Jacqueline K. Hewitt

Metastatic Pheochromocytoma/Paraganglioma Related to Primary Tumor Development in Childhood or Adolescence: Significant Link to SDHB Mutations

PURPOSE To present data on the high rate of SDHB mutations in patients with metastatic pheochromocytoma/paraganglioma whose initial tumor presentation began in childhood or adolescence. PATIENTS AND METHODS From 2000 to 2010, 263 patients with pheochromocytoma/paraganglioma were evaluated through the National Institutes of Health (NIH), Bethesda, MD. Of the 263 patients, 125 patients were found to have metastatic disease; of these 125 patients, 32 patients presented with a tumor before 20 years of age. An additional 17 patients presented with a tumor before 20 years of age but demonstrated no development of metastatic disease. Genetic testing for mutations in the VHL, MEN, and SDHB/C/D genes was performed on patients without previously identified genetic mutations. RESULTS Of the 32 patients who presented with metastatic disease and had their primary tumor in childhood or adolescence, sequence analysis of germline DNA showed SDHB mutations in 23 patients (71.9%), SDHD mutations in three patients (9.4%), VHL mutations in two patients (6.3%), and an absence of a known mutation in four patients (12.5%). The majority of these 32 patients (78.1%) presented with primary tumors in an extra-adrenal location. CONCLUSION The majority of patients with metastatic pheochromocytoma/paraganglioma who presented with a primary tumor in childhood/adolescence had primary extra-adrenal tumors and harbored SDHB mutations. Except for primary tumors located in the head and neck where SDHD genetic testing is advised, we recommend that patients who present with metastatic pheochromocytoma/paraganglioma with primary tumor development in childhood or adolescence undergo SDHB genetic testing before they undergo testing for other gene mutations, unless clinical presentation or family history suggests a different mutation.

Ethical Principles for the Management of Infants with Disorders of Sex Development

The Fifth World Congress on Family Law and Children’s Rights (Halifax, August 2009) adopted a resolution endorsing a new set of ethical guidelines for the management of infants and children with disorders of sex development (DSD) [www.lawrights.asn.au/index.php?option=com_content&view=article&id=76&Itemid=109]. The ethical principles developed by our group were the basis for the Halifax Resolution. In this paper, we outline these principles and explain their basis. The principles are intended as the ethical foundation for treatment decisions for DSD, especially decisions about type and timing of genital surgery for infants and young children. These principles were formulated by an analytic review of clinician reasoning in particular cases, in relation to established principles of bioethics, in a process consistent with the Rawlsian concept of reflective equilibrium as the method for building ethical theory. The principles we propose are: (1) minimising physical risk to child; (2) minimising psychosocial risk to child; (3) preserving potential for fertility; (4) preserving or promoting capacity to have satisfying sexual relations; (5) leaving options open for the future, and (6) respecting the parents’ wishes and beliefs.

Disorders of sex development: Insights from targeted gene sequencing of a large international patient cohort

BackgroundDisorders of sex development (DSD) are congenital conditions in which chromosomal, gonadal, or phenotypic sex is atypical. Clinical management of DSD is often difficult and currently only 13% of patients receive an accurate clinical genetic diagnosis. To address this we have developed a massively parallel sequencing targeted DSD gene panel which allows us to sequence all 64 known diagnostic DSD genes and candidate genes simultaneously.ResultsWe analyzed DNA from the largest reported international cohort of patients with DSD (278 patients with 46,XY DSD and 48 with 46,XX DSD). Our targeted gene panel compares favorably with other sequencing platforms. We found a total of 28 diagnostic genes that are implicated in DSD, highlighting the genetic spectrum of this disorder. Sequencing revealed 93 previously unreported DSD gene variants. Overall, we identified a likely genetic diagnosis in 43% of patients with 46,XY DSD. In patients with 46,XY disorders of androgen synthesis and action the genetic diagnosis rate reached 60%. Surprisingly, little difference in diagnostic rate was observed between singletons and trios. In many cases our findings are informative as to the likely cause of the DSD, which will facilitate clinical management.ConclusionsOur massively parallel sequencing targeted DSD gene panel represents an economical means of improving the genetic diagnostic capability for patients affected by DSD. Implementation of this panel in a large cohort of patients has expanded our understanding of the underlying genetic etiology of DSD. The inclusion of research candidate genes also provides an invaluable resource for future identification of novel genes.

