Leonardo Vieira Neto

Quantitative analysis of somatostatin receptor subtypes (1-5) gene expression levels in somatotropinomas and correlation to in vivo hormonal and tumor volume responses to treatment with octreotide LAR

OBJECTIVE To determine whether the somatostatin receptor subtype (SSTR) expression profile correlates with hormonal and tumor volume responses to postsurgical octreotide long acting repeatable (OCT LAR) treatment. DESIGN AND METHODS Quantitative real-time RT-PCR was used to evaluate the absolute mRNA copy numbers for all five SSTR subtypes in 22 somatotropinomas. Response to OCT LAR was studied by hormone levels (GH and IGF-I) and tumor volume (sella turcica magnetic resonance imaging). RESULTS SSTR5 was present at the highest level followed by SSTR2, SSTR3, SSTR1, and SSTR4 (2327 (1046-5555), 2098 (194-23 954), 97 (0-460), 14 (0-29 480), and 0 (0-652) copies respectively). Positive correlations were found between SSTR2 levels and the percentage decrease of GH and IGF-I after 3 (r=0.49, P<0.027 and r=0.49, P<0.029 respectively) and 6 (r=0.59, P<0.006 and r=0.58, P<0.008 respectively) months of OCT LAR. A negative correlation was found between SSTR5 mRNA levels and the percentage decrease of GH after 3 months of OCT LAR (r=-0.52, P=0.016, n=21). A higher SSTR2/SSTR5 ratio was observed among patients who obtained hormonal control with OCT LAR, when compared with those uncontrolled (2.4 (0.7-10) vs 0.3 (0.1-7.7), P=0.001). A ROC curve analysis showed a SSTR2/SSTR5 ratio of 1.3 as the best predictor of disease control, with a sensitivity of 88% and a specificity of 92% - area under curve, 0.9. A positive correlation was also found between SSTR2 mRNA levels and the percentage decrease in tumor volume after 6 months of OCT LAR (r=0.79, P=0.002, n=12). CONCLUSIONS Somatostatin receptor subtype 2 mRNA expression levels in somatotropinomas correlate positively with in vivo hormonal and tumor volume responses to OCT LAR.

Expression Analysis of Dopamine Receptor Subtypes in Normal Human Pituitaries, Nonfunctioning Pituitary Adenomas and Somatotropinomas, and the Association between Dopamine and Somatostatin Receptors with Clinical Response to Octreotide-LAR in Acromegaly

CONTEXT Dopamine receptor (DR) and somatostatin receptor subtype expression in pituitary adenomas may predict the response to postsurgical therapies. OBJECTIVES Our objectives were to assess and compare the mRNA levels of DR1-5 and somatostatin receptors 1-5 in normal pituitaries (NPs), nonfunctioning pituitary adenomas (NFPAs), and somatotropinomas. In addition, we determined whether the level of DR expression correlates with the in vivo response to octreotide-LAR in acromegalic patients. DESIGN AND PATIENTS Eight NPs, 30 NFPAs, and 39 somatotropinomas were analyzed for receptor mRNA levels by real-time RT-PCR. The DR2 short variant was estimated as the DR2 long/DR2 total (DR2T). The relationship between DR expression and the postsurgical response to octreotide-LAR was assessed in 19 of the acromegalic patients. RESULTS DR3 was not detected. The relationship between expression levels of DR subtypes in NPs and somatotropinomas was DR2T>>>DR4>>DR5>DR1, whereas in NFPAs, DR2T>>>DR4>>DR1>DR5. The DR2 short variant was the predominant DR2 variant in the majority of samples. In acromegalics treated with octreotide-LAR, DR1 was negatively correlated with percent GH reduction (3 months: r = -0.67, P = 0.002; and 6 months: r = -0.58, P = 0.009), and DR5 was positively correlated with percent IGF-I reduction (3 months: r = 0.55, P = 0.01; and 6 months: r = 0.47, P = 0.04). CONCLUSIONS DR2 is the predominant DR subtype in NPs, NFPAs, and somatotropinomas. The fact that DR1, DR4, and DR5 are also expressed in many adenomas tested suggests that these receptors might also play a role in the therapeutic impact of postsurgical medical therapies in patients with NFPA and acromegaly. This was supported by the finding that the in vivo response to octreotide-LAR was negatively associated with DR1 and positively associated with DR5.

