Stefan Karger

FOXO3a: a novel player in thyroid carcinogenesis?

The forkhead box transcription factor FOXO3a has recently been identified as central mediator of the cellular response to oxidative stress inducing cell cycle arrest or apoptosis. The aim of our study was to investigate the regulation of FOXO3a in the thyroid and to determine whether alterations in FOXO3a activity occur in thyroid carcinogenesis. In vitro, we demonstrate that FOXO3a activity is negatively regulated by the PI3K/Akt cascade promoting increased phosphorylation and cytoplasmatic accumulation of FOXO3a with decreased transcription of the target genes p27kip (CDKN1B) and Bim (BCL2L11), but increased expression of GADD45A. By contrast, we show that H(2)O(2) exposure activates FOXO3a in thyrocytes with JNK (MAPK8)-mediated nuclear accumulation of FOXO3a and increased expression of the cell cycle arrest genes p27kip and GADD45A. In vivo, we observed a marked cytoplasmatic accumulation of FOXO3a in differentiated thyroid cancers versus an exclusive nuclear accumulation in follicular adenoma and normal thyroid tissue. Moreover, this cytosolic accumulation of FOXO3a correlated with an increased phospho-Akt expression in thyroid malignancies and was accompanied by decreased expression of the FOXO targets p27kip and Bim and an increase in GADD45A mRNA expression in the thyroid cancers. Our data suggest FOXO3a as a novel player of cellular stress response in the thyroid, mediating the thyrocytes fate either to survive or to undergo apoptosis. Furthermore, PI3K-dependent FOXO3a inactivation may be a novel pathomechanism for the escape from apoptosis in thyroid cancer cells, in particular in follicular thyroid carcinoma.

Forkhead box-O transcription factor: critical conductors of cancer's fate

Cells have evolved elaborated mechanisms to coordinate the cellular answer of either survival or apoptosis. Recent concepts of human carcinogenesis have suggested disturbances in these cellular relays as a potential link to cellular dedifferentiation and uncontrolled proliferation. Forkhead box-O transcription factors (FOXOs) play an important role in tumour suppression by regulating the expression of genes involved in stress resistance, DNA damage repair, cell cycle arrest and apoptosis. The specific regulation of FOXO function is tightly controlled by posttranslational modifications such as phosphorylation, acetylation and ubiquitination. Loss of FOXO function has recently been identified in several human cancers. In this review, we will give an overview about recent progress in the understanding of function and regulation of FOXOs, as well as their role in carcinogenesis. Furthermore, we will discuss a potential clinical use of FOXOs by therapeutically restoring their tumour suppressive properties.

TFF3-Based Candidate Gene Discrimination of Benign and Malignant Thyroid Tumors in a Region with Borderline Iodine Deficiency

BACKGROUND With the advent of microarray technology, increasing numbers of marker genes are proposed to distinguish benign and malignant thyroid lesions. However, most markers await confirmation through independent studies. In this paper, we re-evaluate the diagnostic potential of 10 proposed candidate genes in benign and malignant thyroid pathologies in a region with borderline iodine deficiency. METHODS Quantitative real-time PCR was performed for CCND2, PLAB, PCSK2, HGD1, TFF3, B4GALT, LGALS3, ETS1, ADM3, and TG in 150 thyroid specimens, including 52 benign thyroid nodules (28 follicular adenoma and 24 adenomatous nodules), 52 corresponding normal thyroid tissues, 20 follicular carcinomas, 20 papillary carcinomas, and six undifferentiated carcinomas. RESULTS On a single-gene basis, significant differences in mRNA expression were found for TFF3, PLAB, and ADM3 in benign thyroid nodules and thyroid malignancy. Using two-marker gene sets, we identified 11 combinations, which allowed both a distinction of benign and malignant thyroid nodules and a discrimination of follicular adenoma and carcinoma. However, for cancer prediction, analysis of a minimum of six genes per sample was necessary and allowed correct prediction of a benign thyroid lesion and thyroid cancer with 94% accuracy in the most discriminative set (TFF3/PLAB/TG/ADM3/HGD1/LGALS3). CONCLUSION We confirm the applicability of a number of recently proposed marker genes for the distinction of benign and malignant thyroid tumor and suggest that their diagnostic usefulness is independent of the iodide supply. We propose that the most discriminative marker set identified in our validation study together with marker combinations proposed by other investigators should now be evaluated in multicenter trials.

