Though L-amino acids predominate in living organisms, substantial levels of free D-serine and D-aspartate occur in mammals, especially in nervous and endocrine tissues. Using an antibody specific for glutaraldehyde-fixed D-aspartate, we have localized D-aspartate in rat tissues. In the brain we observe discrete neuronal localizations of Daspartate, especially in the external plexiform layer of the olfactory bulb, hypothalamic supraoptic and paraventricular nuclei, the medial habenula, and certain brainstem nuclei. In rats 3-4 weeks old, we observe D-aspartate in septal nuclei and in a subset of stellate and basket cells of the cerebellum. D-aspartate is also concentrated in glands, including the epinephrine cells of the adrenal medulla, the posterior pituitary, and the pineal gland. Levels in the pineal gland are the highest of any mammalian tissue. Although D-amino acids are well known in bacterial physiology, only recently have they been found in mammals (1-3). Substantial levels of D-serine occur in brain tissue with a regional distribution resembling the the N-methyl-D-aspartate (NMDA) subtype of glutamate receptors (4). By immunohistochemistry we have shown that D-serine is selectively localized to a subpopulation of astrocytes in close proximity to NMDA receptors and released by glutamate stimulation (5, 6). Since D-serine potently activates the glycine site on NMDA receptors, these findings indicate that D-serine is a novel messenger molecule that serves as an endogenous ligand for this site.Lajtha and coworkers (1) discovered very high levels of D-aspartate in the brain and other tissues of mammals. Daspartate levels are highest in neonatal tissues, attaining millimolar concentrations in newborn rat cerebral cortex, pituitary gland, and 3-week-old adrenal gland (7,8). Using immunohistochemistry, we now describe selective neuronal localizations of D-aspartate in discrete brain areas and endocrine structures. D-aspartate oxidase (DAPOX), visualized by a novel histochemical technique, is localized inversely to endogenous D-aspartate. MATERIALS AND METHODSMaterials. Sprague-Dawley rats were purchased from Sasco (Wilmington, MA). Hypophysectomized rats were from Charles River Breeding Laboratories. Mice (129͞SvEv Agouti strain) were from Taconic Farms. Antibody to L-aspartate (9) was from Chemicon. Glutaraldehyde (GA) was from Electron Microscopy Sciences (Fort Washington, PA). The peroxidase Elite staining kit was from Vector Laboratories. All other reagents were from Sigma.Antibody Production and Purification. D-aspartate was coupled to BSA with GA and then reduced with NaBH 4 (10). Rabbits were immunized using a colloidal gold technique (5, 11), and stereoselective, high-affinity antiserum was produced after 7 weeks. Before use, batches of D-aspartate antiserum were diluted 1:10 with 0.05% NaN 3 and 10 mM Tris (pH 7.4). Diluted serum (10 ml) was mixed with 1 ml packed agarose beads that had been coupled to GA-BSA at a concentration of 4.3 mg͞ml with respect to BSA. Antiserum and beads were incubate...

Cushing disease is a condition in which the pituitary gland releases excessive adrenocorticotropic hormone (ACTH) as a result of an adenoma arising from the ACTH-secreting cells in the anterior pituitary. ACTH-secreting pituitary adenomas lead to hypercortisolemia and cause significant morbidity and mortality. Pituitarydirected medications are mostly ineffective, and new treatment options are needed. As these tumors express EGFR, we tested whether EGFR might provide a therapeutic target for Cushing disease. Here, we show that in surgically resected human and canine corticotroph cultured tumors, blocking EGFR suppressed expression of proopiomelanocortin (POMC), the ACTH precursor. In mouse corticotroph EGFR transfectants, ACTH secretion was enhanced, and EGF increased Pomc promoter activity, an effect that was dependent on MAPK. Blocking EGFR activity with gefitinib, an EGFR tyrosine kinase inhibitor, attenuated Pomc expression, inhibited corticotroph tumor cell proliferation, and induced apoptosis. As predominantly nuclear EGFR expression was observed in canine and human corticotroph tumors, we preferentially targeted EGFR to mouse corticotroph cell nuclei, which resulted in higher Pomc expression and ACTH secretion, both of which were inhibited by gefitinib. In athymic nude mice, EGFR overexpression enhanced the growth of explanted ACTH-secreting tumors and further elevated serum corticosterone levels. Gefitinib treatment decreased both tumor size and corticosterone levels; it also reversed signs of hypercortisolemia, including elevated glucose levels and excess omental fat. These results indicate that inhibiting EGFR signaling may be a novel strategy for treating Cushing disease.

After validation, this classification may be useful to accurately identify acromegaly patients with distinctive patterns of disease aggressiveness and outcome, as well as to provide an accurate tool for selection criteria in clinical studies.

