Vorinostat, temozolomide or bevacizumab with irradiation and maintenance BEV/TMZ in pediatric high-grade glioma: A Children's Oncology Group Study.

Background: Outcomes for children with high-grade gliomas (HGG) remain poor. This multicenter phase II trial evaluated whether concurrent use of vorinostat or bevacizumab with focal radiotherapy (RT) improved 1-year event-free survival (EFS) compared to temozolomide in children with newly diagnosed HGG who received maintenance temozolomide and bevacizumab. Methods: Patients ≥ 3 and < 22 years with localized, non-brainstem HGG were randomized to receive RT (dose 54-59.4Gy) with vorinostat, temozolomide, or bevacizumab followed by 12 cycles of bevacizumab and temozolomide maintenance therapy. Results: Among 90 patients randomized, the 1-year EFS for concurrent bevacizumab, vorinostat, or temozolomide with RT was 43.8% (±8.8%), 41.4% (±9.2%), and 59.3% (±9.5%), respectively, with no significant difference among treatment arms. Three- and five-year EFS for the entire cohort was 14.8% and 13.4%, respectively, with no significant EFS difference among the chemoradiotherapy arms. IDH mutations were associated with more favorable EFS (P = .03), whereas H3.3 K27M mutations (P = .0045) and alterations in PIK3CA or PTEN (P = .025) were associated with worse outcomes. Patients with telomerase- and alternative lengthening of telomeres (ALT)-negative tumors (n = 4) had an EFS of 100%, significantly greater than those with ALT or telomerase, or both (P = .002). While there was no difference in outcomes based on TERT expression, high TERC expression was associated with inferior survival independent of the telomere maintenance mechanism (P = .0012). Conclusions: Chemoradiotherapy with vorinostat or bevacizumab is not superior to temozolomide in children with newly diagnosed HGG. Patients with telomerase- and ALT-negative tumors had higher EFS suggesting that, if reproduced, mechanism of telomere maintenance should be considered in molecular-risk stratification in future studies.

Characterization of low-grade epilepsy-associated tumor from implanted stereoelectroencephalography electrodes.

Low-grade epilepsy-associated tumors (LEATs) are a common cause of drug-resistant epilepsy in children. Herein, we demonstrate the feasibility of using tumor tissue derived from stereoelectroencephalography (sEEG) electrodes upon removal to molecularly characterize tumors and aid in diagnosis. An 18-year-old male with focal epilepsy and MRI suggestive of a dysembryoplastic neuroepithelial tumor (DNET) in the left posterior temporal lobe underwent implantation of seven peri-tumoral sEEG electrodes for peri-operative language mapping and demarcation of the peri-tumoral ictal zone prior to DNET resection. Using electrodes that passed through tumor tissue, we show successful isolation of tumor DNA and subsequent analysis using standard methods for tumor classification by DNA, including Glioseq targeted sequencing and DNA methylation array analysis. This study provides preliminary evidence for the feasibility of molecular diagnosis of LEATs or other lesions using a minimally invasive method with microscopic tissue volumes. The implications of sEEG electrodes in tumor characterization are broad but would aid in diagnosis and subsequent targeted therapeutic strategies.

Targeted Next-Generation Sequencing Improves the Prognostication of Patients with Disseminated Appendiceal Mucinous Neoplasms (Pseudomyxoma Peritonei).

BACKGROUND: Appendiceal mucinous neoplasms (AMNs) with disseminated disease (pseudomyxoma peritonei) are heterogeneous tumors with variable clinicopathologic behavior. Despite the development of prognostic systems, objective biomarkers are needed to stratify patients. With the advent of next-generation sequencing (NGS), it remains unclear if molecular testing can improve the evaluation of disseminated AMN patients. METHODS: Targeted NGS was performed for 183 patients and correlated with clinicopathologic features to include American Joint Committee on Cancer/World Health Organization (AJCC/WHO) histologic grade, peritoneal cancer index (PCI), completeness of cytoreduction (CC) score, and overall survival (OS). RESULTS: Genomic alterations were identified for 179 (98%) disseminated AMNs. Excluding mitogen-activated protein kinase genes and GNAS due to their ubiquitous nature, collective genomic alterations in TP53, SMAD4, CDKN2A, and the mTOR genes were associated with older mean age, higher AJCC/WHO histologic grade, lymphovascular invasion, perineural invasion, regional lymph node metastasis, and lower mean PCI (p < 0.040). Patients harboring TP53, SMAD4, ATM, CDKN2A, and/or mTOR gene alterations were found to have lower OS rates of 55% at 5 years and 14% at 10 years, compared with 88% at 5 years and 88% at 10 years for patients without the aforementioned alterations (p < 0.001). Based on univariate and multivariate analyses, genomic alterations in TP53, SMAD4, ATM, CDKN2A, and/or the mTOR genes in disseminated AMNs were a negative prognostic factor for OS and independent of AJCC/WHO histologic grade, PCI, CC score, and hyperthermic intraperitoneal chemotherapy treatment (p = 0.006). CONCLUSIONS: Targeted NGS improves the prognostic assessment of patients with disseminated AMNs and identifies patients who may require increased surveillance and/or aggressive management.

