Unravelling pituitary tumours in medically treated patients with acromegaly: the impact of systematic MRI reassessment.

BACKGROUND: In acromegaly, the primary tumor is usually found during magnetic resonance imaging (MRI) of the pituitary gland. A remnant tumor after surgery is, however, harder to depict. When a tumor is missed, the remaining option is usually lifelong pharmacological treatment. PURPOSE: To identify tumors by reassessment of all available MRI scans in pharmacologically treated patients, operated or not, and to compare our results with the routine MRI reports. MATERIAL AND METHODS: Adult patients diagnosed with acromegaly and managed at a tertiary care center between 2005 and 2021 and currently on pharmacological treatment were included. MRI scans were evaluated in a standardized manner and classified independently by a radiologist and an endocrinologist into "certain," "suspected," or "no tumor." In case of disagreement, consensus was achieved with a senior neuroradiologist. The results were compared using the clinical radiologists' routine MRI reports. RESULTS: We identified certain and suspected tumors in 29/74 and 36/74 patients, respectively. No tumor was identified in nine patients. In five of these, no MRI contrast agent was given. Discrepancy between our results and the routine MRI reports was found in 31/74 patients (P = 0.01). In 22 patients, the routine reports described no tumor while we identified certain tumors in 2/22 patients and suspected tumors in 13/22 patients. CONCLUSION: In most patients with pharmacologically treated acromegaly, we identified a certain or suspected pituitary tumor. These findings were more frequent compared to the routine MRI reports. Based on our results, patients will be considered for a change in long-term treatment modality.

TGFBR3L is associated with gonadotropin production in non-functioning gonadotroph pituitary neuroendocrine tumours.

PURPOSE: Transforming growth factor-beta receptor 3-like (TGFBR3L) is a pituitary enriched membrane protein selectively detected in gonadotroph cells. TGFBR3L is named after transforming growth factor-beta receptor 3 (TGFBR3), an inhibin A co-receptor in mice, due to sequence identity to the C-terminal region. We aimed to characterize TGFBR3L detection in a well-characterized, prospectively collected cohort of non-functioning pituitary neuroendocrine tumours (NF-PitNETs) and correlate it to clinical data. METHODS: 144 patients operated for clinically NF-PitNETs were included. Clinical, radiological and biochemical data were recorded. Immunohistochemical (IHC) staining for FSHß and LHß was scored using the immunoreactive score (IRS), TGFBR3L and TGFBR3 were scored by the percentage of positive stained cells. RESULTS: TGFBR3L staining was selectively present in 52% of gonadotroph tumours. TGFBR3L was associated to IRS of LHß (median 2 [IQR 0-3] in TGFBR3L negative and median 6 [IQR 3-9] in TGFBR3L positive tumours, p < 0.001), but not to the IRS of FSHß (p = 0.32). The presence of TGFBR3L was negatively associated with plasma gonadotropin concentrations in males (P-FSH median 5.5 IU/L [IQR 2.9-9.6] and median 3.0 [IQR 1.8-5.6] in TGFBR3L negative and positive tumours respectively, p = 0.008) and P-LH (median 2.8 IU/L [IQR 1.9-3.7] and median 1.8 [IQR 1.1-3.0] in TGFBR3L negative and positive tumours respectively, p = 0.03). TGFBR3 stained positive in 22% (n = 25) of gonadotroph tumours with no correlation to TGFBR3L. CONCLUSION: TGFBR3L was selectively detected in half (52%) of gonadotroph NF-PitNETs. The association to LHß staining and plasma gonadotropins suggests that TGFBR3L may be involved in hormone production in gonadotroph NF-PitNETs.

Distribution of E- and N-cadherin in subgroups of non-functioning pituitary neuroendocrine tumours.

