Pituitary Tumor Subtypes Research Articles

Identification of Prolactinoma in Pituitary Neuroendocrine Tumors Using Radiomics Analysis Based on Multiparameter MRI.

This study aims to investigate the feasibility of preoperatively predicting histological subtypes of pituitary neuroendocrine tumors (PitNETs) using machine learning and radiomics based on multiparameter MRI. Patients with PitNETs from January 2016 to May 2022 were retrospectively enrolled from four medical centers. A cfVB-Net network was used to automatically segment PitNET multiparameter MRI. Radiomics features were extracted from the MRI, and the radiomics score (Radscore) of each patient was calculated. To predict histological subtypes, the Gaussian process (GP) machine learning classifier based on radiomics features was performed. Multi-classification (six-class histological subtype) and binary classification (PRL vs. non-PRL) GP model was constructed. Then, a clinical-radiomics nomogram combining clinical factors and Radscores was constructed using the multivariate logistic regression analysis. The performance of the models was evaluated using receiver operating characteristic (ROC) curves. The PitNET auto-segmentation model eventually achieved the mean Dice similarity coefficient of 0.888 in 1206 patients (mean age 49.3 ± SD years, 52% female). In the multi-classification model, the GP of T2WI got the best area under the ROC curve (AUC), with 0.791, 0.801, and 0.711 in the training, validation, and external testing set, respectively. In the binary classification model, the GP of T2WI combined with CE T1WI demonstrated good performance, with AUC of 0.936, 0.882, and 0.791 in training, validation, and external testing sets, respectively. In the clinical-radiomics nomogram, Radscores and Hardy'grade were identified as predictors for PRL expression. Machine learning and radiomics analysis based on multiparameter MRI exhibited high efficiency and clinical application value in predicting the PitNET histological subtypes.

E-cadherin expression and gene expression profiles in corticotroph pituitary neuroendocrine tumor subtypes.

Corticotroph adenomas/pituitary neuroendocrine tumors (PitNETs) are associated with significant morbidity and mortality. Predictors of tumor behavior have not shown high prognostic accuracy. For somatotroph adenomas/PitNETs, E-cadherin expression correlates strongly with prognosis. E-cadherin expression has not been investigated in other PitNETs. A retrospective chart review of adults with corticotroph adenomas/PitNETs was conducted to assess correlation between E-cadherin expression and tumor characteristics. In addition, gene expression microarray was performed in subset of tumors (n = 16). Seventy-seven patients were identified; 71% were female, with median age of cohort 45.2 years. Seventy-five percent had macroadenomas, of which 22% were hormonally active. Ninety-five percent of microadenomas were hormonally active. Adrenocorticotropic hormone granulation pattern by IHC identified 63% as densely granulated (DG) and 34% as sparsely granulated (SG). All microadenomas were DG (p < .001); 50% of macroadenomas were DG associated with increased tumor invasion compared to SG. E-cadherin IHC was positive in 80%, diminished in 17%, and absent in 20% and did not correlate with corticotroph PitNETs subtype, size, or prognosis. In contrast to the distinct transcriptomes of corticotroph PitNETs and normal pituitaries, a comparison of clinically active and silent corticotroph PitNETs demonstrated similar molecular signatures indicating their common origin, but with unique differences related to their secretory status.

Alternations of Blood Pressure Following Surgical or Drug Therapy for Prolactinomas.

Several subtypes of pituitary neuroendocrine tumors (PitNETs), such as acromegaly and Cushing's disease, can result in hypertension. However, whether prolactinoma is associated with this complication remains unknown. Moreover, the effect of treatment with surgery or drugs on blood pressure (BP) is unknown. Herein, a retrospective study reviewed 162 patients with prolactinoma who underwent transsphenoidal surgery between January 2005 and December 2022. BP measurements were performed 1 day before and 5 days after surgery. Accordingly, patients' medical characteristics were recorded. In addition, in situ rat and xenograft nude-mice prolactinoma models have been used to mimic prolactinoma. In vivo BP and serum prolactin (PRL) levels were measured after cabergoline (CAB) administration in both rats and mice. Our data suggest that surgery can effectively decrease BP in prolactinoma patients with or without hypertension. The BP-lowering effect was significantly associated with several variables, including age, sex, disease duration, tumor size, invasion, dopamine agonists (DAs)-resistance, recurrence, and preoperative PRL levels. Moreover, in situ and xenograft prolactinomas induced BP elevation, which was alleviated by CAB treatment without and with a statistical difference in rats and mice, respectively. Thus, surgery or CAB can decrease BP in prolactinoma, indicating that pre- and postoperative BP management becomes essential.

