Renuka Dias

The discriminatory value of the low-dose dexamethasone suppression test in the investigation of paediatric Cushing's syndrome.

Background: Low- and high-dose dexamethasone suppression tests (LDDST, HDDST) are used in the investigation of Cushing’s syndrome (CS). In adults with Cushing’s disease (CD), cortisol suppression during LDDST predicts suppression during the HDDST. Methods: We reviewed the results of the LDDST (0.5 mg 6 hourly × 48 h), HDDST (2.0 mg 6 hourly × 48 h) and corticotrophin-releasing hormone (CRH) test in 32 paediatric patients with CS: 24 had CD, 1 ectopic ACTH syndrome, 5 nodular adrenal hyperplasia and 2 adrenocortical tumours. Results: In CD, LDDST suppressed cortisol from 590.7 ± 168.8 (mean ± SD) to 333.7 ± 104.0 nmol/l after 48 h (0 vs. 48 h, p < 0.05; mean suppression, 45.1%; CI (30.8, 59.4%); 16/24 (66%) suppressed >30%; mean suppression 68.1%, CI (58.1, 77.9%)). The HDDST suppressed cortisol from 596.3 ± 174.5 to 47.1 ± 94.8 nmol/l after 48 h (0 vs. 48 h, p < 0.05; mean suppression, 93.5%; CI (88.2, 98.8%) with 17/24 (71%) suppressing to <50 nmol/l and 100% to <50% of baseline). In the LDDST, suppression correlated with that during the HDDST (r = +0.45, p < 0.05) with >30% suppression predicting that in the HDDST and hence CD. CRH increased cortisol by +100.3% (CI 62, 138.5%), 22/24 (91.7%) showing a >20% increase. In the other CS pathologies (n = 8) the LDDST induced no significant decrease in cortisol. Conclusion: The LDDST was of diagnostic value by discriminating between CD and other CS aetiologies. In our view the HDDST is redundant in the investigation of paediatric CS.

Multiple Segmental Uniparental Disomy Associated with Abnormal DNA Methylation of Imprinted Loci in Silver-Russell Syndrome

BACKGROUNDnSilver-Russell syndrome (SRS; online inheritance in man 180860) is a low-birth-weight syndrome characterized by postnatal growth restriction and variable dysmorphic features. Although maternal uniparental disomy (UPD) of chromosome 7 and hypomethylation of H19 have been reported in up to 50% of all cases, no unifying mechanism is apparent.nnnSUBJECTS AND METHODSnTen patients and their parents were studied using the Illumina GoldenGate methylation array and the Illumina 370K HumHap single-nucleotide polymorphism array to identify aberrations in DNA methylation as well as genomic changes including copy number changes and uniparental disomy events.nnnRESULTSnWe found evidence of UPD events outside chromosome 7 in all patients. In up to 30% of patients with SRS, DNA methylation changes occur in imprinted gene loci outside 11p15.5 (PEG3, PLAGL1, and GRB10), not previously consistently linked with SRS. Furthermore, hypermethylation of GRB10 was associated with increased mRNA expression. In addition, 20% of patients appear to have DNA methylation abnormalities within multiple loci. Not all the imprinted loci with methylation defects were affected directly by UPD.nnnCONCLUSIONSnThe association of widespread UPD associated with abnormal methylation and mRNA expression in imprinted genes in SRS is consistent with the concept of UPD as an initial genomic abnormality leading to unstable DNA methylation within the regulatory network of imprinted genes. Furthermore, disruption of any one of these genes may contribute to the heterogeneous clinical spectrum of SRS.