PURPOSE: Clinically non-functioning pituitary neuroendocrine tumours (NF-PitNETs) present a varying degree of aggressiveness, and reliable prognostic markers are lacking. We aimed to characterise the distribution of E- and N-cadherin in corticotroph, PIT1 and null-cell NF-PitNETs, and link it to the course of the tumours. METHODS: The distribution of E- and N-cadherin was investigated by immunohistochemistry in a retrospective cohort of 30 tumours of the less common NF-PitNETs (corticotroph (N = 18), PIT1 (N = 8) and null-cell PitNETs (N = 4)). Immunoreactive scores (IRS) were compared to previously presented cohorts of gonadotroph NF-PitNETs (N = 105) and corticotroph functioning PitNETs (N = 17). RESULTS: We found a low IRS for the extra-cellular domain of E-cadherin (median 0 (IQR 0-0, N = 135)), a medium to high IRS for the intra-cellular domain of E-cadherin (median 6 (IQR 4-9)) and a high IRS for N-cadherin (median 12 (IQR 10.5-12)) throughout the cohort of NF-PitNETs. The corticotroph NF-PitNETs presented a higher IRS for both the extra- and intra-cellular domain of E-cadherin (median 0 (IQR 0-1) and median 9 (IQR 6-12), respectively) than the gonadotroph NF-PitNETs (p < 0.001 for both comparisons). Presence of nuclear E-cadherin was associated with a weaker staining for the intra-cellular domain of E-cadherin (median 4 (IQR 0.5-6) and median 9 (IQR 9-12), for tumours with and without nuclear E-cadherin, respectively), and with a lower rate of re-intervention (p = 0.03). CONCLUSIONS: Considering our results and the benign course of NF-PitNETs, we suggest that a high N-cadherin and downregulation of membranous E-cadherin are not associated with a more aggressive tumour behaviour in these subgroups of NF-PitNETs.

Feasibility of a Digital Patient-Provider Communication Intervention to Support Shared Decision-Making in Chronic Health Care, InvolveMe: Pilot Study.

BACKGROUND: Enhanced communication with health care providers (HCPs) can improve symptom management and health-related quality of life (HRQoL) for patients with chronic health conditions. Access to appropriate communication venues is needed to improve communication, however. As such, digital communication interventions mediated by patient portals carry the potential to support patient-provider communication and interaction and through this, also facilitate shared decision-making (SDM). The InvolveMe intervention was designed to provide patients with the opportunity to communicate symptoms and informational needs prior to consultation via digital assessment, including prioritizing what is most important to discuss with their HCPs, as well as to interact with HCPs through secure messages between outpatient visits. OBJECTIVE: The aim of this study was to assess the feasibility of the InvolveMe intervention by investigating acceptability, demand (ie, system use), and limited efficacy. METHODS: The study was designed as a single-arm, pre-post feasibility study combining quantitative and qualitative methods for data collection. Patients from an endocrine outpatient clinic were invited to use the InvolveMe intervention for 3 months, and HCPs administering InvolveMe were invited to participate in a focus group. Guided by descriptions of how to design feasibility studies by Bowen et al, feasibility was tested by exploring (1) acceptability, using data collected during recruitment from patient participants and nonparticipants (ie, declined to participate or did not meet study requirements), HCP experiences with recruitment, and the System Usability Scale (SUS); (2) demand via exploration of system use through extraction of system log data and HCP experiences with system use; and (3) limited efficacy testing, via exploration of potential effects from the Short-Form Health Survey (RAND 36), Hospital Anxiety and Depression Scale, and Health Literacy Questionnaire. RESULTS: Patient participants (N=23) were a median 54 (range 26-78) years old and primarily male (14/23, 61%). Nonparticipants (N=16) were a median 73 (range 55-80) years old and primarily male (12/16, 75%). The average SUS score was 72.2, indicating good system usability. Assessments were completed by 8 participants from home prior to outpatient visits. The assessments entailed various bodily symptoms and needs for information. Participants sent 17 secure messages related to patient administrative matters, symptoms, and challenges. Focus group participants (N=4) were all female and registered nurses. Data were analyzed in 2 predefined themes: Acceptability and Demand. Acceptability included the subthemes intervention attractiveness and intervention suitability. Demand included the subthemes elements of SDM and intervention challenges and opportunities. All patient participants completed outcome measures at baseline, and 19 (19/23, 83%) completed outcome measures at 3 months. These preliminary efficacy findings were mixed and inconclusive. CONCLUSIONS: The study design provided findings from both patient and HCP perspectives and supported feasibility of the InvolveMe intervention. The investigation of acceptability and demand supported the potential for remote SDM mediated by patient portals using assessments and secure messages. TRIAL REGISTRATION: ClinicalTrials.gov NCT NCT04218721; https://www.clinicaltrials.gov/ct2/show/NCT04218721.

