Pediatric Cushing disease: disparities in disease severity and outcomes in the Hispanic and African-American populations.

BackgroundLittle is known about the contribution of racial and socioeconomic disparities to severity and outcomes in children with Cushing disease (CD).MethodsA total of 129 children with CD, 45 Hispanic/Latino or African-American (HI/AA) and 84 non-Hispanic White (non-HW), were included in this study. A 10-point index for rating severity (CD severity) incorporated the degree of hypercortisolemia, glucose tolerance, hypertension, anthropomorphic measurements, disease duration, and tumor characteristics. Race, ethnicity, age, gender, local obesity prevalence, estimated median income, and access to care were assessed in regression analyses of CD severity.ResultsThe mean CD severity in the HI/AA group was worse than that in the non-HW group (4.9±2.0 vs. 4.1±1.9, P=0.023); driving factors included higher cortisol levels and larger tumor size. Multiple regression models confirmed that race (P=0.027) and older age (P=0.014) were the most important predictors of worse CD severity. When followed up a median of 2.3 years after surgery, the relative risk for persistent CD combined with recurrence was 2.8 times higher in the HI/AA group compared with that in the non-HW group (95% confidence interval: 1.2-6.5).ConclusionOur data show that the driving forces for the discrepancy in severity of CD are older age and race/ethnicity. Importantly, the risk for persistent and recurrent CD was higher in minority children

Rare Germline DICER1 Variants in Pediatric Patients With Cushing's Disease: What Is Their Role?

Context: The DICER1 syndrome is a multiple neoplasia disorder caused by germline mutations in the DICER1 gene. In DICER1 patients, aggressive congenital pituitary tumors lead to neonatal Cushing's disease (CD). The role of DICER1 in other corticotropinomas, however, remains unknown. Objective: To perform a comprehensive screening for DICER1 variants in a large cohort of CD patients, and to analyze their possible contribution to the phenotype. Design, setting, patients, and interventions: We included 192CD cases: ten young-onset (age <30 years at diagnosis) patients were studied using a next generation sequencing panel, and 182 patients (170 pediatric and 12 adults) were screened via whole-exome sequencing. In seven cases, tumor samples were analyzed by Sanger sequencing. Results: Rare germline DICER1 variants were found in seven pediatric patients with no other known disease-associated germline defects or somatic DICER1 second hits. By immunohistochemistry, DICER1 showed nuclear localization in 5/6 patients. Variant transmission from one of the parents was confirmed in 5/7 cases. One patient had a multinodular goiter; another had a family history of melanoma; no other patients had a history of neoplasms. Conclusions: Our findings suggest that DICER1 gene variants may contribute to the pathogenesis of non-syndromic corticotropinomas. Clarifying whether DICER1 loss-of-function is disease-causative or a mere disease-modifier in this setting, requires further studies.This work was supported by the Intramural Research Programs of Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD) and National Institute for Neurological Diseases and Stroke, National Institutes of Health, a grant from the Basque Department of Education (IT795-13), a grant from the Basque Department of Health (GV2018111082), the Merck Serono Research award from Fundacion Salud 2000 (15-EP-004) and the Jose Igea 2018 grant, sponsored by Pfizer, from Fundacion Sociedad Espanola de Endocrinologia Pediatrica (SEEP)

Canonical and Non-Canonical Roles of PFKFB3 in Brain Tumors

PFKFB3 is a bifunctional enzyme that modulates and maintains the intracellular concentrations of fructose-2,6-bisphosphate (F2,6-P2), essentially controlling the rate of glycolysis. PFKFB3 is a known activator of glycolytic rewiring in neoplastic cells, including central nervous system (CNS) neoplastic cells. The pathologic regulation of PFKFB3 is invoked via various microenvironmental stimuli and oncogenic signals. Hypoxia is a primary inducer of PFKFB3 transcription via HIF-1alpha. In addition, translational modifications of PFKFB3 are driven by various intracellular signaling pathways that allow PFKFB3 to respond to varying stimuli. PFKFB3 synthesizes F2,6P2 through the phosphorylation of F6P with a donated PO4 group from ATP and has the highest kinase activity of all PFKFB isoenzymes. The intracellular concentration of F2,6P2 in cancers is maintained primarily by PFKFB3 allowing cancer cells to evade glycolytic suppression. PFKFB3 is a primary enzyme responsible for glycolytic tumor metabolic reprogramming. PFKFB3 protein levels are significantly higher in high-grade glioma than in non-pathologic brain tissue or lower grade gliomas, but without relative upregulation of transcript levels. High PFKFB3 expression is linked to poor survival in brain tumors. Solitary or concomitant PFKFB3 inhibition has additionally shown great potential in restoring chemosensitivity and radiosensitivity in treatment-resistant brain tumors. An improved understanding of canonical and non-canonical functions of PFKFB3 could allow for the development of effective combinatorial targeted therapies for brain tumors