Circulating FK506 binding protein 51 mRNA expression in patients with pituitary adenomas

BackgroundFK506 binding protein 51 (FKBP5) is a co-chaperone regulator of the glucocorticoid receptor (GR). Recent studies have reported increased FKBP5 mRNA in the circulation from patients with Cushing disease (CD) which returned to comparable levels seen in healthy controls following successful trans-nasal trans-sphenoidal (TNTS) surgical corticotroph tumor removal. However, the expression of circulating FKBP5 mRNA levels in other pituitary tumor subtypes and its specificity to corticotroph tumors is unknown. MethodsPre-operative blood was collected from consecutive patients undergoing TNTS for pituitary tumors (n = 57) at our center between 2015 and 2019. Total RNA was isolated from whole blood using RiboPure blood RNA isolation kit and real-time qPCR was used to quantitate circulating FKBP5 mRNA expression. ResultsConsistent with the prior report, higher circulating FKBP5 mRNA levels were observed in 20 patients with CD prior to surgical tumor removal, compared to 21 healthy controls (p < 0.0005) and compared to 8 patients harboring gonadotroph pituitary tumors (p < 0.05) and 6 patients with silent corticotroph pituitary tumors (p < 0.05). However, circulating FKBP5 mRNA levels were higher in 10 patients with prolactin (PRL)-secreting pituitary tumors compared to healthy controls (p < 0.05), and did not differ between patients with CD and patients with growth hormone secreting tumors (GH-omas). ConclusionsAlthough we confirm that circulating FKBP5 mRNA is higher in patients with corticotroph tumors compared to healthy subjects, measurement of circulating FKBP5 does not appear to be helpful to distinguish corticotroph tumors from other pituitary tumor sub-types.

EPID-14. EPIDEMIOLOGY OF ADULT PITUITARY TUMORS IN THE U.S. ACCORDING TO THE 2017 WHO CLASSIFICATION OF ENDOCRINOLOGY TUMOURS

Abstract INTRODUCTION Among adult intracranial tumors, the pituitary represents a frequent site of origin. We examine their contemporary epidemiology with a particular focus on uncommon pituitary tumor types. METHODS Adult patients presenting with pituitary or sellar tumors between 2004-2017 were identified from the U.S. National Cancer Database (comprising &amp;gt;70% of newly-diagnosed cancers). Their epidemiology was assessed in the context of the 2017 WHO Classification of Endocrine Tumours. RESULTS 12.5% of adult intracranial tumors arose in the pituitary region. 113,352 adults with pituitary tumors were identified. Histopathological diagnosis was obtained in only 59% of cases, in which 93% were pituitary adenomas and 6% were craniopharyngiomas. Among craniopharyngiomas, 71% were adamantinomatous and 29% were papillary, between which there were no differences in age, sex, or tumor size—however, papillary craniopharyngiomas were less common among Black nonHispanic patients (p&amp;lt; 0.001). Among the remaining 1% (n=680) of pituitary tumors, posterior pituitary tumors comprised 21%, chordomas 16%, meningiomas 15%, pituitary carcinomas 11%, GCTs 10%, hematolymphoid 8%, other mesenchymal and stromal 7%, neuronal/paraneuronal 6%, and schwannoma 4%. Meningiomas (84%), mesenchymal/stromal (64%), and neuronal/paraneuronal (64%) tumors displayed a female predominance, whereas GCTs (75%) and pituitary carcinomas (62%) exhibited a male predominance. Age at diagnosis, tumor size, and race/ethnicity varied widely across uncommon tumor types. We further examined the subtypes of uncommon pituitary tumors: for sellar chordomas, 19% were chordoid and none were dedifferentiated; for sellar meningiomas, 94% were grade I; for pituitary GCTs, 79% were pure germinomas; for hematolymphoid, 52% were DLBCL, 11% were plasmacytomas, and 9% were Langerhans cell histiocytosis; for neuronal/paraneuronal, 64% were gangliocytomas; and for mesenchymal tumors, 53% were vascular and 16% were SFTs/HPCs. CONCLUSIONS Using national registry data, we provide a detailed dissection of the epidemiology of adult pituitary tumors, with a particular focus on examining uncommon pituitary tumor types in the context of WHO2017.