Prognostic Value of Preoperative Molecular Testing and Implications for Initial Surgical Management in Thyroid Nodules Harboring Suspected (Bethesda V) or Known (Bethesda VI) Papillary Thyroid Cancer.

Importance: Molecular testing is commonly used in the diagnosis of thyroid nodules with indeterminate cytology. The role of molecular testing in prognosticating oncologic outcomes in thyroid nodules with suspicious or malignant cytology is unclear. Objective: To determine whether molecular profiling of Bethesda V (suspicious for thyroid cancer) and VI (thyroid cancer) nodules is associated with improved prognostication and whether it may inform initial treatment. Design, Setting, and Participants: This retrospective cohort study included consecutive patients with Bethesda V or VI nodules who underwent surgery, with histopathology showing differentiated thyroid cancer, between May 1, 2016, and July 31, 2019 in the University of California, Los Angeles health system. Data were analyzed between April 2, 2021, and January 18, 2023. Exposures: Masked ThyroSeq, version 3 molecular analysis after completion of initial treatment and acquisition of follow-up data. Main Outcomes and Measures: Structural disease persistence or recurrence, distant metastasis, and recurrence-free survival were assessed using ThyroSeq Cancer Risk Classifier (CRC) molecular risk groups (low, RAS-like; intermediate, BRAF-like; high, combination of BRAF/RAS plus TERT or other high-risk alterations) using Cox proportional hazards regression models. Results: In 105 patients with papillary thyroid cancer (median [IQR] follow-up, 3.8 [3.0-4.7] years), ThyroSeq identified genomic alterations in 100 (95%) samples (6 [6%] low risk, 88 [88%] intermediate risk, and 6 [6%] high risk; median [IQR] age, 44 [34-56] years; 68 [68%] female and 32 [32%] male). No patients with low-risk or negative results experienced recurrence. Of the 88 patients with intermediate risk, 6 (7%) experienced local recurrence, with 1 of them also developing distant metastasis. The 6 patients with high risk (all with BRAF V600E plus TERT mutation) underwent total thyroidectomy followed by radioactive iodine (RAI) ablation. Four patients with high risk (67%) experienced local recurrence, with 3 of them also developing distant metastasis. Thus, patients with high-risk alterations were more likely to experience persistence or recurrence and distant metastasis than patients with intermediate risk. In a multivariable analysis incorporating patient age, sex, cancer size, ThyroSeq molecular risk group, extrathyroidal extension, lymph node positivity, American Thyroid Association risk, and RAI ablation, only cancer size (hazard ratio, 1.36; 95% CI, 1.02-1.80) and ThyroSeq CRC molecular risk group (high vs intermediate and low: hazard ratio, 6.22; 95% CI, 1.04-37.36) were associated with structural recurrence. Conclusions and Relevance: Among the 6% of patients with high-risk ThyroSeq CRC alterations in this cohort study, the majority experienced recurrence or distant metastasis despite initial treatment with total thyroidectomy and RAI ablation. In contrast, patients with low- and intermediate-risk alterations had a low recurrence rate. Preoperative knowledge of molecular alteration status at diagnosis may allow for deescalation of initial surgery and refining of the intensity of postoperative surveillance in patients presenting with Bethesda V and VI thyroid nodules.

A Combined DNA/RNA-based Next-Generation Sequencing Platform to Improve the Classification of Pancreatic Cysts and Early Detection of Pancreatic Cancer Arising From Pancreatic Cysts.