A Digital Patient-Provider Communication Intervention (InvolveMe): Qualitative Study on the Implementation Preparation Based on Identified Facilitators and Barriers.

BACKGROUND: Chronic health conditions are affecting an increasing number of individuals, who experience various symptoms that decrease their quality of life. Digital communication interventions that enable patients to report their symptoms have been shown to positively impact chronic disease management by improving access to care, patient-provider communication, clinical outcomes, and health-related quality of life. These interventions have the potential to prepare patients and health care providers (HCPs) before visits and improve patient-provider communication. Despite the recent rapid development and increasing number of digital communication interventions that have shown positive research results, barriers to realizing the benefits offered through these types of interventions still exist. OBJECTIVE: The aim of this study is to prepare for the implementation of a digital patient-provider communication intervention in the daily workflow at 2 outpatient clinics by identifying potential determinants of implementation using the Consolidated Framework for Implementation Research (CFIR) to tailor the use of digital communication intervention to the intended context and identify key aspects for an implementation plan. METHODS: A combination of focus groups, workshops, and project steering committee meetings was conducted with HCPs (n=14) and patients (n=2) from 2 outpatient clinics at a university hospital. The CFIR was used to guide data collection and analysis. Transcripts, written minutes, and notes were analyzed and coded into 5 CFIR domains using thematic analysis. RESULTS: Data were examined and analyzed into 18 CFIR constructs relevant to the study purpose. On the basis of the identified determinants, important intervention tailoring includes adjustments to the digital features and adjustments to fit the clinical workflow and a decision to conduct a future pilot study. Furthermore, it was decided to provide the intervention to patients as early as possible in their disease trajectory, with tailored information about its use. Key aspects for the implementation plan encompassed maintaining the identified engagement and positive attitude, involving key stakeholders in the implementation process, and providing the needed support and training. CONCLUSIONS: This study offers insight into the involvement of stakeholders in the tailoring and implementation planning of a digital communication intervention in clinical practice. Stakeholder involvement in the identification of implementation facilitators and barriers can contribute to the tailoring of digital communication interventions and how they are used and can also inform systematic and targeted implementation planning.

Case report: a patient with thyroid storm, refractory cardiogenic shock, and cardiac arrest treated with Lugol's iodine solution and veno-arterial extra corporal membrane oxygenation support.

BACKGROUND: Thyroid storm is a life-threatening condition. Refractory cardiogenic shock and cardiac arrest are rare complications of thyroid storm and the treatment options are limited. CASE SUMMARY: A 35-year- old woman treated for Grave's disease was admitted with thyrotoxicosis complicated by infection and neutropenia caused by thionamide treatment. After treatment including beta-blockers, steroids, and Lugol's iodine solution, she went into cardiac arrest. Echocardiography after resuscitation demonstrated severe biventricular heart failure. The patient was in refractory cardiogenic shock with recurrent cardiac arrest and mechanical circulatory support with a veno-arterial extra corporal membrane oxygenation (V-A ECMO) circuit was established. After 2 days on V-A ECMO and supportive treatment with iodine solution, glucocorticosteroids, and levosimendan, her myocardial function recovered and thyroid hormone levels were normalized. Veno-arterial extra corporal membrane oxygenation was discontinued, and the patient was treated with early total thyroidectomy. The patient made a full recovery with no neurological/cognitive impairment, as assessed after 4 weeks. DISCUSSION: Adverse reactions to standard treatment of hyperthyroidism contributed to this patient's development of thyroid storm and the following refractory cardiogenic shock. When she was critically unstable, levosimendan improved myocardial function while inotropic support with dobutamine was ineffective, likely due to prolonged beta-antagonist administration. Temporary support with V-A ECMO, until effective lowering of thyroid hormone levels and improvement in myocardial function were obtained, was life-saving in this young patient and may be considered in refractory cardiogenic shock caused by thyroid storm.

Cut-off values for sufficient cortisol response to low dose Short Synacthen Test after surgery for non-functioning pituitary adenoma.

