Cushing's Disease Whole Exome Sequencing Study

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We are currently analyzing whole exome sequencing (WES) data with appropriate follow up to identify important genetic factors associated with Cushing's disease (CD) and related abnormal physical features. The ultimate goal is to identify a genetic variant or variants that cause CD. CD is a condition in which the pituitary gland produces inappropriately high levels of adrenocorticotropic hormone (ACTH). The ACTH stimulates the adrenal gland to produce excess cortisol, leading to clinical disease. CD is caused by ACTH secreting pituitary tumors. CD is a serious condition. It requires surgery to remove the tumor. The tumors sometimes recur in which case radiation or medical therapy is required which is not always successful. CD can cause a wide range of problems due to the high cortisol levels. These include diabetes, fractures, poor growth, and hypertension. CD can be fatal. Whole exome sequencing (WES) is a powerful tool for identifying important genetic variants associated with medical conditions. It is an efficient method of determining the genetic code (sequence) of all the regions in the genome that are translated into protein, the exons. The exons constitute about 1% of DNA, thus sequencing exons provides a large amount of information at a lower cost than sequencing the entire genome. Pediatric aged patients seen at NICHD with a confirmed diagnosis of CD are evaluated for this study. Those who have histopathologically confirmed disease in conjunction with DNA, hormonal documentation of the disease and complete clinical data are potential cases. Analysis comparing variants found in the cases with large control populations has been done. In addition, the data are being examined for copy number variants in the exons to determine if they play a role in Cushing's disease. Whole exome sequencing has now been performed on tumor tissue in patients where tumor tissue is available to determine if there are specific tumor-associated variants in the exome. These data have been analyzed in the same manner as the blood samples. In addition, samples from patients who have ectopic posterior pituitaries have been obtained and whole exome sequenced. The data have been screened using standard data sets to identify rare variants. 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. We performed a comprehensive screening for DICER1 variants in a large cohort of CD patients, and analyzed their possible contribution to the phenotype. We included 192 CD 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. 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. 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. Germline loss-of-function CDKN1B gene variants cause the autosomal dominant syndrome of multiple endocrine neoplasia type 4 (MEN4). Even though pituitary neuroendocrine tumors are a well-known component of the syndrome, only 2 cases of Cushing's disease (CD) have so far been described in this setting. We screened a large cohort of CD patients for CDKN1B gene defects and to determine their functional effects. We screened 211 CD patients (94.3% pediatric) by germline whole-exome sequencing (WES) only (n = 157), germline and tumor WES (n = 27), Sanger sequencing (n = 6), and/or germline copy number variant (CNV) analysis (n = 194). Sixty cases were previously unpublished. Variant segregation was investigated in the patients' families, and putative pathogenic variants were functionally characterized. Five variants of interest were found in 1 patient each: 1 truncating (p.Q107Rfs*12) and 4 nontruncating variants, including 3 missense changes affecting the CDKN1B protein scatter domain (p.I119T, p.E126Q, and p.D136G) and one 5' untranslated region (UTR) deletion (c.-29_-26delAGAG). No CNVs were found. All cases presented early (10.5 1.3 years) and apparently sporadically. Aside from colon adenocarcinoma in 1 carrier, no additional neoplasms were detected in the probands or their families. In vitro assays demonstrated protein instability and disruption of the scatter domain of CDKN1B for all variants tested. In summary, five patients with CD and germline CDKN1B variants of uncertain significance (n = 2) or pathogenic/likely pathogenic (n = 3) were identified, accounting for 2.6% of the patients screened. Our finding that germline CDKN1B loss-of-function may present as apparently sporadic, isolated pediatric CD has important implications for clinical screening and genetic counselling. Pituitary developmental defects lead to partial or complete hormone deficiency and significant health problems. The majority of cases are sporadic and of unknown cause. We screened 28 patients with pituitary stalk interruption syndrome (PSIS) for mutations in the FAT/DCHS family of protocadherins that have high functional redundancy. We identified seven variants, four of which putatively damaging, in FAT2 and DCHS2 in six patients with pituitary developmental defects recruited through a cohort of patients with mostly ectopic posterior pituitary gland and/or pituitary stalk interruption. All patients had growth hormone deficiency and two presented with multiple hormone deficiencies and small glands. FAT2 and DCHS2 were strongly expressed in the mesenchyme surrounding the normal developing human pituitary. We analyzed Dchs2-/- mouse mutants and identified anterior pituitary hypoplasia and partially penetrant infundibular defects. Overlapping infundibular abnormalities and distinct anterior pituitary morphogenesis defects were observed in Fat4-/- and Dchs1-/- mouse mutants but all animal models displayed normal commitment to the anterior pituitary cell type. Together our data implicate FAT/DCHS protocadherins in normal hypothalamic-pituitary development and identify FAT2 and DCHS2 as candidates underlying pituitary gland developmental defects such as ectopic pituitary gland and/or pituitary stalk interruption. Carney complex (CNC), is an autosomal dominant multiple neoplasia and lentiginosis syndrome. We aimed to identify risk factors associated with the occurrence and recurrence of cardiac myxomas, the predominant cause of death in CNC patients.Patients with CNC were monitored prospectively between 1995 and 2020 for the development of cardiac myxomas.Of the 319 patients studied, 136 (42.6%) developed myxomas. The mean age at diagnosis was 28.7 16.6 years in females and 25.0 16.4 years in males. By age 30, 35% of females and 45% of males had at least one myxoma. The CNC-related lesions, lentigines, cutaneous, mucosal, or breast myxomas, thyroid nodules, pituitary adenoma, and schwannoma were significantly more frequent (all p < 0.05) among patients with myxomas. Forty-four percent of patients had recurrences; nearly all within the first 8 and 16 years for males and females, respectively. Recurrences were more common in females.This is the l