AIP expression in sporadic somatotropinomas is a predictor of the response to octreotide LAR therapy independent of SSTR2 expression.

mutation and/or a history of medical treatment with SRL before surgery. Biochemical assessment was performed 3 months after surgery by oral glucose tolerance test (OGTT) and evaluations of IGF1 levels. Patients were considered non-cured based on nonsuppressible GH levels on OGTT and plasma IGF1 levels higher than those of age-matched normal subjects. Medical therapy with OCT-LAR was started at a dose of 20 mg every 4 weeks, and the dose was increased to 30 mg in patients with non-controlled disease after 3 months of therapy. The efficacy of medical therapy was evaluated during the last patient visit. Patients were considered to have non-controlled disease if they had plasma IGF1 levels greater than those of age-matched normal subjects or basal GH levels O1.0 ng/ml after at least 6 months of treatment with OCT-LAR at a dosage of 30 mg. The AIP and SSTR2 expressions were analyzed through immunohistochemistry in paraffin-embedded

Low Aryl Hydrocarbon Receptor-Interacting Protein Expression Is a Better Marker of Invasiveness in Somatotropinomas than Ki-67 and p53

Background: Some pituitary adenomas exhibit fast growth and invade surrounding structures. To date, there is no robust marker to predict invasiveness. Aim: To evaluate Ki-67, p53 and aryl hydrocarbon receptor-interacting protein (AIP) expression and compare these between invasive and noninvasive somatotropinomas and nonfunctioning pituitary adenomas (NFPAs). Methods: Protein expression was determined by immunohistochemistry. Tumors were classified according to percentage of immunolabeled nuclei for Ki-67 and p53. AIP immunopositivity was graded according to a score encompassing pattern and intensity. Invasiveness was defined according to radiological and surgical criteria. Results: Thirty-eight sporadic somatotropinomas were studied. Median Ki-67 labeling index in invasive and noninvasive tumors was 1.6 (range 0–20.6) and 0.26 (0–2.2), respectively (p = 0.01). With a 2.3% cut-off point obtained by ROC curve analysis, invasive adenomas were distinguished with 100% specificity, 39% sensitivity, and 63% accuracy. Low AIP expression was also correlated with tumor invasiveness (p = 0.001), with sensitivity, specificity and accuracy of 78, 80, and 79%, respectively. Expression of p53 was not different among tumors. Twenty-nine NFPAs were studied, with no significant difference between Ki-67, p53 and AIP expression in invasive and noninvasive tumors. High AIP expression was more frequent in NFPAs, with Ki-67 >3% (p = 0.051), especially when only gonadotrope cell adenomas (n = 25) were considered (p = 0.012). Conclusions: These data suggest, for the first time, that AIP is a better marker of invasiveness in somatotropinomas than Ki-67 and p53. In addition, low AIP expression is observed in invasive somatotropinomas, in contrast with high AIP expression in NFPAs (mainly gonadotrope cell tumors) with high proliferative indices.

Efficacy of medical treatment in Cushing's disease: a systematic review

Reported rates of response to medical therapies used in Cushings disease (CD) vary widely. The aim of this review is to analyse systematically the efficacy of medical therapies for CD and to assess the strength of the supporting evidence.

Acromegaly Has a Negative Influence on Trabecular Bone, But Not on Cortical Bone, as Assessed by High-Resolution Peripheral Quantitative Computed Tomography

INTRODUCTION Acromegaly is one of the causes of secondary osteoporosis, although studies of bone mineral density (BMD) have yielded conflicting results and none of them have evaluated the bone properties. OBJECTIVES AND PATIENTS Our objective was to correlate, in a cohort of 82 acromegalic patients, BMD and bone microarchitecture, using dual-energy x-ray absorptiometry and high-resolution peripheral quantitative computed tomography, with the presence of type 2 diabetes mellitus (T2DM), disease activity, and gonadal status and to compare these bone parameters between 45 eugonadal acromegalic patients and 45 healthy controls. RESULTS Acromegalic patients with T2DM had lower trabecular density and trabecular bone volume to tissue volume ratio in the distal tibia. Patients with active acromegaly exhibited a higher BMD and T-score in the lumbar spine (P = .02 for both) and a higher cortical density in the distal tibia when compared with those with controlled acromegaly (P = .001). After multiple linear regression (including age, presence of T2DM, acromegaly activity, and gonadal status), eugonadism remained the main determinant of bone parameters. The 45 acromegalic patients with eugonadism were compared with 45 age- and sex-matched controls and exhibited lower trabecular densities and impaired microstructures. CONCLUSIONS Acromegaly appears to have a deleterious effect on trabecular bone microarchitecture, and in this specific population, the gonadal status might be more important than T2DM or acromegaly activity in determining bone health. High-resolution peripheral quantitative computed tomography seems promising for evaluating acromegalic bone properties and for addressing the limitations posed by dual-energy x-ray absorptiometry.

