Why don’t corticotroph tumors always produce Cushing’s disease?

Objective Silent corticotroph tumors are a pituitary neuroendocrine tumor subtype of corticotroph lineage that do not clinically express Cushing’s disease. The silencing of this type of tumor is not fully understood. The aim of the present study was to delve into the lack of secretory activity, studying the post-transcriptional and post-translational regulation of POMC/ACTH in a series of molecularly identified functioning and silent corticotroph tumors. Design We analyzed 24 silent corticotroph, 23 functioning corticotroph and 25 silent gonadotroph tumors. Methods We used Sanger sequencing, quantitative real-time PCR and Western blot to analyze genetic alterations in POMC, gene expression of TBX19, NEUROD1, POMC, PCSK1, PCSK2, CPE and PAM and protein expression of POMC, PC1/3, PC2, CPE and PAM. Results We found different polymorphisms in the POMC gene of corticotroph tumors, some of them related to deficiency of proopiomelanocortin. Silent corticotroph tumors showed lower PC1/3 gene and protein expression than functioning ones, especially compared to micro-functioning corticotroph tumors (all P < 0.05). Moreover, we found a positive correlation between PC2 and CPE gene and protein expression (rho ≥ 0.670, P < 0.009) in silent corticotroph tumors compared with functioning ones. Conclusions By studying the post-transcriptional and post-translational processing of POMC and ACTH, respectively, in a large series of silent and functioning corticotroph tumors, we found that the lack of secretory activity of these tumors is related to an impaired processing of POMC and a high degradation of ACTH, with the macro-functioning corticotroph tumor behaving as an intermediate state between micro-functioning and silent corticotroph tumors.

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Epigenetic regulation of somatostatin and somatostatin receptors in neuroendocrine tumors and other types of cancer

Reviews in Endocrine and Metabolic Disorders ◽

10.1007/s11154-020-09607-z ◽

2020 ◽

Author(s):

M.J. Klomp ◽

S.U. Dalm ◽

M. de Jong ◽

R.A. Feelders ◽

J. Hofland ◽

...

Keyword(s):

Protein Expression ◽

Neuroendocrine Tumors ◽

Epigenetic Regulation ◽

Somatostatin Receptors ◽

Secretory Activity ◽

Expression Levels ◽

Epigenetic Machinery ◽

Gene And Protein Expression

Abstract Both somatostatin (SST) and somatostatin receptors (SSTRs) are proteins with important functions in both physiological tissue and in tumors, particularly in neuroendocrine tumors (NETs). NETs are frequently characterized by high SSTRs expression levels. SST analogues (SSAs) that bind and activate SSTR have anti-proliferative and anti-secretory activity, thereby reducing both the growth as well as the hormonal symptoms of NETs. Moreover, the high expression levels of SSTR type-2 (SSTR2) in NETs is a powerful target for therapy with radiolabeled SSAs. Due to the important role of both SST and SSTRs, it is of great importance to elucidate the mechanisms involved in regulating their expression in NETs, as well as in other types of tumors. The field of epigenetics recently gained interest in NET research, highlighting the importance of this process in regulating the expression of gene and protein expression. In this review we will discuss the role of the epigenetic machinery in controlling the expression of both SSTRs and the neuropeptide SST. Particular attention will be given to the epigenetic regulation of these proteins in NETs, whereas the involvement of the epigenetic machinery in other types of cancer will be discussed as well. In addition, we will discuss the possibility to target enzymes involved in the epigenetic machinery to modify the expression of the SST-system, thereby possibly improving therapeutic options.

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OR06-01 The Role of Germline Defects in Cushing’s Disease

Journal of the Endocrine Society ◽

10.1210/jendso/bvaa046.323 ◽

2020 ◽

Vol 4 (Supplement_1) ◽

Author(s):

Laura C Hernández-Ramírez ◽

Nathan Pankratz ◽

John Lane ◽

Fabio R Faucz ◽

Prashant Chittiboina ◽

...