OBJECTIVE: The aim was to study the prevalence of secondary adrenal insufficiency before and after surgery for non-functioning pituitary adenomas, as well as determine risk factors for developing secondary adrenal insufficiency. A secondary aim was to determine adequate p-cortisol response to a 1-µg Short Synacthen Test after surgery. DESIGN: Longitudinal cohort study. METHODS: One hundred seventeen patients (52/65 females/males, age 59 years) undergoing primary surgery for clinically non-functioning pituitary adenomas were included. P-cortisol was measured in morning blood samples. Three months after surgery, a Short Synacthen Test was performed. RESULTS: All tumours were macroadenomas (mean size 26.9 mm, range 13-61 mm). The surgical indications were visual impairment (93), tumour growth (16), pituitary apoplexy (6) and headache (2). Before surgery, 17% of the patients had secondary adrenal insufficiency (SAI), decreasing to 15% 3 months postoperatively. Risk of SAI was increased in patients operated for pituitary apoplexy (p < 0.001), while age, sex, tumour size and complication rate were not different from the remaining cohort. Three months after surgery, all patients with baseline p-cortisol ≥ 172 nmol/l (6.2 µg/dl) and peak p-cortisol during Short Synacthen Test ≥ 320 nmol/l (11.6 µg/dl) tapered cortisone unproblematically. In patients with intact hypothalamic-pituitary-adrenal axis, p-cortisol peaked < 500 nmol/l (18.1 µg/dl) during Short Synacthen Test in 48% of patient. CONCLUSION: Pituitary surgery is safe and transsphenoidal surgery rarely causes new SAI. Relying solely on morning p-cortisol for diagnosing secondary adrenal insufficiency gives false positives and the Short Synacthen Test remains useful. A peak p-cortisol ≥ 320 during (11.6 µg/dl) Short Synacthen Test indicates a sufficient response, while < 309 nmol/l (11.2 µg/dl) indicates secondary adrenal insufficiency.

Hip Structure Analyses in Acromegaly: Decrease of Cortical Bone Thickness After Treatment: A Longitudinal Cohort Study.

Long-standing growth hormone (GH) excess causes the skeletal clinical signs of acromegaly with typical changes in bone geometry, including increased cortical bone thickness (CBT). However, a high prevalence and incidence of vertebral fractures has been reported. The aim of this study was to assess the course of cortical bone dimensions in the hip by comparing patients with acromegaly and clinically nonfunctioning pituitary adenomas (NFPAs) at baseline and 1 year after pituitary surgery (1-year PO) in a longitudinal cohort study. DXA was performed in patients with acromegaly (n = 56) and NFPA (n = 47). CBT in the femoral neck (CBTneck), calcar (CBTcalcar), and shaft (CBTshaft) were determined by hip structural analysis (HSA). CBT at baseline and the change to 1-year PO were compared. Test results were adjusted for differences in gender distribution, age, and gonadal status. Cortical thickness analyses showed higher values [mm] at baseline in patients with acromegaly compared with NFPA: CBTneck median [25th; 75th] 6.2 [4.7; 8.0] versus 5.1 [4.1; 6.4] (p = 0.006), CBTcalcar 4.8 [4.2, 5.7] versus 4.0 [3.2, 4.5] (p < 0.001), CBTshaft 6.2 [5.1, 7.2] versus 5.2 [4.6, 6.0], (p = 0.003). In acromegaly, GH was correlated with CBTneck (r = 0.31, p = 0.020), whereas IGF-1 was correlated with CBTcalcar (r = 0.39, p = 0.003) at baseline. In acromegaly, CBTneck decreased by 11.2%, p = 0.002 during follow-up. Finally, the decrease in CBTneck and CBTcalcar in acromegaly was significant compared with NFPA (p = 0.023 and p = 0.017, respectively). Previous observations of increased CBT in acromegaly were confirmed with DXA-derived HSA in a large, well-defined cohort. The decline in CBT in acromegaly could contribute to the increased fracture risk in acromegaly despite increased bone dimensions and disease control. © 2019 The Authors. JBMR Plus published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research.

Are volume measurements of non-functioning pituitary adenomas reliable?

