MGMT immunohistochemistry in pituitary tumors: controversies with clinical implications.

INTRODUCTION: Temozolomide (TMZ) is currently considered as a rational therapeutic option for patients with progressively aggressive pituitary adenomas and carcinomas not responding to conventional therapies. Administration of TMZ results in clinical response and improvement in survival of many of these patients depending upon the expression of the DNA repair enzyme O-6 methylguanine DNA transferase (MGMT). Low or negative MGMT immunoreactivity predicts responsiveness to TMZ therapy. Therefore, MGMT serves as a criterion to select candidate patients anticipating response to treatment. MATERIALS AND METHODS: The MGMT expression was investigated in 25 pituitary adenomas with Ki-67 labeling index more that 3% and p53 expression, using various antigen retrieval protocols. After direct application of the antibody, only one adenoma yielded positive for MGMT. However, after pretreatment of tissue sections with antigen retrieval protocols, another 3 adenomas, initially negative turned to positive. CONCLUSIONS: These findings could explain lack of response to TMZ treatment in patients with false negative MGMT immunohistochemistry. Evaluation of tumor samples for MGMT expression should carefully be carried-out using the optimum immunohistochemical protocol to obtain consistent and reliable results that help to identify patients that could respond to TMZ therapy.

Somatostatin and dopamine receptor expression in neuroendocrine neoplasms: correlation of immunohistochemical findings with somatostatin receptor scintigraphy visual scores.

CONTEXT: The expression of somatostatin (sstr1-5) and dopamine (DR) receptors in neuroendocrine neoplasms (NENs) facilitates diagnosis by tumour visualization with somatostatin receptor scintigraphy (SRS) and directs towards specific treatment with peptide receptor radionuclide therapy (PRRT) with radiolabelled somatostatin analogues. OBJECTIVE: To investigate the co-expression of sstrs, D2R in relation to pre-operative SRSs in NENs. DESIGN: Prospective two-centre study. PATIENTS AND MEASUREMENTS: We analysed pre-operative SRS of 60 patients [44 with gastrointestinal (GI) NENs and 16 with lung NENs] and compared SRS results with immunohistochemical (IHC) reactivity for sstr2, sstr3, sstr5 in sample tissues from primary (n = 54) and metastatic (n = 27) lesions and IHC reactivity for D2R in 23 samples from primary GI-NENs lesions. RESULTS: Sstr2 was the commonest sstr expressed (65·4%) and was co-expressed with sstr3 and sstr5 in 32·1% and 24·7% of the specimens, respectively. In 67 of 81 specimens (82·7%), there was concordance of sstr2 immunohistochemistry with SRS findings (P < 0·001). D2R was expressed in only 8 of 23 (34·8%) GI-NENs while was co-expressed with sstr2 in all cases. SRS grade, as per Krenning scale, was higher in metastatic foci, large-size (>2 cm) tumours and GI-NENs, whereas sstr2 intensity was greater in GI compared to lung NENs. SRS grade showed higher correlation with sstr2 (r = 0·6, P < 0·001) and D2R (r = 0·5, P < 0·001) IHC intensity scores than tumour size (r = 0·4, P < 0·001) and sstr3 (r = 0·4, P < 0·001) intensity score. CONCLUSIONS: Sstr2 IHC expression and SRS are useful tools for the diagnosis and management of NENs because they display a high concordance. IHC expression of DR2 seems to be of potential clinical significance in GI-NENs tumours.

High-risk pituitary adenomas and strategies for predicting response to treatment.

