3rd International Symposium on Disorders of Sex Development 20 – 22 May 2011 · Universität zu Lübeck
Programme & Abstracts From Gene to Gender - What we’ve learned and what we need to learn - New Prospects in DSD Research
Sessions on Genetics, Hormones & Action, Morphologic Variation in DSD and Clinical Aspects · Round Table · Poster Session and Oral Sessions
20 – 22 May 2011, Lübeck, Germany 1
Contents
Contents...... 1
Welcome Remarks ...... 3
About EuroDSD...... 4
General Information...... 5
Scientific Programme ...... 6
Social Programme...... 9
Speakers’ List...... 10
Poster Presentation...... 12
Abstracts Keynote Lectures ...... 15
Abstracts Oral Presentations...... 31
Abstracts Poster Presentations ...... 42
Maps...... 86
Imprint ...... 90
Sponsors ...... 91
SHORT STATURE PORTFOLIO
Gezielt zum Erfolg Differenzierte Therapie von Wachstumsstörungen
für die GH-Therapie für die IGF-I-Therapie
NutropinAq® 10 mg/2 ml (30 I.E.), Injektionslösung. Wirkstoff: Somatropin. Zusammensetzung: Eine Zylinderampulle enthält: 10 mg (30 I.E.) Somatropin (humanes Wachstumshormon) aus mittels rekombinanter DNA-Technologie herge- stellt. Sonstige Bestandteile: Natriumchlorid, Phenol, Polysorbat 20, Natriumcitrat, Wasserfreie Citronensäure, Wasser für Injektionszwecke. Anwendungsgebiete: Langzeitbehandlung von Kindern mit Wachstumsstörungen infolge ungenügender Sekretion von endogenem Wachstumshormon (WH). Langzeitbehandlung von Wachstumsstörungen infolge Turner-Syndroms. Behandlung von präpubertären Kindern mit Wachstumsstörungen infolge chronischer Niereninsuffizienz bis zum Zeitpunkt einer Nieren- transplantation. Substitution von WH bei Erwachsenen, die entweder bereits seit der Kindheit o. seit dem Erwachsenenalter an einem WH-Mangel leiden. Gegenanzeigen: Überempfindlichkeit gegen Somatropin o. einen der sonstigen Bestandteile. Patienten mit geschlossener Epiphyse. Patienten mit aktivem Neoplasma. Die NutropinAq®-Therapie muss abgebrochen werden, falls es Anzeichen für ein Tumorwachstum gibt. Patienten mit akuter, schwerer Erkrankung infolge Komplikationen nach einer Herz- o. Bauch- operation, multiplen Unfallverletzungen. Patienten mit akuter Ateminsuffizienz. Nebenwirkungen: Sehr häufig bei Erwachsenen, häufig bei Kindern: Arthralgie, Myalgie, Ödem, peripheres Ödem. Häufig: Antikörper-Bildung, Hypothyreose, verminderte Glukose- toleranz, Kopfschmerzen, Hypertonie, Reaktionen an der Inj.-stelle, Asthenie. Gelegentlich: Malignom, Neoplasma, Anämie, Hypoglykämie, Hyperphosphatämie, Persönlichkeitsstörung, Somnolenz, Nystagmus, Papillenödem, Diplopie, Schwindel, Tachykardie, Hypertonie, Erbrechen, Bauchschmerzen, Blähungen, Übelkeit, Lipodystrophie, Hautatrophie, exfoliative Dermatitis, Urtikaria, Hirsutismus, Hauthypertrophie, Muskelatrophie, Knochenschmerzen, Karpaltunnelsyndrom, Harninkontinenz, Hämaturie, Polyurie, erhöhte Harnlassfrequenz/Pollakisurie, Urinanomalie, Genitalfluor, an der Inj.-stelle: Atrophie, Blutung u. Schwellung, Hypertrophie. Selten: Diabetes mellitus, Neuropathie, intrakranieller Hochdruck, Diarrhoe, abnorme Nierenfunktionswerte. Häufig: ZNS-Neoplasma. Häufig: Menorrhagie. Häufig: Nierenversagen, Peritonitis, Knochennekrose, erhöhter Kreatininspiegel. Sehr häufig: Parästhesie. Häufig: Hyperglykämie, Hyperlipidämie, Schlaflosigkeit, Störung der Synovialis, Arthrose, Muskelschwäche, Rückenschmerzen, Brustschmerzen, Gynäkomastie. Verstärktes Wachstum von Muttermalen o. Leberflecken möglich. Bei Kindern mit WH-Mangel Risiko für intrakraniellen Hochdruck. Patienten mit endokrin. Störung Risiko für Epiphysiolyse. Leukämie bei einer kleinen Anzahl Patienten mit WH-Mangel. Ursächlich. Zshg. mit Therapie unwahrscheinlich. Die Informationen zu Warnhinweisen und Vorsichtsmaßnahmen sowie Wechselwirkungen mit anderen Mitteln entnehmen Sie bitte der Fachinformation. Verschreibungspflichtig (NR). Ipsen Pharma GmbH, 76275 Ettlingen. Stand der Information: Oktober 2006. Zul.-Nr.: EU/1/00/164/003, EU/1/00/164/005
