: 78.9% (2 CR, 13 PR)

      Eli Lilly Nederaland B.V. Retsevmo Summary of Product Characterisitcs, https://www.ema.europa.eu/en/documents/product-information/retsevmo-epar-product-information_en.pdf; 2021 [accessed 25 November 2021].



      Treatment naïve ORR: 100%

      (1 CR, 7 PR)

      Eli Lilly and Company. Retevmo Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2020/213246s000lbl.pdf; 2020 [accessed 25 November 2021].



      RET mutation, previously treated: 18.4

      Eli Lilly Nederaland B.V. Retsevmo Summary of Product Characterisitcs, https://www.ema.europa.eu/en/documents/product-information/retsevmo-epar-product-information_en.pdf; 2021 [accessed 25 November 2021].





      Treatment naïve: NR

      RET mutation, previously treated: 27.4
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      • Sherman E.
      • Robinson B.
      • Solomon B.
      • Kang H.
      • Lorch J.
      • et al.
      Efficacy of selpercatinib in RET-altered thyroid cancers.




      Treatment naïve ORR: NR

      NR
      ALK rearrangementCrizotinib
      • Godbert Y.
      • Henriques de Figueiredo B.
      • Bonichon F.
      • Chibon F.
      • Hostein I.
      • Pérot G.
      • et al.
      Remarkable response to crizotinib in woman with anaplastic lymphoma kinase-rearranged anaplastic thyroid carcinoma.
      ATC: 1PR (90% across all pulmonary lesions)NENENE
      BRAF V600E mutationDabrafenib

      Novartis Pharmaceuticals Corporation. Dabrafenib Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2018/202806s010lbl.pdf; 2018 [accessed 25 November 2021].

      DTC: 14
      • Falchook G.S.
      • Millward M.
      • Hong D.
      • Naing A.
      • Piha-Paul S.
      • Waguespack S.G.
      • et al.
      BRAF inhibitor dabrafenib in patients with metastatic BRAF-mutant thyroid cancer.
      ORR: 29%(4 PR)

      • Falchook G.S.
      • Millward M.
      • Hong D.
      • Naing A.
      • Piha-Paul S.
      • Waguespack S.G.
      • et al.
      BRAF inhibitor dabrafenib in patients with metastatic BRAF-mutant thyroid cancer.
      NR
      • Falchook G.S.
      • Millward M.
      • Hong D.
      • Naing A.
      • Piha-Paul S.
      • Waguespack S.G.
      • et al.
      BRAF inhibitor dabrafenib in patients with metastatic BRAF-mutant thyroid cancer.
      NR
      • Falchook G.S.
      • Millward M.
      • Hong D.
      • Naing A.
      • Piha-Paul S.
      • Waguespack S.G.
      • et al.
      BRAF inhibitor dabrafenib in patients with metastatic BRAF-mutant thyroid cancer.
      11.3
      • Falchook G.S.
      • Millward M.
      • Hong D.
      • Naing A.
      • Piha-Paul S.
      • Waguespack S.G.
      • et al.
      BRAF inhibitor dabrafenib in patients with metastatic BRAF-mutant thyroid cancer.
      Dabrafenib

      Novartis Pharmaceuticals Corporation. Dabrafenib Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2018/202806s010lbl.pdf; 2018 [accessed 25 November 2021].

      PTC: 26 (22 assessable)
      • Shah M.
      • Wei L.
      • Wirth L.
      • Daniels G.
      • De Souza J.
      • Timmers C.
      • et al.
      Results of randomized phase II trial of dabrafenib versus dabrafenib plus trametinib in BRAF-mutated papillary thyroid carcinoma.
      ORR: 50%
      • Shah M.
      • Wei L.
      • Wirth L.
      • Daniels G.
      • De Souza J.
      • Timmers C.
      • et al.
      Results of randomized phase II trial of dabrafenib versus dabrafenib plus trametinib in BRAF-mutated papillary thyroid carcinoma.
      15.6
      • Shah M.
      • Wei L.
      • Wirth L.
      • Daniels G.
      • De Souza J.
      • Timmers C.
      • et al.
      Results of randomized phase II trial of dabrafenib versus dabrafenib plus trametinib in BRAF-mutated papillary thyroid carcinoma.
      NR
      • Shah M.
      • Wei L.
      • Wirth L.
      • Daniels G.
      • De Souza J.
      • Timmers C.
      • et al.
      Results of randomized phase II trial of dabrafenib versus dabrafenib plus trametinib in BRAF-mutated papillary thyroid carcinoma.
      11.4
      • Shah M.
      • Wei L.
      • Wirth L.
      • Daniels G.
      • De Souza J.
      • Timmers C.
      • et al.
      Results of randomized phase II trial of dabrafenib versus dabrafenib plus trametinib in BRAF-mutated papillary thyroid carcinoma.
      Dabrafenib and RAI
      • Rothenberg S.M.
      • McFadden D.G.
      • Palmer E.L.
      • Daniels G.H.
      • Wirth L.J.
      Redifferentiation of iodine-refractory BRAF V600E-mutant metastatic papillary thyroid cancer with dabrafenib.
      PTC: 10PR: 2

      SD: 4
      NRNRNR
      Dabrafenib and trametinib (ATC only)
      FDA approved but not EMA approved.

      Novartis Pharmaceuticals Corporation. Dabrafenib Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2018/202806s010lbl.pdf; 2018 [accessed 25 November 2021].

      ATC: 16
      • Subbiah V.
      • Kreitman R.J.
      • Wainberg Z.A.
      • Cho J.Y.
      • Schellens J.H.M.
      • Soria J.C.
      • et al.
      Dabrafenib and trametinib treatment in patients with locally advanced or metastatic BRAF V600-mutant anaplastic thyroid cancer.
      ORR: 69%(1 CR, 10 PR)

      • Subbiah V.
      • Kreitman R.J.
      • Wainberg Z.A.
      • Cho J.Y.
      • Schellens J.H.M.
      • Soria J.C.
      • et al.
      Dabrafenib and trametinib treatment in patients with locally advanced or metastatic BRAF V600-mutant anaplastic thyroid cancer.
      NE;



      12-month DoR: 90%
      • Subbiah V.
      • Kreitman R.J.
      • Wainberg Z.A.
      • Cho J.Y.
      • Schellens J.H.M.
      • Soria J.C.
      • et al.
      Dabrafenib and trametinib treatment in patients with locally advanced or metastatic BRAF V600-mutant anaplastic thyroid cancer.
      NE;



      12-month OS: 80%
      • Subbiah V.
      • Kreitman R.J.
      • Wainberg Z.A.
      • Cho J.Y.
      • Schellens J.H.M.
      • Soria J.C.
      • et al.
      Dabrafenib and trametinib treatment in patients with locally advanced or metastatic BRAF V600-mutant anaplastic thyroid cancer.
      NE;



      12-month PFS: 79%
      • Subbiah V.
      • Kreitman R.J.
      • Wainberg Z.A.
      • Cho J.Y.
      • Schellens J.H.M.
      • Soria J.C.
      • et al.
      Dabrafenib and trametinib treatment in patients with locally advanced or metastatic BRAF V600-mutant anaplastic thyroid cancer.
      Dabrafenib and trametinib
      FDA approved but not EMA approved.

      Novartis Pharmaceuticals Corporation. Dabrafenib Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2018/202806s010lbl.pdf; 2018 [accessed 25 November 2021].

      PTC: 27 (24 assessable)
      • Shah M.
      • Wei L.
      • Wirth L.
      • Daniels G.
      • De Souza J.
      • Timmers C.
      • et al.
      Results of randomized phase II trial of dabrafenib versus dabrafenib plus trametinib in BRAF-mutated papillary thyroid carcinoma.
      ORR: 54%
      • Shah M.
      • Wei L.
      • Wirth L.
      • Daniels G.
      • De Souza J.
      • Timmers C.
      • et al.
      Results of randomized phase II trial of dabrafenib versus dabrafenib plus trametinib in BRAF-mutated papillary thyroid carcinoma.
      13.3
      • Shah M.
      • Wei L.
      • Wirth L.
      • Daniels G.
      • De Souza J.
      • Timmers C.
      • et al.
      Results of randomized phase II trial of dabrafenib versus dabrafenib plus trametinib in BRAF-mutated papillary thyroid carcinoma.
      NR
      • Shah M.
      • Wei L.
      • Wirth L.
      • Daniels G.
      • De Souza J.
      • Timmers C.
      • et al.
      Results of randomized phase II trial of dabrafenib versus dabrafenib plus trametinib in BRAF-mutated papillary thyroid carcinoma.
      15.1
      • Shah M.
      • Wei L.
      • Wirth L.
      • Daniels G.
      • De Souza J.
      • Timmers C.
      • et al.
      Results of randomized phase II trial of dabrafenib versus dabrafenib plus trametinib in BRAF-mutated papillary thyroid carcinoma.
      Vemurafenib
      • Brose M.S.
      • Cabanillas M.E.
      • Cohen E.E.
      • Wirth L.J.
      • Riehl T.
      • Yue H.
      • et al.
      Vemurafenib in patients with BRAF(V600E)-positive metastatic or unresectable papillary thyroid cancer refractory to radioactive iodine: a non-randomised, multicentre, open-label, phase 2 trial.
      PTC: 51



      Treatment naïve: 26

      Previous VEGFR: 25

      Treatment naïve ORR: 39% (10 PR)Previous VEGFR ORR: 27%

      (6 PR)
      Treatment naïve: 16.5

      Previous VEGFR: 7.4
      Treatment naïve: NR

      Previous VEGFR: 14.4
      Treatment naïve: 18.2

      Previous VEGFR: 8.9
      Vemurafenib and RAI
      • Dunn L.A.
      • Sherman E.J.
      • Baxi S.S.
      • Tchekmedyian V.
      • Grewal R.K.
      • Larson S.M.
      • et al.
      Vemurafenib redifferentiation of BRAF mutant, RAI-refractory thyroid cancers.
      DTC: 12PR: 2

      SD: 2
      NRNRNR
      Selumetinib
      No clinical benefit with selumetinib monotherapy prior to RAI observed in the ASTRA trial [106].
      and RAI
      • Ho A.L.
      • Grewal R.K.
      • Leboeuf R.
      • Sherman E.J.
      • Pfister D.G.
      • Deandreis D.
      • et al.
      Selumetinib-enhanced radioiodine uptake in advanced thyroid cancer.
      DTC: 20

      (BRAFV600E: 9;

      NRAS: 5;

      RET/PTC: 3;

      wild-type: 3)
      Confirmed PR: 5 (4 NRAS; 1 BRAF)