High-risk pituitary adenomas are aggressive. They show clinical and imaging features similar to those of carcinomas, including infiltration of the surrounding brain structures, but lack cerebrospinal or systemic metastases. In addition, they display distinct behavior, including tendency for fast growth and frequent recurrences, which are difficult to control. The term "high-risk" adenoma was first introduced in the 4th edition of the World Health Organization Classification of Endocrine Tumors in 2017. Five defined adenoma types belong to this category, including sparsely granulated somatotroph, lactotroph in men, Crooke cell, silent corticotroph, and plurihormonal PIT-1 positive adenomas. The morphological and immunohistochemical characteristics of high-risk adenomas are herein described in detail. In addition, the clinical features and the treatment options are presented. This review focuses on predictive markers assessed by immunohistochemistry, which help clinicians to design the appropriate treatment strategies for high-risk adenomas. Somatostatin receptor status predicts effectiveness of postsurgical treatment with somatostatin analogs, and MGMT expression predicts response to treatment with temozolomide. This comprehensive review presents the clinical and pathological features of high-risk pituitary adenomas, underlines the contribution of immunohistochemistry, and emphasizes the leading role of pathology in the design of optimal clinical management.

Thyroid Paraganglioma With Medullary Carcinoma: A Unique Combination in a Patient in Association With Multiple Endocrine Neoplasia Type 2B Syndrome With Prolonged Survival.

Head and neck paragangliomas (PGLs) most commonly derive from the carotid body, jugulotympanic, vagal, and laryngeal paraganglia. Thyroid PGLs originate in the inferior laryngeal paraganglion, which may lie inside the thyroid parenchyma. Intrathyroid PGLs are rare with approximately 75 cases reported to date, mostly as solitary lesions. The coexistence of thyroid PGL with medullary thyroid carcinoma (MTC) has not been reported. Here, we report a unique case of intrathyroid PGL concomitant with MTC in the context of multiple endocrine neoplasia type 2B syndrome. Interestingly, the patient showed a prolonged survival with good clinical response to tyrosine kinase inhibitors, despite her advanced metastatic MTC. We discuss the challenges in pathology, differential diagnosis, and genetic background for the development of these thyroid lesions.

Association of Pathology Markers with Somatostatin Analogue Responsiveness in Acromegaly.

Background: Somatotroph adenomas (SAs) exhibit a variable responsiveness to somatostatin analogue (SS-a) treatment, a process that is not well understood. We investigated established and novel histological markers as predictors of SS-a responsiveness. Methods: We retrospectively investigated pathology samples from 36 acromegalic patients that underwent transsphenoidal surgery. Clinical, hormonal, and imaging data were available in 24/36 patients, before and after SS-a treatment. Specimens were semiquantitatively analyzed with immunocytochemistry for Ki-67, KER, SSTR-2, SSTR-5, ZAC-1, E-cadherin, and AIP. Results: Collectively, 18 (50%) adenomas were each classified as densely/sparsely granulated somatotroph adenomas (DGSAs/SGSAs), respectively. Patients that received preoperative SS-a had lower expression of SSTR-2 compared to those that did not (2.0 (1.0, 3.0) vs. 3.0 (3.0, 3.0), p = 0.042). Compared with DGSAs, SGSAs had higher Ki-67 labeling index (LI) (1.0 (0.5, 1.0) vs. 2.0 (1.0, 3.5), p = 0.013), and a higher proportion of high MR T2 signal (1 (6%) vs. 6 (33%), p = 0.035), and tended to express less ZAC-1 (p = 0.061) and E-cadherin (p = 0.067). In linear regression corrected for baseline growth hormone (GH), ZAC-1 immunostaining was significantly associated with a decrease in GH levels after SS-a treatment (beta (95% confidence interval): -1.53 (-2.80, -0.26), p = 0.021). No markers were associated with changes in circulating insulin-like growth factor-I (IGF-I) after treatment with SS-a. Conclusion: The novel marker ZAC-1 was associated with GH response to medical treatment with SS-a. The SGSA cases were characterized by higher Ki-67 values and MR T2 signals indicative of an inferior response to SS-a. These findings improve our understanding of the mechanisms underlying SA response to medical treatment.

Update on pituitary adenomas in the 2017 World Health Organization classification: innovations and perspectives.

