Current State of Molecular Cytology in Thyroid Nodules: Platforms and Their Diagnostic and Theranostic Utility.

The high prevalence of thyroid nodules and increased availability of neck ultrasound have led to an increased incidence of diagnostic thyroid fine needle aspirations, with approximately 20% yielding indeterminate results. The recent availability of molecular tests has helped guide the clinical management of these cases. This paper aims to review and compare three main commercially available molecular cytology platforms in the U.S.-Afirma GSC, Thyroseq GC, and ThyGeNEXT + ThyraMIR. Sequential improvements of the Afirma GSC and Thyroseq GC tests have increased positive and negative predictive values, sensitivity, and specificity. Comparative studies revealed similar diagnostic performance between these tests, with considerations for factors such as cost and processing time. Thyroseq GC provides detailed genomic information and specific management recommendations. ThyGeNEXT + ThyraMIR, though less studied, presents promising results, particularly in miRNA analysis for weak driver mutations. Challenges in interpreting results include variations in reporting and the evolving nature of testing platforms. Questions persist regarding cost-effectiveness and the utility of ultrasound characteristics in selecting candidates for molecular testing. While molecular testing has primarily served diagnostic purposes, advancements in understanding genetic alterations now offer therapeutic implications. FDA-approved options target specific genetic alterations, signaling a promising future for tailored treatments.

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.

Novel DIO1 Gene Mutation Acting as Phenotype Modifier for Novel Compound Heterozygous TPO Gene Mutations Causing Congenital Hypothyroidism.

A family with congenital hypothyroidism was identified with two novel deleterious compound heterozygous thyroid peroxidase (TPO) mutations (c.962C>A, and c.1577C>T). Serum thyroid tests showed higher-than-expected serum-free thyroxine (T4) relative to TT3, while reverse triiodothyronine (rT3) was also elevated. Two siblings manifested a more severe phenotype of developmental delay compared with another sibling and were found to harbor an additional novel heterozygous deleterious iodothyronine deiodinase 1 (DIO1) mutation (c.395G>A). In the context of L-T4 replacement, the decreased D1 activity results in abnormal thyroid hormone metabolism with decreased triiodothyronine (T3) generation from L-T4 and may result in decreased T3 bioavailability during critical stages of development.

Anti-Ma-1 and Anti-Ma-2 Antibodies in Isolated Fatal Hypothalamitis.

Lymphocytic hypothalamitis (LHT) is a rare disease characterized by pituitary dysfunction, autonomic instability, metabolic disturbances, and neuropsychiatric disorders. We report the case of a 30-year-old man found to have LHT that progressed despite treatment with methylprednisolone and intravenous immunoglobulin (IVIG). A literature review was conducted to identify prior studies pertaining to LHT. Our patient presented with several weeks of fatigue, cold intolerance, weight loss, confusion, and headache. Laboratory tests were consistent with panhypopituitarism. Brain magnetic resonance imaging revealed an infiltrative enhancing mass in the area of the hypothalamus, and stereotactic biopsy findings showed reactive inflammatory changes. A course of hormone replacement (levothyroxine), methylprednisolone, and IVIG was initiated. The patient's course was complicated by a fatal tonsillar herniation. Autopsy revealed LHT. The diagnosis and management of autoimmune LHT are challenging. The entity should be considered in the setting of panhypopituitarism with a hypothalamic mass. Differentiating paraneoplastic and nonparaneoplastic hypothalamitis should be established for management and prognosis. Definitive treatment remains unclear; treatment with corticosteroids should be attempted, followed by consideration of other immunosuppressive agents, such as rituximab. If a paraneoplastic syndrome is suspected, management should also be directed toward the primary tumor.

BIOCHEMICAL CONTROL DURING LONG-TERM FOLLOW-UP OF 230 ADULT PATIENTS WITH CUSHING DISEASE: A MULTICENTER RETROSPECTIVE STUDY.