OBJECTIVE: We report the development and validation of a combined DNA/RNA next-generation sequencing (NGS) platform to improve the evaluation of pancreatic cysts. BACKGROUND AND AIMS: Despite a multidisciplinary approach, pancreatic cyst classification, such as a cystic precursor neoplasm, and the detection of high-grade dysplasia and early adenocarcinoma (advanced neoplasia) can be challenging. NGS of preoperative pancreatic cyst fluid improves the clinical evaluation of pancreatic cysts, but the recent identification of novel genomic alterations necessitates the creation of a comprehensive panel and the development of a genomic classifier to integrate the complex molecular results. METHODS: An updated and unique 74-gene DNA/RNA-targeted NGS panel (PancreaSeq Genomic Classifier) was created to evaluate 5 classes of genomic alterations to include gene mutations (e.g., KRAS, GNAS, etc.), gene fusions and gene expression. Further, CEA mRNA ( CEACAM5 ) was integrated into the assay using RT-qPCR. Separate multi-institutional cohorts for training (n=108) and validation (n=77) were tested, and diagnostic performance was compared to clinical, imaging, cytopathologic, and guideline data. RESULTS: Upon creation of a genomic classifier system, PancreaSeq GC yielded a 95% sensitivity and 100% specificity for a cystic precursor neoplasm, and the sensitivity and specificity for advanced neoplasia were 82% and 100%, respectively. Associated symptoms, cyst size, duct dilatation, a mural nodule, increasing cyst size, and malignant cytopathology had lower sensitivities (41-59%) and lower specificities (56-96%) for advanced neoplasia. This test also increased the sensitivity of current pancreatic cyst guidelines (IAP/Fukuoka and AGA) by >10% and maintained their inherent specificity. CONCLUSIONS: PancreaSeq GC was not only accurate in predicting pancreatic cyst type and advanced neoplasia but also improved the sensitivity of current pancreatic cyst guidelines.

Molecular Profiling of 50 734 Bethesda III-VI Thyroid Nodules by ThyroSeq v3: Implications for Personalized Management.

CONTEXT: Comprehensive genomic analysis of thyroid nodules for multiple classes of molecular alterations detected in a large series of fine needle aspiration (FNA) samples has not been reported. OBJECTIVE: To determine the prevalence of clinically relevant molecular alterations in Bethesda categories III-VI (BCIII-VI) thyroid nodules. METHODS: This retrospective analysis of FNA samples, tested by ThyroSeq v3 using Genomic Classifier and Cancer Risk Classifier at UPMC Molecular and Genomic Pathology laboratory, analyzed the prevalence of diagnostic, prognostic, and targetable genetic alterations in a total of 50 734 BCIII-VI nodules from 48 225 patients. RESULTS: Among 50 734 informative FNA samples, 65.3% were test-negative, 33.9% positive, 0.2% positive for medullary carcinoma, and 0.6% positive for parathyroid. The benign call rate in BCIII-IV nodules was 68%. Among test-positive samples, 73.3% had mutations, 11.3% gene fusions, and 10.8% isolated copy number alterations. Comparing BCIII-IV nodules with BCV-VI nodules revealed a shift from predominantly RAS-like alterations to BRAF V600E-like alterations and fusions involving receptor tyrosine kinases (RTK). Using ThyroSeq Cancer Risk Classifier, a high-risk profile, which typically included TERT or TP53 mutations, was found in 6% of samples, more frequently BCV-VI. RNA-Seq confirmed ThyroSeq detection of novel RTK fusions in 98.9% of cases. CONCLUSION: In this series, 68% of BCIII-IV nodules were classified as negative by ThyroSeq, potentially preventing diagnostic surgery in this subset of patients. Specific genetic alterations were detected in most BCV-VI nodules, with a higher prevalence of BRAF and TERT mutations and targetable gene fusions compared to BCIII-IV nodules, offering prognostic and therapeutic information for patient management.

Molecular Testing Predicts Incomplete Response to Initial Therapy in Differentiated Thyroid Carcinoma Without Lateral Neck or Distant Metastasis at Presentation: Retrospective Cohort Study.

Background: Molecular testing (MT) is emerging as a potential prognostic factor that can be available before treatment of differentiated thyroid carcinoma begins. Among patients eligible for either lobectomy or total thyroidectomy as their initial therapy, our study aims were to assess (1) if conventionally available preoperative factors are associated with incomplete response to initial therapy, and (2) if MT results can be a surrogate for the ATA Risk Stratification System (RSS) to estimate risk of recurrence. Methods: The data of consecutive thyroid cancer patients without preoperative lateral neck disease or distant metastasis who underwent index thyroidectomy between November 1, 2017 and October 31, 2021 were reviewed. Logistic regression models including preoperative variables such as MT and/or the postoperatively available RSS were constructed to predict disease recurrence, either structural or biochemical. Model discrimination using the c-statistic and goodness-of-fit test were compared. Results: Among 945 patients studied, 50 (5.2%) recurred with 18-month median follow-up. Recurrences were detected in 17 (2.9%), 20 (6.7%), and 13 (22.8%) patients with RSS-low, -intermediate, and -high cancers, respectively (p < 0.001). In multivariable analysis, only tumor size was associated with recurrence (odds ratio [OR] 1.3, 95% confidence interval [CI] 1.1-1.5). In a different model analyzing 440 (46.6%) patients with available MT results, recurrence was associated with both larger tumor size (OR 1.4 [95% CI 1.1-1.8]) and MT results (p < 0.001). Including MT improved the c-statistic by 27%, which was statistically no different than the model incorporating only the RSS (p = 0.15). Conclusions: Disease recurrence was observed across all ATA RSS categories in short-term follow-up, and tumor size was the only conventional preoperative factor associated with recurrence. When MT results were incorporated, they not only improved predictive ability beyond tumor size alone, but also yielded similar ability as the gold standard ATA RSS. Thus, MT results might aid the development of novel preoperative risk stratification algorithms.