      SD: 3

      (1 RET/PTC; 1 NRAS; 1 wild-type)
      NRNRNR
      Dabrafenib, trametinib, and RAI
      • Leboulleux S.
      • Cao C.
      • Zerdoud S.
      • Attard M.
      • Bournaud C.
      • Benisvy D.
      • et al.
      MERAIODE: A redifferentiation phase II trial with trametinib and dabrafenib followed by radioactive iodine administration for metastatic radioactive iodine refractory differentiated thyroid cancer patients with a BRAFV600E mutation (NCT03244956).
      DTC: 21PR: 38%

      SD: 52%

      NRNRNR
      TSC2 mutation/

      MTOR mutation
      Everolimus
      • Wagle N.
      • Grabiner B.C.
      • Van Allen E.M.
      • Amin-Mansour A.
      • Taylor-Weiner A.
      • Rosenberg M.
      • et al.
      Response and acquired resistance to everolimus in anaplastic thyroid cancer.
      ATC: 1Near CR18NRNR
      ALK, anaplastic lymphoma kinase; ATC, anaplastic thyroid cancer; BRAF, B-Raf proto-oncogene; CR, complete response; DoR, duration of response; DTC, differentiated thyroid cancer; EMA, European Medicines Agency; FDA, US Food and Drug Administration; FTC, follicular thyroid cancer; MTOR, mechanistic target of rapamycin; NE, not estimable; NR, not reported; NRAS, neuroblastoma RAS viral oncogene homolog; NTRK, neurotrophic tyrosine receptor kinase; ORR, objective response rate; OS, overall survival; PDTC, poorly differentiated thyroid cancer; PFS, progression-free survival; PR, partial response; PTC, papillary thyroid cancer; RAI, radioactive iodine; RET, rearranged during transfection; SD, stable disease; TC, thyroid cancer; TSC2, tuberous sclerosis complex 2; VEGFR, vascular endothelial growth factor receptor.
      *Investigator assessed unless otherwise specified.
      FDA approved but not EMA approved.
      No clinical benefit with selumetinib monotherapy prior to RAI observed in the ASTRA trial

      U.S. National Library of Medicine, ClinicalTrials.gov. Comparing complete remission after treatment with selumetinib/placebo in patient with differentiated thyroid cancer (ASTRA), https://clinicaltrials.gov/ct2/show/NCT01843062; 2013 [accessed 25 November 2021].

      .

      Targeted treatment landscape for advanced disease

      BRAF

      BRAF is the most common driver mutation in thyroid cancer, with a transversion mutation leading to the substitution of valine by glutamic acid at amino acid position 600 (V600E) the most frequent alteration (reported in ∼ 45% of adult patients with DTC, and ∼ 58% of patients with PTC) [
      • Murugan A.K.
      • Qasem E.
      • Al-Hindi H.
      • Shi Y.
      • Alzahrani A.S.
      Classical V600E and other non-hotspot BRAF mutations in adult differentiated thyroid cancer.
      ,

      Cancer Genome Atlas Research Network. Integrated genomic characterization of papillary thyroid carcinoma Cell 2014;159:676–90.

      ]. BRAF gene fusions with diverse gene partners have been identified in PTC [

      Cancer Genome Atlas Research Network. Integrated genomic characterization of papillary thyroid carcinoma Cell 2014;159:676–90.

      ]. The MKIs lenvatinib and sorafenib both inhibit oncogenic BRAF among other cellular targets [
      • Crispo F.
      • Notarangelo T.
      • Pietrafesa M.
      • Lettini G.
      • Storto G.
      • Sgambato A.
      • et al.
      BRAF inhibitors in thyroid cancer: clinical impact, mechanisms of resistance and future perspectives.
      ]. Treatments under investigation include those that specifically inhibit BRAF (e.g. dabrafenib and vemurafenib) or mitogen-activated protein/extracellular signal-regulated kinase kinase (MEK), a protein downstream of BRAF in the mitogen-activated protein kinase (MAPK) pathway (e.g. selumetinib and trametinib).
      Dabrafenib in combination with the MEK inhibitor trametinib is approved by the US Food and Drug Administration (FDA) for patients with locally advanced or metastatic ATC with BRAF V600E mutations and no satisfactory locoregional treatment options [

      Novartis Pharmaceuticals Corporation. Dabrafenib Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2018/202806s010lbl.pdf; 2018 [accessed 25 November 2021].

      ]. The European Society for Medical Oncology (ESMO) recommends dabrafenib in combination with trametinib for patients with BRAF V600E-positive ATC (if available); however, this treatment is not yet approved by the European Medicines Agency (EMA) for thyroid cancer [
      • Filetti S.
      • Durante C.
      • Hartl D.
      • Leboulleux S.
      • Locati L.D.
      • Newbold K.
      • et al.
      Thyroid cancer: ESMO clinical practice guidelines for diagnosis, treatment and follow-up.
      ]. In a small series of patients with ATC, an overall response rate (ORR) of 69% was achieved [
      • Subbiah V.
      • Kreitman R.J.
      • Wainberg Z.A.
      • Cho J.Y.
      • Schellens J.H.M.
      • Soria J.C.
      • et al.
      Dabrafenib and trametinib treatment in patients with locally advanced or metastatic BRAF V600-mutant anaplastic thyroid cancer.
      ]; in a different study, the ORR in PTC was 54% with dabrafenib plus trametinib [
      • Shah M.
      • Wei L.
      • Wirth L.
      • Daniels G.
      • De Souza J.
      • Timmers C.
      • et al.
      Results of randomized phase II trial of dabrafenib versus dabrafenib plus trametinib in BRAF-mutated papillary thyroid carcinoma.
      ] (Table 1). With regards to safety, in the study in patients with ATC, 69% experienced adverse events (AEs) suspected to be related to treatment [
      • Subbiah V.
      • Kreitman R.J.
      • Wainberg Z.A.
      • Cho J.Y.
      • Schellens J.H.M.
      • Soria J.C.
      • et al.
      Dabrafenib and trametinib treatment in patients with locally advanced or metastatic BRAF V600-mutant anaplastic thyroid cancer.
      ], and in PTC, treatment-related AEs were similar to those previously reported in phase III trials in melanoma [
      • Shah M.
      • Wei L.
      • Wirth L.
      • Daniels G.
      • De Souza J.
      • Timmers C.
      • et al.
      Results of randomized phase II trial of dabrafenib versus dabrafenib plus trametinib in BRAF-mutated papillary thyroid carcinoma.
      ].
      Vemurafenib is approved by the EMA for the treatment of patients with unresectable or metastatic melanoma with BRAF V600E mutations [

      Genentech. Zelboraf Summary of Product Characteristics, https://www.ema.europa.eu/en/documents/product-information/zelboraf-epar-product-information_en.pdf; 2016 [accessed 25 November 2021].

      ]. It is currently not approved for use in patients with thyroid cancer. In a phase II study of patients with metastatic PTC refractory to RAI and positive for the BRAF V600E mutation, vemurafenib showed anti-tumour activity in patients who had never received an MKI targeting vascular endothelial growth factor receptor (VEGFR) as well as in patients who had been previously treated with a VEGFR MKI (Table 1). Serious AEs were experienced by nearly two-thirds of patients receiving vemurafenib [
      • Brose M.S.
      • Cabanillas M.E.
      • Cohen E.E.
      • Wirth L.J.
      • Riehl T.
      • Yue H.
      • et al.
      Vemurafenib in patients with BRAF(V600E)-positive metastatic or unresectable papillary thyroid cancer refractory to radioactive iodine: a non-randomised, multicentre, open-label, phase 2 trial.
      ].
      BRAF and MEK inhibitors are currently under investigation in combination with RAI, to understand whether these drugs can act to restore RAI uptake (so-called ‘RAI redifferentiation’) in RAI-R tumours (Table 1) [
      • Dunn L.A.
      • Sherman E.J.
      • Baxi S.S.
      • Tchekmedyian V.
      • Grewal R.K.
      • Larson S.M.
      • et al.
      Vemurafenib redifferentiation of BRAF mutant, RAI-refractory thyroid cancers.
      ,
      • Ho A.L.
      • Grewal R.K.
      • Leboeuf R.
      • Sherman E.J.
      • Pfister D.G.
      • Deandreis D.
      • et al.
      Selumetinib-enhanced radioiodine uptake in advanced thyroid cancer.
      ,
      • Rothenberg S.M.
      • McFadden D.G.
      • Palmer E.L.
      • Daniels G.H.
      • Wirth L.J.
      Redifferentiation of iodine-refractory BRAF V600E-mutant metastatic papillary thyroid cancer with dabrafenib.
      ,
      • Leboulleux S.
      • Cao C.
      • Zerdoud S.
      • Attard M.
      • Bournaud C.
      • Benisvy D.
      • et al.
      MERAIODE: A redifferentiation phase II trial with trametinib and dabrafenib followed by radioactive iodine administration for metastatic radioactive iodine refractory differentiated thyroid cancer patients with a BRAFV600E mutation (NCT03244956).
      ]. BRAF inhibitors are also under investigation in combination with immuno-oncology treatments [

      U.S. National Library of Medicine, ClinicalTrials.gov. Pembrolizumab, dabrafenib, and trametinib before surgery for the treatment of BRAF-mutated anaplastic thyroid cancer, https://clinicaltrials.gov/ct2/show/NCT04675710; 2020 [accessed 25 November 2021].

      ,

      U.S. National Library of Medicine, ClinicalTrials.gov. Study of cemiplimab combined with dabrafenib and trametinib in people with anaplastic thyroid cancer, https://clinicaltrials.gov/ct2/show/NCT04238624; 2020 [accessed 25 November 2021].

      ] (Table 2) and phosphatidylinositol 3-kinase (PI3K) inhibitors [

      U.S. National Library of Medicine, ClinicalTrials.gov. Vemurafenib plus copanlisib in radioiodine-refractory (RAIR) thyroid cancers, https://clinicaltrials.gov/ct2/show/NCT04462471; 2020 [accessed 25 November 2021].