Enhanced 5-ALA Induced Fluorescence in Hormone Secreting Pituitary Adenomas

Introduction Cushing’s Disease (CD) is caused by millimeter-sized corticotropinomas (microadenomas) that lead to supraphysiological levels of glucocorticoid. Up to 40% of microadenomas are not visualized on gold-standard MR imaging. Pituitary adenomas metabolize exogenous 5-ALA (an endogenous metabolite) to protoporphyrin IX (PpIX) at rates 20-50 times higher compared with normal tissues. PpIX intensely fluoresces red (635nm) when excited with blue light (375-440nm), enabling its use as an intraoperative fluorescence imaging agent. 5-ALA is now an FDA approved prodrug. We examined the efficacy of ALA-induced-PpIX fluorescence in human derived adenomatous and normal pituitary samples. We explored the modulation of PpIX conversion with CRH or dexamethasone (DEX), and subcellular localization of PpIX. Methods We used flow cytometry for PpIX intensity analysis. A human-derived corticotropinoma, it’s adjacent normal gland, murine normal pituitary cells, and AtT20 cells were incubated with 5-ALA (300 nM) with/without DEX (1µM) or CRH (50nM). For confocal microscopy, live cells imaged for PpIX (405nm/615nm) and mitochondrial (550nm/615nm) fluorescence. Results We found a 10-fold-increase in 5-ALA induced PpIX fluorescence intensity in human-derived adenomatous compared to adjacent normal pituitary tissue (n=1, p\u3c0.05). AtT-20 cell lines (n=6, p\u3c0.05) fluoresced 7-fold more intensely compared to normal murine pituitary tissue (n=3, p\u3c0.05). The addition of DEX, before or after 5-ALA exposure, increased the fluorescence intensity by 31% (n=4, p\u3c0.05). The addition of CRH did not have a significant effect on 5-ALA fluorescence (n=3, p\u3c0.05). We saw localization of 5-ALA to mitochondria, and mitochondrial disruption in 5-ALA treated At-T20s. Conclusions Our results support the use of 5-ALA for fluorescence guided resection in hormone secreting microadenomas. The supraphysiological levels of glucocorticoids, as seen in CD, may enhance the 5-ALA fluorescence in corticotropinomas. We confirm the mitochondrial localization and disruption by 5-ALA, a basis for photodynamic therapy

Petrous Carotid Exposure with Eustachian Tube Preservation: A Morphometric Elucidation

Inadvertent injury to eustachian tube leading to cerebrospinal fluid rhinorrhea is a known complication associated with drilling of Glasscock's triangle to expose the horizontal petrous internal carotid artery (ICA) for management of difficult tumors (especially malignant) or aneurysms at the cranial base. Contrary to the usual approach, we hypothesize that a “medial-to-lateral” approach to Glasscock's triangle drilling will minimize eustachian tube injury. Four formalin-fixed human cadaveric heads were dissected, and underwent appropriate morphometric analysis; yielding a total of eight datasets. The diameter of the horizontal petrous ICA exposed was 4.7 ± 0.9 mm (range, 3.8 to 5.6 mm).The mean distance from the medial carotid wall midpoint to the medial-most point on the eustachian tube was 6.35 ± 0.58 mm (range, 5.4 to 7.1 mm), yielding a “safety zone” for eustachian tube, ranging 0.2 to 1.9 mm lateral to the lateral carotid wall. With the medial-to-lateral approach, the eustachian tube remained preserved in all the specimens. The results of our study provide a practical, consistent, and safe method of maximizing horizontal petrous carotid artery exposure while minimizing the eustachian tube injury