A thyroid EIF1AX story: how clinical, cytologic, and molecular surveillance led to appropriate management.

INTRODUCTION: Indeterminate thyroid cytology diagnoses are associated with intermediate risks of malignancy. Application of molecular testing (MT) to indeterminate specimens provides additional diagnostic and prognostic information. While a positive or suspicious MT result may prompt surgery, a negative MT result is associated with a low probability of cancer or noninvasive follicular thyroid neoplasm with papillary-like nuclear features and approximates that of a benign cytology diagnosis. Furthermore, ThyroSeq v3 MT has a "currently negative" result for findings with the probability of cancer or noninvasive follicular thyroid neoplasm with papillary-like nuclear feature that is slightly greater than that for the negative ThyroSeq v3 MT result but less than 10%, suggesting active surveillance. In this report, we discuss a case of a patient for whom clinical, cytologic, and molecular surveillance led to timely surgery and management. CLINICAL DETAILS: A 53-year-old man with a thyroid isthmus nodule had a fine-needle aspiration cytology diagnosis of atypia of undetermined significance and a subsequent ThyroSeq v3 MT, which revealed an EIF1AX mutation and a "currently negative" MT result. Surveillance with additional fine-needle aspiration samples demonstrated concerning genomic alterations (fluctuating EIF1AX allelic frequency and a non-V600E BRAF mutation), culminating in the conversion to a positive MT result 3 years later. Resection revealed an encapsulated noninvasive, oncocytic solid subtype of papillary thyroid carcinoma with increased mitotic activity. CONCLUSION: The case is notable for clinical, cytologic, and molecular surveillance demonstrating sequential pathologic alterations in an indeterminate thyroid nodule with EIF1AX mutation, leading to timely resection of the neoplasm before invasion manifested.

Association of comprehensive thyroid cancer molecular profiling with tumor phenotype and cancer-specific outcomes.

BACKGROUND: Molecular testing improves the diagnostic accuracy of thyroid cancer. Whether specific molecular testing results are associated with tumor phenotype or provide prognostic information needs further delineation. METHODS: Consecutive thyroid cancer patients after index thyroidectomy with ThyroSeq version 3 (Rye Brook, NY) molecular testing obtained on preoperative fine-needle aspiration or thyroidectomy specimens from patients with thyroid cancer were categorized into 3 molecular risk groups based on detected mutations, fusions, copy number alterations, and/or gene expression alterations and correlated with histopathology and recurrence, defined as biochemical or structural. RESULTS: Of 578 patients, 49.9%, 37.5%, and 12.6% had molecular risk group-low, molecular risk group-intermediate, and molecular risk group-high cancers, respectively. With a median 19-month follow-up, 9.1% patients recurred. Compared with molecular risk group-low, molecular risk group-intermediate cancers were diagnosed in younger patients and more often had microscopic extrathyroidal extension, involved margins, and nodal disease. Compared with molecular risk group-intermediate, molecular risk group-high cancers were diagnosed in older patients and more often had gross extrathyroidal extension and vascular invasion. In multivariable analysis, recurrence was more likely in molecular risk group-high cancers than in molecular risk group-intermediate (hazard ratio = 4.0; 95% confidence interval, 1.9-8.6; P < .001) and more likely in molecular risk group-intermediate than in molecular risk group-low (hazard ratio = 5.0; 95% confidence interval, 2.0-12.5; P < .001). CONCLUSION: Using modern comprehensive genotyping, the genetic profile of thyroid cancers can be categorized into 3 novel molecular risk groups that were associated with histopathologic phenotype and recurrence in short-term follow-up.

Identifying metastatic renal cell carcinoma in thyroid fine-needle aspirates by molecular testing.