Keyword(s):

Genetic Testing ◽

Family History ◽

Cushing’S Disease ◽

Sanger Sequencing ◽

Disease Onset ◽

Cushing's Disease ◽

Incomplete Penetrance ◽

Genetic Causes ◽

Sporadic Cases ◽

Hotspot Mutations

Abstract J.L.M. and C.A.S. contributed equally. Introduction: Cushing’s Disease (CD) has been described as a component of a number of familial genetic syndromes. Yet, such cases may go largely unrecognized due to phenotypic variability, incomplete penetrance, and the rarity of the disease. We determined the frequency and type of germline genetic causes of CD and characterized their clinical phenotype in a large cohort of CD patients. Methods: We studied 245 unrelated CD patients (139 females, 60.4%), referred to our Center between 1997–2018, including 230 pediatric (≤18 years at disease onset, 93.9%) and 15 adult patients (6.1%). Germline genetic causes were identified by whole exome sequencing in 184 patients and by Sanger sequencing of specific genes in 39 patients; 22 patients did not undergo genetic testing, due to low quality or insufficient DNA. When available (n=66), corticotropinoma DNA was screened for USP8 hotspot variants using Sanger sequencing. Results: Eighteen patients (7.3%) had positive family history: nine presented as FIPA with unknown genetic cause, eight presented as MEN1 (seven had confirmed MEN1 variants), and one had a family history of pheochromocytoma/paraganglioma and pituitary adenoma with unknown genetic cause. Among the 227 sporadic patients (92.7%), 13 (5.7%) simplex cases had putative pathogenic variants in the following genes: CDKN1B (n=5), CABLES1 (n=3), AIP (n=1), PRKAR1A (n=1),TP53 (n=1),TSC2 (n=1), and USP8 (n=1). Altogether, cases with potentially inheritable genetic causes(familial and simplex) accounted for 12.7% (31/245) of all patients. There were no statistically significant differences in age at disease onset, age at diagnosis or tumor diameter between patients with potentially inheritable genetic defects and the rest of the cohort. In the pediatric subset, however, there was a non-significant higher frequency of macroadenomas among familial and simplex patients (21.4%), compared with sporadic patients (12.4%, P=0.19). Somatic USP8 hotspot mutations were found in 33.3% (12/36) of sporadic patients, but only in 3.3% (1/30) of familial and simplex (e.g. patients with a disease-associated germline defect, but no affected relatives) cases (P=0.0038). The global frequency of USP8 defects was 19.7% (13/66). Conclusions: Potentially inheritable cases of CD accounted for one-eighth of the patients in our cohort: 64.5% (20/31) of them are associated with defects in genes with a known involvement in CD. Patients with germline genetic causes of CD might present as apparently sporadic cases, due variability in disease penetrance. Unlike sporadic cases, somatic USP8 hotspot mutations are rare in those with inheritable causes of CD, suggesting different drivers for tumorigenesis in each group. Identifying the genetic causes of CD should lead to a more precise genetic testing and counselling and might aid in developing targeted therapeutic strategies.

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Analysis of Gene and Protein Expression in Atherosclerotic Mouse Aorta by Western Blot and Quantitative Real-Time PCR

Methods in Molecular Biology - Methods in Mouse Atherosclerosis ◽

10.1007/978-1-4939-2929-0_21 ◽

2015 ◽

pp. 309-322 ◽

Cited By ~ 3

Author(s):

José Rivera-Torres

Keyword(s):

Protein Expression ◽

Western Blot ◽

Real Time ◽

Real Time Pcr ◽

Quantitative Real Time Pcr ◽

Gene And Protein Expression ◽

Mouse Aorta

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The activation of somatostatinergic receptors by either somatostatin-14 or cortistatin-17 often inhibits ACTH hypersecretion in patients with Cushing’s disease

Acta Endocrinologica ◽

10.1530/eje-07-0147 ◽

2007 ◽

Vol 157 (4) ◽

pp. 393-398 ◽

Cited By ~ 15

Author(s):

R Giordano ◽

A Picu ◽

L Bonelli ◽

F Broglio ◽

F Prodam ◽

...