The World Health Organization (WHO) classifications of tumors offer invaluable support in the diagnosis of tumors, with every new edition including novel information and diagnostic updates. The new 2017 WHO Classification of Tumors of Endocrine Organs, 4th edition, includes innovations in both terminology and diagnostic guidelines for pituitary adenomas, along with new entities, molecular information, and novel treatment modalities. The recommended reporting system of pituitary adenomas is based on morphology and assessment of the hormonal content by immunohistochemistry. Electron microscopy and immunohistochemistry for Ki-67 and p53 and transcription factors, while presenting additional information, are not recommended for routine diagnosis. Other markers may also yield information of prognostic and predictive significance. In sum, the 2017 WHO classification provides pathologists and clinicians with new and comprehensive information of great use for the diagnosis and treatment of pituitary tumors.

Collision sellar lesions: experience with eight cases and review of the literature.

The concomitant presence of a pituitary adenoma with a second sellar lesion in patients operated upon for pituitary adenoma is an uncommon entity. Although rare, quite a great variety of lesions have been indentified coexisting with pituitary adenomas. In fact, most combinations have been described before, but an overview with information on the frequency of combined pathologies in a large series has not been published. We present a series of eight collision sellar lesions indentified among 548 transsphenoidally resected pituitary adenomas in two Neurosurgical Departments. The histological studies confirmed a case of sarcoidosis within a non-functioning pituitary adenoma, a case of intrasellar schwannoma coexisting with growth hormone (GH) secreting adenoma, two Rathke's cleft cysts combined with pituitary adenomas, three gangliocytomas associated with GH-secreting adenomas, and a case of a double pituitary adenoma. The pertinent literature is discussed with emphasis on pathogenetic theories of dual sellar lesions. Although there is no direct evidence to confirm the pathogenetic relationship of collision sellar lesions, the number of cases presented in literature makes the theory of an incidental occurrence rather doubtful. Suggested hypotheses about a common embryonic origin or a potential interaction between pituitary adenomas and the immune system are presented.

Non-adenomatous sellar lesions: experience of a single centre and review of the literature.

Non-adenomatous sellar lesions represent a diagnostic and occasionally a therapeutic challenge. The purpose of the study was to provide an overview of the clinical and radiographic characteristics of non-adenomatous sellar lesions operated at our department, with emphasis on treatment options. In our transsphenoidal surgical series of 300 cases, 29 non-adenomatous sellar lesions (9.7%) were identified by histology. Medical files and imaging findings were retrospectively analysed. Clinical presentation was usually a result of local mass effect. Imaging features were not always diagnostic. Of 29 non-adenomatous sellar lesions, total tumour excision with transsphenoidal surgery was achieved in 17, and no supplemental treatment was needed. Lesions partially resected were further treated with reoperation, radiation therapy, or medical therapy, tailored to individuals. In our experience, non-adenomatous sellar lesions represent a more prevalent clinical entity than expected. Pituitary surgeons should be aware of their existence to reconstruct treatment strategy, proceeding to adjuvant therapy when necessary.

Thyrotoxic periodic paralysis as the first manifestation of a thyrotropin-secreting pituitary adenoma.

Thyrotoxic Periodic Paralysis (TPP) is a rare manifestation of hyperthyroidism characterized by muscle weakness and hypokalemia. Thyroid-Stimulating Hormone (TSH)-secreting pituitary adenoma is a rare cause of hyperthyroidism. Even more rare is the occurrence of TPP as the first manifestation of a TSH-secreting pituitary adenoma. We report a 31-year-old Asian male patient suffering from TPP caused by a TSH-secreting adenoma, who was evaluated for persistent episodes of muscle paralysis. Laboratory investigation revealed hypokalemia as well as elevated levels of both thyroid hormones and TSH. The Magnetic Resonance Imaging (MRI) of the pituitary gland revealed a microadenoma, thus suggesting the presence of a TSH-secreting adenoma. The patient underwent transphenoidal resection and the pathological investigation confirmed the diagnosis of TSH-secreting pituitary adenoma. After the adenomectomy and the restoration of euthyroidism, the patient did not experience any episode of hypokalemic paralysis or weakness. Despite its rarity, TSH-secreting pituitary adenoma should be included in the differential diagnosis of TPP.

Somatostatin receptor profile in pituitary thyrotroph adenomas.