Renal cell carcinoma (RCC) is the most common type of cancer found to metastasize to the thyroid gland. These tumors may represent a diagnostic challenge in cytology. However, most RCC tumors carry VHL alterations, which are rare in primary thyroid tumors. The aim of this study was to evaluate the utility of molecular testing in detecting metastatic RCC in thyroid fine-needle aspiration (FNA) samples. From November 2017 until March 2022, thyroid FNA samples with ThyroSeq v3 results showing both VHL alterations and low/absent expression of thyroid cell markers were analyzed. Eighteen samples from 15 patients met the inclusion criteria. On molecular analysis, deleterious VHL mutations were found in nine (50%) nodules, VHL copy number alteration (CNA) in two (11%), and both mutations and CNA in seven (39%). None of the cases showed mutations commonly found in thyroid tumors. The mean age of these patients was 68 (range, 49-89) years with a male to female ratio of 2:1. Eight (53%) patients had multiple thyroid nodules on ultrasound. On cytology, 14 (78%) nodules were diagnosed as Bethesda III, 2 (11%) as Bethesda IV, and 2 (11%) as Bethesda V. At the time of cytology review, the history of RCC, sometimes remote, was available for ten patients. Of the 14 patients with medical history or surgical follow-up available, all had history of RCC or renal mass or revealed metastatic RCC on thyroidectomy. This study demonstrates that molecular testing can reliably identify metastatic RCC in thyroid nodules with indeterminate cytology, which could improve patient management.

Prognostic of recurrence and survival in poorly differentiated thyroid cancer.

The prognosis of poorly differentiated thyroid carcinomas (PDTC) defined by the Turin criteria is variable. The aim of this study on 51 PDTC patients was to determine clinical, histological and molecular prognostic factors associated with recurrence in patients with localized disease at initial treatment and with overall survival in patients with distant metastases. Of 40 patients for whom next-generation sequencing (NGS) by ThyroSeq v3 was able to be performed on historical samples, we identified high-risk molecular signature (TERT, TP53 mutations) in 24 (60%) cases, intermediate risk signature in 9 (22.5%) cases and low-risk signature in 7 (17.5%) cases. Potentially actionable mutations were identified in 10% of cases. After a median follow-up of 57.5 months, recurrence occurred in 11 (39%) of the 28 patients with localized disease. The American Thyroid Association (ATA) high risk of relapse, high mitotic count, high molecular risk signature and CD163 expression were associated with recurrence (P = 0.009, 0.01, 0.049, 0.03 respectively). After a median follow-up of 49.5 months, thyroid cancer-related death occurred in 53% of the patients with distant metastases. There was no significant prognostic factor associated with death in univariate analysis. However, none of the patients with intermediate ATA risk of recurrence and none of the patients with low-risk molecular signature died from the disease. In addition, high molecular-risk signature was associated with the presence of synchronous or metachronous distant metastasis (P = 0.007) and with poor overall survival (P = 0.01). In conclusion, ATA risk of relapse and high mitotic count was associated with higher rate of recurrence in localized PDTC. High molecular-risk signature was associated with the presence of distant metastasis and poor overall survival. Further studies are needed to determine if molecular testing adds to ATA risk stratification or response to therapy in predicting outcomes.

Performance of a Multigene Genomic Classifier in Thyroid Nodules with Suspicious for Malignancy Cytology.