Increlex® 10 mg/ml Injektionslösung. Wirkstoff: Mecasermin. Zusammensetzung: Jeder ml enthält 10 mg Mecasermin. Jede Durchstechflasche enthält 40 mg Mecasermin. Mecasermin ist ein aus Zellen mittels rekombinanter DNA-Technologie ge- wonnener humaner IGF-1. Sonstige Bestandteile: Benzylalkohol, Natriumchlorid, Polysorbat 20, Essigsäure 99 %, Natriumacetat, Wasser für Injektionszwecke. Wirkstoffgruppe: Somatropin und Somatropin-Agonisten. Anwendungsgebiete: Für die Langzeitbehand- lung von Wachstumsstörungen bei Kindern und Jugendlichen mit schwerem primärem Mangel an IGF-1. Schwerer primärer IGF-1-Mangel wird definiert durch: Körpergrößen-SDS≤ -3,0 und basale IGF-1-Konzentration unterhalb der 2,5. Perzentile für Alter und Geschlecht und Wachstumshormonsuffizienz. Ausschluss der sekundären Formen des IGF-1-Mangels wie Unterernährung, Schilddrüsenunterfunktion oder chronische Behandlung mit pharmakologischen Dosen von anti-inflammatorischen Steroiden. Schwerer primärer IGF-1-Mangel umfasst Patienten mit Mutationen im GH-Rezeptor (GHR), mit Mutationen im Post-GHR-Signalweg und mit IGF-1-Gendefekten; sie haben keinen Wachstumshormonmangel, weshalb nicht zu erwarten ist, dass sie auf eine Behandlung mit exogen gegebenem Wachstums- hormon ausreichend ansprechen. Eine Bestätigung der Diagnose durch einen IGF-1-Generationstest wird empfohlen. Gegenanzeigen: Überempfindlichkeit gegen den Wirkstoff oder einen der sonstigen Bestandteile. Intravenöse Anwendung. Aktives Neoplasma oder Verdacht auf Neoplasma. Falls es Anzeichen für ein Tumorwachstum gibt, muss die Behandlung abgebrochen werden. Benzylalkohol darf bei Früh- oder Neugeborenen nicht angewendet werden. Nebenwirkungen: Sehr häufig: Thymushypertrophie, Kopfschmerzen, Hypakusis, tonsilläre Hypertrophie, Schnarchen, Hypoglykämie, Hypertrophie an der Injektionsstelle. Häufig: Herzgeräusche, abnormale Tympanometrie, abnormales EKG, erhöhte Alaninaminotransferase*, erhöhte Aspartataminotransferase*, Gewichtszunahme, Kardio- megalie, Ventrikelhypertrophie, atriale Hypertrophie*, Tachykardie, paroxysmale Tachykardie*, Mitralklappeninsuffizienz*, Trikuspidalklappeninsuffizienz*, kongenitale Kieferfehlbildung, Pigmentnävus*, Lymphadenopathie*, Krämpfe, fieberhafte Krämpfe*, benigner intrakranieller Hochdruck, Bewusstlosigkeit*, Schlafapnoe, Schwindel, Zittern*, Restless legs*, Papillenödem, reduzierte Sehleistung*, Myopie*, Otorrhoe, Ohrenerkrankung*, Mittelohrerkrankung*, Trommelfellerkrankung*, Ohrenschmerzen, verstopfte Ohren*, Flüssigkeit im Mittelohr, Hypertrophie der Adenoide, Hypertrophie der Nasenmuschel*, Dyspnoe*, Nasenschleimhauterkrankungen*, obstruktive Atemwegserkrankungen*, abnormale Atmung*, verstopfte Nase, Mundatmung, Erbrechen, Brechreiz*, Bauchschmerzen*, Schmerzen im Oberbauch*, abdominale Distension*, Dysphagie*, Nierensteine*, Hydronephrose*, Nierenkolik*, Hauthypertrophie, Akrochordon*, abnormale Haarstruktur*, Arthralgie, Schmerzen in den Extremitäten, Myalgie, Skoliose*, Wirbelsäulendeformität*, Weichteilerkran- kungen*, Muskelkrämpfe*, Flankenschmerz*, Steifheit des Bewegungsapparats*, hypoglykämischer Anfall, Hyperglykämie, Hyperlipidämie*, Adipositas*, fieberhafte Infektion*, Infektion der