PURPOSE: Precise radiological assessment of tumour volume is important in the follow-up of non-functioning pituitary adenomas (NFPAs). We compared the reliability of two methods for tumour volume measurements in the pre- and postoperative setting. METHODS: We assessed the volume of 22 NFPAs at magnetic resonance imaging (MRI) scans before surgery and the first and third postoperative MRI obtained after submission from hospital. Volumetric assessments were performed both by summation of slices (SOS) and by diameter measures. All volumes were calculated independently by two readers. RESULTS: The preoperative intra- and inter-rater reliability was good for both the SOS and the diameter method (intraclass correlation coefficient (ICC) 0.996 and 0.990, and ICC: 0.982 and 0.967, respectively). The first postoperative investigation showed poorer intra- and inter-rater reliability for both methods (ICC: 0.872 and 0.791 and ICC: 0.792 and 0.810, respectively). The third postoperative MRI showed good intra-rater reliability (ICC: 0.961 and 0.962, respectively), but poorer inter-rater reliability for both methods (ICC: 0.759 and 0.703, respectively). Volume assessment by SOS presented overall slightly higher reliability than the diametric method. Overall, the reliability between the two methods was good when measured by the same reader (ICC: 0.988, 0.945 and 0.962, for the preoperative, first and third postoperative MRI, respectively). CONCLUSION: The preoperative intra- and inter-rater reliabilities were satisfactory for both the SOS and diametric method. Postoperative MRI scans showed poorer reliability, suggesting that measurements at these time points should be interpreted with care. For each MRI scan, reliability between methods was satisfactory when investigated by the same reader.

The role of E and N-cadherin in the postoperative course of gonadotroph pituitary tumours.

PURPOSE: Gonadotroph tumours are the most abundant of the clinically silent pituitary tumours. There is a lack of reliable prognostic markers predicting their clinical course. Our aim was to determine the level of E-cadherin and N-cadherin in a cohort of clinically silent gonadotroph pituitary tumours, and compare them to the rate of reintervention. METHODS: Tumour tissue from primary surgery was retrospectively investigated and compared with clinical data. Immunohistochemical (N = 105) and real time-qPCR (N = 85) analyses for the levels of N-cadherin and the extra- and intracellular domains of E-cadherin were performed. The immunoreactive scores (IRS) and mRNA relative quantity were compared to the rate of reintervention. RESULTS: The tumours presented a high IRS for N-cadherin (Median 12 (IQR 12-12)) and almost no immunoreactivity for the extracellular domain of E-cadherin (Median 0 (IQR 0-0)). The membranous staining for the intracellular domain of E-cadherin varied (Median 6 (IQR 4-6). Reduced membranous expression of the intracellular domain of E-cadherin was associated with nuclear presence of the same domain. Nuclear staining for the intracellular domain of E-cadherin was associated with a lower rate of reintervention (p = 0.01). CONCLUSION: We found that silent gonadotroph tumours presented high IRS for N-cadherin and low IRS for the extracellular domain of E-cadherin. A substantial proportion of the tumours presented nuclear staining for the intracellular domain of E-cadherin, accompanied by a reduced membranous expression of the intracellular domain of E-cadherin. Absence of nuclear staining for the intracellular domain of E-cadherin served as an independent predictor of reintervention.

Gene expression profiling of fast- and slow-growing non-functioning gonadotroph pituitary adenomas.

OBJECTIVE: Reliable biomarkers associated with aggressiveness of non-functioning gonadotroph adenomas (GAs) are lacking. As the growth of tumor remnants is highly variable, molecular markers for growth potential prediction are necessary. We hypothesized that fast- and slow-growing GAs present different gene expression profiles and reliable biomarkers for tumor growth potential could be identified, focusing on the specific role of epithelial-mesenchymal transition (EMT). DESIGN AND METHODS: Eight GAs selected for RNA sequencing were equally divided into fast- and slow-growing group by the tumor volume doubling time (TVDT) median (27.75 months). Data were analyzed by tophat2, cufflinks and cummeRbund pipeline. 40 genes were selected for RT-qPCR validation in 20 GAs based on significance, fold-change and pathway analyses. The effect of silencing MTDH (metadherin) and EMCN (endomucin) on in vitro migration of human adenoma cells was evaluated. RESULTS: 350 genes were significantly differentially expressed (282 genes upregulated and 68 downregulated in the fast group, P-adjusted <0.05). Among 40 selected genes, 11 showed associations with TVDT (-0.669

Estrogen Receptor α, a Sex-Dependent Predictor of Aggressiveness in Nonfunctioning Pituitary Adenomas: SSTR and Sex Hormone Receptor Distribution in NFPA.