3D Volumetric Measurements of GH Secreting Adenomas Correlate with Baseline Pituitary Function, Initial Surgery Success Rate, and Disease Control

There is scarce data on the clinical utility of volume measurement for growth hormone (GH)-secreting pituitary adenomas. The current study objective was to assess the association between pituitary adenoma volumes and baseline endocrine evaluation, initial surgical success rate, and disease control among patients with acromegaly. A retrospective cohort study was conducted at a clinical research center including patients with acromegaly due to GH-secreting pituitary adenomas. Baseline hormonal evaluation and adenoma characteristics according to MRI were collected. Volumetric measurements of pituitary adenomas were performed using a semi-automated lesion segmentation and tumor-volume assessment tools. Rates of post-operative medical treatment, radiation therapy, and re-operation were gathered from the patients' medical records. Twenty seven patients (11 females) were included, median age 21.0 years (interquartile range 29 years, range 3-61 years). Patients harboring adenomas with a volume &lt;2 000 mm3 had higher chance to achieve disease remission [94.1% (n=16) vs. 50.0% (n=4), p&lt;0.05]. Adenoma volumes positively correlated with baseline plasma GH levels before and after oral glucose administration, and with plasma IGF-I and PRL levels. Adenoma volume had negative correlation with morning plasma cortisol levels. Finally, patients harboring larger adenomas required 2nd surgery and/or medical treatment more often compared with subjects with smaller adenomas. Accurate 3D volume measurement of GH-secreting pituitary adenomas may be used for the prediction of initial surgery success and for disease control rates among patients with a GH-secreting pituitary adenomas and performs better than standard size assessments

Cushing disease in a patient with multiple endocrine neoplasia type 2B

Context: Multiple endocrine neoplasia type 2B (MEN2B) is a rare autosomal-dominant cancer syndrome characterized in part by metastatic medullary thyroid cancer (MTC) and pheochromocytoma. Cushing disease is a rare cause of endogenous hypercortisolism in children. Case description: We describe a 21-year-old African-American male who was diagnosed at age 10 with an ACTH-secreting pituitary microadenoma. At age 16 he developed medullary thyroid cancer and was found to have multiple endocrine neoplasia type 2B with the characteristic M918T mutation of the RET proto-oncogene. Following thyroidectomy, he was initiated on Vandetanib, a tyrosine kinase inhibitor, and has since had stable disease over the last 5 years. Conclusions: Our patient is the first individual with MEN2B to be described with Cushing disease. The RET oncogene may play a role in pituitary tumorigenesis; alternatively, the coexistence of these two entities may represent an extremely rare coincidence. Keywords: Hypercortisolemia, Neuroendocrine tumor, Genetic syndrome, RE

Screening for GPR101 defects in pediatric pituitary corticotropinomas.

Cushing disease (CD) in children is caused by adrenocorticotropic hormone (ACTH)-secreting pituitary adenomas. Germline or somatic mutations in genes such as MEN1, CDKIs, AIP, and USP8 have been identified in pediatric CD, but the genetic defects in a significant percentage of cases are still unknown. We investigated the orphan G protein-coupled receptor GPR101, a gene known to be involved in somatotropinomas, for its possible involvement in corticotropinomas. We performed GPR101 sequencing, expression analyses by RT-qPCR and immunostaining, and functional studies (cell proliferation, pituitary hormones secretion, and cAMP measurement) in a series of patients with sporadic CD secondary to ACTH-secreting adenomas in whom we had peripheral and tumor DNA (N=36). No increased GPR101 expression was observed in tumors compared to normal pituitary (NP) tissues, nor did we find a correlation between GPR101 and ACTH expression levels. Sequence analysis revealed a very rare germline heterozygous GPR101 variant (p.G31S) in one patient with CD. Overexpression of the p.G31S variant did not lead to increased growth and proliferation, although modest effects on cAMP signaling were seen. GPR101 is not overexpressed in ACTH-secreting tumors compared to NPs. A rare germline GPR101 variant was found in one patient with CD but in vitro studies did not support a consistent pathogenic effect. GPR101 is unlikely to be involved in the pathogenesis of CD