Patient-reported outcomes in adequately treated hypothyroidism: Insights from the German versions of ThyDQoL, ThySRQ and ThyTSQ

BackgroundDisease-specific patient-reported outcome measures (PROMs) have been developed as important research tools in the study of various diseases. For hypothyroidism there exist three validated disease-specific questionnaires in English: the Thyroid-Dependent Quality of Life Questionnaire (ThyDQoL), the Underactive Thyroid Symptom Rating Questionnaire (ThySRQ) and the Thyroid Treatment Satisfaction Questionnaire (ThyTSQ). We report psychometric properties of new German versions of the questionnaires including construct validity from two independent samples.Methods230 envelopes with ThyDQoL, ThySRQ and ThyTSQ were given out to patients receiving levothyroxine for diagnosed hypothyroidism. Reliability and factor analyses were performed, correlations and hypothesised subgroup differences calculated to assess psychometric properties. Independently, 18 patients with treated hypothyroidism for autoimmune thyroiditis (Hashimoto’s disease) and 18 healthy control subjects were enrolled in a clinical study. Participants filled in the above questionnaires alongside well-known generic PROMs, e.g. the Beck Depression Inventory, the 12-item Well-Being Questionnaire and the Short-Form-36. Two blood samples were taken. Groups were compared and correlations between disease-specific and generic instruments analysed. Relationships between PROMs and biochemically determined thyroid hormone status were investigated.Results102 patients returned completed questionnaires (response rate 44%). The newly translated questionnaires had satisfactory psychometric properties. Cronbach’s alpha was 0.92 for ThyDQoL, 0.81 for ThySRQ and 0.86 for ThyTSQ. For each of the questionnaires, a single factor structure explained the data best. Adequately treated patients with thyroid stimulating hormone levels in the upper normal range reported more symptoms in the ThySRQ. Those with autoimmune hypothyroidism reported being more bothered by depressive symptoms. Within the clinical sample, correlation with well-known generic instruments revealed good construct validity. In the clinical sample patients reported more symptoms in the ThySRQ, being more bothered by tiredness, higher depression and reduced well-being despite biochemically adequate treatment. Correlations between PROMs and biochemical thyroid hormone status revealed moderate though consistent associations.ConclusionsPsychometric properties including construct validity of German versions of the ThyDQoL, ThySRQ and ThyTSQ are satisfactory. Feasibility and sensitivity in a clinical sample could be shown. We encourage the use of disease-specific PROMs in future studies as important additions to generic instruments in clinical research on hypothyroidism.ZusammenfassungHintergrundKrankheitsspezifische Selbsteinschätzungsfragebögen, so genannte „patient-reported outcome measures“(PROMs), wurden als wichtige Forschungsinstrumente für verschiedene Erkrankungen entwickelt. Für die Hypothyreose wurden drei englischsprachige Fragebögen entwickelt: der Lebensqualitätfragebogen zur Schilddrüsenunterfunktion (ThyDQoL), der Symptomfragebogen zur Schilddrüsenunterfunktion (ThySRQ) sowie ein Fragebogen zur Behandlungszufriedenheit bei Schilddrüsenunterfunktion (ThyTSQ). Wir stellen psychometrische Eigenschaften der neuen deutschen Versionen dieser Fragebögen vor, inklusive Daten zur Konstruktvalidität aus zwei unabhängigen Stichproben.Methoden230 Umschläge mit ThyDQoL, ThySRQ und ThyTSQ wurden an Patienten ausgegeben, die wegen einer gesicherten Hypothyreose mit Levothyroxin substituiert wurden. Mit Hilfe von Reliabilitäts- und Faktoranalysen, Korrelationsanalysen und hypothesengeleiteten Subgruppenvergleichen wurden die psychometrischen Eigenschaften der Fragebögen analysiert. 18 Patienten mit behandelter Hypothyreose autoimmuner Genese (Hashimoto Thyreoiditis) sowie 18 gesunde Kontrollprobanden nahmen an einer unabhängigen klinischen Studie teil. Die Teilnehmer füllten zu den oben genannten auch bekannte generische PROMs aus, darunter das Beck Depressions Inventar, den 12-Item Well-Being Questionnaire und den Short Form-36. Zwei Blutproben wurden entnommen. Die Gruppen wurden verglichen und Korrelationen zwischen krankheitsspezifischen und generischen Instrumenten sowie Zusammenhänge zwischen Fragebogendaten und Laborwerten berechnet.Ergebnisse102 Patienten schickten ausgefüllte Fragebögen an die Autoren zurück (Rücklaufquote 44%). Die neu übersetzten Fragebögen zeigten gute psychometrische Eigenschaften. Cronbach’s Alpha betrug 0.92 für den ThyDQoL, 0.81 für den ThySRQ und 0.86 für den ThyTSQ. Alle Fragebögen wurden am besten durch eine Einfaktorenlösung beschrieben. Patienten mit Thyroidea-stimulierendem Hormon (TSH) im oberen Normbereich berichteten mehr Symptome im ThySRQ und diejenigen mit autoimmuner Genese stärkere Beeinträchtigung durch depressive Symptome. Die Konstruktvalidität der Fragebögen konnte durch erwartungskonforme Korrelationen zu generischen Fragebögen in der klinischen Studie nachgewiesen werden. Die Patientengruppe berichtete trotz normwertigem TSH mehr Symptome im ThySRQ, mehr Beeinträchtigung durch Müdigkeit, sowie höhere Depressionswerte und geringeres Wohlbefinden als die gesunde Kontrollgruppe. Wir konnten moderate aber konsistente Zusammenhänge zwischen den Fragebogendaten und den Laborwerten finden.SchlussfolgerungDie psychometrischen Eigenschaften der Fragebögen ThyDQol, ThySRQ und ThyTSQ sowie ihre Konstruktvalidität sind zufriedenstellend. Machbarkeit und Sensitivität der Fragebögen in einer klinischen Studie konnten gezeigt werden. Die krankheitsspezifischen PROMs stellen eine wichtige Ergänzung zu den generischen Instrumenten dar, mit potentiellem Nutzen für die zukünftige klinische Forschung auf dem Gebiet der Hypothyreose.