oberen Atemwege*, Otitis media, seröse Otitis media, chronisch-seröse Otitis media*, Otitis externa*, Pharyngitis*, Tonsillitis, Ohrenentzündung, Mundsoor*, Adenotonsillektomie*, Adenoidektomie, Einsetzen von Ohrröhrchen, Schleimhauthyperplasie, Hypertrophie, Schmerzen an der Injektionsstelle, Bluterguss an der Injektionsstelle, Fibrose an der Injektionsstelle*, Reaktion an der Injektionsstelle*, Schwellung an der Injektionsstelle*, Verhärtung an der Injektionsstelle*, Pigmentveränderung an der Injektionsstelle*, Schleimhautödem*, Asthenie*, Lethargie*, Beschwerden in der Brust*, Gynäkomastie, Eierstock- zyste*, Depression*, Nachtangst, Nervosität, abnormales Verhalten*, Verwirrung* (* = trat bei nur 1 Patienten (1 %) auf). Die nachfolgenden Nebenwirkungen wurden nach Markteinführung von INCRELEX beobachtet. Da diese Nebenwirkungen auf freiwilliger Basis aus einer Patientenpopulation unbekannter Größe berichtet werden, ist es nicht möglich, ihre Häufigkeit zuverlässig abzuschätzen: Anaphylaxie, generalisierte Urtikaria, Angioödem und Dyspnoe. Zu den Symptomen der auf Anaphylaxie hinweisenden Fälle gehörten Nesselausschlag, Angioödem und Dyspnoe. Für einige Patienten war ein Krankenhausaufenthalt erforderlich. Bei allen Patienten traten die Symptome bei wiederholter Anwendung nicht wieder auf. Juckreiz, Urtikaria. Die Informationen zu Warnhinweisen und Vorsichtsmaßnahmen für die Anwendung, Wechselwirkungen mit anderen Arzneimitteln und sonstige Wechselwirkungen sowie Schwangerschaft und Stillzeit entnehmen Sie bitte der Fachinformation. Verschreibungspflichtig NR( ). Ipsen Pharma GmbH, D-76275 Ettlingen. Stand der Information: Juli 2010. Zul.-Nr.: EU/1/07/402/001
Ipsen Pharma GmbH, Einsteinstraße 30, D-76275 Ettlingen, Tel. + 49 7243 184 - 80, Fax + 49 7243 184 ‑ 39, www.ipsen-pharma.de
Portfolio_Anzeige_A4_0411.indd 1 21.04.2011 14:05:25 Uhr
20 – 22 May 2011, Lübeck, Germany 3
Welcome Remarks
Dear Friends and Colleagues,
On behalf of the EuroDSD consortium and the local organising committee, it is a great pleasure to welcome you to Lübeck for our 3rd International Symposium on Disorders of Sex Development.
The EuroDSD consortium was formed following to the previous “Lübeck Meetings on DSD” in 2004 and 2006 as an international collaboration intended to study the natural history and pathophysiology of DSD. Exciting results have been achieved which will be presented at this meetings. Additionally, interesting abstracts have been submitted, which will be presented in the Oral Sessions and in the Poster Session. The Programme has been prepared to focus on the impending changes in our appreciations of Disorders of Sex Development, from a biological, epidemiological, psychological, surgical and last but not least clinical point of view. As there are no parallel sessions, you will be able to visit all presentations. The posters will be presented during the whole meeting, the Poster Session to be formally held on Friday evening.
We are very grateful to all distinguished speakers and poster presenters for sharing their data with us. Also we heartedly thank the experts, who will chair plenary lectures, the oral sessions and the poster presentations. We have invited experts from outside the DSD-consortium to chair the sessions, so we can discuss the forthcoming strategy for future networking research in field of DSD. The formal discussion will be held at the round table towards the end of the meeting on Sunday. This discussion should summarize the results of the main sessions and build the roadmap for a future consortium and a research network.