Epidemiology of common and uncommon adult pituitary tumors in the U.S. according to the 2017 World Health Organization classification.

To examine the contemporary epidemiology of adult pituitary tumors with a particular focus on uncommon tumor types, using the 2017 WHO Classification of pituitary tumors. Adult patients presenting with a pituitary or sellar tumor between 2004 and 2017 were identified from the U.S. National Cancer Database, with tumor type categorized according to the 2017 WHO classification. Descriptive epidemiological statistics were evaluated and reported for all pituitary tumor types and subtypes. 113,349 adults with pituitary tumors were identified, 53.0% of whom were female. The majority of pituitary tumors were pituitary adenomas (94.0%), followed by craniopharyngiomas (3.8%). Among pituitary adenomas, whereas 71.6% of microadenomas presented in females, only 46.7% of macroadenomas and 41.3% of giant adenomas did (p < 0.001). For craniopharyngiomas, 71.2% were adamantinomatous and 28.8% were papillary, with adamantinomatous tumors associated with Black non-Hispanic race/ethnicity (ORadj=2.44 vs. White non-Hispanic, 99.9%CI = 1.25-4.75, p < 0.001) in multivariable analysis.The remaining 0.7% (n = 676) of pathology-confirmed pituitary tumor types were composed of: 21% tumors of the posterior pituitary, 16% chordomas, 11% pituitary carcinomas (i.e. adenohypophyseal histology with metastasis; herein most frequently to bone), 10% meningiomas, 8% germ cell tumors, 7% hematolymphoid (largely DLBCLs), and 4% neuronal/paraneuronal (largely gangliogliomas). Pituitary carcinomas and posterior pituitary tumors demonstrated a male predilection (62.2% and 56.0%, respectively), whereas sellar meningiomas predominated in females (84.1%). Age, race/ethnicity, tumor size, and overall survival further varied across uncommon pituitary tumor types. Our findings provide a detailed contemporary dissection of the epidemiology of common and uncommon adult pituitary tumors in the context of WHO2017.

Expression of programmed death-ligand 1 (PD-L1) in human pituitary neuroendocrine tumor.

To explore the programmed death-ligand 1 (PD-L1) expression in varied subtypes of pituitary neuroendocrine tumors with assessment of their clinical behavior at diagnosis and follow-up. We conducted a retrospective monocentric study, including all patients operated in the Academic Hospital of Angers (France) for a pituitary neuroendocrine tumor between 2012 and 2018. PDL-1 immunostaining was performed using a European Conformity-In Vitro Diagnostic-labeled anti-PDL1 antibody (clone 22C3). PD-L1 immunostaining was evaluated as the percentage of tumor cells showing positive membrane staining, into four grades: grade 0 = < 1%, grade 1 = 1 to 5%, grade 2 = 6 to 49% and grade 3 = ≥ 50%. PD-L1 expression was compared with tumor features (secretion, proliferation, invasion) and outcome. The study included 139 pituitary neuroendocrine tumors, including 84 (60%) nonfunctioning adenomas. Twenty-five pituitary neuroendocrine tumors were PD-L1 positive (18%), including 3 grade 3, 8 grade 2 and 14 grade 1. PD-L1 expression was not different between functioning and nonfunctioning adenomas (p = 0.26). Among 16 tumors with proliferative markers (Ki-67 ≥ 3% and p53 positive), only one was PD-L1 positive. In our series, PD-L1 was expressed in a rather small proportion of PitNET (18%), and this immune marker was not associated with any biological characteristic or behavior of the pituitary tumors. Thus, PD-L1 staining may be necessary before considering PD-L1 blockage in pituitary neuroendocrine tumors, in case of therapeutic impasse.

Elevated levels of circulating betahydroxybutyrate in pituitary tumor patients may differentiate prolactinomas from other immunohistochemical subtypes