OBJECTIVES: Thyrotroph adenomas are the most infrequent adenohypophysial tumors. Somatostatin (SST) inhibits hormone secretion and suppresses cell proliferation. SST receptors (sstr) belong to a family of 5 types of G-coupled membrane proteins, which show high binding affinity to SST. Currently, SST analogs used to treat pituitary adenomas, have a preferential binding activity to sstr2 and sstr5. The aim of this study was to evaluate the status of all active sstrs on cell membrane of thyrotroph adenomas. PATIENTS: Nine cases of thyrotroph adenomas were studied for all types of sstrs. All patients were clinically associated with hyperthyroidism. The adenomas were initially diagnosed and classified by histology and immunohistochemistry for all pituitary hormones and two of them were examined by electron microscopy. METHODS: For sstr immunohistochemistry, antisera against all sst types (1, 2A, 2B, 3, 4 and 5) were used. To enhance sensitivity, the tyramide amplification technique was applied. This is the first report investigating the full spectrum of sstrs in thyrotroph adenomas by immunohistochemistry. RESULTS: All tumors were immunoreactive for ß-subunit of thyroid-stimulating hormone and for α-subunit of glycoprotein hormones. The sst2A, sst2B and sstr5 were co-expressed in all adenomas. The sstr1 and sstr3 were noted in 8 and sstr4 in 7 adenomas respectively. High scores 2+ and 3+ were prominent in sstr2A, sstr2B, sstr3 and sstr5. High score 3+ for sstr4 was also noted in one tumor, while score 3+ for sstr1 was not observed. CONCLUSIONS: Knowledge of the sstr status may contribute to a better selection of patients, anticipating benefit from treatment with SST analogs. Given that multiligand SST analogs have a broader ability to bind other sstrs, such as sstr1 and sstr3, patients with thyrotroph adenomas expressing these receptors may benefit from novel sstr targeting therapy.

Investigation of somatostatin receptor profile of neuroendocrine carcinomas of the breast.

Carcinomas of the breast with neuroendocrine features are rare primary neoplasms positive for neuroendocrine markers. According to the WHO classification of tumours of the breast they are divided into three morphologically distinct categories. They comprise neuroendocrine tumour (NET), neuroendocrine carcinoma (NEC) and carcinoma with neuroendocrine differentiation (NED). The purpose of this study was to investigate for the first time the full spectrum of sstr expression status in breast carcinomas with neuroendocrine features. Fifteen primary breast carcinomas with histological and immunohistochemical neuroendocrine features were studied. Four of them were classified as NETs and two as NECs, and the remaining 9 as carcinomas with NED. All six types of somatostatin receptor (sstr) types (sstr1, sstr 2A, sstr2B, sstr3, sstr4 and sstr5) were investigated by immunohistochemistry. To assess the distribution and intensity of membranous receptor immunoreactivity, a four-scale scoring system was used. Overall predominant receptors were sstr2A, sstr2B, sstr3 and sstr5 showing the highest membranous staining scores 3+ and 2 + . The sstr1 was not detected. Given that carcinomas with neuroendocrine features represent distinct entities, patients with such tumours may benefit from sstr targeting therapies. Immunohistochemistry for sstrs can predict the effectiveness of administration of SST analogues to those patients, thus contributing to achieve the maximum therapeutic outcome, particularly in NETs and NECs with scores 2+ and 3 + .

TRAIL receptor upregulation and the implication of KRAS/BRAF mutations in human colon cancer tumors.