Background: Molecular testing is increasingly used to refine the probability of cancer and assess recurrence risk in thyroid nodules with Bethesda III/IV fine needle aspiration (FNA) cytology. However, limited data exist for Bethesda V (suspicious for malignancy [SFM]) samples. This study evaluated the performance of ThyroSeq v3 (TSv3) in thyroid nodules with SFM cytology. Methods: In this single-institution retrospective cohort study, consecutive thyroid FNA samples diagnosed as SFM with TSv3 testing and known surgical outcome were identified. Clinical, pathology, and molecular findings were reviewed. The TSv3 Cancer Risk Classifier was used to determine molecular risk groups (MRGs). For test-negative cases diagnosed as cancer/noninvasive follicular thyroid neoplasm with papillary-like nuclear features, TSv3 was performed on the resected tumors. Results: Among 128 SFM samples studied, 100 (78.1%) were TSv3 positive, and 28 (21.9%) were negative. The cancer prevalence on surgery was 82.8%. Among test-positive samples, 95% were malignant and 5% benign. Among test-negative samples, 17 (60.7%) were benign and 11 (39.3%) malignant. Overall, TSv3 had a sensitivity of 89.6% (confidence interval; CI 82.4-94.1) and a specificity of 77.3% (CI 56.6-89.9). For a cancer prevalence of 50-75% expected in SFM cytology by the Bethesda system, the negative predictive value was expected to range from 71.2% to 88.1% and the positive predictive value from 79.8% to 92.2%. Among test-positive nodules, 20% were MRG-Low (mostly RAS-like alterations), 66% MRG-Intermediate (mostly BRAF-like alterations), and 14% MRG-High. Among patients with cancer, 65 (61.3%) were American Thyroid Association low risk, 25 (23.6%) intermediate risk, and 6 (5.7%) high risk. During the mean follow-up of 51.2 months (range: <1 to 470 months), 12 (13.0%) patients had disease recurrence, which was more common in MRG-High (54.6%) compared with MRG-Intermediate (9.5%) and MRG-Low (0%) cancers (p < 0.001). Upon reexamining tumors with false-negative results, half of evaluable cases had alterations likely missed due to limiting FNA sampling, and the remainder represented low-risk tumors. Potentially targetable alterations were identified in 10 samples. Conclusions: In this large series of SFM thyroid nodules, TSv3 further improved cancer prediction and detected RAS-like, BRAF-like, high-risk, and potentially targetable alterations, all of which may inform more optimal patient management. MRGs were associated with recurrence-free survival, offering potential preoperative cancer risk stratification.

Clinicopathologic Characteristics and Postsurgical Follow-Up of Noninvasive Follicular Thyroid Neoplasm with Papillary-Like Nuclear Features in the Postnomenclature Revision Era.

Background: Noninvasive encapsulated follicular variant papillary thyroid carcinoma (EFVPTC) was reclassified as "noninvasive follicular thyroid neoplasm with papillary-like nuclear features" (NIFTP) in 2016. Most existing studies that examined outcomes included patients managed as EFVPTC and only retrospectively reclassified as NIFTP. This is the first study to evaluate the clinicopathologic, molecular, and surveillance characteristics of patients diagnosed with NIFTP at the time of surgery and managed based on this diagnosis. Methods: We performed a retrospective cohort study of consecutive cases diagnosed as NIFTP from June 2016 to October 2021 identified from electronic medical records at a large tertiary care institution. Patients with coexisting low-risk thyroid cancers ≥1.0 cm in size or any size aggressive histology were excluded, and review of demographic, clinical, imaging, cytologic, and molecular genetic data was performed. Initial care was delivered according to existing clinical guidelines, with a consensus institutional plan for five-year follow-up after surgery. Results: Among 79 patients with 84 nodules diagnosed as NIFTP after surgery, 83.5% (66/79) were women and the mean age was 51 years (range, 21-84). Mean NIFTP size was 2.4 cm (range 0.15-8.0). On ultrasound, the majority of nodules were categorized as thyroid imaging, reporting and data system TI-RADS 3 (55.3%, 42/76), and TI-RADS 4 (36.8%, 28/76). On cytology, they were typically diagnosed as Bethesda III (69.1%, 47/68) or Bethesda IV (23.5%, 16/68). Molecular testing was performed on 62 nodules, and molecular alterations were found in 93.5% (58/62). The most common alterations identified in NIFTP were RAS mutation (75.4%, 43/57), THADA fusion (12.3%, 7/57), and BRAFK601E mutation (7.0%, 4/57). Fifty-two (65.8%) patients underwent lobectomy and 27 (34.2%) total thyroidectomy, and no patient received completion thyroidectomy. Twenty-one patients (26.5%) had coexisting papillary or follicular microcarcinoma. None of the patients received radioiodine ablation. On a mean follow-up of 28.5 months (range, 6-69 months), no structural or biochemical recurrences were observed. Conclusions: In this large cohort of patients with NIFTP diagnosed at the time of surgery and managed typically by lobectomy with no radioiodine ablation, no evidence of tumor recurrence was identified on a limited follow-up. This finding supports indolent clinical course of NIFTP.

Clinicopathologic features of thyroid nodules with PTEN mutations on preoperative testing.