OBJECTIVE: Cushing disease (CD) results from excessive exposure to glucocorticoids caused by an adrenocorticotropic hormone-secreting pituitary tumor. Inadequately treated CD is associated with significant morbidity and elevated mortality. Multicenter data on CD patients treated in routine clinical practice are needed to assess treatment outcomes in this rare disorder. The study purpose was to describe the burden of illness and treatment outcomes for CD patients. METHODS: Eight pituitary centers in four U.S. regions participated in this multicenter retrospective chart review study. Subjects were CD patients diagnosed at ≥18 years of age within the past 20 years. Descriptive statistical analyses were conducted to examine presenting signs, symptoms, comorbidities, and treatment outcomes. RESULTS: Of 230 patients, 79% were female (median age at diagnosis, 39 years; range, 18 to 78 years). Length of follow-up was 0 to 27.5 years (median, 1.9 years). Pituitary adenomas were 0 to 51 mm. The most common presenting comorbidities included hypertension (67.3%), polycystic ovary syndrome (43.5%), and hyperlipidemia (41.5%). Biochemical control was achieved with initial pituitary surgery in 41.4% patients (91 of 220), not achieved in 50.0% of patients (110 of 220), and undetermined in 8.6% of patients (19 of 220). At the end of follow-up, control had been achieved with a variety of treatment methods in 49.1% of patients (110 of 224), not achieved in 29.9% of patients (67 of 224), and undetermined in 21.0% of patients (47 of 224). CONCLUSION: Despite multiple treatments, at the end of follow-up, biochemical control was still not achieved in up to 30% of patients. These multicenter data demonstrate that in routine clinical practice, initial and long-term control is not achieved in a substantial number of patients with CD. ABBREVIATIONS: BLA = bilateral adrenalectomy CD = Cushing disease CS = Cushing syndrome eCRF = electronic case report form MRI = magnetic resonance imaging PCOS = polycystic ovary syndrome.

Defects of Thyroid Hormone Synthesis and Action.

Congenital hypothyroidism (CH) is the most common inborn endocrine disorder and causes significant morbidity. To date, we are only aware of the molecular basis responsible for the defects in a small portion of patients with CH. A better understanding of the pathophysiology of these cases at the genetic and molecular basis provides useful information for proper counseling to patients and their families a well as for the development of better targeted therapies. This article provides a succinct outline of the pathophysiology and genetics of the known causes of thyroid dysgenesis, dyshormonogenesis, and syndrome of impaired sensitivity to thyroid hormone.

A NOVEL MUTATION CAUSING COMPLETE THYROID BINDING GLOBULIN DEFICIENCY (TBG-CD MIA) IN A MALE WITH COEXISTING GRAVES DISEASE.

OBJECTIVE: An asymptomatic male was found on screening to have a low serum TSH and total T4. The diagnosis of Graves' disease was made with positive thyroid stimulating immunoglobulin (TSI) and elevated free T4 in the presence of complete TBG deficiency (TBG-CD). Genetic testing of the patient and family members revealed a novel frameshift mutation in the TBG (SERPINA7) gene resulting in a complete deficiency of the protein. METHODS: The laboratory testing included total T4, free T4 by analog method and direct dialysis and TBG measurements. Sequencing of genomic DNA was performed from peripheral blood. RESULTS: A 35-year-old East Indian male was referred to endocrinology because of abnormal thyroid function tests (TFTs): TSH 0.01 mIU/L (0.4-3.6), total T4 3.0 µg/dl (5.5-10.5) done as part of a "routine office visit". Upon further testing, the serum free T4 2.0 ng/dl (0.8-1.8) and TSI 355% (<140% baseline) were elevated and the diagnosis of Graves' disease was made. TBG deficiency was suspected because the total T4 concentration was inconsistent with hyperthyroidism and further testing confirmed TBG was undetectable. Sequencing of the TBG gene revealed a novel hemizygous frameshift mutation: p.Ala64ProfsTer106, TBG-CD Mia (numbering excludes 20 a.a. signal peptide) associated with the complete deficiency of TBG in a patient with Graves' disease. CONCLUSION: Patients with Graves' disease harboring a TBG mutation have conflicting TFTs. If a clinically hyperthyroid patient presents with normal or low total T4, serum TBG should be measured to identify an abnormality and prevent unnecessary testing.

False Positive Findings on I-131 WBS and SPECT/CT in Patients with History of Thyroid Cancer: Case Series.