Truncated somatostatin receptor variant sst5TMD4 confers aggressive features (proliferation, invasion and reduced octreotide response) to somatotropinomas

The GH/IGF1 response of somatotropinomas to somatostatin analogues (SSA) is associated with their pattern of somatostatin receptor (sst1-sst5) expression. Recently, we demonstrated that expression of a truncated sst5-variant (sst5TMD4) can influence the secretory response of somatotropinomas to SSA-therapy; however, its potential relationship with aggressive features (e.g. invasion/proliferation) is still unknown. Here, we show that sst5TMD4 is present in 50% of non-functioning pituitary-adenomas (NFPA) (n = 30) and 89% of somatotropinomas (n = 36), its expression levels being highest in somatotropinomas > > NFPAs > > > normal pituitaries (negligible expression; n = 8). In somatotropinomas, sst5TMD4 mRNA and protein levels correlated positively, and its expression was directly associated with tumor invasiveness (cavernous/sphenoid sinus), and inversely correlated with age and GH/IGF1 reduction after 3-6 months with octreotide-LAR therapy. GNAS+ somatotropinomas expressed lower sst5TMD4 levels. ROC analysis revealed sst5TMD4 expression as the only marker, within all sst-subtypes, capable to predict tumor invasiveness in somatotropinomas. sst5TMD4 overexpression increased cell viability in cultured somatotropinoma (n = 5). Hence, presence of sst5TMD4 associates with increased aggressive features and worse prognosis in somatotropinomas, thereby providing a potentially useful tool to refine somatotropinoma diagnosis, predict outcome of clinical response to SSA-therapy and develop new therapeutic targets.

Dopamine receptor subtype 2 expression profile in nonfunctioning pituitary adenomas and in vivo response to cabergoline therapy

To determine the dopamine receptor subtype 2 (DR2) mRNA levels and protein expression and to evaluate the effect of adjuvant cabergoline therapy on tumour volume (TV) in patients with postoperative residual nonfunctioning pituitary adenoma (NFPA).

Adverse effects of glucocorticoids: coagulopathy.

Hypercortisolism is associated with various systemic manifestations, including central obesity, arterial hypertension, glucose intolerance/diabetes mellitus, dyslipidemia, nephrolithiasis, osteoporosis, gonadal dysfunction, susceptibility to infections, psychiatric disorders, and hypercoagulability. The activation of the hemostatic system contributes to the development of atherosclerosis and subsequent cardiovascular morbidity and mortality. Previous studies have identified an increased risk of both unprovoked and postoperative thromboembolic events in patients with endogenous and exogenous Cushings syndrome (CS). The risk for postoperative venous thromboembolism in endogenous CS is comparable to the risk after total hip or knee replacement under short-term prophylaxis. The mechanisms that are involved in the thromboembolic complications in hypercortisolism include endothelial dysfunction, hypercoagulability, and stasis (Virchows triad). It seems that at least two factors from Virchows triad must be present for the occurrence of a thrombotic event in these patients. Most studies have demonstrated that this hypercoagulable state is explained by increased levels of procoagulant factors, mainly factors VIII, IX, and von Willebrand factor, and also by an impaired fibrinolytic capacity, which mainly results from an elevation in plasminogen activator inhibitor 1. Consequently, there is a shortening of activated partial thromboplastin time and increased thrombin generation. For these reasons, anticoagulant prophylaxis might be considered in patients with CS whenever they have concomitant prothrombotic risk factors. However, multicenter studies are needed to determine which patients will benefit from anticoagulant therapy and the dose and time of anticoagulation.

The Role of Temozolomide in the Treatment of a Patient with a Pure Silent Pituitary Somatotroph Carcinoma

OBJECTIVE To describe a case of a pure silent somatotroph pituitary carcinoma. METHODS We describe a 54-year-old female with a clinically nonfunctioning pituitary macroadenoma diagnosed 15 years earlier. RESULTS The patient underwent transsphenoidal surgery and no visible tumor remnant was observed for 6 years. A magnetic resonance imaging (MRI) detected the recurrence of a 1.2 × 1.5 cm macroadenoma. The patient was submitted to conventional radiotherapy (4500 cGy), and the tumor volume remained stable for 7 years. Then, an MRI revealed a slight increase in tumor size, and 2 years later, a subsequent MRI detected a very large, invasive pituitary mass. The patient was resubmitted to transsphenoidal surgery, and the histopathological examination showed diffuse positivity for growth hormone (GH). The nadir GH level during an oral glucose tolerance test was 0.06 ng/mL, and the pre- and postoperative insulin like growth factor type I (IGF-I) levels were within the normal range. Abdominal, chest, brain, and spine MRI showed multiple small and hypervascular liver and bone lesions suggestive of metastases. Liver biopsy confirmed metastasis of GH-producing pituitary carcinoma. The patient has been treated with temozolomide and zoledronic acid for 7 months and with octreotide long-acting release (LAR) for 4 months. The primary tumor and metastases are stable. CONCLUSION Despite being an extremely rare event, pituitary carcinoma may develop several years after the successful treatment of even a silent GH-producing pituitary adenoma, which suggests that close long-term follow-up is necessary.