The incidence of cancer in thyroid nodules carrying germline or somatic phosphatase and tensin homolog (PTEN) mutations is not well-defined. This study characterizes the clinical and histopathologic features of thyroid nodules with preoperatively detected PTEN mutations and their impact on management. Thyroid nodules with PTEN mutations on molecular testing of fine-needle aspiration (FNA) specimens from November 2017 to July 2020 at our institution were included. Demographic and clinicopathologic data were obtained through retrospective chart review. We identified 49 PTEN mutation-positive nodules from 48 patients. Surveillance was pursued for 28 patients and surgery for 20 patients. There were 14 follicular adenomas (FA), 4 oncocytic adenomas, 1 oncocytic hyperplastic nodule, and 1 encapsulated follicular variant papillary thyroid carcinoma (EFVPTC). The EFVPTC had two somatic PTEN mutations, an NRAS mutation, and was a low-risk tumor with capsular but no angiolymphatic invasion. Four patients, all with multiple nodules, had PTEN hamartoma syndrome (PHTS) with germline mutations or a clinical diagnosis of Cowden syndrome (CS); two had surgery finding FAs, and one previously had follicular carcinoma removed. Among surveillance patients, 1/20 had a significant increase in the size of the thyroid nodule and underwent repeat FNA, and no thyroid malignancy was found with a mean of 1.77 years of follow-up (range 1.00-2.78). Thyroid nodules with isolated somatic PTEN mutations are primarily benign and unlikely to grow at a high rate, at least on short-term follow-up. About 8% of patients with PTEN mutations may have PHTS or CS, which should be suspected in younger patients with multiple thyroid nodules.

Evaluation of the Molecular Landscape of Pediatric Thyroid Nodules and Use of a Multigene Genomic Classifier in Children.

Importance: Definitive diagnosis of a thyroid nodule in a child is obtained through diagnostic surgery. This is problematic because pediatric thyroid surgery is associated with higher rates of complications. In adults, preoperative molecular testing improves the management of thyroid nodules, but this has not been validated in children. Objective: To determine whether the molecular landscape of pediatric thyroid nodules is amenable to detection by a multigene genomic classifier (GC) test (ThyroSeq v3; Sonic Healthcare USA). Design, Setting, and Participants: This was a retrospective consecutive case series and GC testing of fine-needle aspiration (FNA) and formalin-fixed paraffin-embedded (FFPE) tissues from sequential pediatric thyroidectomies performed between January 2003 and December 2019 at a single tertiary academic medical center. The study included 95 patients (median [range] age, 16.3 [4.8 to 21.1] years; 75 [79%] female) who underwent surgery for a thyroid nodule. Interventions: A total of 118 thyroid nodule samples (95 FFPE, 23 companion FNAs) yielded informative next-generation sequencing data and multigene GC. Main Outcomes and Measures: The primary outcome was the determination of the pediatric thyroid molecular landscape. The secondary outcome was the diagnostic accuracy of the GC test for pediatric thyroid nodules. Results: Of the 95 patients, 75 (79%) were female, and the median (IQR) age was 16.3 (14.0-17.3) years. Next-generation sequencing confirmed the unique molecular landscape of malignant pediatric thyroid nodules (compared with adults), which is dominated by gene fusions (most commonly RET and NTRK), rare BRAF/RAS alterations, and no TP53 or TERT promoter pathogenic variants. Several poorly differentiated thyroid cancers harbored DICER1 variants. Benign nodules appeared to be almost exclusively associated with TSHR and DICER1 alterations. The test demonstrated a 96% sensitivity (95% CI, 87%-99%) and 78% specificity (95% CI, 64%-88%). The negative predictive value was 95% (95% CI, 88%-98%) and the positive predictive value was 83% (95% CI, 74-89%). The concordance of GC between 23 pairs of matched FFPE and FNA tissues was 96%. Conclusions and Relevance: The study results of this retrospective consecutive case series suggest that the molecular landscape of pediatric nodules is unique but remains amenable to molecular classification. The multigene GC test, with high sensitivity and reasonably high specificity, represents a potential addition to the diagnostic workup of children with thyroid nodules and may decrease the use of diagnostic surgery.

Clinicopathological features and outcomes of thyroid nodules with EIF1AX mutations.