We welcome you at the new Audimax of the Universität zu Lübeck, where the meeting will not only give scientific knowledge, but also pave the grounds for personal discussions and exchange. You will have ample time for this at our welcoming reception in the Audimax on Friday evening. The Movie “Orchids – My intersex adventure” will be shown for those interested, thanks to Prof. Garry Warne. On Saturday evening, we invite you to the old city of Lübeck to our social evening, which will be held at the “Gemeinnützige”. This institution was founded at the end of the 18th century by educated citizens who organized charitable events. The Restaurant with its historical ballrooms and lovely garden is situated in a patrician house of the early 13th century vis-à-vis the Sankt Jakobi Kirche zu Lübeck.
We wish you a pleasant meeting and we hope that you enjoy your stay with us.
With warmest regards,
Olaf Hiort Lutz Wünsch
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
4 3rd International Symposium on Disorders of Sex Development
About EuroDSD
The EuroDSD is a collaboration of doctors and scientists from all over Europe. It is part of the 7th Framework Programme and its call for a collaborative project on the natural course and pathophysiology of rare diseases,
The consortium of EuroDSD consists of 13 European organisations
• Universität zu Lübeck • University of Glasgow • Insitut Pasteur, Paris • University of Cambridge • The University of Birmingham • Erasmus Universitair Medisch Centrum, Rotterdam • University College London • University of Pisa • Karolinska Institutet, Stockholm • Westfälische-Wilhelms-Universität Münster • Hospices Civils de Lyon • GABO: milliarium mbH & Co. KG, München • Christian-Albrechts-Universität zu Kiel
The EuroDSD consortium began its work on May 1st, 2008, funded as a collaborative project by the 7th framework programme of the European Union. It hypothesized that a stringent stepwise analysis of individuals with Disorders of Sex Development (DSD) will result in a systematic and reliable discovery of DSD- relevant biochemical, genetic and functional profiles. It was our aim to create a European data series on clinical features of patients with DSD of known and unknown cause. To achieve this, we created a registry incorporating pseudomized patient data and constructed a virtual research environment according to the consortium needs and with a very high security level. We have set out to meet the highest ethical standards, and all data are entered with respect to the individual consent level given by the patients. Since 2009, the consortium has opened up the registry to other international institutions that fullfil high security and ethical guidelines that were established in EuroDSD. Furthermore, the consortium has worked on the detection of new diagnostic markers, both in steroidogenesis as well as in genetics and it wants to provide the basis for explaining the natural course of these disorders. Characterization of the functional aspects of androgen action as the main basis for sex-related phenotype is seen to improve the understanding of the pathophysiology of DSD utilizing novel models. These approaches and the results are presented at the meeting. It will be a goal of the Symposium, to formulate new issues based on the current knowledge that can be addressed in a global scientific network on DSD research.
20 – 22 May 2011, Lübeck, Germany 5
General Information
Meeting Venue Universität zu Lübeck Building 65: Audimax, AM 3, AM-Foyer
Congress Language English, no translation facilities
Scientific and technical Scientific Poster Presentation and exhibition Exhibition of companies in the Foyer of the Audimax, Universität zu Lübeck
Internet WLAN
Badges Name badge upon registration Please wear it during all the symposium activities and social events
Onsite Registration Start 20 May 2011, 15:00 Foyer of the Audimax, Universität zu Lübeck
Conference Office All matters regarding registration, hotel booking, abstract handling, exhibition management and general information are managed by Kristin Bätzel, event lab., Leipzig
Meals included in the Coffee and lunch breaks according to the Scientific Programme registration fee Poster Party/Get-together on Friday, 20 May 2011
Certification Accredited by Ärztekammer Schleswig-Holstein (13 credit points)
Programme Organizing Olaf Hiort, Committee Lutz Wünsch, Ute Thyen, Ralf Werner, Paul-Martin Holterhus