      ].
      Table 2Planned and ongoing clinical trials of immuno-oncology treatments for thyroid cancer.
      Study numberTreatmentsThyroid cancer typePhaseEstimated enrolment (n)StatusEstimated study completion date
      NCT03181100Atezolizumab with chemotherapy*ATC/PDTCII50RecruitingJuly 2023
      NCT03914300Cabozantinib, nivolumab, and ipilimumabAdvanced DTCII24Suspended (scheduled interim monitoring)January 2023
      NCT04061980Encorafenib and binimetinib and/or nivolumabBRAF V600E-positive DTCII40RecruitingAugust 2024
      NCT04171622Lenvatinib and pembrolizumabATCII25Not yet recruitingAugust 2022
      NCT04675710Pembrolizumab, dabrafenib, and trametinibATC, PDTCII30RecruitingJune 2024
      NCT02973997Lenvatinib and pembrolizumabDTC, PDTCII60Active, not recruitingSeptember 2022
      NCT04731740Pembrolizumab and lenvatinib or chemotherapyPDTC, ATCII36Suspended (financial problems)December 2023
      NCT03246958Nivolumab and ipilimumab
      Nivolumab or ipilimumabalone for 2 weeks, followed by ipilimumabor nivolumab (respectively) 2 weeks after initial treatment.
      DTC, MTC, ATCII53Active, not recruitingMarch 2025
      NCT04524884Toripalimab and surufatinibMTC, DTCII10Not yet recruitingSeptember 2022
      NCT04521348Camrelizumab and famitinibMTC, ATC, DTCII115RecruitingJune 2023
      NCT03360890Pembrolizumab and chemotherapyATCII46RecruitingSeptember 2022
      NCT04238624Cemiplimab with dabrafenib and trametinibATCII15RecruitingJune 2022
      NCT03753919Durvalumab with tremelimumabATC, DTC, MTCII46RecruitingJuly 2022
      NCT04400474Cabozantinib with atezolizumabATCII144RecruitingMarch 2024
      ATC, anaplastic thyroid cancer; BRAF, B-Raf proto-oncogene; DTC, differentiated thyroid cancer; MTC, medullary thyroid cancer; PDTC, poorly differentiated thyroid cancer.
      *Four experimental treatment groups: Cohort I (vemurafenib, cobimetinib, atezolizumab); Cohort II (atezolizumab, cobimetinib); Cohort III (atezolizumab, bevacizumab); Cohort IV (nab-paclitaxel, atezolizumab, paclitaxel).
      Nivolumab or ipilimumab alone for 2 weeks, followed by ipilimumab or nivolumab (respectively) 2 weeks after initial treatment.

      RET

      More than 20 fusions between the 3′ portion of RET (containing the tyrosine kinase domain) and the 5′ portion of partner genes have been described in PTC [
      • Santoro M.
      • Moccia M.
      • Federico G.
      • Carlomagno F.
      RET gene fusions in malignancies of the thyroid and other tissues.
      ]. Although only found in < 10% of PTC/PDTC cases [
      • Landa I.
      • Ibrahimpasic T.
      • Boucai L.
      • Sinha R.
      • Knauf J.A.
      • Shah R.H.
      • et al.
      Genomic and transcriptomic hallmarks of poorly differentiated and anaplastic thyroid cancers.
      ,
      • Pozdeyev N.
      • Gay L.M.
      • Sokol E.S.
      • Hartmaier R.
      • Deaver K.E.
      • Davis S.
      • et al.
      Genetic analysis of 779 advanced differentiated and anaplastic thyroid cancers.
      ], RET represents an actionable target in advanced thyroid cancer. Both mutations and rearrangements of RET lead to constitutive RET protein activation and aberrant stimulation of both the MAPK and PI3K pathways [
      • Tirrò E.
      • Martorana F.
      • Romano C.
      • Vitale S.R.
      • Motta G.
      • Di Gregorio S.
      • et al.
      Molecular alterations in thyroid cancer: from bench to clinical practice.
      ]. Both lenvatinib and sorafenib inhibit RET; however, as MKIs they also have activity against other tyrosine kinases (such as VEGFR), which has implications for non-RET-mediated effects and off-target side effects [
      • Cabanillas M.E.
      • Ryder M.
      • Jimenez C.
      Targeted therapy for advanced thyroid cancer: kinase inhibitors and beyond.
      ,

      Bayer HealthCare Pharmaceuticals I. Nexavar (sorafenib) PI; 2018.

      ,

      Eisai GmbH. Lenvima summary of product characteristics, https://www.ema.europa.eu/en/documents/product-information/lenvima-epar-product-information_en.pdf; 2020 [accessed 25 November 2021].

      ,
      • Seoane J.
      • Capdevila J.
      The right compound for the right target: tackling RET.
      ].
      Treatments that selectively target RET may mitigate these broad target issues while maintaining anti-tumour activity [
      • Seoane J.
      • Capdevila J.
      The right compound for the right target: tackling RET.
      ]. Selpercatinib is a selective RET kinase inhibitor approved by the EMA for adult patients with advanced RET fusion-positive thyroid cancer who require systemic therapy following prior treatment with sorafenib and/or lenvatinib. It is also approved in patients aged 12 years and older with advanced RET-mutant MTC who require systemic therapy following prior treatment with cabozantinib and/or vandetanib [

      Eli Lilly Nederaland B.V. Retsevmo Summary of Product Characterisitcs, https://www.ema.europa.eu/en/documents/product-information/retsevmo-epar-product-information_en.pdf; 2021 [accessed 25 November 2021].

      ]. This approval is based on the phase I/II LIBRETTO-001 trial, in which patients with MTC and PTC with RET mutations or fusions were enrolled [
      • Wirth L.J.
      • Sherman E.
      • Robinson B.
      • Solomon B.
      • Kang H.
      • Lorch J.
      • et al.
      Efficacy of selpercatinib in RET-altered thyroid cancers.
      ]. Selpercatinib showed efficacy among patients who were treatment naïve, and those who had received previous treatment (Table 1). Most (94%) patients experienced at least one treatment-related AE, with 28% of patients experiencing a grade 3 treatment-related AE, and 2% experiencing a grade 4 treatment-related AE [
      • Wirth L.J.
      • Sherman E.
      • Robinson B.
      • Solomon B.
      • Kang H.
      • Lorch J.
      • et al.
      Efficacy of selpercatinib in RET-altered thyroid cancers.
      ].
      Pralsetinib is another selective RET inhibitor approved by the FDA for patients ≥ 12 years of age with advanced or metastatic RET-mutant MTC who require systemic therapy, or with advanced or metastatic RET fusion-positive thyroid cancer who require systemic therapy and whose disease is RAI-R [

      Roche. Gavreto Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2020/214701s000lbl.pdf; 2020 [accessed 25 November 2021].

      ]. The efficacy and safety of pralsetinib was evaluated in the ARROW trial: ORR was 60% for patients with MTC (complete response [CR], 1.8%; partial response [PR], 58%) and 89% (PR, 89%) for patients with PTC (Table 1). Overall, 15% of patients experienced a serious treatment-related AE [

      Roche. Gavreto Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2020/214701s000lbl.pdf; 2020 [accessed 25 November 2021].

      ,
      • Subbiah V.
      • Hu M.I.
      • Wirth L.J.
      • Schuler M.
      • Mansfield A.S.
      • Curigliano G.
      • et al.
      Pralsetinib for patients with advanced or metastatic RET-altered thyroid cancer (ARROW): a multi-cohort, open-label, registrational, phase 1/2 study.
      ].
      A third selective RET kinase inhibitor, BOS172738, has shown promising results in a phase I dose-escalation and dose-expansion trial, exhibiting a favourable safety profile and an ORR of 44% (n = 7/16) in patients with MTC [
      • Schoffski P.
      • Cho B.
      • Italiano A.
      • Loong H.
      • Massard C.
      • Rodriguez L.
      • et al.
      BOS172738, a highly potent and selective RET inhibitor, for the treatment of RET-altered tumors including RET-fusion+ NSCLC and RET-mutant MTC: Phase 1 study results.
      ].

      NTRK gene fusions

      Gene fusions involving NTRK1, -2, or -3 with various fusion partners represent an important, actionable biomarker in thyroid cancer [
      • Tirrò E.
      • Martorana F.
      • Romano C.
      • Vitale S.R.
      • Motta G.
      • Di Gregorio S.
      • et al.
      Molecular alterations in thyroid cancer: from bench to clinical practice.
      ]. As a result of in-frame gene rearrangements, the tyrosine kinase domain of the NTRK gene comes under control of the active promoter of the partner gene, leading to enhanced expression and constitutive activation of the fusion gene. NTRK1–3 gene fusions have been identified among adult and paediatric patients across multiple tumour types, and are considered primary oncogenic drivers [
      • Cocco E.
      • Scaltriti M.
      • Drilon A.
      NTRK fusion-positive cancers and TRK inhibitor therapy.
      ]. While particularly prevalent in several rare tumour types (e.g. secretory carcinoma of the salivary gland and infantile fibrosarcoma), NTRK gene fusions have also been identified in more common tumour types, including thyroid cancer [
      • Gatalica Z.
      • Xiu J.
      • Swensen J.
      • Vranic S.
      Molecular characterization of cancers with NTRK gene fusions.
      ]. The frequency of NTRK gene fusions in thyroid cancer is reported to be 2.5–25.9% [
      • Prasad M.L.
      • Vyas M.
      • Horne M.J.
      • Virk R.K.
      • Morotti R.
      • Liu Z.
      • et al.
      NTRK fusion oncogenes in pediatric papillary thyroid carcinoma in northeast United States.
      ,
      • Stransky N.
      • Cerami E.
      • Schalm S.
      • Kim J.L.
      • Lengauer C.
      The landscape of kinase fusions in cancer.
      ,

      Roche Registration GmBH. Rozlytrek Summary of Product Characteristics, https://www.ema.europa.eu/en/documents/product-information/rozlytrek-epar-product-information_en.pdf; 2020 [accessed 25 November 2021].

      ].
      Larotrectinib is a selective and specific inhibitor of tropomyosin receptor kinase A (TRKA), TRKB, and TRKC approved for use in Europe and the USA [

      Bayer AG. VITRAKVI Summary of Product Characteristics, https://www.ema.europa.eu/en/documents/product-information/vitrakvi-epar-product-information_en.pdf; 2022 [accessed 1 March 2022].

      ,

      Bayer HealthCare Pharmaceuticals Inc. VITRAKVI Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2018/211710s000lbl.pdf; 2018 [accessed 25 November 2021].

      ]. Larotrectinib is indicated for treatment of adult and paediatric patients with solid tumours that display an NTRK gene fusion who have locally advanced or metastatic disease, or where surgical resection is likely to result in severe morbidity, and who have no satisfactory treatment options [

      Bayer AG. VITRAKVI Summary of Product Characteristics, https://www.ema.europa.eu/en/documents/product-information/vitrakvi-epar-product-information_en.pdf; 2022 [accessed 1 March 2022].