Introduction. Although whole body scan (WBS) with I-131 is a highly sensitive tool for detecting normal thyroid tissue and metastasis of differentiated thyroid cancer (DTC), it is not specific. Additional information, provided by single photon emission computed tomography combined with X-ray computed tomography (SPECT/CT) and by the serum thyroglobulin level, is extremely useful for the interpretation of findings. Case Presentation. We report four cases of false positive WBS in patients with DTC: ovarian uptake corresponding to an endometrioma, scrotal uptake due to a spermatocele, rib-cage uptake due to an old fracture, and hepatic and renal uptake secondary to a granuloma and simple cyst, respectively. Conclusions. Trapping, organification, and storage of iodine are more prominent in thyroid tissue but not specific. Physiologic sodium-iodine symporter expression in other tissues explains some, but not all, of the WBS false positive cases. Other proposed etiologies are accumulation of radioiodine in inflamed organs, metabolism of radiodinated thyroid hormone, presence of radioiodine in body fluids, and contamination. In our cases nonthyroidal pathologies were suspected since the imaging findings were not corroborated by an elevated thyroglobulin level, which is considered a reliable tumor marker for most well-differentiated thyroid cancers. Clinicians should be aware of the potential pitfalls of WBS in DTC to avoid incorrect management.

New JAG1 Mutation Causing Alagille Syndrome Presenting With Severe Hypercholesterolemia: Case Report With Emphasis on Genetics and Lipid Abnormalities.

Context: Alagille syndrome is a rare autosomal-dominant genetic disorder caused by defects in the Notch signaling pathway, which involves multiple organ systems. Bile duct paucity is the main characteristic feature of the disease, which often leads to cholestatic hypercholesterolemia. Case Description: We report the case of a male infant who had developed failure to thrive, jaundice, intermittent pruritus, and multiple diffuse symmetrical skin xanthomas at 1 year of age. He was diagnosed with pulmonary stenosis, butterfly vertebrae of T4, T6, and T8; horseshoe kidney, and embryotoxon in the left eye. Laboratory workup revealed severe hypercholesterolemia. Alagille syndrome was suspected and confirmed by genetic testing, which identified a previously undescribed frameshift pathogenic heterozygous variant in the JAG1 gene, p.Arg486Lysfs*5. Conclusions: Here, we report a unique case of a patient diagnosed with Alagille syndrome who was found to have a previously undescribed frameshift pathogenic mutation in the JAG1 gene and who presented with xanthomatosis and levels of hypercholesterolemia more than 2 times higher than those previously reported in the literature. We also provide a review of the different pathophysiologic mechanisms associated with the increase in serum cholesterol and low-density lipoprotein cholesterol concentrations seen in cholestatic liver disease in general and in Alagille syndrome in particular.

Thyroid Hormone Replacement in Patients Following Thyroidectomy for Thyroid Cancer.

Thyroid hormone replacement therapy in patients following thyroidectomy for thyroid cancer, although a potentially straightforward clinical problem, can present the clinician and patient with a variety of challenges. Most often the problems are related to the dose and preparation of thyroid hormone (TH) to use. Some patients feel less well following thyroidectomy and/or radioiodine ablation than they did before their diagnosis. We present evidence that levothyroxine (L-T4) is the preparation of choice, and keeping the thyroid-stimulating hormone (TSH) between detectable and 0.1 mU/L should be the standard of care in most cases. In unusual circumstances, when the patient remains clinically hypothyroid despite a suppressed TSH, we acknowledge there may be as yet unidentified factors influencing the body's response to TH, and individualized therapy may be necessary in such patients.

A Novel non-sense Mutation in Keratin 10 Causes a Familial Case of Recessive Epidermolytic Ichthyosis.

Epidermolytic ichthyosis (EI) is a rare skin disorder characterized by generalized erythroderma and cutaneous blistering at birth, which is substituted by hyperkeratosis later in life. It is caused by autosomal dominant mutations in highly conserved regions of KRT1 and KRT10. To date, only 4 mutations with autosomal recessive inheritance of EI have been described in consanguineous families. All of them affect the 2B domain of KRT10. In the present study we describe four patients with EI (including one lethal case) born from unaffected parents in a consanguineous family of a native Venezuelan community. The objective of this study was to characterize the clinical, genetic and morphological aspects of the disease in this family, as well as understand its functional implications. Genomic DNA was sequenced for KRT10 and KRT1. Immunofluoresence for keratin expression was performed on cutaneous biopsies. After examination of cutaneous biopsies histology, our results showed hyperkeratosis and acantholysis with an expanded granular layer. Sequencing of KRT10 demonstrated a non-sense mutation (p.Tyr282Ter.) corresponding to the 1B domain of the protein in patients and a heterozygous pattern in other family members, resulting in complete absence of K10. The loss of K10 was compensated by upregulation of K14 and K17. In conclusion, this novel mutation in KRT10 is the first recessive genetic variation that is not located in the so called "hot spot" for recessive EI, suggesting that other areas of the gene are also susceptible for such mutations.