The diagnosis of various histological subtypes of pituitary tumors is made using serum based hormone panel test. However, certain subtypes secrete more than one hormone, making the diagnosis ambiguous. Here, we performed 1H-NMR based metabolomic analysis of serum and whole-blood from luteinizing/follicle-stimulating (LH/FSH)-secreting (n = 24), prolactinomas (n = 14), and non-functional (NF) (n = 9) tumors. We found elevated levels of betahydroxybutyrate (BHB) in serum and whole-blood (WB) of prolactinomas (0.481 ± 0.211/0.329 ± 0.228 mM in serum/WB), but it was statistically significant (p ≤ 0.0033, Bonferroni correction) only in serum when compared with LH/FSH-secreting tumor patients (0.269 ± 0.139/0.167 ± 0.113 mM in serum/WB). Phenylalanine in NF tumors was found to be elevated in both serum and WB when compared with prolactinomas but it met the statistical significance criteria (p ≤ 0.0028) only in the serum. Alanine (p ≤ 0.011), tyrosine (p ≤ 0.014) and formate (p ≤ 0.011) were also elevated in NF tumors but none showed statistically significance when compared with prolactinomas. Quantification of BHB and the above amino acids in the circulation may aid in the development of blood-based in vitro diagnostic methods which can supplement the currently used serum hormone panel in the diagnosis of various subtypes of pituitary tumors.

How Valuable Is the RT-qPCR of Pituitary-Specific Transcription Factors for Identifying Pituitary Neuroendocrine Tumor Subtypes According to the New WHO 2017 Criteria?

The classification of pituitary neuroendocrine tumors (PitNETs) subtypes continues generating interest. In 2017, the World Health Organization (WHO) proposed considering the immunohistochemical (IHC) analysis of pituitary-specific transcription factors (TF) for their typification. The present study targeted the quantification of pituitary-specific TF (TPIT, PIT-1, SF-1, GATA2, ESR1) gene expression by RT-qPCR to overcome the shortcomings of IHC and to complement it. We analyzed 251 tumors from our collection of PitNETs and performed additional IHC studies in a subset of 56 samples to analyze the concordance between gene and protein expression of the TF. The molecular and IHC studies allowed us to significantly reduce the percentage of null cell tumors in our series, most of which were reclassified as gonadotroph tumors. The concordance between the molecular and the immunohistochemical studies was good for tumors coming from the corticotroph and Pit-1 lineages but worsened for the rest of the tumors. Indeed, the RT-qPCR helped to improve the typification of plurihormonal Pit-1 and unusual tumors. Overall, our results suggest that the RT-qPCR of pituitary-specific TF and hormone genes could help pathologists, endocrinologists, and neurosurgeons to improve the management of patients with pituitary tumors.

Exosome-Transmitted lncRNA H19 Inhibits the Growth of Pituitary Adenoma.

Our previous study demonstrated that the expression of long noncoding RNA (lncRNA) H19 was frequently downregulated in human primary pituitary adenomas and negatively correlated with tumor progression. However, the role of exosomal lncRNA H19 in the inhibition of pituitary tumor growth remains unclear. To investigate whether exosomal H19 could be transported across the cell membrane to exert its inhibitory effect on pituitary tumor growth. Empty lentivirus GH3 cells with or without H19 overexpression were used to establish a xenograft model. Isolated exosomes were identified by transmission electron microscopy, nanoparticle tracking, and Western blotting. The expression levels of serum exosomal H19 from 200 healthy subjects and 206 patients with various subtypes of pituitary tumors were detected by ultracentrifugation and quantitative real-time PCR. The growth of distal tumor cells was inhibited by transferring exosomal H19, which could be transported through cell membrane and exert its inhibitory effect. Cabergoline increased H19 expression and played a synergic therapeutic effect with exosomal H19. Exosomal H19 inhibited phosphorylation of the mTORC1 substrate 4E-BP1. Of note, the expression level of exosomal H19 in the patients with all subtypes of pituitary tumors was significantly lower than that in the healthy subjects. The change of plasma exosomal H19 level may be correlated with the prognosis or drug response of the patients. Exosomal H19 inhibits the growth of distal pituitary tumors through inhibiting 4E-BP1 phosphorylation. Plasma exosomal H19 may serve as an important biomarker for predicting medical responses of patients with prolactinomas.

Is the molecular study of pituitary transcription factors useful in the identification of Pituitary Neuroendocrine Tumor subtypes according to the new WHO 2017 criteria?