Steroid Sulfatase Deficiency and Androgen Activation Before and After Puberty

Context: Steroid sulfatase (STS) cleaves the sulfate moiety off steroid sulfates, including dehydroepiandrosterone (DHEA) sulfate (DHEAS), the inactive sulfate ester of the adrenal androgen precursor DHEA. Deficient DHEA sulfation, the opposite enzymatic reaction to that catalyzed by STS, results in androgen excess by increased conversion of DHEA to active androgens. STS deficiency (STSD) due to deletions or inactivating mutations in the X-linked STS gene manifests with ichthyosis, but androgen synthesis and metabolism in STSD have not been studied in detail yet. Patients and Methods: We carried out a cross-sectional study in 30 males with STSD (age 6–27 y; 13 prepubertal, 5 peripubertal, and 12 postpubertal) and 38 age-, sex-, and Tanner stage-matched healthy controls. Serum and 24-hour urine steroid metabolome analysis was performed by mass spectrometry and genetic analysis of the STS gene by multiplex ligation-dependent probe amplification and Sanger sequencing. Results: Genetic analysis showed STS mutations in all patients, comprising 27 complete gene deletions, 1 intragenic deletion and 2 missense mutations. STSD patients had apparently normal pubertal development. Serum and 24-hour urinary DHEAS were increased in STSD, whereas serum DHEA and testosterone were decreased. However, total 24-hour urinary androgen excretion was similar to controls, with evidence of increased 5α-reductase activity in STSD. Prepubertal healthy controls showed a marked increase in the serum DHEA to DHEAS ratio that was absent in postpubertal controls and in STSD patients of any pubertal stage. Conclusions: In STSD patients, an increased 5α-reductase activity appears to compensate for a reduced rate of androgen generation by enhancing peripheral androgen activation in affected patients. In healthy controls, we discovered a prepubertal surge in the serum DHEA to DHEAS ratio that was absent in STSD, indicative of physiologically up-regulated STS activity before puberty. This may represent a fine tuning mechanism for tissue-specific androgen activation preparing for the major changes in androgen production during puberty.

Familial Isolated Primary Pigmented Nodular Adrenocortical Disease Associated with a Novel Low Penetrance PRKAR1A Gene Splice Site Mutation

Background/Aims: Primary pigmented nodular adrenocortical disease (PPNAD) is associated with inactivating germline protein kinase A regulatory subunit type 1-α (PRKAR1A) mutations and loss of heterozygosity at the 17q22–24 locus in approximately 50% patients. PRKAR1A mutations are observed in both isolated PPNAD (iPPNAD) and Carney complex (CNC). Most mutations result in a functionally null-allele and exhibit high penetrance. We genotyped members of an extended family for a novel PRKAR1A mutation and undertook detailed phenotyping for CNC in the affected individuals. Methods: A 10.5-year-old male was diagnosed with PPNAD; the patient’s mother also had iPPNAD. A 13-year-old sibling and 7 other relatives (mean age 58.2, range 29.1–80.2 years) were referred for PRKAR1A mutation analysis. Results: DNA analysis of the index case and parent revealed a novel germline heterozygous PRKAR1A mutation at the +1 position of the acceptor site of intron 3 [c.349 G>T]. The same heterozygous splice site mutation was present in the sibling with no PPNAD or CNC manifestations and 2 other individuals aged 54.9 and 57.1 years who had subclinical Cushing’s syndrome but no features of CNC. Conclusion: We conclude that c.349 G>T, a novel splice site germline PRKAR1A defect, has low penetrance resulting in incomplete clinical expression in this kindred.

Isolated Addison's disease is unlikely to be caused by mutations in MC2R, MRAP or STAR, three genes responsible for familial glucocorticoid deficiency

Background Familial glucocorticoid deficiency (FGD) is a rare autosomal recessive disease caused by ACTH resistance and leads to isolated glucocorticoid deficiency. Although FGD patients typically have normal mineralocorticoid secretion, subtle alterations in the renin–angiotensin–aldosterone axis have been reported in a subset of patients at presentation. Anecdotally, some patients with FGD have been initially diagnosed as having Addisons disease (AD), with implications for treatment and genetic counselling. Currently, mutations in three genes: the ACTH receptor (MC2R); the melanocortin 2 receptor accessory protein (MRAP); and the steroidogenic acute regulatory protein (STAR) are known to give rise to FGD types 1–3. We investigated a cohort of autoantibody-negative AD patients for mutations in these genes. Methods Forty patients with known AD without evidence of autoimmune disease were screened for mutations in MC2R, MRAP and STAR. In addition, patients were genotyped for the MC2R promoter polymorphism previously associated with reduced responsiveness to ACTH. Results No mutations in MC2R, MRAP or STAR were identified in any patient. The frequencies of the MC2R promoter polymorphism were similar to those reported in healthy controls. Conclusions FGD does not appear to be underdiagnosed in the AD population. However, in ∼50% of patients with FGD, no genetic cause has yet been identified and it is possible that the other, as yet unidentified, genes giving rise to FGD may be implicated in AD.