Keyword(s):

Cushing’S Disease ◽

Secretory Activity ◽

Cushing's Disease ◽

Receptor Subtypes ◽

Inhibitory Effects ◽

Glucose Levels ◽

Central Actions ◽

The Individual ◽

Somatostatin 14

Object: Somatostatin (SS) is known to inhibit GH and insulin, while its effect on corticotrope secretion is controversial: inhibition of ACTH secretion by agonists activating somatostatinergic receptors (sst)-2 and sst-5 was reported in vitro. Cortistatin (CST) not only binds all sst receptor subtypes but also possesses central actions that are not shared by SS. Design: In nine patients with Cushing’s disease (CD), ACTH, cortisol, GH, insulin, and glucose levels were studied during 120-min i.v. infusion of SS-14 (2.0 μg/kg per h), CST-17 (2.0 μg/kg per h) or saline. Results: Both SS or CST significantly affected the hypothalamic–pituitary–adrenal axis. Cortisol was decreased to the same extent by either SS or CST (P < 0.05). Both SS and CST decreased ACTH, although statistical difference was reached only during CST (P < 0.05). Analyzing the individual responses as Δareas under curve (ΔAUCs), a clear and consensual inhibition of ACTH and cortisol under either SS or CST was recorded in five out of nine patients. Both SS or CST inhibited (P < 0.05) insulin, that even showed a rebound (P < 0.01) at the end of infusion. GH was not modified by either peptide. Conclusion: SS and CST often display similar inhibitory effects on the HPA axis in CD. The activation of sst receptors by both peptides is followed in almost 50% of patients by a remarkable inhibition of ACTH and cortisol hypersecretion. These findings reinforce the view that sst receptors are involved in the control of the secretory activity of tumoral corticotropic cells.

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SAT-300 Evading Death: Noxa in Cushing’s Disease Pituitary Adenomas

Journal of the Endocrine Society ◽

10.1210/jendso/bvaa046.1312 ◽

2020 ◽

Vol 4 (Supplement_1) ◽

Author(s):

Reinier Alvarez ◽

Konstantinos Floros ◽

Kory Johnson ◽

Abdel Elkahloun ◽

Weiwei Wu ◽

...

Keyword(s):

Cell Death ◽

Protein Expression ◽

Cushing’S Disease ◽

Fold Increase ◽

Cushing's Disease ◽

Benign Tumors ◽

Human Pituitary ◽

Human Pituitary Adenoma ◽

Altered Expression ◽

Normal Gland

Abstract Introduction: Recurrence of Cushing’s disease (CD) caused by benign pituitary microadenomas are challenging clinical problems. Mechanisms underlying adenoma formation and recurrence remain unknown. PMAIP1 gene codes for Noxa, a Bcl-2 homology 3 (BH3) pro-apoptotic protein frequently downregulated in malignant human tumors.1-6 The role of dysregulated apoptosis remains largely unknown in benign tumors and in CD. We hypothesized that altered expression of Noxa protein is a pro-survival adaptation employed by CD adenomas. Methods: Syngeneic human pituitary adenoma and adjacent normal gland pairs (n=2), and an additional CD adenoma were analyzed with RNAseq. 10 CD, 1 growth hormone (GH) and 1 non-functioning adenoma (NFPA) underwent immunohistochemical (IHC) analysis for Noxa expression, which was graded by a neuropathologist as 0=none, 1=light, 2=medium, 3=strong. Staining grade represents relative protein expression. Results: Compared to adjacent normal pituitary tissue, we found that adenomas (n = 3) had a 3.76 fold increase in PMAIP1 mRNA. However, there was attenuated Noxa IHC staining in adenomas compared to normal pituitary in 8 of 10 CD patients (2:3, respectively), but similar staining in 2 of 10 CD patients (2:2 and 2-3:2-3). In GH and NFPA, we found similar patterns of Noxa suppression in the adenomas compared to the normal gland. Conclusion: Despite elevated PMAIP1 (Noxa) gene expression in adenomas compared to adjacent normal gland in CD, protein expression was reduced in adenomas. This downregulation of Noxa protein expression may contribute to reduced apoptosis of tumor cells. These findings suggest that CD adenomas gain pro-survival advantage by downregulating Noxa protein at post-transcriptional or post-translational level. References 1. Escobar, D. et al. Cell Death Dis.6, 1-14 (2015).2. Brinkmann, K. et al. Cell Rep.3, 881-891 (2013).3. Liu, Y. L. et al. Oncotarget5, 11237-11251 (2014).4. Dengler, M. A. et al. Cell Death Dis.5, 1-10 (2014).5. Liang, L. et al. J. Oral Pathol. Med.48, 52-59 (2019).6. Tahir, S. K. et al. Cancer Res.67, 1176-1183 (2007).

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