      ]. Recently, data from 29 patients with TRK fusion thyroid cancer (PTC n = 20; FTC n = 2; ATC n = 7) treated with larotrectinib (data cut-off July 2020) were published. Of the 28 patients with evaluable disease, the ORR was 71% (95% CI 51–87), with 2 CRs, 18 PRs, and 4 cases of stable disease [

      Waguespack S, Drilon A, Lin J, Brose M, McDermott R, Almubarak M, et al. Long-term efficacy and safety of larotrectinib in patients with advanced TRK fusion-positive thyroid carcinoma. ATA, 2021, Oral presentation 15.

      ]. The response rate was 86% in patients with DTC and 29% in those with ATC. Larotrectinib had a favourable safety profile, with 7% of patients experiencing a grade 3 treatment-related AE [

      Waguespack S, Drilon A, Lin J, Brose M, McDermott R, Almubarak M, et al. Long-term efficacy and safety of larotrectinib in patients with advanced TRK fusion-positive thyroid carcinoma. ATA, 2021, Oral presentation 15.

      ].
      Entrectinib is another inhibitor of TRKA, TRKB, and TRKC that also inhibits ALK (anaplastic lymphoma kinase), ROS1 (c-ros oncogene 1) tyrosine kinase, JAK2 (janus kinase 2), and TNK2 (tyrosine kinase non receptor 2) [

      Roche Registration GmBH. Rozlytrek Summary of Product Characteristics, https://www.ema.europa.eu/en/documents/product-information/rozlytrek-epar-product-information_en.pdf; 2020 [accessed 25 November 2021].

      ,

      Genentech USA, Inc. ROZLYTREK Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2019/212725s000lbl.pdf, 2019 [accessed 25 November 2021].

      ]. Entrectinib is an MKI first approved by the FDA and later by the EMA [

      Roche Registration GmBH. Rozlytrek Summary of Product Characteristics, https://www.ema.europa.eu/en/documents/product-information/rozlytrek-epar-product-information_en.pdf; 2020 [accessed 25 November 2021].

      ,

      Genentech USA, Inc. ROZLYTREK Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2019/212725s000lbl.pdf, 2019 [accessed 25 November 2021].

      ]. It is indicated by the EMA for the treatment of patients aged 12 years and older with TRK fusion solid tumours who have a disease that is locally advanced, metastatic, or where surgical resection is likely to result in severe morbidity, who have no other satisfactory treatment options, and who have not received a prior TRK inhibitor [

      Roche Registration GmBH. Rozlytrek Summary of Product Characteristics, https://www.ema.europa.eu/en/documents/product-information/rozlytrek-epar-product-information_en.pdf; 2020 [accessed 25 November 2021].

      ]. Detailed thyroid-specific data are not yet available for entrectinib. An integrated analysis of clinical trial data for patients aged ≥ 18 years with NTRK fusion-positive cancers treated with ≥ 1 dose of entrectinib included 14 different tumour types, 11% (n = 13) of patients had thyroid cancer. The ORR in patients with thyroid cancer was 54% (95% CI 25–81) and the duration of response was 13 months (95% CI 8 months–not estimable). In the overall safety population (N = 626), most treatment-related AEs were reversible and resolved via dose modifications or reductions. Rates of the most common treatment-related AEs were broadly similar between the overall safety population and the NTRK fusion-positive safety-evaluable population (n = 193) [
      • Bazhenova L.
      • Liu S.
      • Lin J.
      • Lu S.
      • Drilon A.
      • Chawla S.
      • et al.
      Efficacy and safety of entrectinib in patients with locally advanced/metastatic NTRK fusion-positive solid tumours.
      ].

      ALK and MTOR

      Mutations of ALK (L1198F and G1201E) have been identified in up to 11% of ATCs, but not in the generally indolent DTC subtypes, suggesting they may play a role in the aggressive phenotype of ATC [
      • Murugan A.K.
      • Xing M.
      Anaplastic thyroid cancers harbor novel oncogenic mutations of the ALK gene.
      ]. ALK gene fusions, however, have been identified in both PTC and PDTC [
      • Landa I.
      • Ibrahimpasic T.
      • Boucai L.
      • Sinha R.
      • Knauf J.A.
      • Shah R.H.
      • et al.
      Genomic and transcriptomic hallmarks of poorly differentiated and anaplastic thyroid cancers.
      ,

      Cancer Genome Atlas Research Network. Integrated genomic characterization of papillary thyroid carcinoma Cell 2014;159:676–90.

      ]. The most common ALK rearrangement involves the striatin (STRN) gene, but several additional gene fusion partners have been identified [
      • Landa I.
      • Ibrahimpasic T.
      • Boucai L.
      • Sinha R.
      • Knauf J.A.
      • Shah R.H.
      • et al.
      Genomic and transcriptomic hallmarks of poorly differentiated and anaplastic thyroid cancers.
      ,

      Cancer Genome Atlas Research Network. Integrated genomic characterization of papillary thyroid carcinoma Cell 2014;159:676–90.

      ,
      • Kelly L.M.
      • Barila G.
      • Liu P.
      • Evdokimova V.N.
      • Trivedi S.
      • Panebianco F.
      • et al.
      Identification of the transforming STRN-ALK fusion as a potential therapeutic target in the aggressive forms of thyroid cancer.
      ]. A case report of a patient with ATC harbouring an ALK fusion who had an exceptional response to crizotinib provides promising but limited evidence of efficacy in thyroid cancer (Table 1) [
      • Godbert Y.
      • Henriques de Figueiredo B.
      • Bonichon F.
      • Chibon F.
      • Hostein I.
      • Pérot G.
      • et al.
      Remarkable response to crizotinib in woman with anaplastic lymphoma kinase-rearranged anaplastic thyroid carcinoma.
      ].
      The mechanistic target of rapamycin (mTOR) pathway is constitutively activated in some thyroid cancers [
      • Tavares C.
      • Eloy C.
      • Melo M.
      • Gaspar da Rocha A.
      • Pestana A.
      • Batista R.
      • et al.
      mTOR pathway in papillary thyroid carcinoma: different contributions of mTORC1 and mTORC2 complexes for tumor behavior and SLC5A5 mRNA expression.
      ], and represents another potentially actionable target for advanced thyroid cancer. A case report of a patient with ATC with mutations in tuberous sclerosis complex 2 (TSC2) and MTOR illustrated potential activity of everolimus, an mTOR inhibitor, with a near CR achieved for 18 months [
      • Wagle N.
      • Grabiner B.C.
      • Van Allen E.M.
      • Amin-Mansour A.
      • Taylor-Weiner A.
      • Rosenberg M.
      • et al.
      Response and acquired resistance to everolimus in anaplastic thyroid cancer.
      ]. Further evidence for the use of ALK inhibitors and mTOR inhibitors in larger patient cohorts is needed.

      Techniques to detect molecular alterations

      To decide whether targeted treatments are appropriate for individual patients with advanced thyroid cancer, screening for specific gene mutations and rearrangements is essential [
      • Fugazzola L.
      • Elisei R.
      • Fuhrer D.
      • Jarzab B.
      • Leboulleux S.
      • Newbold K.
      • et al.
      2019 European Thyroid Association guidelines for the treatment and follow-up of advanced radioiodine-refractory thyroid cancer.
      ]. Several techniques are available to detect molecular alterations in clinical samples (summarized in Table 3). Factors that should be considered when selecting the optimal approach include turnaround time, required expertise, and cost [
      • Hsiao S.J.
      • Zehir A.
      • Sireci A.N.
      • Aisner D.L.
      Detection of tumor NTRK gene fusions to identify patients who may benefit from tyrosine kinase (TRK) inhibitor therapy.
      ]. Immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) have high sensitivity and specificity [
      • Marchiò C.
      • Scaltriti M.
      • Ladanyi M.
      • Iafrate A.
      • Bibeau F.
      • Dietel M.
      • et al.
      ESMO recommendations on the standard methods to detect NTRK fusions in daily practice and clinical research.
      ]. Reverse transcription polymerase chain reaction (RT-PCR) is also specific, but is less sensitive than IHC and FISH [
      • Caria P.
      • Dettori T.
      • Frau D.V.
      • Borghero A.
      • Cappai A.
      • Riola A.
      • et al.
      Assessing RET/PTC in thyroid nodule fine-needle aspirates: the FISH point of view.
      ,
      • Letovanec I.
      • Finn S.
      • Zygoura P.
      • Smyth P.
      • Soltermann A.
      • Bubendorf L.
      • et al.
      Evaluation of NGS and RT-PCR methods for ALK rearrangement in European NSCLC patients: results from the European Thoracic Oncology Platform Lungscape Project.
      ,
      • Zhao J.
      • Liu P.
      • Yu Y.
      • Zhi J.
      • Zheng X.
      • Yu J.
      • et al.
      Comparison of diagnostic methods for the detection of a BRAF mutation in papillary thyroid cancer.
      ]. Pan-TRK IHC is inexpensive with rapid turnaround times, but has limitations. This assay detects both wild-type and aberrant TRK protein expression, which means that it can be challenging to interpret results in tissue with physiological expression (e.g. central nervous system and smooth muscle). Furthermore, reduced sensitivity for NTRK3 fusions has been observed [
      • Solomon J.P.
      • Hechtman J.F.
      Detection of NTRK fusions: merits and limitations of current diagnostic platforms.
      ,
      • Zito Marino F.
      • Pagliuca F.
      • Ronchi A.
      • Cozzolino I.
      • Montella M.
      • Berretta M.
      • et al.
      NTRK fusions, from the diagnostic algorithm to innovative treatment in the era of precision medicine.
      ]. FISH is considered the gold standard for assessing chromosomal abnormalities, but for genetic alterations that involve non-canonical sites or novel genes, false negatives can be common [
      • Solomon J.P.
      • Hechtman J.F.
      Detection of NTRK fusions: merits and limitations of current diagnostic platforms.
      ]. Overall, RT-PCR has been successful in diagnosing fusion-driven malignancies (e.g. leukaemia); however, for genes with diverse fusion partners and breakpoints (as seen in NTRK gene fusions), RT-PCR may have limited utility [
      • Solomon J.P.
      • Hechtman J.F.
      Detection of NTRK fusions: merits and limitations of current diagnostic platforms.
      ].
      Table 3Clinical laboratory techniques used to identify molecular alterations in thyroid cancer.
      MethodTurnaround time*Required tissue*Detection of fusions and/or mutationsSensitivity/specificity
      Sensitivity and specificity of particular assays/molecular targets can vary according to tumour type.
      AdvantagesDisadvantages
      IHC
      • Hsiao S.J.
      • Zehir A.
      • Sireci A.N.
      • Aisner D.L.
      Detection of tumor NTRK gene fusions to identify patients who may benefit from tyrosine kinase (TRK) inhibitor therapy.
      ,
      • Zhao J.
      • Liu P.
      • Yu Y.
      • Zhi J.
      • Zheng X.
      • Yu J.
      • et al.
      Comparison of diagnostic methods for the detection of a BRAF mutation in papillary thyroid cancer.
      ,
      • Zito Marino F.
      • Pagliuca F.
      • Ronchi A.
      • Cozzolino I.
      • Montella M.
      • Berretta M.
      • et al.
      NTRK fusions, from the diagnostic algorithm to innovative treatment in the era of precision medicine.
      ,
      • Murphy D.A.
      • Ely H.A.
      • Shoemaker R.
      • Boomer A.
      • Culver B.P.
      • Hoskins I.
      • et al.
      Detecting gene rearrangements in patient populations through a 2-step diagnostic test comprised of rapid IHC enrichment followed by sensitive next-generation sequencing.
      ,
      • Belli C.
      • Penault-Llorca F.
      • Ladanyi M.
      • Normanno N.
      • Scoazec J.Y.
      • Lacroix L.
      • et al.
      ESMO recommendations on the standard methods to detect RET fusions and mutations in daily practice and clinical research.
      ,
      • Agarwal S.
      • Bychkov A.
      • Jung C.K.
      Emerging biomarkers in thyroid practice and research.
      ,
      • Teixidó C.
      • Karachaliou N.
      • Peg V.
      • Gimenez-Capitan A.
      • Rosell R.
      Concordance of IHC, FISH and RT-PCR for EML4-ALK rearrangements.
      ,
      • Wong D.
      • Yip S.
      • Sorensen P.H.
      Methods for identifying patients with tropomyosin receptor kinase (TRK) fusion cancer.
      1–2 days1–2 slidesFusions only
      • BRAF: 97–99%/86–98%
        RET: 55–65%/40–85%
        TRK: 95%/100%
        TRK1 sensitivity: 88–96%
        TRK2 sensitivity: 89–100%
        TRK3 sensitivity: 55–79%
        ALK: 67–100%/93–100%
      • Rapid and inexpensive process
        Established approach, widely available
        Mutation-specific antibodies enable rapid single-gene testing
      • Specificity for gene rearrangements may be insufficient to confirm a diagnostic result
        Not easily multiplexed for other biomarkers
        No reliable markers for RET fusions
        Does not discriminate between fusions and overexpression of wild-type TRK
      FISH
      • Hsiao S.J.
      • Zehir A.
      • Sireci A.N.
      • Aisner D.L.
      Detection of tumor NTRK gene fusions to identify patients who may benefit from tyrosine kinase (TRK) inhibitor therapy.
      ,
      • Caria P.
      • Dettori T.
      • Frau D.V.
      • Borghero A.
      • Cappai A.
      • Riola A.
      • et al.
      Assessing RET/PTC in thyroid nodule fine-needle aspirates: the FISH point of view.
      ,
      • Belli C.
      • Penault-Llorca F.
      • Ladanyi M.
      • Normanno N.
      • Scoazec J.Y.
      • Lacroix L.
      • et al.
      ESMO recommendations on the standard methods to detect RET fusions and mutations in daily practice and clinical research.
      ,
      • Kirchner M.
      • Glade J.
      • Lehmann U.
      • Merkelbach-Bruse S.
      • Hummel M.
      • Lehmann A.
      • et al.
      NTRK testing: first results of the QuiP-EQA scheme and a comprehensive map of NTRK fusion variants and their diagnostic coverage by targeted RNA-based NGS assays.
      ,
      • Vielh P.
      • Balogh Z.
      • Suciu V.
      • Richon C.
      • Job B.
      • Meurice G.
      • et al.
      DNA FISH diagnostic assay on cytological samples of thyroid follicular neoplasms.