Diagnosis and Treatment of Cushing’s Disease in Children

Cushings disease (CD) is rare in the paediatric age range, but presents a diagnostic and therapeutic challenge. Most paediatric endocrinologists have limited experience managing children or adolescents with CD and thus benefit from close consultation with adult colleagues. A diagnostic protocol for investigation is required which broadly follows the model for adult patients. Treatment strategies for CD are described and critically appraised. The management of paediatric CD patients after cure also presents challenges for optimizing growth, bone health, reproduction and body composition from childhood into and during adult life.

Phenotypic spectrum and responses to recombinant human IGF1 (rhIGF1) therapy in patients with homozygous intronic pseudoexon growth hormone receptor mutation

BACKGROUNDnPatients with homozygous intronic pseudoexon GH receptor (GHR) mutations (6Ψ) have growth hormone insensitivity (GHI) (growth failure, IGF1 deficiency and normal/elevated serum GH). We report 9 patients in addition to previously described 11 GHR 6Ψ patients and their responses to rhIGF1 therapy.nnnMETHODSn20 patients (12 males, 11 families, mean age 4.0u2009±u20092.2 years) were diagnosed genetically in our centre. Phenotypic data and responses to rhIGF1 treatment were provided by referring clinicians. Continuous parametric variables were compared using Student t-test or ANOVA.nnnRESULTSn10/20 (50%) had typical facial features of GHI, 19/20 (95%) from consanguineous families and 18/20 (90%) of Pakistani origin. At diagnosis, mean height SDS: -4.1u2009±u20090.95, IGF1 SDS: -2.8u2009±u20091.4; IGFBP3 SDS: -3.0u2009±u20092.1 and mean basal and peak GH levels: 11.9u2009µg/L and 32.9u2009µg/L, respectively. 1/12 who had IGF1 generation test, responded (IGF1: 132-255u2009ng/mL). 15/20 (75%; 11M) received rhIGF1 (mean dose: 114u2009µg/kg twice daily, mean duration: 5.3u2009±u20092.5 years). Mean baseline height velocity of 4.7u2009±u20091.1u2009cm/year increased to 7.4u2009±u20091.8u2009cm/year (Pu2009=u20090.001) during year 1 of therapy. Year 3 mean height SDS (-3.2u2009±u20091.0) was higher than pre-treatment height SDS (-4.3u2009±u20090.8) (Pu2009=u20090.03). Mean cumulative increase in height SDS after year 5 was 1.4u2009±u20090.9. Difference between target height (TH) SDS and adult or latest height SDS was less than that of TH SDS and pre-treatment height SDS (2.1u2009±u20091.2 vs 3.0u2009±u20090.8; Pu2009=u20090.02).nnnCONCLUSIONnIn addition to phenotypic heterogeneity in the cohort, there was mismatch between clinical and biochemical features in individual patients with 6Ψ GHR mutations. rhIGF1 treatment improved height outcomes.

Special Aspects of Cushing’s Syndrome: Childhood

Cushing’s syndrome (CS) is a rare pediatric problem on which few pediatric endocrinologists have extensive experience. Hence, close collaboration with an adult endocrinologist colleague is strongly recommended. This chapter reviews the main types of CS occurring in the pediatric age range. Results are presented from the authors’own experience of management of over 50 cases of pediatric CS during the last 25 years. These include McCune–Albright syndrome in infancy, adrenocortical tumors, primary nodular adrenal hyperplasia, ectopic ACTH syndrome, and Cushing’s disease. The major clinical features are described, including linear growth and puberty, and diagnostic techniques are discussed. From a personal database of 38 cases of Cushing’s disease (CD), the relative benefits of pituitary MR imaging versus bilateral inferior petrosal sinus sampling for ACTH are appraised. Therapeutic strategy is discussed for primary adrenal CS and pituitary-dependent CD. The results of transsphenoidal surgery and pituitary radiotherapy are described together with the challenge of optimizing postcure linear growth and body composition.