Searchable abstracts of presentations at key conferences in endocrinology ISSN 1470-3947 (print) | ISSN 1479-6848 (online)

Dopamine Agonists for Pituitary Adenomas

Dopamine agonists (DA) are well established as first-line therapy for prolactinomas. These tumors express high levels of dopamine 2 receptors (D2R), leading to the strong efficacy of DA in reducing tumor size and hormonal secretion. Other pituitary tumor subtypes express D2R to varying degrees, leading to an extensive body of research into potential off-label use of DA in non-prolactinoma pituitary tumors. Preclinical models of Cushing's disease, acromegaly, and nonfunctioning pituitary tumors (NFPT) demonstrate D2R expression in cell lines and cultured tumors as well as effectiveness of DA in reducing hormonal secretion in functioning tumors and arresting tumor proliferation. Clinical studies have shown some efficacy of DA in treatment of these tumors. In Cushing's disease, DA therapy results in normalization of urinary cortisol levels in approximately 25% of patients, but reported rates of tumor shrinkage are very low; in acromegaly, DA therapy leads to normalization of insulin-like growth factor I and tumor shrinkage in approximately one-third of patients, and improved responses when used in combination with somatostatin receptor ligands. Among patients with NFPT, pooled results show 30% experience reduction of tumor size and 58% show stabilization of disease. DA therapy appears to have some clinical benefit in patients with non-prolactinoma pituitary tumors, and may be an option for medical therapy in some clinical scenarios.

Overview of the 2017 WHO Classification of Pituitary Tumors.

This review focuses on discussing the main changes on the upcoming fourth edition of the WHO Classification of Tumors of the Pituitary Gland emphasizing histopathological and molecular genetics aspects of pituitary neuroendocrine (i.e., pituitary adenomas) and some of the non-neuroendocrine tumors involving the pituitary gland. Instead of a formal review, we introduced the highlights of the new WHO classification by answering select questions relevant to practising pathologists. The revised classification of pituitary adenomas, in addition to hormone immunohistochemistry, recognizes the role of other immunohistochemical markers including but not limited to pituitary transcription factors. Recognizing this novel approach, the fourth edition of the WHO classification has abandoned the concept of "a hormone-producing pituitary adenoma" and adopted a pituitary adenohypophyseal cell lineage designation of the adenomas with subsequent categorization of histological variants according to hormone content and specific histological and immunohistochemical features. This new classification does not require a routine ultrastructural examination of these tumors. The new definition of the Null cell adenoma requires the demonstration of immunonegativity for pituitary transcription factors and adenohypophyseal hormones Moreover, the term of atypical pituitary adenoma is no longer recommended. In addition to the accurate tumor subtyping, assessment of the tumor proliferative potential by mitotic count and Ki-67 index, and other clinical parameters such as tumor invasion, is strongly recommended in individual cases for consideration of clinically aggressive adenomas. This classification also recognizes some subtypes of pituitary neuroendocrine tumors as "high-risk pituitary adenomas" due to the clinical aggressive behavior; these include the sparsely granulated somatotroph adenoma, the lactotroph adenoma in men, the Crooke's cell adenoma, the silent corticotroph adenoma, and the newly introduced plurihormonal Pit-1-positive adenoma (previously known as silent subtype III pituitary adenoma). An additional novel aspect of the new WHO classification was also the definition of the spectrum of thyroid transcription factor-1 expressing pituitary tumors of the posterior lobe as representing a morphological spectrum of a single nosological entity. These tumors include the pituicytoma, the spindle cell oncocytoma, the granular cell tumor of the neurohypophysis, and the sellar ependymoma.

Clinical Identification of Oncogenic Drivers and Copy-Number Alterations in Pituitary Tumors.

Pituitary tumors are the second most common adult primary brain tumor, with a variable clinical course. Recent work has identified a number of genetic determinants of pituitary tumor subtypes, which may augment traditional histopathologic classification schemes. We sought to determine whether pituitary tumors could be stratified based on objective molecular characteristics using a clinical genomics assay. We performed a retrospective analysis of patients operated on at the Brigham and Women's Hospital from 2012 to 2016 whose pituitary tumors were profiled using multiplexed next-generation sequencing. We analyzed 127 pituitary tumors, including 114 adenomas, 5 craniopharyngiomas, and 8 tumors of other histologies. We observed recurrent BRAFV600E mutations in papillary craniopharyngiomas, CTNNB1 mutations in adamantinomatous craniopharyngiomas, and activating GNAS mutations in growth hormone-secreting adenomas. Furthermore, we validated the presence of two distinct genomic subclasses in adenomas (i.e., those with disrupted or quiet copy-number profiles) and the significant association of disruption with functional hormone status (P < 0.05). We report the clinical implementation of next-generation sequencing of pituitary tumors. We confirmed previously identified molecular subclasses for these tumors and show that routine screening as part of clinical practice is both feasible and informative. This large-scale proof-of-principle study may help to guide future institutional efforts for pituitary tumor classification as well as the incorporation of such techniques into prospective analysis as part of clinical trials.