TRAIL raises hopes as a promising anti-tumor agent due to its selectivity toward cancer cells. Higher expression of its pro-death receptors TRAIL-R1 (DR4) and TRAIL-R2 (DR5) attenuates higher sensitivity to TRAIL-induced apoptosis, and represents a marker for better cancer prognosis and treatment. Since receptor availability can be analogous to ligand efficacy, we performed RT-PCR analysis of DR4 and DR5 in 51 colon cancer biopsy specimens and respective normal mucosa, while 11 of these tumors were determined immunohistochemically for protein expression. Transcriptional analysis showed that DR4 and DR5 were significantly upregulated in 37 and 47% of the tumor samples respectively, while both DR4 and DR5 were coinstantaneously upregulated in 31% of the samples analyzed. Positive transcriptional regulation of DRs was recorded as early as Dukes' A stage. Furthermore, protein expression analysis yielded results comparable to DR4 and DR5 increased mRNA levels. Possible contributing events to DR upregulation involve presence of frequent oncogenic mutations in the MAPK pathway, and was investigated by direct sequencing in all 51 tumors. Samples (6/8) hosting either a KRAS(G12V) or BRAF(V600E) mutation, significantly amplified the upregulated expression of DR4 and DR5, showing strong inter-relation between overexpression and presence of oncogenic KRAS/ BRAF mutations. In the light of recent data concerning TRAIL receptor distribution, we contribute further by presenting DR5 as the most frequently upregulated DR in colon cancer. Furthermore, oncogenic mutations may directly or indirectly enhance DR expression, potentially sensitizing these tumors to TRAIL-based therapies.

Intra-sellar schwannoma co-existing with GH-secreting pituitary adenoma.

BACKGROUND: We present a unique example of an intra-sellar schwannoma co-existing with a growth hormone (GH)-secreting pituitary adenoma. METHOD AND FINDINGS: The patient presented with acromegaly and magnetic resonance imaging (MRI) revealed an intra-sellar mass. The tumour was totally resected via a sub-labial trans-sphenoidal approach. Histopathology demonstrated the presence of a GH-secreting adenoma as well as a schwannoma at the periphery of the adenoma. After surgical excision, remission of the acromegaly occurred. Follow-up monitoring showed no evidence of recurrence of the adenoma two years after surgery. CONCLUSION: To the best of our knowledge, this is the first example of an intra-sellar schwannoma co-existing with a GH-secreting pituitary adenoma.

Sphenoid wing meningioma and glioblastoma multiforme in collision - case report and review of the literature.

The coexistence of two histologically different primary tumours in the same brain region is relatively rare. The situation where these tumours in collision appear in an area of a previously excised meningioma is even more rare and only two cases have been reported so far. We present the third case of a 73-year-old woman who underwent an uneventful excision of a right sphenoid wing meningioma. She was re-admitted 3 years later due to reappearance of a tumour in the area adjacent to the previously excised meningioma. Histological diagnosis revealed a collision tumour of a glioblastoma multiforme and a fibrillary meningioma. The coincidence of these two different neoplasms in the same location at the same time 3 years after surgical removal of a meningioma leads us to speculate on the pathogenesis, and to review the literature regarding this particular issue.

Double adenomas of the pituitary: an imaging, pathological, and clinical diagnostic challenge.

Double and multiple adenomas of the pituitary are composed of two or more distinct tumors located in the same gland. They represent uncommon lesions measuring less than 1 cm, reported as having a low incidence in autopsies and occurring even more infrequently in surgical series. The histological diagnosis of double adenomas in surgical material is often extremely difficult, and confirmation requires immunohistochemistry and, occasionally, electron microscopy. Fragmented tissue material submitted for histology after transsphenoidal resection complicates the diagnosis. Difficulties in demonstrating double or multiple adenomas by imaging techniques contribute to diagnostic failure. Magnetic resonance imaging (MRI) techniques may disclose two separate adenomas located in the same pituitary gland. Intraoperative MRI and imaging ultrasonography, together with positron emission computed tomography, more accurately identify sites of residual tumors. These techniques might also detect postoperatively a residual tumor belonging to the second component of double adenoma. Double adenomas may also create extreme clinical diagnostic challenges. It is almost impossible to suspect functioning double adenomas with combined hormone secretion, each one secreting a different hormone, and distinguish them from an isolated plurihormonal adenoma, simultaneously secreting more than one hormone. Double adenomas may underlie surgical failure when one adenoma is removed while the other is left behind. Despite the low frequency of double adenomas, identification and resection of both of them is of major importance for the achievement of biochemical cure.