Impact of pregnancy on prevalence of goitre and nodular thyroid disease in women living in a region of borderline sufficient iodine supply.

An interplay of genetic, epigenetic, and environmental factors contributes to thyroid disease. In a cross-sectional study, we aimed to determine the influence of parity in combination with other risk factors on the prevalence of goitre and nodular thyroid disease (NTD) in women living in a region of previous overt iodine deficiency, which experienced a continuous improvement in alimentary iodine supply in the last two decades. Thyroid ultrasonography (7.5 MHz; Merck Thyromobil) was performed by the same investigator in 736 women living in Thuringia and Saxony. Age and BMI were documented and a comprehensive history on pregnancies, family history of thyroid disease, and past or present smoking was obtained. Goitre prevalence was 19.1%. Solitary thyroid nodules were detected in 21.5%, and multiple nodules in 23.8% of women. In a multivariate analysis, neither age nor parity was positively correlated with goitre prevalence and NTD. A significant correlation was detected between BMI and goitre and multinodular disease. Goitre was found in 25.3% of women with a positive family history for thyroid disease, as opposed to 16.1% goitre in women with a negative family history. Neither goitre nor NTD were associated with a history of smoking in the whole study population. Thyroid nodules and/or goitre are present in up to 45% of women in an area of previous overt iodine deficiency. Whereas BMI and family history are positively correlated with the presence of NTD and goitre, no such correlation could be detected for pregnancy and smoking after processing our data with multivariate analyses.