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
6 3rd International Symposium on Disorders of Sex Development
Scientific Programme
Friday, May 20 Time Topic Speaker
15:00 Registration Audimax Foyer
16:30 Opening Ceremony Audimax 3
17:00 Opening Lecture Audimax 3 Chairs: O. Hiort, L. Wünsch
K-01 From Gene to Gender - What we’ve learned and what we need to learn - New Prospects in DSD Research G. Warne
18:00 Poster Session Audimax Foyer
19:00 Poster Party Audimax Foyer
Saturday, May 21 Time Topic Speaker
09:00 Genetics Audimax 3 Chairs: R. Siebert, G. Scherer
K-02 New genes for DSD - CGH Microarray P. Wieacker
K-03 Old genes, much faster explored-DSDChip K. McElreavy
K-04 Beyond the genes: Epigenetic control P. M. Holterhus
10:30 Coffee Break Audimax Foyer
11:00 Hormones & Action Audimax 3 Chairs: C. Flück, A. Juul
K-05 Co-Factors of androgen action R. Werner
K-06 New approaches to measuring hormones F. Riepe
K-07 Urinary steroids: Sense and sensitivity W. Arlt
K-08 Pubertal outcome: Predicting changes I.A. Hughes
13:00 Lunch Break Audimax Foyer
20 – 22 May 2011, Lübeck, Germany 7
Scientific Programme
Saturday, May 21 Time Topic Speaker
14:00 Oral Session 1 Chairs: F. Eckholt, A. Nordenskjold Audimax 3
O-01 Disclosure in disorders of sex development: Preliminary analysis using parent report V. Pasterski
O-02 From ethical guidelines to clinical practice – Assessing the impact of value changes in clinical practice J. Streuli
O-03 Management of vaginal hypoplasia in disorders of sex development: A retrospective study comparing surgical and non-surgical options N. Callens
O-04 Long term outcomes in XY DSD raised as females treated in a single institution K. Gueniche
O-05 Satisfaction with treatment, genital surgery and sexual life in adults with XY disorders of sex development – Results from the German clinical evaluation study B. Köhler
O-06 Voice characteristics in adult CAH-Women A. Nordenskjold
O-07 Bone Mineral Density in Women with complete androgen insensitivity syndrome E. Dati
15:30 Coffee Break Audimax Foyer
16:00 Morphologic Variation in DSD Audimax 3 Chairs: J. Schober, P. Cuckow
K-09 Estrogen and sexual differentiation: Lessions learned from the spottetd hyena, normal and mutant mice L. Baskin
K-10 Gonadal Development and Tumour Risk M. Cools
K-11 Evaluation of DSD with abnormal genitalia: a surgical approach P. Mouriquand
K-12 Checklist for structural description L. Wünsch
18:00 End
19:30 Evening Programme
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
8 3rd International Symposium on Disorders of Sex Development
Scientific Programme
Sunday, May 22 Time Topic Speaker
09:00 Clinical Aspects Audimax 3 Chairs: H. Meyer-Bahlburg, L. Audi
K-13 Outcome Studies U. Thyen
K-14 The logistics and ethics of data bases S. F. Ahmed
K-15 Application of an ESPE DSD e-learning webportal S.L.S. Drop
K-16 Research in DSD – beyond descriptive studies O. Hiort
11:00 Coffee Break Audimax Foyer
11:30 Oral Session 2 Audimax 3 Chairs: J. Achermann, M. Niedziela
O-08 Development of a novel UPLC-MS/MS-Assay for eight C21 steroids, reference data for children, and its application on congenital adrenal hyperplasia A. Kulle
O-09 Recurrent aberrations identified by array-CGH in patients with Mayer-Rokitansky-Küster-Hauser-Syndrome S. Ledig
O-10 Exome sequencing reveals novel genetic factors involved in DSD A. Bashamboo
O-11 Functional analysis of novel androgen receptor variants predict phenotypes in patients with androgen insensitivity syndrome A. Brinkmann
12:30 Round Table Audimax 3
K-16 Future Perspectives of DSD Research R. Siebert C. Flück J. Schober H. Meyer-Bahlburg C. Quigley I. Mazen O. Hiort
13:15 Closing Remarks and Fare Well
20 – 22 May 2011, Lübeck, Germany 9
Social Programme
Friday, 20 May 2011
Time Topic 19:00 Poster Party / Get-together Audimax Foyer Including finger food After the Poster Session we would like to invite you to our Poster Party for personal discussions and exchange. 20:00 Orchids – My intersex adventure Audimax 3 The Society Netzwerk DSD /Intersexualität e.V. will present the Australian Movie “Orchids – My intersex adventure” from the Documentary filmmaker, Phoebe Hart.
We would like to thank Prof. Garry Warne for providing this film.
Saturday, 21 May 2011
Time Topic 19:30 Dinner and Party Gemeinnützige Onsite registration possible (35 €)
On Saturday evening, we would like to invite you to the old city of Lübeck for our social evening, which will be held at the “Gemeinnützige”.