Ectopic Cushing's syndrome due to CRH secreting liver metastasis in a patient with medullary thyroid carcinoma.

Ectopic production of CRH by a medullary thyroid carcinoma or its metastases is a rare cause of ectopic Cushing's syndrome (ECS). We report a 45-year old male with medullary thyroid carcinoma (MTC), who, 24 years following the initial diagnosis, presented with clinical and biochemical evidence of an ACTH dependent Cushing's syndrome. Rapid deterioration of his clinical condition and elevated cortisol levels were observed. Computed tomographic imaging of the abdomen revealed extensive liver metastases. The patient underwent fine needle aspiration biopsy of a liver lesion and immunohistochemistry showed that the cells expressed calcitonin, carcino-embryonic antigen and synaptophysin. Further analysis revealed that the material also expressed CRH. This is an unusual case of a CRH-secreting liver metastasis from a medullary thyroid carcinoma 24 years after the initial diagnosis of MTC.

Carcinoid syndrome and carcinoid crisis secondary to a metastatic carcinoid tumour of the lung: a therapeutic challenge.

We describe a 53-year-old male patient, with a known history of metastatic carcinoid tumour of the lung, who developed a variety of symptoms of the carcinoid syndrome and subsequently a carcinoid crisis. Although bronchial carcinoid tumours are very rarely associated with symptoms of the carcinoid syndrome, a subset may develop a severe hypersecretory syndrome and exhibit an aggressive behaviour. In cases with excessive tumour load and difficult-to-control hypersecretory syndrome, management by a specialized multidisciplinary team using evidence-based regimens is mandatory to deal with the life-threatening carcinoid crisis, to improve patients' outcome and quality of life.

Galectin-3 as a marker distinguishing functioning from silent corticotroph adenomas.

OBJECTIVE: Galectin-3 (Gal-3) belongs to the family of carbohydrate-binding proteins with high affinity for galactoside and is involved in many biological processes including cell growth and differentiation, cell adhesion, tumor progression, apoptosis and metastasis. The aim of this study was to disclose differences in the expression of Gal-3 in silent and functioning corticotroph pituitary adenomas. DESIGN: We examined 30 pituitary adenomas (19 functioning corticotroph, 11 silent corticotroph adenomas). Two prolactinomas and 2 functioning somatotroph adenomas served as positive controls. Antigen retrieval was done by three-minute incubation via pressure boiler in citrate buffer solution, pH 6.0. A polymer was used as a secondary link to DAB chromogen. The independent variables t-test was used for comparison of the mean expression of Gal-3 in the two different corticotroph adenoma subgroups. RESULTS: Eighteen of the functioning corticotroph adenomas (94.73%) were positive for Gal-3 with a cytoplasmic and focally membranous distribution; two cases also exhibited nuclear expression, whereas 9 of the silent corticotroph adenomas (81.81%) had zero or<1% expression of Gal-3 (p=0.001). CONCLUSIONS: Gal-3 is highly expressed in functioning corticotroph adenomas of the pituitary gland, while silent adenomas exhibit very focal to null expression of Gal-3. This observation can be used in the pathological diagnosis to separate functioning from silent corticotroph adenomas of the pituitary.

The gonadotroph origin of null cell adenomas.

OBJECTIVE: The term "null cell" adenoma was first proposed in 1980 to designate pituitary adenomas lacking clinical, biochemical and morphological markers to disclose their cell origin. DESIGN: The aim of this study was to investigate the presence of α- and ß-gonadotropin subunits in clinically nonfunctioning pituitary tumors, which were initially immunonegative and thus diagnosed as null cell adenomas. For this reason, we reapplied immunohistochemistry using a more sensitive method comprising a tyramide signal amplification technique, combined with a polymer antibody immunohistochemical detection system. RESULTS: With this approach, all these previously negative tumors became positive for α- and ß-gonadotropin hormone subunits. CONCLUSIONS: Our results prove that so-called "null cell" adenomas produce α-SU or/and ß-FSH or ß-LH and therefore are gonadotrph adenomas in origin.