      1–2 days1–2 slidesFusions only
      • BRAF: 56%/98%
        RET: 73–100%/55–100%
        NTRK: 94%/96%
        ALK: NA
      • Established approach, widely available
        Ability to detect and analyse the distribution of molecular alterations at a single-cell level
        Break-apart FISH detects rearrangements without knowledge of partner gene
      • Labour intensive and costly, particularly for rare biomarkers
        Detects DNA fusions; does not confirm alteration is functionally expressed
        Not easily multiplexed with other biomarkers
        Limited scalability for high volume testing
        Sensitivity and specificity variable depending on break-apart assay design and parameters
      Sanger sequencing
      • Zhao J.
      • Liu P.
      • Yu Y.
      • Zhi J.
      • Zheng X.
      • Yu J.
      • et al.
      Comparison of diagnostic methods for the detection of a BRAF mutation in papillary thyroid cancer.
      ,
      • Cha Y.J.
      • Koo J.S.
      Next-generation sequencing in thyroid cancer.
      ,
      • Kim J.K.
      • Seong C.Y.
      • Bae I.E.
      • Yi J.W.
      • Yu H.W.
      • Kim S.J.
      • et al.
      Comparison of immunohistochemistry and direct sequencing methods for identification of the BRAF(V600E) mutation in papillary thyroid carcinoma.
      10–15 days10 slidesFusions and mutations
      • BRAF: 97%/95%
        RET: NA
        NTRK: NA
        ALK: NA
      • Highly specific
      • Less sensitive than NGS
        High costs
      RT-PCR
      • Hsiao S.J.
      • Zehir A.
      • Sireci A.N.
      • Aisner D.L.
      Detection of tumor NTRK gene fusions to identify patients who may benefit from tyrosine kinase (TRK) inhibitor therapy.
      ,
      • Caria P.
      • Dettori T.
      • Frau D.V.
      • Borghero A.
      • Cappai A.
      • Riola A.
      • et al.
      Assessing RET/PTC in thyroid nodule fine-needle aspirates: the FISH point of view.
      ,
      • Letovanec I.
      • Finn S.
      • Zygoura P.
      • Smyth P.
      • Soltermann A.
      • Bubendorf L.
      • et al.
      Evaluation of NGS and RT-PCR methods for ALK rearrangement in European NSCLC patients: results from the European Thoracic Oncology Platform Lungscape Project.
      ,
      • Zhao J.
      • Liu P.
      • Yu Y.
      • Zhi J.
      • Zheng X.
      • Yu J.
      • et al.
      Comparison of diagnostic methods for the detection of a BRAF mutation in papillary thyroid cancer.
      5–10 days2–3 slidesFusions and mutations
      • BRAF: 99%/91%
        RET: 14%/100%
        NTRK: NA
        ALK: 70%/87%
      • Rapid and inexpensive
        Well-established technique in molecular genetics laboratories
        Detects rearrangements without knowledge of partner gene
      • PCR primer pairs must be designed and validated for each biomarker
        Does not confirm protein expression
      NGS
      • Hsiao S.J.
      • Zehir A.
      • Sireci A.N.
      • Aisner D.L.
      Detection of tumor NTRK gene fusions to identify patients who may benefit from tyrosine kinase (TRK) inhibitor therapy.
      ,
      • Letovanec I.
      • Finn S.
      • Zygoura P.
      • Smyth P.
      • Soltermann A.
      • Bubendorf L.
      • et al.
      Evaluation of NGS and RT-PCR methods for ALK rearrangement in European NSCLC patients: results from the European Thoracic Oncology Platform Lungscape Project.
      ,
      • Murphy D.A.
      • Ely H.A.
      • Shoemaker R.
      • Boomer A.
      • Culver B.P.
      • Hoskins I.
      • et al.
      Detecting gene rearrangements in patient populations through a 2-step diagnostic test comprised of rapid IHC enrichment followed by sensitive next-generation sequencing.
      ,
      • Hechtman J.F.
      NTRK insights: best practices for pathologists.
      ,
      • Nikiforova M.N.
      • Wald A.I.
      • Roy S.
      • Durso M.B.
      • Nikiforov Y.E.
      Targeted next-generation sequencing panel (ThyroSeq) for detection of mutations in thyroid cancer.
      ,
      • Yang S.
      • Aypar U.
      • Rosen E.
      • Mata D.
      • Benayed R.
      • Mullaney K.
      • et al.
      A performance comparison of commonly used assays to detect RET fusions.
      10–15 days5 slidesFusions and mutations
      • BRAF (DNA): 95–98%/up to 100%
        RET (DNA): 100%/100%
        NTRK (RNA): 95%/100%
        NTRK (DNA): 97%/100%
        NTRK3 sensitivity: 77%
        ALK (RNA): 85%/79%
      • Enables broad screening of numerous gene loci
        Commercial kits available
        Can detect unknown fusion partners
        Readily multiplexed
        RNA-based NGS
        Ensures only transcriptionally active alterations are detected, and allows in-frame versus out-of-frame confirmation
      • May require high level of infrastructure investment
        Requires high level bioinformatics capability
        Dependent on quality of isolated DNA/RNA and on the content of tumoural DNA
        DNA-based NGS
        Does not confirm protein expression
        Detected alterations may not be expressed or in-frame
        Large introns can prove problematic
        RNA-based NGS
        Detection of transcripts expressed at low levels may be challenging
      ALK, anaplastic lymphoma kinase; BRAF, B-Raf proto-oncogene; FISH, fluorescence in situ hybridization; IHC, immunohistochemistry; NA, not available; NGS, next-generation sequencing; NTRK, neurotrophic tyrosine receptor kinase; RET, rearranged during transfection; RT-PCR, reverse transcription polymerase chain reaction; TRK, tropomyosin receptor kinase.
      *Turnaround times and tissue requirements are illustrative and will vary according to individual centres.
      Sensitivity and specificity of particular assays/molecular targets can vary according to tumour type.
      Despite certain limitations, IHC, FISH, and RT-PCR are established approaches for assessing genetic alterations, and are generally widely available. They are, however, relatively low-throughput techniques. The growing number of genomic biomarkers driving treatment decision-making means that broad testing methods are becoming increasingly important. Next-generation sequencing (NGS) of DNA and RNA are specific and sensitive technique [
      • Marchiò C.
      • Scaltriti M.
      • Ladanyi M.
      • Iafrate A.
      • Bibeau F.
      • Dietel M.
      • et al.
      ESMO recommendations on the standard methods to detect NTRK fusions in daily practice and clinical research.
      ] (Table 3) that allow simultaneous detection of multiple genomic alterations, and can obtain the most information from the least amount of tissue [
      • Hsiao S.J.
      • Zehir A.
      • Sireci A.N.
      • Aisner D.L.
      Detection of tumor NTRK gene fusions to identify patients who may benefit from tyrosine kinase (TRK) inhibitor therapy.
      ]. Commercially available platforms have been reviewed elsewhere [
      • Marchiò C.
      • Scaltriti M.
      • Ladanyi M.
      • Iafrate A.
      • Bibeau F.
      • Dietel M.
      • et al.
      ESMO recommendations on the standard methods to detect NTRK fusions in daily practice and clinical research.
      ,
      • Suresh P.S.
      • Venkatesh T.
      • Tsutsumi R.
      • Shetty A.
      Next-generation sequencing for endocrine cancers: Recent advances and challenges.
      ]. To accurately detect relatively low-frequency molecular alterations (e.g. of ALK and NTRK) that occur in thyroid cancer, it is important to also consider the analytical sensitivity and specificity of mutation detection to minimize false-negative and false-positive results. NTRK gene fusions have not been extensively characterized, and therefore DNA-NGS has limitations, especially for rearrangements involving NTRK2 and NTRK3 owing to involvement of large intronic regions. Additionally, many of the fusions detected by DNA-NGS are of unknown functional significance and require confirmation by RNA-based assay, or IHC [
      • Marchiò C.
      • Scaltriti M.
      • Ladanyi M.
      • Iafrate A.
      • Bibeau F.
      • Dietel M.
      • et al.
      ESMO recommendations on the standard methods to detect NTRK fusions in daily practice and clinical research.
      ]. Tissue preparation is also an important consideration, particularly for the preservation of RNA required for RT-PCR and RNA-NGS.
      A two-step testing approach may provide a solution to balance cost, availability, and specificity: rapid, low-cost screening by IHC, and referral of only putatively positive samples for higher resolution multiplexed NGS-based testing [
      • Murphy D.A.
      • Ely H.A.
      • Shoemaker R.
      • Boomer A.
      • Culver B.P.
      • Hoskins I.
      • et al.
      Detecting gene rearrangements in patient populations through a 2-step diagnostic test comprised of rapid IHC enrichment followed by sensitive next-generation sequencing.
      ]. Costs associated with NGS are high, particularly when screening for rare alterations, such as NTRK gene fusions, but cost-effectiveness can be improved by using a triage-based strategy to enrich the population likely to harbour a NTRK gene fusion [
      • Beresford L.
      • Murphy P.
      • Dias S.
      • Claxton L.
      • Walton M.
      • Metcalf R.
      • et al.
      Appraising the costs of genomic testing for histology-independent technologies: an illustrative example for NTRK fusions.
      ]. Furthermore, NGS allows simultaneous analysis of multiple genetic alterations, thus offering a more efficient and tissue-saving analysis than serial single biomarker analysis [
      • Malone E.R.
      • Oliva M.
      • Sabatini P.J.B.
      • Stockley T.L.
      • Siu L.L.
      Molecular profiling for precision cancer therapies.
      ].