This institution was founded at the end of the 18th century by educated citizens who organized charitable events. Address: The restaurant with its historical ballrooms and lovely garden is Die Gemeinnützige th situated in a patrician house of the early 13 century vis-à-vis Restaurant Heinrichs the Sankt Jakobi Kirche zu Lübeck. Königstraße 5-7 23552 Lübeck
Programme: JAZZ FOR FUN
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
10 3rd International Symposium on Disorders of Sex Development
Speakers’ List
Keynote Speakers (in alphabetic order)
Speaker/Institution Speaker/Institution Session/Abstract No Session/Abstract No Faisal Ahmed Ken McElreavy University of Glasgow, UK Institut Pasteur, Paris, France Clinical Aspects Genetics K-14 K-03
Wiebke Arlt Pierre Mouriquand University of Birmingham, UK Université de Lyon, France Hormones & Action Morphologic Variation in DSD K-07 K-11
Laurence Baskin Felix Riepe University of California, San Francisco, Christian-Albrechts-Universtät Kiel, USA Germany Morphologic Variation in DSD Hormones & Action K-09 K-06
Martine Cools Ute Thyen Erasmus University of Rotterdam, NL Universität zu Lübeck, Germany Morphologic Variation in DSD Clinical Aspects K-10 K-13
Stenvert Drop Garry Warne Erasmus University of Rotterdam, NL University of Melbourne, Australia Clinical Aspects Opening Lecture K-15 K-01
Olaf Hiort Ralf Werner Universität zu Lübeck, Germany Universität zu Lübeck, Germany Clinical Aspects Hormones & Action K-16 K-05
Paul-Martin Holterhus Peter Wieacker Christian-Albrechts-Universität Kiel, Westfälische Wilhelms-Universität Germany Münster, Germany Genetics Genetics K-04 K-02
Ieuan Hughes Lutz Wünsch University of Cambridge, UK Universität zu Lübeck, Germany Hormones & Action Morphologic Variation in DSD K-08 K-12
20 – 22 May 2011, Lübeck, Germany 11
Speakers’ List
Chairs (in alphabetic order)
Speaker/Institution Session John Achermann Oral Session 2 University College London, UK Ivo Arnhold Poster Session Hospital das Clinicas, Brasil Clinical Aspects 2 Laura Audi Clinical Aspects Hospital Vall d'Hebron, Spain Silvano Bertelloni Poster Session University Hospital Pisa, Italy Genetics Albert Brinkmann, Poster Session ErasmusMC, The Netherlands Hormones & Action 1 Peter Cuckow Morphological Variation in DSD Great Ormond Street Hospital for Children, UK Felicitas Eckholt Oral Session 1 Friedrich Schiller University Jena, Germany Christa Flück Hormones & Action Inselspital, Universitätsklinik für Kinderheilkunde, Switzerland Anders Juul Hormones & Action University Department of Growth and Reproduction, Denmark Nils Krone Poster Session University of Birmingham, UK Hormones & Action 2 Dagmar l’Allemand Poster Session Ostschweizer Kinderspital, St. Gallen, Switzerland Clinical Aspects 1 Leendert Looijenga Poster Session Erasmus MC, The Netherlands Morphologic Variation in DSD Heino Meyer-Bahlburg Clinical Aspects NYS Psychiatric Institute, USA Marek Niedziela Oral Session 2 Poznan University of Medical Sciences, Poland Agneta Nordenskjold Oral Session 1 Karolinska Institutet, Sweden Charmian Quigley Round Table Lilly Corporate Center, USA Gerd Scherer Genetics Universitätsklinikum Freiburg, Germany Justine Schober Morphological Variation in DSD Hamot Medical Centre, USA Reiner Siebert Genetics UK S-H, Campus Kiel, Germany
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
12 3rd International Symposium on Disorders of Sex Development
Poster Presentation
Genetics Chair: S. Bertelloni No Presenting Author Titel P-01 Baldazzi, Lilia Seven novel NR5A1 gene mutations in a cohort of 16 Italian patients with 46,XY disorders of sex development (DSD) without adrenal insufficiency P-02 Norling, Ameli Array-CGH to identify novel genes involved in 46,XY and 46,XX gonadal dysgenesis: how many are still to be found? P-03 Miles, Harriet Louise Identical twins of opposite sex P-04 McElreavey, Ken Mutation analysis of NR5A1 encoding steroidogenic factor 1 in 81 patients with 46,XY disorders of sex development (DSD) including hypospadias P-05 van der Zwan, Yvonne A novel SRY missense mutation in a 46,XY female patient with bilateral gonadoblastoma and dysgerminoma P-06 Delle Chiaie, Barbara Array-CGH analysis in the Ghent University Hospital disorders of sexual development population