Comprehensive genotyping and clinical characterisation reveal 27 novel NKX2-1 mutations and expand the phenotypic spectrum

Background NKX2-1 encodes a transcription factor with large impact on the development of brain, lung and thyroid. Germline mutations of NKX2-1 can lead to dysfunction and malformations of these organs. Starting from the largest coherent collection of patients with a suspected phenotype to date, we systematically evaluated frequency, quality and spectrum of phenotypic consequences of NKX2-1 mutations. Methods After identifying mutations by Sanger sequencing and array CGH, we comprehensively reanalysed the phenotype of affected patients and their relatives. We employed electrophoretic mobility shift assay (EMSA) to detect alterations of NKX2-1 DNA binding. Gene expression was monitored by means of in situ hybridisation and compared with the expression level of MBIP, a candidate gene presumably involved in the disorders and closely located in close genomic proximity to NKX2-1. Results Within 101 index patients, we detected 17 point mutations and 10 deletions. Neurological symptoms were the most consistent finding (100%), followed by lung affection (78%) and thyroidal dysfunction (75%). Novel symptoms associated with NKX2-1 mutations comprise abnormal height, bouts of fever and cardiac septum defects. In contrast to previous reports, our data suggest that missense mutations in the homeodomain of NKX2-1 not necessarily modify its DNA binding capacity and that this specific type of mutations may be associated with mild pulmonary phenotypes such as asthma. Two deletions did not include NKX2-1, but MBIP, whose expression spatially and temporarily coincides with NKX2-1 in early murine development. Conclusions The high incidence of NKX2-1 mutations strongly recommends the routine screen for mutations in patients with corresponding symptoms. However, this analysis should not be confined to the exonic sequence alone, but should take advantage of affordable NGS technology to expand the target to adjacent regulatory sequences and the NKX2-1 interactome in order to maximise the yield of this diagnostic effort.

A multi-exon deletion within WWOX is associated with a 46,XY disorder of sex development

Disorders of sex development (DSD) are congenital conditions where chromosomal, gonad or genital development is atypical. In a significant proportion of 46,XY DSD cases it is not possible to identify a causative mutation, making genetic counseling difficult and potentially hindering optimal treatment. Here, we describe the analysis of a 46,XY DSD patient that presented at birth with ambiguous genitalia. Histological analysis of the surgically removed gonads showed bilateral undifferentiated gonadal tissue and immature testis, both containing malignant germ cells. We screened genomic DNA from this patient for deletions and duplications using an Illumina whole-genome SNP microarray. This analysis revealed a heterozygous deletion within the WWOX gene on chromosome 16, removing exons 6–8. Analysis of parental DNA showed that the deletion was inherited from the mother. cDNA analysis confirmed that the deletion maintained the reading frame, with exon 5 being spliced directly onto exon 9. This deletion is the first description of a germline rearrangement affecting the coding sequence of WWOX in humans. Previously described Wwox knockout mouse models showed gonadal abnormalities, supporting a role for WWOX in human gonad development.

Hormone treatment of gender identity disorder in a cohort of children and adolescents.

Objective: To describe the experience of hormone treatment of gender identity disorder (GID) in children and adolescents within a specialist clinic.

Ethical Principles: An Essential Part of the Process in Disorders of Sex Development Care