      Recommendations: Optimal molecular testing strategy for follicular cell-derived thyroid cancer

      The decision to initiate systemic therapy for patients with RAI-R advanced/metastatic thyroid cancer should be made with individual patient and disease characteristics in mind, and with due consideration of patient preferences [
      • Filetti S.
      • Durante C.
      • Hartl D.
      • Leboulleux S.
      • Locati L.D.
      • Newbold K.
      • et al.
      Thyroid cancer: ESMO clinical practice guidelines for diagnosis, treatment and follow-up.
      ]. To fully understand disease characteristics and clinical behaviour, it is important that the molecular profile of the tumour is documented. However, understanding who, and when, to test and for which specific molecular aberrations is under debate.
      Following recent developments with targeted therapies for advanced thyroid cancer, we have new treatment options for patients harbouring actionable genetic alterations. Recent testing algorithms suggest that the TRK fusion-positive population may be enriched by first excluding the presence of other known driver mutations [
      • Marchiò C.
      • Scaltriti M.
      • Ladanyi M.
      • Iafrate A.
      • Bibeau F.
      • Dietel M.
      • et al.
      ESMO recommendations on the standard methods to detect NTRK fusions in daily practice and clinical research.
      ,
      • Solomon J.P.
      • Benayed R.
      • Hechtman J.F.
      • Ladanyi M.
      Identifying patients with NTRK fusion cancer.
      ]. However, ESMO recommends up-front screening with NGS panels as the least expensive and most efficient approach to identify actionable driver mutations [
      • Marchiò C.
      • Scaltriti M.
      • Ladanyi M.
      • Iafrate A.
      • Bibeau F.
      • Dietel M.
      • et al.
      ESMO recommendations on the standard methods to detect NTRK fusions in daily practice and clinical research.
      ]. We therefore recommend testing all patients with RAI-R advanced/metastatic follicular cell-derived thyroid cancer, and all patients with ATC, for actionable genomic alterations, including NTRK and RET gene fusions, BRAF V600E mutations, and ALK fusions. We also recommend that the most specific test that can generate results in a timely manner be performed. This may mean that different tests may be performed first in different centres. For patients with ATC, we emphasize the need for immediate testing for NTRK and RET fusions to minimize delays in treating this very aggressive and deadly subtype.
      Clinical detection of NTRK gene fusions has been based predominantly on comprehensive nucleic acid-based profiling (DNA-based NGS and targeted RNA sequencing) [
      • Cocco E.
      • Scaltriti M.
      • Drilon A.
      NTRK fusion-positive cancers and TRK inhibitor therapy.
      ]. ESMO recommends FISH, RT-PCR, or RNA-based sequencing panels to confirm the presence of an NTRK fusion in tumours with a high prevalence of TRK fusions (e.g. secretory carcinoma of the breast or salivary glands, and congenital fibrosarcoma). For tumours where NTRK gene fusions are less common, such as in thyroid cancer, RNA-NGS or pre-screening by pan-TRK IHC followed by confirmatory RNA-NGS testing is recommended [
      • Marchiò C.
      • Scaltriti M.
      • Ladanyi M.
      • Iafrate A.
      • Bibeau F.
      • Dietel M.
      • et al.
      ESMO recommendations on the standard methods to detect NTRK fusions in daily practice and clinical research.
      ]. In cases where a gene fusion is suspected, but has returned a negative result by upfront NGS, confirmatory testing by RT-PCR or FISH could be considered. More recently, a comparison of molecular techniques to detect NTRK1 and -3 rearrangements cautioned that pan-TRK IHC had high specificity, but only moderate sensitivity in NTRK3 fusion cases. Results should also be interpreted with caution because of heterogeneity in staining [
      • Lee Y.-C.
      • Chen J.-Y.
      • Huang C.-J.
      • Chen H.-S.
      • Yang A.-H.
      • Hang J.-F.
      Detection of NTRK1/3 rearrangements in papillary thyroid carcinoma using immunohistochemistry, fluorescent in situ hybridization, and next-generation sequencing.
      ]. Any TRK-positive cases detected by IHC therefore need to be confirmed by NGS, FISH, or RT-PCR.
      ESMO has not recommended IHC as a method to detect RET gene fusions in follicular cell-derived thyroid cancer because of the lack of sensitivity and, in particular, specificity of the test. Instead, NGS should be used. If NGS is not available, FISH or RT-PCR is recommended. If a formalin-fixed, paraffin-embedded (FFPE) sample is not available, a liquid biopsy should be performed [
      • Belli C.
      • Penault-Llorca F.
      • Ladanyi M.
      • Normanno N.
      • Scoazec J.Y.
      • Lacroix L.
      • et al.
      ESMO recommendations on the standard methods to detect RET fusions and mutations in daily practice and clinical research.
      ]. Circulating tumour DNA (ctDNA) analysis can be effective and useful for patients who do not have sufficient biopsy materials. However, most widely used NGS platforms for ctDNA analysis may not be effective for detecting gene rearrangements [
      • Solomon J.P.
      • Hechtman J.F.
      Detection of NTRK fusions: merits and limitations of current diagnostic platforms.
      ,
      • Suh Y.
      • Kwon M.
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      Limited clinical and diagnostic utility of circulating tumor DNA detection in patients with early-stage well-differentiated thyroid cancer: comparison with benign thyroid nodules and healthy individuals.
      ]. Furthermore, although ctDNA analysis is specific, sensitivity can be moderate as compared with analysis of tissue samples, and technical performance of ctDNA analysis varies according to tumour type [
      • Ye P.
      • Cai P.
      • Xie J.
      • Zhang J.
      Reliability of BRAF mutation detection using plasma sample: a systematic review and meta-analysis.
      ] and commercial assay used [
      • Solomon J.P.
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      Detection of NTRK fusions: merits and limitations of current diagnostic platforms.
      ].
      The American Thyroid Association recommends that in patients diagnosed with possible ATC, assessment of BRAF V600E mutations can be performed by IHC [
      • Bible K.C.
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      • Brierley J.
      • Brito J.P.
      • Cabanillas M.E.
      • Clark Jr, T.J.
      • et al.
      2021 American Thyroid Association guidelines for management of patients with anaplastic thyroid cancer.
      ]. ESMO guidelines also encourage the use of molecular profiling in ATC to determine whether there are any actionable mutations for targeted therapies [
      • Filetti S.
      • Durante C.
      • Hartl D.
      • Leboulleux S.
      • Locati L.D.
      • Newbold K.
      • et al.
      Thyroid cancer: ESMO clinical practice guidelines for diagnosis, treatment and follow-up.
      ]. In a phase II study of vemurafenib in patients with metastatic RAI-R PTC, BRAF V600E mutation status was detected by IHC and confirmed by real-time PCR [
      • Brose M.S.
      • Cabanillas M.E.
      • Cohen E.E.
      • Wirth L.J.
      • Riehl T.
      • Yue H.
      • et al.
      Vemurafenib in patients with BRAF(V600E)-positive metastatic or unresectable papillary thyroid cancer refractory to radioactive iodine: a non-randomised, multicentre, open-label, phase 2 trial.
      ]. Other studies have demonstrated that this mutation can be detected with high specificity and selectivity in patients with PTC, PDTC, and ATC [
      • Abd Elmageed Z.Y.
      • Sholl A.B.
      • Tsumagari K.
      • Al-Qurayshi Z.
      • Basolo F.
      • Moroz K.
      • et al.
      Immunohistochemistry as an accurate tool for evaluating BRAF-V600E mutation in 130 samples of papillary thyroid cancer.
      ,
      • Ghossein R.A.
      • Katabi N.
      • Fagin J.A.
      Immunohistochemical detection of mutated BRAF V600E supports the clonal origin of BRAF-induced thyroid cancers along the spectrum of disease progression.
      ].
      Lee et al. propose that all patients with PTC should first be tested for BRAF V600E mutations, to exclude this common oncogenic driver [
      • Lee Y.-C.
      • Chen J.-Y.
      • Huang C.-J.
      • Chen H.-S.
      • Yang A.-H.
      • Hang J.-F.
      Detection of NTRK1/3 rearrangements in papillary thyroid carcinoma using immunohistochemistry, fluorescent in situ hybridization, and next-generation sequencing.
      ]. We also recommend testing for RET in this population as this is commonly altered in PTC [
      • Santoro M.
      • Moccia M.
      • Federico G.
      • Carlomagno F.
      RET gene fusions in malignancies of the thyroid and other tissues.
      ]. The mutations that can lead to the development of PTC are generally mutually exclusive; however, it has been shown in rare cases that BRAF V600E and RET are possible dual mutations in PTC, especially in patients with recurrent disease [
      • Fagin J.A.
      Genetics of papillary thyroid cancer initiation: implications for therapy.
      ,
      • Guerra A.
      • Zeppa P.
      • Bifulco M.
      • Vitale M.
      Concomitant BRAF(V600E) mutation and RET/PTC rearrangement is a frequent occurrence in papillary thyroid carcinoma.
      ,
      • Henderson Y.C.
      • Shellenberger T.D.
      • Williams M.D.
      • El-Naggar A.K.
      • Fredrick M.J.
      • Cieply K.M.
      • et al.
      High rate of BRAF and RET/PTC dual mutations associated with recurrent papillary thyroid carcinoma.
      ]. If BRAF V600E-negative, pan-TRK IHC is recommended, followed by confirmatory testing by NGS or FISH (next-generation DNA sequencing can be performed if enough sample is available, followed by RNA sequencing if negative for an oncogenic driver mutation). Owing to the heterogeneity of pan-TRK staining, which does not detect the fusion, but rather the overexpression of TRK proteins commonly seen in tumours harbouring NTRK gene fusions, a negative result does not necessarily mean that a TRK fusion is not present. This is particularly true when detecting NTRK3 fusions by IHC because these alterations are rare and the technique is less sensitive than when testing for NTRK1/2 fusions. In the case of a negative pan-TRK IHC pre-screening test, morphological analysis of tissue samples is recommended: non-infiltrative borders, clear cell change, and reduced nuclear elongation and irregularity are indicative of a potential TRK fusion that warrants confirmatory testing by NGS or FISH [
      • Lee Y.-C.
      • Chen J.-Y.
      • Huang C.-J.
      • Chen H.-S.
      • Yang A.-H.
      • Hang J.-F.
      Detection of NTRK1/3 rearrangements in papillary thyroid carcinoma using immunohistochemistry, fluorescent in situ hybridization, and next-generation sequencing.
      ]. Histology-based triage may be particularly useful for rare cancer types that commonly harbour NTRK fusions (e.g. paediatric PTC), and has been recommended by others to enrich for such alterations [
      • Solomon J.P.
      • Benayed R.
      • Hechtman J.F.
      • Ladanyi M.
      Identifying patients with NTRK fusion cancer.
      ]. The prevalence of RET and NTRK gene fusions increases in paediatric patients with PTC, and in individuals with radiation-induced PTC [
      • Prasad M.L.
      • Vyas M.
      • Horne M.J.
      • Virk R.K.
      • Morotti R.
      • Liu Z.
      • et al.
      NTRK fusion oncogenes in pediatric papillary thyroid carcinoma in northeast United States.
      ,
      • Ricarte-Filho J.C.
      • Li S.
      • Garcia-Rendueles M.E.
      • Montero-Conde C.
      • Voza F.
      • Knauf J.A.
      • et al.
      Identification of kinase fusion oncogenes in post-Chernobyl radiation-induced thyroid cancers.
      ,
      • Santoro M.
      • Carlomagno F.
      Central role of RET in thyroid cancer.
      ,
      • Pekova B.
      • Sykorova V.
      • Dvorakova S.
      • Vaclavikova E.
      • Moravcova J.
      • Katra R.
      • et al.
      RET, NTRK, ALK, BRAF, and MET fusions in a large cohort of pediatric papillary thyroid carcinomas.
      ]. Our proposed testing strategy for RAI-R PTC, FTC, PDTC, and all patients with ATC can be found in Fig. 2.
      Figure thumbnail gr2
      Fig. 2Proposed testing strategy for follicular cell-derived thyroid cancer. (A) Testing strategy when starting with FISH, RT-PCR, or NGS. (B) Testing strategy when starting with IHC. *Test to be performed is dependent on individual centre’s experience. Chosen test should be the most sensitive, specific, and fastest test available. If possible, tests should be run simultaneously to minimize delays to treatment. FFPE NGS preferred for RET. Dabrafenib in combination with trametinib is approved by the FDA for use among patients with locally advanced or metastatic ATC with BRAF V600E mutations, and no satisfactory locoregional treatment options. It is not approved by the EMA
      [