Context: Nonfunctioning pituitary adenomas (NFPAs) are fairly common and require a multidisciplinary approach. Reliable markers of a clinically aggressive course are lacking. Medical treatment is not available, and transsphenoidal surgery is the preferred primary treatment. Objective: We aimed to characterize the somatostatin, estrogen, and progesterone receptor distribution for NFPAs and compare it with factors of tumor aggressiveness. Design: Tumor samples for immunohistochemistry (n = 145) and quantitative reverse transcription polymerase chain reaction (n = 106) analyses of somatostatin receptor (SSTR) 1, SSTR2, SSTR3, SSTR5, estrogen receptor α (ERα), and progesterone receptor (PR) were measured by immunoreactive score (IRS) and messenger RNA relative quantity and retrospectively compared with variables of aggressiveness. Setting: All patients were operated at the same tertiary referral center. Participants: A total of 164 patients with NFPA and tumor tissue from the primary operation were included. Results: SSTR3 was expressed abundantly by immunohistochemistry in all NFPAs. The IRS of ERα correlated with that of SSTR2 in male patients only (males, P < 0.001; females, P = 0.8). Low ERα level was linked to a higher reintervention rate (P = 0.001) and earlier reintervention (P = 0.004) in male patients only (females, P = 0.95 and P = 0.65, respectively). Absence of ERα together with age provided a good prediction model for reintervention in male patients with gonadotroph adenomas. Conclusions: SSTR3 is expressed abundantly in NFPAs and is therefore a possible target for medical treatment. Absence of ERα together with young age may predict tumor recurrence in groups of NFPAs. Further validation in systematic prospective studies is needed.

Early postoperative growth in non-functioning pituitary adenomas; A tool to tailor safe follow-up.

PURPOSE: Non-functioning pituitary adenomas are common, and the treatment and follow-up of these patients represent a multidisciplinary challenge. First line treatment is transphenoidal surgery, with debulking or total removal of tumour. A substantial portion of the tumours relapse after surgery, and there is no consensus of how to follow these patients postoperatively. Our aim was to characterize the postoperative growth of non-functioning pituitary adenomas and correlate it to clinical and paraclinical data. METHODS: We retrospectively registered 52 patients operated for non-functioning pituitary adenomas, with four or more consecutive MR-investigations not interrupted by secondary treatment. Adenoma volumes were estimated by the Cavalieri principle with summation of manually drawn areas multiplied by slice interval. Growth curves were modelled and tumour volume doubling time was calculated for 39 tumours with regrowth after surgery. RESULTS: A total of 13 tumours showed exponential growth, 10 linear growth and 16 logistic growth after surgery. The remaining 13 did not show regrowth of tumour. Seven of the exponential growing tumours underwent secondary surgery, compared to one and two of linear and logistic growing tumours (p = 0.03), respectively. Initial tumour volume doubling time was significantly lower in logistic growing tumours than in exponential growing tumours (p < 0.01). Men had tumours with lower tumour volume doubling time than women (p = 0.03). None of the tumours demonstrated signs of accelerated growth. CONCLUSION: Residual tumours following surgery frequently grow. The logistic growing tumours had the fastest initial growth in our cohort. We found no indication of accelerated growth, whereby the tumour volume doubling time might be used to predict a "worst-case" scenario when planning follow-up of these patients.

Selection and validation of reliable reference genes for RT-qPCR analysis in a large cohort of pituitary adenomas.