child. All of these decisions are ethically important, as they all have significant, far-reaching consequences for the infant. The issue is not whether a particular type of decision happens to be controversial at a particular time; it is about making sound, ethically appropriate and justifiable decisions for every patient. That having been said, it is worthwhile to note that issues over genital surgery have not gone away [3] . There is still considerable difference in opinion and practice. Taking congenital adrenal hyperplasia as an example, the current guideline [3] is for consideration of feminising genitoplasty in girls with congenital adrenal hyperplasia at Prader stage 3 and above. However, some centres recommend that surgery in this group should occur in early infancy while others argue that surgery should be deferred until puberty, when it may be better understood and accepted [4] . The debate about surgery is far from over. Articulating a clear set of principles for the management of DSD is not merely a theoretical exercise. The principles are intended for use. In a multidisciplinary environment, where the ethical aim is shared decisionmaking with parents, the principles provide the essential starting point for any discussions and decision-making. They focus discussion, highlight information or perspecIn their editorial commentary on our paper on ethical principles in the management of infants with disorders of sex development (DSD) [1] , Dreger et al. [2] assert that there is no longer any need for statements of ethical principles in relation to management of DSD. The field has moved on from controversy about genital surgery, they claim, and the issue is now implementation of what is agreed to be best practice in care of these children and their families. They see no value in our formulation of these principles since they are nothing new, being ‘essentially the same’ as what clinicians, ethicists and activists already think. This is an underestimation of both the importance of actually articulating a set of clear specific ethical principles, and the crucial role that these principles can play in achieving implementation of holistic comprehensive care. The principles which we articulated apply not just to decisions about genital surgery; as we noted in our article, they apply to all management decisions that need to be made about infants with DSD, including sex of rearing, hormonal therapy and all forms of surgery, not only genitoplasty. The principles also apply to less ‘medical’, but equally important decisions, such as what, when and how to tell young children about their condition, as well as how to advise parents to think about and relate to their Received: March 21, 2011 Accepted: April 4, 2011 Published online: August 4, 2011 HORMONE RESEARCH IN PÆDIATRICS

Preventing female virilisation in congenital adrenal hyperplasia: The controversial role of antenatal dexamethasone.

Congenital adrenal hyperplasia (CAH) refers to a group of recessively inherited disorders of cortisol production, which in the classical form results in virilisation of female fetuses. Since the 1980s, antenatal treatment with dexamethasone has been recommended in high‐risk pregnancies to minimise the risk of virilising the female genitalia of affected fetuses. To be effective, this treatment requires implementation in early pregnancy, prior to the commencement of autonomous fetal adrenal androgen synthesis. Using this approach, seven of eight high‐risk pregnancies are treated unnecessarily, prior to establishing the fetal gender or the confirmed diagnosis of a genetically affected pregnancy. In the face of ongoing concerns regarding potential adverse maternal–fetal effects of antenatal dexamethasone exposure, a review of this practice has been advocated by expert advisory groups. In this review, we summarise current controversies, potential improvements and future directions in the management of pregnancies at risk of CAH. In high‐risk families, recent genomic advances include early prenatal diagnosis utilising noninvasive genetic techniques to minimise unnecessary dexamethasone exposure to unaffected fetuses. In affected pregnancies when families elect for antenatal treatment, optimal antenatal dosing regimens need to be defined and a standardised treatment and follow‐up protocol are recommended. Establishment of a national registry with standardised follow‐up will allow future families to be better informed of the risks and benefits of both treated and untreated fetal CAH.

Hormone replacement in disorders of sex development: Current thinking

Congenital disruptions of sex hormone production lead to wide-ranging developmental and physiological effects in individuals who have atypical chromosomal, gonadal or anatomic sex. Aberrant developmental sex hormone exposure causes disorders of genital anatomy, attainment of secondary sexual characteristics and has long-term effects on metabolism, fertility and psychological functioning. Principles in the management of disorders of sex development (DSD) aim to improve physiological health and long-term outcome, as well as development of male or female sexual anatomy. Concerns raised by DSD patient advocacy groups about beneficence and autonomy with respect to prescribed hormone treatments and avoidance of unnecessary genital and gonadal surgery have demanded greater informed consent and attention to long-term outcome. Hormone treatment is influenced by underlying clinical diagnosis and by factors such as sex of rearing and gender identity of the affected individual. We describe diagnostic criteria for different DSDs, clinical considerations in management protocols, together with current concepts and detailed practical hormone treatments for male and female individuals with DSD. Gender identity issues requiring multidisciplinary consensus, ethical consideration and informed consent or assent from the young person are also addressed.

Management of disorders of sex development

Disorders of sex development comprise a group of diagnoses that represent significant and controversial management difficulties, such as those relating to gender assignment, genital surgery and risk of gonadal malignancy or future gender dysphoria. These issues arise on the background of a diagnosis that is often fraught with psychological trauma for patients and their families. There have been many changes in the medical approach to management in recent years; however, more research is required. This should focus on achieving accurate diagnosis in the first case, and on following outcomes to advise best practice. Finally, rational dialogue between multidisciplinary care providers, ethicists and patients and their families will only lead to improved care for these children.