      Novartis Pharmaceuticals Corporation. Dabrafenib Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2018/202806s010lbl.pdf; 2018 [accessed 25 November 2021].

      ]
      . §Selpercatinib is EMA approved for the treatment of adults with advanced RET fusion-positive thyroid cancer who require systemic therapy following prior treatment with sorafenib and/or lenvatinib and for the treatment of adult and paediatric patients 12 years of age and older with advanced RET-mutant MTC who require systemic therapy following prior treatment with cabozantinib and/or vandetanib
      [

      Eli Lilly Nederaland B.V. Retsevmo Summary of Product Characterisitcs, https://www.ema.europa.eu/en/documents/product-information/retsevmo-epar-product-information_en.pdf; 2021 [accessed 25 November 2021].

      ]
      . **Pralsetinib is FDA approved for the treatment of adult and paediatric patients 12 years of age and older with advanced or metastatic RET-mutant MTC who require systemic therapy or with advanced or metastatic RET fusion-positive thyroid cancer who require systemic therapy and whose disease is RAI-R (if RAI is appropriate). It is not approved by the EMA
      [

      Roche. Gavreto Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2020/214701s000lbl.pdf; 2020 [accessed 25 November 2021].

      ]
      . ††Larotrectinib is indicated by the EMA as a monotherapy for the treatment of adult and paediatric patients with solid tumours that display an NTRK gene fusion, who have a disease that is locally advanced, metastatic, or where surgical resection is likely to result in severe morbidity, and who have no satisfactory treatment options
      [

      Bayer AG. VITRAKVI Summary of Product Characteristics, https://www.ema.europa.eu/en/documents/product-information/vitrakvi-epar-product-information_en.pdf; 2022 [accessed 1 March 2022].

      ]
      . ‡‡Entrectinib is indicated by the EMA as a monotherapy for the treatment of adult and paediatric patients 12 years of age and older with solid tumours expressing an NTRK gene fusion, who have a disease that is locally advanced, metastatic, or where surgical resection is likely to result in severe morbidity, who have not received a prior TRK inhibitor, and who have no satisfactory treatment options
      [

      Roche Registration GmBH. Rozlytrek Summary of Product Characteristics, https://www.ema.europa.eu/en/documents/product-information/rozlytrek-epar-product-information_en.pdf; 2020 [accessed 25 November 2021].

      ]
      . §§IHC is not ESMO recommended as a method to detect RET fusions in follicular cell-derived thyroid cancer because of the lack of sensitivity and specificity of the test. ***A negative IHC result does not necessarily mean that a TRK fusion is not present due to the heterogeneity of pan-TRK staining, and histological assessment is recommended, with confirmatory testing performed if an NTRK gene fusion is suspected based on morphology. ALK, anaplastic lymphoma kinase; ATC, anaplastic thyroid cancer; BRAF, B-Raf proto-oncogene; EMA, European Medicines Agency; ESMO, European Society for Medical Oncology; FDA, US Food and Drug Administration; FFPE, formalin-fixed, paraffin-embedded; FISH, fluorescence in situ hybridization; FTC, follicular thyroid cancer; IHC, immunohistochemistry; MTC, medullary thyroid cancer; NGS, next-generation sequencing; NTRK, neurotrophic tyrosine receptor kinase; PDTC, poorly differentiated thyroid cancer; PTC, papillary thyroid cancer; RAI-R, radioactive iodine-refractory; RET, rearranged during transfection; RT-PCR, reverse transcription polymerase chain reaction; TRK, tropomyosin receptor kinase.
      One of the greatest barriers to accessing targeted treatment is implementation of effective screening programmes to ensure that all patients who could potentially benefit from treatment are identified. This requires education on targetable molecular alterations in advanced thyroid cancer, and the potential clinical benefits that can be achieved once targeted therapy is initiated. Regular communication between endocrinologists, nuclear medicine specialists, pathologists, surgeons, and medical oncologists, in the form of a multidisciplinary tumour team, is essential to ensure that all patients with aggressive thyroid cancers have access to the latest advances in treatment, including clinical trials. We recommend that all patients who may benefit from targeted treatment are referred to a centre that has this multidisciplinary team set-up. We also recommend that a reference pathologist is consulted for any thyroid/endocrine pathology.

      Future perspectives

      The identification of actionable alterations in thyroid cancer has presented new opportunities for patients to receive targeted treatments. Clinical trials have indicated that RET and TRK inhibitors may be more tolerable for patients than MKIs and BRAF inhibitors. In the LIBRETTO trial, 2% of patients discontinued selpercatinib because of treatment-related AEs [
      • Wirth L.J.
      • Sherman E.
      • Robinson B.
      • Solomon B.
      • Kang H.
      • Lorch J.
      • et al.
      Efficacy of selpercatinib in RET-altered thyroid cancers.
      ], and 9% of patients with RET-altered thyroid cancer in the ARROW trial discontinued pralsetinib treatment due to AEs [

      Roche. Gavreto Prescribing Information, https://www.accessdata.fda.gov/drugsatfda_docs/label/2020/214701s000lbl.pdf; 2020 [accessed 25 November 2021].