Distinct pattern of oxidative DNA damage and DNA repair in follicular thyroid tumours

Increased oxidative stress has been linked to thyroid carcinogenesis. In this paper, we investigate whether oxidative DNA damage and DNA repair differ in follicular adenoma (FA) and follicular thyroid carcinoma (FTC). 7,8-Dihydro-8-oxoguanine (8-OxoG) formation was analysed by immunohistochemistry in 46 FAs, 52 FTCs and 18 normal thyroid tissues (NTs). mRNA expression of DNA repair genes OGG1, Mut Y homologue (MUTYH) and endonuclease III (NTHL1) was analysed by real-time PCR in 19 FAs, 25 FTCs and 19 NTs. Induction and repair of oxidative DNA damage were studied in rat FRTL-5 cells after u.v. irradiation. Moreover, activation of DNA damage checkpoints (ataxia telangiectasia mutated (ATM) and H2A histone family, member X (H2AFX (H2AFX))) and proliferation index (MIB-1) were quantified in 28 non-oxyphilic and 24 oxyphilic FTCs. Increased nuclear and cytosolic 8-OxoG formation was detected in FTC compared with follicular adenoma, whereby cytosolic 8-OxoG formation was found to reflect RNA oxidation. Significant downregulation of DNA repair enzymes was detected in FTC compared with FA. In vitro experiments mirrored the findings in FTC with oxidative stress-induced DNA checkpoint activation and downregulation of OGG1, MUTYH and NTHL1 in FRTL-5 cells, an effect that, however, was reversible after 24  h. Further analysis of FTC variants showed decreased oxidative DNA damage, sustained checkpoint activation and decreased proliferation in oxyphilic vs non-oxyphilic FTC. Our data suggest a pathophysiological scenario of accumulating unrepaired DNA/RNA damage in FTC vs counterbalanced DNA/RNA damage and repair in FA. Furthermore, this study provides the first evidence for differences in oxidative stress defence in FTC variants with possible implications for therapeutic response and prognostic outcome.

Structural and functional MRI study of the brain, cognition and mood in long-term adequately treated Hashimoto's thyroiditis

The current study investigated neuropsychological and underlying structural and functional brain alterations in long-term adequately treated patients with Hashimotos thyroiditis in order to examine much discussed residual complaints in patients in relation to possible long-term neural alterations with a specific interest in the underlying autoimmune process. Eighteen patients with treated hypothyroidism due to Hashimotos thyroiditis (mean age 32, range 18-54 years; two males; mean treatment duration 4.4 years) and 18 healthy matched control subjects underwent 3-Tesla magnetic resonance imaging (MRI). Voxel-based morphometry was used to investigate grey matter density, resting-state functional MRI to analyse the brain connectivity of areas known to be altered in hypothyroidism and event-related functional MRI to examine brain activity during associative memory encoding. Neuropsychological assessment included memory, working memory, psychomotor speed and attention. We previously reported subclinically reduced mood in this study population and investigated its neural correlates here. Thyroid stimulating hormone, free triiodthyronine, free thyroxine and thyroid peroxidase antibodies were measured in serum. We did not find cognitive deficits or alterations in grey matter density, functional connectivity or associative memory-related brain activity in comparison to the control group and cognition was unrelated to thyroid serum measures in the patient group. Thyroid peroxidase antibodies in the patient group correlated with increased grey matter density in right amygdala and enhanced connectivity between subcallosal and parahippocampal areas. Treatment duration was associated with brain structure in frontal and occipital cortex and connectivity between left amygdala and frontal cortex. Mood correlated with brain areas associated with distinct functional networks, but not with those most prominently affected in depression. In conclusion, no cognitive or neural alterations were detected in this young and otherwise healthy cohort of patients in comparison to a healthy control group and current mood status could not be related to depression-related networks. However, autoimmune activity and treatment duration showed a relationship with depression and hypothyroidism-related brain structure and function. They are thus promising factors to further investigate residual complaints despite biochemically adequate treatment in patients with Hashimotos thyroiditis. Given the small sample size, all findings require replication.