EIF1AX gene mutations are reported in both benign and malignant thyroid tumors, with unclear outcomes when detected preoperatively. The aim of this study was to determine the features and outcomes of thyroid nodules with various types of mutation identified in cytologic (fine-needle aspiration) samples on preoperative ThyroSeq testing and with surgical outcomes. In this single-institution retrospective study of 31 consecutive patients, 77% were female and nodule size ranged from 1.5 to 9.4 cm with widely varying cytologic and TI-RADS ultrasound categorizations. Among two main mutational hotspots, 55% were located in exon 2 and 45% at the intron 5/exon 6 splice site. On histology, 45% of -positive nodules were cancer/noninvasive follicular thyroid neoplasm with papillary-like nuclear features (NIFTP) including 19% encapsulated follicular variant papillary thyroid carcinoma, 10% follicular carcinoma, 10% anaplastic carcinoma (ATC), and 7% NIFTP. Almost half (48%) of patients had one or more coexisting mutations, most frequently RAS. The prevalence of cancer/NIFTP was 80% for mutation with coexisting molecular alteration vs 13% with an isolated mutation (P = 0.0002). Cancer probability was associated with mutation type and was 64% for splice-site mutation and 29% for non-splice mutation (P = 0.075). All 3 nodules with EIF1AX+RAS+TERT+TP53 mutations were ATC. In summary, in this study, all nodules with an isolated non-splice mutation were benign, one-third of those with an isolated splice mutation were cancer, and most nodules with coexisting with RAS or other alterations were malignant. These findings suggest that clinical management decisions for patients with EIF1AX-mutant nodules should consider both the type of mutation and its co-occurrence with other genetic alterations.

Molecular profiling of papillary thyroid carcinomas in healthcare workers exposed to low dose radiation at the workplace.

PURPOSE: Exposure to ionizing radiation, especially during childhood, is a well-established risk factor for thyroid cancer. The vast majority of radiation-induced cancers are papillary carcinomas (PTCs). These tumors typically have gene fusions in contrast to point mutations prevalent in sporadic PTCs. The aim of this study was to investigate the molecular profiles of PTC patients with workplace exposure to ionizing radiation. METHODS: A retrospective review of 543 patients who underwent surgery with diagnosis of PTC was performed. A cohort of nine healthcare specialists previously exposed to radiation sources during their professional practice was selected and analyzed using the ThyroSeq mutation panel for point mutations and gene fusions associated with thyroid cancer. RESULTS: The molecular analysis of surgical samples of PTCs was informative and revealed genetic alterations in five patients. BRAF V600E was found in four (67%) cases whereas RET/PTC1 fusion in one (17%) and one sample (17%) was wild type for point mutations and fusions. One sample completely failed molecular analysis while two others were negative for genes fusions but failed DNA analysis; these three samples were excluded. CONCLUSIONS: In this limited cohort of healthcare workers exposed to low dose of ionizing radiation at the workplace and developed PTC, the molecular profiling determined BRAF V600E point mutation as the most common event, arguing against the role of workplace radiation exposure in the etiology of these tumors.

Non-functional pancreatic neuroendocrine tumours: ATRX/DAXX and alternative lengthening of telomeres (ALT) are prognostically independent from ARX/PDX1 expression and tumour size.

OBJECTIVE: Recent studies have found aristaless-related homeobox gene (ARX)/pancreatic and duodenal homeobox 1 (PDX1), alpha-thalassemia/mental retardation X-linked (ATRX)/death domain-associated protein (DAXX) and alternative lengthening of telomeres (ALT) to be promising prognostic biomarkers for non-functional pancreatic neuroendocrine tumours (NF-PanNETs). However, they have not been comprehensively evaluated, especially among small NF-PanNETs (≤2.0 cm). Moreover, their status in neuroendocrine tumours (NETs) from other sites remains unknown. DESIGN: An international cohort of 1322 NETs was evaluated by immunolabelling for ARX/PDX1 and ATRX/DAXX, and telomere-specific fluorescence in situ hybridisation for ALT. This cohort included 561 primary NF-PanNETs, 107 NF-PanNET metastases and 654 primary, non-pancreatic non-functional NETs and NET metastases. The results were correlated with numerous clinicopathological features including relapse-free survival (RFS). RESULTS: ATRX/DAXX loss and ALT were associated with several adverse prognostic findings and distant metastasis/recurrence (p<0.001). The 5-year RFS rates for patients with ATRX/DAXX-negative and ALT-positive NF-PanNETs were 40% and 42% as compared with 85% and 86% for wild-type NF-PanNETs (p<0.001 and p<0.001). Shorter 5-year RFS rates for ≤2.0 cm NF-PanNETs patients were also seen with ATRX/DAXX loss (65% vs 92%, p=0.003) and ALT (60% vs 93%, p<0.001). By multivariate analysis, ATRX/DAXX and ALT status were independent prognostic factors for RFS. Conversely, classifying NF-PanNETs by ARX/PDX1 expression did not independently correlate with RFS. Except for 4% of pulmonary carcinoids, ATRX/DAXX loss and ALT were only identified in primary (25% and 29%) and NF-PanNET metastases (62% and 71%). CONCLUSIONS: ATRX/DAXX and ALT should be considered in the prognostic evaluation of NF-PanNETs including ≤2.0 cm tumours, and are highly specific for pancreatic origin among NET metastases of unknown primary.