      ]. No AEs leading to discontinuation of treatment were observed in individuals with thyroid cancer treated with larotrectinib [
      • Cabanillas M.
      • Drilon A.
      • Farago A.
      • Brose M.
      • McDermott R.
      • Sohal D.
      • et al.
      Larotrectinib treatment of advanced TRK fusion thyroid cancer.
      ]. In the pivotal phase III trials for MKIs, the proportion of patients who discontinued treatment because of treatment-related AEs was 14% for lenvatinib and 19% for sorafenib [
      • Schlumberger M.
      • Tahara M.
      • Wirth L.J.
      • Robinson B.
      • Brose M.S.
      • Elisei R.
      • et al.
      Lenvatinib versus placebo in radioiodine-refractory thyroid cancer.
      ,
      • Brose M.S.
      • Nutting C.M.
      • Jarzab B.
      • Elisei R.
      • Siena S.
      • Bastholt L.
      • et al.
      Sorafenib in radioactive iodine-refractory, locally advanced or metastatic differentiated thyroid cancer: a randomised, double-blind, phase 3 trial.
      ]. Over a quarter of patients discontinued treatment due to AEs related to vemurafenib [
      • Brose M.S.
      • Cabanillas M.E.
      • Cohen E.E.
      • Wirth L.J.
      • Riehl T.
      • Yue H.
      • et al.
      Vemurafenib in patients with BRAF(V600E)-positive metastatic or unresectable papillary thyroid cancer refractory to radioactive iodine: a non-randomised, multicentre, open-label, phase 2 trial.
      ], while 8% of patients discontinued dabrafenib treatment for the same reason [
      • Subbiah V.
      • Kreitman R.J.
      • Wainberg Z.A.
      • Cho J.Y.
      • Schellens J.H.M.
      • Soria J.C.
      • et al.
      Dabrafenib and trametinib treatment in patients with locally advanced or metastatic BRAF V600-mutant anaplastic thyroid cancer.
      ].
      Immuno-oncology is another area of precision medicine that is also gaining momentum in advanced thyroid cancer, including immune checkpoint blockade (Table 2). Inhibition of cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) and programmed cell death protein (PD-1), among other proteins in the checkpoint cascade, is currently being investigated as new molecular targets in patients with advanced thyroid cancers [
      • Naoum G.E.
      • Morkos M.
      • Kim B.
      • Arafat W.
      Novel targeted therapies and immunotherapy for advanced thyroid cancers.
      ,
      • Capdevila J.
      • Wirth L.J.
      • Ernst T.
      • Ponce Aix S.
      • Lin C.C.
      • Ramlau R.
      • et al.
      PD-1 blockade in anaplastic thyroid carcinoma.
      ,
      • Dierks C.
      • Seufert J.
      • Aumann K.
      • Ruf J.
      • Klein C.
      • Kiefer S.
      • et al.
      Combination of lenvatinib and pembrolizumab is an effective treatment option for anaplastic and poorly differentiated thyroid carcinoma.
      ]. Depending on the outcomes of these trials, updated guidelines that incorporate testing for immuno-oncology markers may need to be developed. Furthermore, evidence is emerging of the prognostic significance of tumour RNA-expression profiles in addition to DNA alterations [
      • Capdevila J.
      • Matos I.
      • Mancuso F.
      • Iglesias C.
      • Nuciforo P.
      • Zafon C.
      • et al.
      Identification of expression profiles defining distinct prognostic subsets of radioactive-Iodine refractory differentiated thyroid cancer from the DECISION Trial.
      ], which may further expand our understanding of thyroid tumour biology, prognosis, and approaches to treatment.
      A key area of uncertainty in the treatment of thyroid cancer is the optimal sequence of targeted treatments. One limitation of targeted therapies is potentially transient efficacy due to the development of escape mechanisms by tumours [
      • Al-Jundi M.
      • Thakur S.
      • Gubbi S.
      • Klubo-Gwiezdzinska J.
      Novel targeted therapies for metastatic thyroid cancer: a comprehensive review.
      ]. In TRK fusion-positive cancers, resistance can be mediated by the acquisition of NTRK kinase domain mutations, including solvent-front and gatekeeper mutations. Mutations resulting in amino acid substitutions at the kinase solvent-front of fusion proteins have been identified in RET-mutated and TRK fusion tumours following targeted treatment [
      • Russo M.
      • Misale S.
      • Wei G.
      • Siravegna G.
      • Crisafulli G.
      • Lazzari L.
      • et al.
      Acquired resistance to the TRK inhibitor entrectinib in colorectal cancer.
      ,
      • Cocco E.
      • Schram A.M.
      • Kulick A.
      • Misale S.
      • Won H.H.
      • Yaeger R.
      • et al.
      Resistance to TRK inhibition mediated by convergent MAPK pathway activation.
      ,
      • Solomon B.J.
      • Tan L.
      • Lin J.J.
      • Wong S.Q.
      • Hollizeck S.
      • Ebata K.
      • et al.
      RET solvent front mutations mediate acquired resistance to selective RET inhibition in RET-driven malignancies.
      ]. Certain resistance mutations may be overcome by second-generation TRK inhibitors that are being explored in clinical trials. For example, selitrectinib is a second-generation selective TRK inhibitor that has demonstrated clinical activity in patients with TRK fusion solid tumours that have become resistant to first-generation TRK inhibitors; preliminary findings from a phase I study (n = 20) and an FDA expanded-access single patient protocol (n = 11) indicate an ORR of up to 50% [

      Hyman D, Kummar S, Farago AF, Geoerger B, Mau-Sorensen M, Taylor M, et al. Phase I and expanded access experience of LOXO-195 (BAY 2731954), a selective next-generation TRK inhibitor (TRKi). AACR, Poster CT127; 2019.

      ]. Repotrectinib, a second-generation ROS1, TRK, and ALK inhibitor, has demonstrated anti-tumour activity in patients with ROS1- or NTRK3-rearranged tumours that harbour resistant solvent-front mutations in an ongoing first-in-human dose-escalation trial [
      • Drilon A.
      • Ou S.I.
      • Cho B.C.
      • Kim D.W.
      • Lee J.
      • Lin J.J.
      • et al.
      Repotrectinib (TPX-0005) is a next-generation ROS1/TRK/ALK inhibitor that potently inhibits ROS1/TRK/ALK solvent- front mutations.
      ]. It has been shown that RET-positive cancers do not develop gatekeeper mutations after treatment with selpercatinib and pralsetinib. However, non-gatekeeper mutations can confer resistance to RET inhibitors, indicating the need for RET-specific treatments that can evade both gatekeeper and non-gatekeeper mutations [
      • Subbiah V.
      • Shen T.
      • Terzyan S.S.
      • Liu X.
      • Hu X.
      • Patel K.P.
      • et al.
      Structural basis of acquired resistance to selpercatinib and pralsetinib mediated by non-gatekeeper RET mutations.
      ]. We are only beginning to understand molecular mechanisms of tumour resistance; further research is key to fully appreciating the implications for practice. This could include testing novel combinations of treatments to avoid resistance and repeated tumour or liquid biopsies to monitor genetic changes. A new model of clinical trials methodology and management is needed to study this new generation of drugs in resistant tumours that emerge following the exposure to first-generation drugs.
      The optimal therapeutic sequencing of MKIs and targeted therapy, or their combination with immunotherapy, is not yet known, but knowledge of the molecular profile of the tumour allows informed treatment decisions to be made. More data from ongoing clinical trials will help to document the best sequence of available molecular therapies. Of note, the LIBRETTO-531 phase III study of selpercatinib in patients with RET-positive MTC is currently recruiting [

      U.S. National Library of Medicine, ClinicalTrials.gov. A study of selpercatinib (LY3527723) in participants with RET-mutant medullary thyroid cancer (LIBRETTO-531), https://clinicaltrials.gov/ct2/show/NCT04211337; 2019 [accessed 25 November 2021].

      ]. In this trial, selpercatinib will be compared to cabozantinib or vandetanib. A phase III study of pralsetinib versus standard of care (AcceleRET-MTC) is also planned to start in January 2022 [

      U.S. National Library of Medicine, ClinicalTrials.gov. A study of pralsetinib versus standard of care (SOC) for treatment of RET-mutated medullary thyroid cancer (MTC). (AcceleRET-MTC), https://clinicaltrials.gov/ct2/show/NCT04760288; 2021 [accessed 25 November 2021].

      ].

      Conclusions

      Recent leaps in our understanding of the molecular basis of follicular cell-derived thyroid cancers have led to advances in treatment approaches for patients with advanced disease. A shift in the management paradigm has begun [
      • Al-Jundi M.
      • Thakur S.
      • Gubbi S.
      • Klubo-Gwiezdzinska J.
      Novel targeted therapies for metastatic thyroid cancer: a comprehensive review.
      ], with the molecular landscape, rather than tumour histology/morphology, driving the need for an individualized treatment approach. The availability of drugs that target specific molecular alterations highlights the importance of optimal molecular testing to identify suitable candidates for such therapies. Until recently, patients with advanced RAI-R DTC and PDTC have had poor prognoses. However, treatment options with clinical benefit are now available for patients with tumours harbouring an actionable target. As such, we recommend routine testing for genetic alterations to be included as part of the clinical work-up for patients who have RAI-R advanced/metastatic PTC, FTC, and PDTC, and for all patients with ATC. Close interactions between endocrinologists, surgeons, nuclear medicine specialists, pathologists, and medical oncologists, as part of a multidisciplinary team, would help to ensure that patients are tested in a timely manner, and that when an actionable target is identified the appropriate treatment is offered.

      CRediT authorship contribution statement

      Jaume Capdevila: Conceptualization, Writing – original draft, Writing – review & editing. Ahmad Awada: Conceptualization, Writing – original draft, Writing – review & editing. Dagmar Führer-Sakel: Conceptualization, Writing – original draft, Writing – review & editing. Sophie Leboulleux: Conceptualization, Writing – original draft, Writing – review & editing. Patrick Pauwels: Conceptualization, Writing – original draft, Writing – review & editing.

      Declaration of Competing Interest

      The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Jaume Capdevila has acted as a scientific consultant (speaker and advisory roles) for Novartis, Pfizer, Ipsen, Exelixis, Bayer, Eisai, Advanced Accelerator Applications, Amgen, Lilly, Sanofi, and Merck Serono; he has received research support from Novartis, Pfizer, AstraZeneca, Advanced Accelerator Applications, Eisai, and Bayer. Ahmad Awada has acted as a scientific consultant (speaker and advisory roles) for and received research grants from Roche, Lilly, Amgen, Eisai, BMS, Pfizer, Novartis, MSD, Genomic Health, Ipsen, AstraZeneca, Bayer, Leo Pharma, Merck, and Daiichi. Dagmar Führer-Sakel has acted as a scientific consultant (speaker and advisory roles) for Eisai, Sanofi, and MedUpdate; she has received research support from Novartis and Lilly. Sophie Leboulleux has acted as a scientific consultant (speaker and advisory roles) for Bayer, Eisai, and Lilly; she has received research support from Sanofi and Novartis. Patrick Pauwels has acted as a scientific consultant (speaker and advisory roles) for Pfizer, Bayer, Lilly, Roche, Novartis, Incyte, Amgen, and AstraZeneca; he has received research support from Bayer, MSD, Roche, and AstraZeneca.

      Acknowledgement

      Medical writing support was provided by Anna Bakewell, PhD, Liz Hartfield, PhD, and Cameron Ward, BSc, of 7.4 Limited, and funded by Bayer AG according to Good Publication Practice guidelines. All authors contributed equally to the development of the manuscript. Responsibilities for opinions, conclusions, and data interpretation lie with the authors. This work was not funded by any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

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