Schilddrüsenerkrankungen und Schwangerschaft

Thyroid diseases in pregnancy must be recognized as a specific challenge for the clinician. Any pregnancy is causing alterations in thyroid hormone metabolism which have to be differentiated from pathologic states of thyroid function. Any thyroid disease of the mother with disturbances in the functional state of the gland could induce an adverse influence on the course of pregnancy. Furthermore, it can be associated with adverse consequences on fetal development. Especially hypothyroidism has to be avoided during pregnancy due to a danger of affected neurocognitive development of the offspring. Yet also maternal hyperthyroidism can lead to impairments in the course of pregnancy and to fetal thyroid dysfunction. Further clinical attention should be given to thyroid autoimmunity. There is a clear relationship between autoimmune thyroid disease and decreased fertility and an increase in the rate of spontaneous miscarriages. Furthermore, it displays an increased risk for the manifestation of postpartum thyroiditis. The management of nodular thyroid disease and malignancy does not differ from that of nonpregnant women/patients. Thyroid scintiscan and radioiodine therapy must be avoided during pregnancy and lactation. This review deals with the broad variety of thyroid disorders and function disturbances during and after pregnancy. All described diagnostic and therapeutic procedures are based upon the recent Clinical Practice Guideline of the Endocrine Society published in August 2007.ZusammenfassungDer Umgang mit Schilddrüsenerkrankungen während der Schwangerschaft stellt eine besondere klinische Herausforderung dar. Jede Schwangerschaft verursacht physiologische Veränderungen des Schilddrüsenhormonmetabolismus, die von pathologischen Funktionszuständen abgegrenzt werden müssen. Eine Schilddrüsenerkrankung der Mutter mit tatsächlicher Störung der Schilddrüsenfunktion kann einen nachteiligen Einfluss auf den Schwangerschaftsverlauf ausüben und negative Auswirkungen auf die Entwicklung des Fetus bedingen. Insbesondere die Hypothyreose ist während einer Schwangerschaft wegen beeinträchtigter neurokognitiver Entwicklung des Ungeborenen unter allen Umständen zu vermeiden. Auch eine mütterliche Hyperthyreose kann zur Beeinträchtigung des Schwangerschaftsverlaufs und zur fetalen Schilddrüsendysfunktion führen.Von klinischer Bedeutung sind per se auch Autoimmunerkrankungen der Schilddrüse, da sie Ursache von Fertilitätsstörungen und einer erhöhten Rate von Spontanaborten sein können und ein erhöhtes Risiko für die Entwicklung einer Post-partum-Thyreoiditis darstellen.Das Management von Schilddrüsenknoten einschließlich Malignität unterscheidet sich nur unwesentlich vom Vorgehen beim allgemeinen Patienten. Nuklearmedizinische Diagnostik und Therapie verbieten sich allerdings während der Schwangerschaft und ebenso in der Stillperiode.Dieser Übersichtsartikel befasst sich mit den unterschiedlichen Facetten thyreoidaler Erkrankungen und Funktionsstörungen in und nach der Schwangerschaft. Die hier dargestellten diagnostischen und therapeutischen Vorgehensweisen basieren auf der aktuellen Clinical Practice Guideline der Endocrine Society vom August 2007.AbstractThyroid diseases in pregnancy must be recognized as a specific challenge for the clinician. Any pregnancy is causing alterations in thyroid hormone metabolism which have to be differentiated from pathologic states of thyroid function. Any thyroid disease of the mother with disturbances in the functional state of the gland could induce an adverse influence on the course of pregnancy. Furthermore, it can be associated with adverse consequences on fetal development. Especially hypothyroidism has to be avoided during pregnancy due to a danger of affected neurocognitive development of the offspring. Yet also maternal hyperthyroidism can lead to impairments in the course of pregnancy and to fetal thyroid dysfunction.Further clinical attention should be given to thyroid autoimmunity. There is a clear relationship between autoimmune thyroid disease and decreased fertility and an increase in the rate of spontaneous miscarriages. Furthermore, it displays an increased risk for the manifestation of postpartum thyroiditis.The management of nodular thyroid disease and malignancy does not differ from that of nonpregnant women/patients. Thyroid scintiscan and radioiodine therapy must be avoided during pregnancy and lactation.This review deals with the broad variety of thyroid disorders and function disturbances during and after pregnancy. All described diagnostic and therapeutic procedures are based upon the recent Clinical Practice Guideline of the Endocrine Society published in August 2007.