Objective: Clinically nonfunctioning pituitary adenoma (NFPA) remains the only pituitary tumor subtype for which no effective medical therapy is available or recommended. We evaluated dopamine agonist (DA) therapy for preventing growth of postsurgical pituitary tumor remnants. Design: The study design included historical cohort analysis of clinical results at two pituitary referral centers with different standard practices for postoperative NFPA management: DA therapy or conservative follow-up. Methods: Seventy-nine patients followed for 8.8 ± 6.5 years were treated with DA, initiated upon residual tumor detection on postoperative MRI (preventive treatment (PT) group, n = 55), or when tumor growth was subsequently detected during follow-up (remedial treatment (RT) group, n = 24). The control group (n = 60) received no medication. Tumoral dopamine and estrogen receptor expression assessed by quantitative RT-PCR and immunostaining were correlated with response to treatment. Results: Tumor mass decreased, remained stable, or enlarged, respectively, in 38, 49, and 13% of patients in the PT group, and in 0, 53, and 47% of control subjects; shrinkage or stabilization was achieved in 58% of enlarging tumors in the RT group, P < 0.0001. Fifteen-year progression-free survival rate was 0.805, 0.24, and 0.04, respectively, for PT, RT, and control groups (P < 0.001). About 42% of patients in the control group required additional surgery or radiotherapy, compared with 38 and 13% subjects in the RT and PT groups, respectively (P = 0.002). Outcome measures were not related to NFPA D2R abundance. Conclusions: Dopamine agonist therapy in patients with NFPA is associated with decreased prevalence of residual tumor enlargement after transsphenoidal surgical resection.

Silent corticotrophins adenomas (SCAs) are clinically silent and non-secreting but immunostain positively for ACTH. We hypothesize that SCAs comprise both corticotroph and gonadotroph characteristics. Cohort analysis from 1994-2008 with follow-up time ranging from 1-15 years in a tertiary referral center. We compared preoperative and postoperative clinical results and tumor cytogenesis in 25 SCAs and 84 nonfunctioning adenomas in 109 consecutive patients diagnosed pre-operatively with nonfunctioning pituitary adenomas. Clinical outcomes were radiologic and hormonal measures. Pathologic outcomes were expression of relevant pituitary hormones, tissuespecific transcription factors, and electron microscopy features. Preoperative SCA presentation was similar to that observed for nonfunctioning adenomas. However, SCAs recurred postoperatively at a median of 3 years vs. 8 years for nonfunctioning adenomas (p<0.0001). Fiftyfour percent of patients with SCAs had new onset postoperative hypopituitarism vs. 17% of nonfunctioning adenomas (p<0.025). SCAs (n=18) were immunopositive for ACTH, cytoplasmic and nuclear SF-1, NeuroD1, DAX-1, and alpha-gonadotropin subunit, but Tpit negative, and coexpression of tumor ACTH with either SF-1 or LH was detected. In contrast, functional corticotroph adenomas (n=11) were immunopositive for ACTH, nuclear SF-1, NeuroD1, and Tpit, but negative for DAX-1, a gonadotroph cell transcription factor. Gonadotroph adenomas (n=23) were immunonegative for ACTH and Tpit but positive for nuclear SF-1, NeuroD1, and DAX-1. SCA electron microscopy demonstrated ultrastructural features consistent with corticotroph and

Purpose-Silent corticotroph adenomas (SCAs) comprise 20% of all corticotroph adenomas and 3-19% of nonfunctioning adenomas (NFAs). As they do not manifest clinical or biochemical hypercortisolism, they are diagnosed after pathologic examination of resected tumor tissue demonstrates positive ACTH expression. While preoperative features are similar to those of NFAs, SCAs may have more cavernous sinus invasion. Further, patients with SCAs tend to have more frequent and earlier recurrences than those with NFAs, often necessitating multiple surgeries and other modalities of treatment. This article reviews the incidence, pathogenesis, and clinical behavior of SCAs.Methods-A systematic literature review was performed using PubMed for information regarding silent corticotroph adenomas.Results-Up to date findings regarding epidemiology, pathogenesis, pathology, clinical presentation, postoperative course, and management of patients with SCAs are presented.Conclusions-This review highlights the necessity of rigorous monitoring for recurrences and hypopituitarism in patients with SCAs.

Purpose As ErbB signaling is a determinant of prolactin synthesis, role of ErbB receptors was tested for prolactinoma outcomes and therapy. The objective of this study was to characterize ErbB receptor expression in prolactinomas and then perform a pilot study treating resistant prolactinomas with a targeted tyrosine kinase inhibitor (TKI). Methods Retrospective analysis of prolactinomas and pilot study for dopamine agonist resistant prolactinomas in tertiary referral center. We performed immunofluorescent staining of a tissue array of 29 resected prolactinoma tissues for EGFR, ErbB2, ErbB3, and ErbB4 correlated with clinical features. Two patients with aggressive resistant prolactinomas enrolled and completed trial. They received lapatinib 1250 mg daily for 6 months with tumor and hormone assessments. Main outcome measures were positive tumor staining of respective ErbB receptors, therapeutic reduction of prolactin levels and tumor shrinkage. Results Treated PRL levels and tumor volumes were suppressed in both subjects treated with TKI. EGFR expression was positive in 82% of adenomas, ErbB2 in 92%, ErbB3 in 25%, and ErbB4 in 71%, with ErbB2 score > EGFR>ErbB4>ErbB3. Higher ErbB3 expression was associated with optic chiasm compression (p = 0.03), suprasellar extension (p = 0.04), and carotid artery encasement (p= 0.01). Higher DA response rates were observed in tumors with higher ErbB3 expression. Conclusions Prolactinoma expression of specific ErbB receptors is associated with tumor invasion, symptoms, and response to dopamine agonists. Targeting ErbB receptors may be effective therapy in patients with resistant prolactinomas.

No abstract