Differential somatostatin receptor (SSTR) 1–5 expression and downstream effectors in histologic subtypes of growth hormone pituitary tumors

PurposeThe aim of this study was to examine whether differential expression of somatostatin receptors (SSTR) 1–5 and downstream effectors are different in densely (DG) and sparsely (SG) granulated histological growth hormone (GH) pituitary tumor subtypes. MethodsThe study included 33 acromegalic patients with 23 DG and 10 SG tumors. SSTR1–5 were measured by qPCR and immunoblotting. Signaling candidates downstream of SSTR2 were also assessed. ResultsSSTR2 mRNA and protein levels were significantly higher in DG compared to SG tumors. Downstream of SSTR2, p27kip1 was decreased (2.6-fold) in SG compared to DG tumors, suggesting a potential mechanism of SSA resistance in SG tumors with intact SSTR2 expression. Re-expression of E-cadherin in GH pituitary cell increased p27kip1 levels. ConclusionsHistological subtyping correlated with SSTR2, E cadherin and p27kip protein levels and these may serve as useful biomarkers in GH tumors to predict behavior and response to therapy with SSA.

Decoding the genetic basis of Cushing's disease: USP8 in the spotlight.

Cushing's disease (CD) arises from pituitary-dependent glucocorticoid excess due to an ACTH-secreting corticotroph tumor. Genetic hits in oncogenes and tumor suppressor genes that afflict other pituitary tumor subtypes are not found in corticotrophinomas. Recently, a somatic mutational hotspot was found in up to half of corticotrophinomas in the USP8 gene that encodes a protein that impairs the downregulation of the epidermal growth factor receptor (EGFR) and enables its constitutive signaling. EGF is an important regulator of corticotroph function and its receptor is highly expressed in Cushing's pituitary tumors, where it leads to increased ACTH synthesis in vitro and in vivo. The mutational hotspot found in corticotrophinomas hyper-activates USP8, enabling it to rescue EGFR from lysosomal degradation and ensure its stimulatory signaling. This review presents new developments in the study of the genetics of CD and focuses on the USP8-EGFR system as trigger and target of corticotroph tumorigenesis.

Current status on histological classification in Cushing's disease.

Managing Cushing's disease remains a challenge. Surgery is the first option of treatment and it offers a high success rate. Even in cases where biochemical remission is not achieved, it is crucial to obtain surgical tissue for morphological diagnosis because the therapeutic approach can be modified according to the findings. A literature search was performed using PubMed for information regarding pathology and Cushing's disease. The histopathological features found in the pituitary gland of patients with Cushing's disease are presented. Different subtypes of ACTH-producing pituitary tumors are recognized and characterized. The significance of finding a normal pituitary gland with or without Crooke's changes is also discussed.

Components of the Canonical and Non-Canonical Wnt Pathways Are Not Mis-Expressed in Pituitary Tumors

IntroductionCanonical and non-canonical Wnt pathways are involved in the genesis of multiple tumors; however, their role in pituitary tumorigenesis is mostly unknown.ObjectiveThis study evaluated gene and protein expression of Wnt pathways in pituitary tumors and whether these expression correlate to clinical outcome.Materials and MethodsGenes of the Wnt canonical pathway: activating ligands (WNT11, WNT4, WNT5A), binding inhibitors (DKK3, sFRP1), β-catenin (CTNNB1), β-catenin degradation complex (APC, AXIN1, GSK3β), inhibitor of β-catenin degradation complex (AKT1), sequester of β-catenin (CDH1), pathway effectors (TCF7, MAPK8, NFAT5), pathway mediators (DVL-1, DVL-2, DVL-3, PRICKLE, VANGL1), target genes (MYB, MYC, WISP2, SPRY1, TP53, CCND1); calcium dependent pathway (PLCB1, CAMK2A, PRKCA, CHP); and planar cell polarity pathway (PTK7, DAAM1, RHOA) were evaluated by QPCR, in 19 GH-, 18 ACTH-secreting, 21 non-secreting (NS) pituitary tumors, and 5 normal pituitaries. Also, the main effectors of canonical (β-catenin), planar cell polarity (JNK), and calcium dependent (NFAT5) Wnt pathways were evaluated by immunohistochemistry.ResultsThere are no differences in gene expression of canonical and non-canonical Wnt pathways between all studied subtypes of pituitary tumors and normal pituitaries, except for WISP2, which was over-expressed in ACTH-secreting tumors compared to normal pituitaries (4.8x; p = 0.02), NS pituitary tumors (7.7x; p = 0.004) and GH-secreting tumors (5.0x; p = 0.05). β-catenin, NFAT5 and JNK proteins showed no expression in normal pituitaries and in any of the pituitary tumor subtypes. Furthermore, no association of the studied gene or protein expression was observed with tumor size, recurrence, and progressive disease. The hierarchical clustering showed a regular pattern of genes of the canonical and non-canonical Wnt pathways randomly distributed throughout the dendrogram.ConclusionsOur data reinforce previous reports suggesting no activation of canonical Wnt pathway in pituitary tumorigenesis. Moreover, we describe, for the first time, evidence that non-canonical Wnt pathways are also not mis-expressed in the pituitary tumors.