United States and European Multicenter Prospective Study for the Analytical Performance and Clinical Validation of a Novel Sensitive Fully Automated Immunoassay for Calcitonin

BACKGROUND The objective of this study is the validation and proof of clinical relevance of a novel electrochemiluminescence immunoassay (ECLIA) for the determination of serum calcitonin (CT) in patients with medullary thyroid carcinoma (MTC) and in different diseases of the thyroid and of calcium homeostasis. METHODS This was a multicenter prospective study on basal serum CT concentrations performed in 9 US and European referral institutions. In addition, stimulated CT concentrations were measured in 50 healthy volunteers after intravenous calcium administration (2.5 mg/kg bodyweight). RESULTS In total, 1929 patients and healthy controls were included. Limits of blank, detection, and quantification for the ECLIA were 0.3, 0.5, and 1 ng/L, respectively. Highest intra- and interassay coefficients of variation were 7.4% (CT concentration, 0.8 ng/L) and 7.0% (1.1 ng/L), respectively. Medians (interval) of serum CT concentrations in 783 healthy controls were 0.8 ng/L (<0.5-12.7) and 3 ng/L (<0.5-18) for females and males, respectively (97.5th percentile, 6.8 and 11.6 ng/L, respectively). Diagnostic sensitivity and specificity were 100%/97.1% and 96.2%/96.4%, for female/males, respectively. Patients (male/female) with primary hyperparathyroidism, renal failure, and neuroendocrine tumors showed CT concentrations >97.5th percentile in 33%/4.7%, 18.5%/10%, and 8.3%/12%, females/males, respectively. Peak serum CT concentrations were reached 2 min after calcium administration (161.7 and 111.8 ng/L in males and females, respectively; P < 0.001). CONCLUSIONS Excellent analytical performance, low interindividual variability, and low impact of confounders for increased CT concentrations in non-MTC patients indicate that the investigated assay has appropriate clinical utility. Calcium-stimulated CT results suggest good test applicability owing to low interindividual variability.

Der Schilddrüsenknoten – Differentialdiagnostik und Therapiekonzepte

KasuistikEin 25-jähriger Mann stellt sich in unserer endokrinologischen Ambulanz zur weiteren Abklärung eines vom Hausarzt per Palpation diagnostizierten Schilddrüsen-(SD-)Knotens vor. Der Patient berichtet von einem unspezifischen Druckgefühl rechts paralaryngeal, welches er vor etwa einem 1/2 Jahr erstmals bemerkt hat. Weitere Beschwerden werden nicht angegeben, insbesondere kein Gewichtsverlust und kein Nachtschweiß. Der Stuhlgang sei unauffällig. Der Patient wirkt ruhig und ausgeglichen, der Puls ist normofrequent.In der klinischen Untersuchung fällt bereits bei rekliniertem Hals eine rechtsseitige Vorwölbung neben dem Kehlkopf auf. Diese ist nicht druckdolent und weist während des Schluckakts eine gute Verschieblichkeit auf. In der Sonographie der SD zeigt sich neben einem unauffälligen linken SD-Lappen ein nahezu lappenausfüllender, echoarmer, teils zystisch durchsetzter Knoten mit schmalem Halosaum im rechten SD-Lappen. Kalzifikationen lassen sich nicht nachweisen, die Halslymphknoten erscheinen beidseits unauffällig.Laborchemisch erfolgt die Bestimmung des thyroideastimulierenden Hormons (TSH) sowie des Calcitonins. Hier zeigen sich eine Euthyreose bei normwertigem TSH (1,9 mU/l) sowie ein unauffälliger Calcitoninwert.Auf eine Szintigraphie wird bei laborchemischem Ausschluss einer funktionellen SD-Autonomie verzichtet.Aufgrund der Größe des Knotens wird dem Patienten eine weiterführende Diagnostik mittels Feinnadelaspiration empfohlen. In der gewonnenen Zytologie zeigen sich unauffällig strukturierte Thyreozytencluster mit z.T. regressiven Veränderungen. Hinweise auf Malignität ergeben sich nicht. Wegen der lokalen Beschwerdesymptomatik sowie der Sorge des Patienten um eine zukünftige maligne Entartung des Knotens veranlassen wir eine Hemithyreoidektomie rechts. Die Histologie bestätigt den benignen Befund der Feinnadelaspirationszytologie mit Diagnose eines regressiv veränderten Kolloidknotens mit zystischer Degeneration.Der Patient kommt weiterhin ohne eine SD-Hormon-Substitutionsbehandlung aus. Es erfolgt die Empfehlung, auf eine jodreiche Ernährung zu achten und eine jährliche SD-Kontrolle mittels TSH-Bestimmung und Sonographie vornehmen zu lassen.