Hormones & Action 1 Chair: A. Brinkmann P-07 Petroli, Reginaldo José Synergestic effect for p.Q798E and p.C806F mutations in a patient with Complete Androgen Insensitivity Syndrome P-08 Hughes, Ieuan Analysis of DSD patient mutations located in the N-terminal domain of the Androgen Receptor: helping to understand domains with molten globular structure P-09 Mooslehner, Katrin Developing cell assays suitable for characterizing AR mutations associated with Androgen Insensitivity Syndrome P-10 Grötsch, Helga Rwdd1: an androgen receptor coregulator with a putative role in male genital development P-11 Bunch, Trevor Investigating the pathogenicity of androgen receptor (AR) mis-sense mutations in Androgen Insensitivity Syndrome (AIS)
Hormones & Action 2 Chair: N. Krone P-12 Hughes, Ieuan Identification of the FKBP4 mutation c.T1575C (pX460Q) in a DSD patient with aberrant genital skin fibroblast ligand binding, but a normal AR P-13 Miles, Harriet Louise Clinical outcome studies for disorders of androgen production: SRD5A2 and 17ßSD-3 P-14 Ellaithi, Mona Disorders of sexual development in a group of Sudanese: Harmonical diagnosis P-15 Riedl, Stefan Severe virilization of a 46,XX newborn and her mother during pregnancy: A novel case of aromatase deficiency P-16 Doehnert, Ulla Hormone Profiles in Adolescents and Adults with Complete Androgen Insensitivity Syndrome (CAIS) P-17 Rodie, Martina Exploring the utility of urine steroid metabolite ratios in evaluating over 500 suspected cases of disorders of sex hormone synthesis
20 – 22 May 2011, Lübeck, Germany 13
Poster Presentation
Morphologic Variation in DSD Chair: L. Looijenga No Presenting Author Titel P-18 van der Zwan, Yvonne Feminizing surgery in women with Congenital Adrenal Hyperplasia: Long term outcome study on anatomy, cosmetics, sexual- and psychological functioning P-19 Slowikowska-Hilczer, A fate of dysgenetic gonads Jolanta P-20 Schober, Justine Sensorial role of the epithelium of the labia minora in prepubertal girls P-21 Schober, Justine Immunocytochemical characterization of Pacinian-like corpuscles in the labia minora of prepubertal girls P-22 Schober, Justine Sensory Neural Characteristics of Merkle Cells in the Labia Minora of Prepubertal Girls P-23 Wolffenbuttel, Katja Hypospadias repair with preservation of the vagina in boys with DSD P-24 McGowan, Ruth Clinical and molecular genetic investigation of congenital Müllerian abnormalities P-25 Hersmus, Remko Delayed recognition of two Disorder of Sex Development (DSD) cases: a missed possibility for early diagnosis of malignant germ cell tumors P-26 Özbey, Hüseyin Surgical correction of webbed penis associated with penoscrotal transposition
Clinical Aspects 1 Chair: D. l’Allemand P-27 Huber, Kerstin Sexual quality of life in individuals with Complete Androgen Insensitivity (CAIS) and Mayer-Rokitansky-Küster-Hauser Syndrome (MRKHS) P-28 Fliegner, Maike Infertility and the wish for a child: A study on attitudes towards parenthood in people with complete androgen insensitivity syndrome (CAIS) and Mayer- Rokitansky-Küster-Hauser Syndrome (MRKHS) P-29 Prochnow, Caroline Body experience in individuals with different conditions of intersexuality P-30 Birnbaum, Wiebke Comparison of clinical and metobolic effects of testosterone and estradiol in adult gonadectomized patients with 46, XY DSD due to complete androgen insensitivity syndrome (CAIS) P-31 Brunner, Franziska Evaluation of Gonadectomy in Adults with Complete Androgen Insensitivity Syndrome (CAIS): A Patients´ Perspective P-32 Dessens, Arianne B Body image and sexuality in Indonesian treated and untreated subjects with DSD P-33 Meyer-Bahlburg, Cognitive and educational outcome of offspring from dexamethasone- Heino FL treated pregnancies at risk for congenital adrenal hyperplasia due to 21- hydroxylase deficiency P-34 Handford, Christina The pros and cons of a third gender: Attitudes amongst intersex adults with complete (CAIS) or partial (PAIS) androgen insensitivity syndrome P-35 Callens, Nina Multidimensional body image and sexual functioning comparisons among patients with disorders of sex development (DSD) and controls: A multisite study