Reversal of Endogenous Dopamine Receptor Silencing in Pituitary Cells Augments Receptor-Mediated Apoptosis

Dopamine (DA)-agonist targeting of the DA D(2) receptor (D2R) in prolactinomas is the first-line treatment choice for suppression of prolactin and induction of tumor shrinkage. Resistance to DA agonists seems to be related to receptor number. Using the MMQ and GH3 pituitary cell lines, that either do or do not express D2R, respectively, we explored the epigenetic profile associated with the presence or absence of D2R in these cells lines. These studies led us to explore pharmacological strategies designed to restore receptor expression and thereby potentially augment DA agonist-mediated apoptosis. We show in GH3 cells that the D2R harbors increased CpG island-associated methylation and enrichment for histone H3K27me3. Conversely, MMQ cells and normal pituitaries show enrichment for H3K9Ac and barely detectable H3K27me3. Coculture of GH3 cells with the demethylating agent zebularine and the histone deacetylase inhibitor trichostatin A was responsible for a decrease in CpG island methylation and enrichment for the histone H3K9Ac mark. In addition, challenge of GH3 cells with zebularine alone or coculture with both agents led to expression of endogenous D2R in these cells. Induced expression D2R in GH3 cells was associated with a significant increase in apoptosis indices to challenge with either DA or bromocriptine. Specificity of a receptor-mediated response was established in coincubations with specific D2R antagonist and siRNA approaches in GH3 cell and D2R expressing MMQ cell lines. These studies point to the potential efficacy of combined treatment with epigenetic drugs and DA agonists for the medical management of different pituitary tumor subtypes, resistant to conventional therapies.

Expression of functional KISS1 and KISS1R system is altered in human pituitary adenomas: evidence for apoptotic action of kisspeptin-10

KISS1 was originally identified as a metastasis-suppressor gene able to inhibit tumor progression. KISS1 gene products, the kisspeptins, bind to a G-protein-coupled receptor (KISS1R, formerly GPR54), which is highly expressed in placenta, pituitary, and pancreas, whereas KISS1 mRNA is mainly expressed in placenta, hypothalamus, striatum, and pituitary. KISS1/KISS1R pituitary expression profile, coupled to their anti-tumoral capacities, led us to hypothesize that this system may be involved in the biology of pituitary tumors. To explore this notion, expression levels of KISS1R and KISS1 were evaluated in normal and adenomatous pituitaries. Additionally, functionality of this system was assessed by treating dispersed pituitary adenoma cells in primary culture with kisspeptin-10 and evaluating intracellular calcium kinetics and apoptotic rate. Both KISS1 and KISS1R were expressed in normal pituitary, whereas this simultaneous expression was frequently lost in pituitary tumors, where diverse patterns of KISS1/KISS1R expression were observed that differed among distinct types of pituitary adenomas. Measurement of calcium kinetics revealed that kisspeptin-10 elicits a remarkable increase in [Ca(2+)](i) in individual cells from four out of the five GH-producing adenomas studied, whereas cells derived from non-functioning pituitary adenomas (NFPA, n=45) did not respond. In contrast, kisspeptin-10 treatment increased the apoptotic rate in cells derived from both GH-producing and NFPA. These results provide primary evidence that KISS1 and KISS1R expression can be differentially lost in pituitary tumor subtypes, where this system can exert functional, proapoptotic actions, and thereby offer novel insights to investigate the biology and therapeutic options to treat these tumors.