BACKGROUND: Real-time reverse transcription quantitative PCR (RT-qPCR) has become the method of choice for quantification of gene expression changes. The most important limitations of RT-qPCR are inappropriate data normalization and inconsistent data analyses. Pituitary adenomas are common tumours, and the appropriate interpretation of increasingly published data within this field is prevented by the lack of a proper selection and validation of stably expressed reference genes. AIM: To find and validate the optimal reference gene or gene combination for reliable RT-qPCR gene expression in both non-functioning (NFPA) and hormone secreting (GH and ACTH) pituitary adenomas. MATERIAL AND METHODS: Thirty commonly used reference genes (PCR array reference gene panel, BioRad, Hercules, CA) were quantified by RT-qPCR in 24 pituitary adenomas (12 NFPA, 8 GH and 4 ACTH). The data was analysed using three programs: geNorm (Qbase+), Normfinder and BestKeeper having different algorithms to identify the most stable reference gene or combination of reference genes. Three reference genes ALAS1, PSMC4 and GAPDH, were selected for further validation in a larger cohort of 223 adenomas (141 NFPA, 63 GH and 19 ACTH). RESULTS: In all adenomas, ALAS1 and PSMC4 were the most stable reference genes as estimated by geNorm and Normfinder, whereas Bestkeeper ranked RPLP0 and ACTB as the two most stable out of 10 carefully selected genes. The best gene combination was PSMC4 and ALAS1 (geNorm) or PSMC4 and GAPDH (Normfinder). The validation experiment (geNorm) showed that the most stable gene combinations were ALAS1 and GAPDH in NFPA, and PSMC4 and GAPDH in hormone secreting adenomas. CONCLUSIONS: Several of the reference genes expressed good stability yielding several candidate genes. PSMC4 and ALAS1 were overall the most stably expressed genes in pituitary adenoma merely differing in ranking order. PSMC4 and ALAS1 have so far not been reported as reference genes in pituitary adenomas. The various reference gene algorithms showed a mixed selection of top ranked genes, thus suggesting a need for an individualised and rational choice of reference genes.

A high-throughput analysis of the IDH1(R132H) protein expression in pituitary adenomas.

PURPOSE: Inactivating mutations of isocitrate dehydrogenase (IDH) 1 and 2, mitochondrial enzymes participating in the Krebs tricarboxylic acid cycle play a role in the tumorigenesis of gliomas and also less frequently in acute myeloid leukemia and other malignancies. Inhibitors of mutant IDH1 and IDH2 may potentially be effective in the treatment of the IDH mutation driven tumors. Mutations in the succinate dehydrogenase, the other enzyme complex participating in the Krebs cycle and electron transfer of oxidative phosphorylation occur in the paragangliomas, gastrointestinal stromal tumors, and occasionally in the pituitary adenomas. We aimed to determine whether the IDH1(R132H) mutation, the most frequent IDH mutation in human malignancies, occurs in pituitary adenomas. METHODS: We performed immunohistochemical analysis by using a monoclonal anti-IDH1(R132H) antibody on the tissue microarrays containing specimens from the pituitary adenomas of different hormonal types from 246 patients. In positive samples, the status of the IDH1 gene was further examined by molecular genetic analyses. RESULTS: In all but one patient, there was no expression of mutated IDH1(R132H) protein in the tumor cells by immunohistochemistry. Only one patient with a recurring clinically non-functioning gonadotroph adenoma demonstrated IDH1(R132H)-immunostaining in both the primary tumor and the recurrence. However, no mutation in the IDH1 gene was detected using different molecular genetic analyses. CONCLUSION: IDH1(R132H) mutation occurs only exceptionally in pituitary adenomas and does not play a role in their pathogenesis. Patients with pituitary adenomas do not seem to be candidates for treatment with the inhibitors of mutant IDH1.

Non-functioning pituitary adenomas: growth and aggressiveness.

Pituitary adenomas (PAs) are common, comprising approximately one third of all intracranial tumors. Non-functioning pituitary adenomas (NFPAs) are the most common PAs. Although usually benign, the NFPAs represent therapeutic challenges because of their location close to the optic chiasm and nerves, and the proximity to the pituitary gland. The therapeutic alternatives are surgery and radiation. To date there is no effective medical treatment. NFPAs are classified according to different modalities, but there are no reliable marker of aggressiveness to guide the clinician in monitoring the patient. More information on growth patterns with constituent biological markers are needed to tailor the care of this patient group. Studies characterizing the membrane receptors of NFPAs have shown promising results, which may give rise to the development of medical treatment.