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
14 3rd International Symposium on Disorders of Sex Development
Poster Presentation
Clinical Aspects 2 Chair: I. Arnhold No Presenting Author Titel P-36 Russo, Gianni Use of GnRH analogues in the management of disorders of sex development diagnosed at puberty P-37 Tufail, Irum Clinical, hormonal and chromosomal analysis of undervirilized male/46XYDSD - A 3years experience of national institute of child health P-38 Rodie, Martina The European DSD Register - A Platform For International Collaborative Research P-39 Russo, Gianni 46,XY and female phenotype fetus: A challenging diagnosis P-40 Szarras-Czapnik, Maria Concomitant gonadal dysgenesis and non-classic congenital adrenal hyperplasia in a girl with 46,XY DSD P-41 Özbey, Hüseyin Transverse testicular ectopia, persistent Müllerian duct and bilateral duplication of the vas deferens. Embryology, presentation and surgical management P-42 Özbey, Hüseyin Bilateral testicular regression syndrome: Report of 2 years old boy P-43 Mazen, Inas Pattern of DSD and genital anomalies: Egyptian experience P-44 Holmdahl, Gundela Fertility and pregnancy outcome in women with congenital adrenal hyperplasia due to 21-hydroxylase deficiency
Presenting Authors Presenting Author Abstract Page Presenting Author Abstract Page Baldazzi, Lilia...... P-01...... 42 Miles, Harriet Louise...... P-13...... 54 Birnbaum, Wiebke ...... P-30...... 71 Mooslehner, Katrin...... P-09...... 50 Brunner, Franziska ...... P-31...... 72 Norling, Ameli...... P-02...... 43 Bunch, Trevor ...... P-11...... 52 Özbey, Hüseyin...... P-26...... 67 Callens, Nina ...... P-35...... 76 Özbey, Hüseyin...... P-41...... 82 Delle Chiaie, Barbara...... P-06...... 47 Özbey, Hüseyin...... P-42...... 83 Dessens, Arianne B ...... P-32...... 73 Petroli, Reginaldo José...... P-07...... 48 Doehnert, Ulla...... P-16...... 57 Prochnow, Caroline...... P-29...... 70 Ellaithi, Mona...... P-14...... 55 Riedl, Stefan...... P-15...... 56 Fliegner, Maike...... P-28...... 69 Rodie, Martina...... P-17...... 58 Grötsch, Helga...... P-10...... 51 Rodie, Martina...... P-38...... 79 Handford, Christina ...... P-34...... 75 Russo, Gianni...... P-36...... 77 Hersmus, Remko...... P-25...... 66 Russo, Gianni...... P-39...... 80 Holmdahl, Gundela ...... P-44...... 85 Schober, Justine...... P-20...... 61 Huber, Kerstin...... P-27...... 68 Schober, Justine...... P-21...... 62 Hughes, Ieuan ...... P-08...... 49 Schober, Justine...... P-22...... 63 Hughes, Ieuan ...... P-12...... 53 Slowikowska-Hilczer, Jolanta.... P-19...... 60 Mazen, Inas...... P-43...... 84 Szarras-Czapnik, Maria...... P-40...... 81 McElreavey, Ken...... P-04...... 45 Tufail, Irum...... P-37...... 78 McGowan, Ruth ...... P-24...... 65 van der Zwan, Yvonne ...... P-05...... 46 Meyer-Bahlburg, Heino FL ...... P-33...... 74 van der Zwan, Yvonne ...... P-18...... 59 Miles, Harriet Louise...... P-03...... 44 Wolffenbuttel, Katja...... P-23...... 64
20 – 22 May 2011, Lübeck, Germany 15
Abstracts Keynote Lectures
K-01 FROM GENE TO GENDER. WHAT WE'VE LEARNED AND WHAT WE NEED TO LEARN - NEW PROSPECTS IN DSD RESEARCH
# G.L. Warne1 1Royal Children's Hospital Melbourne, Endocrinology and Diabetes, Parkville Victoria, Australia The last decade’s publications relating to DSD show great advances in four fields. In the first, the field of laboratory-based research, the decade began with explorations of gene structure, genotype-phenotype correlations and the application of the human genome project, which was completed in 2003. Through studies of naturally-occurring conditions, such as 46,XY DSD and 46,XX testicular DSD, new genes were added to the pathway for testis development. At last, some genes that regulate ovarian development were discovered. Genetic insights were gained into some old conditions, such as premature ovarian failure, and some new diseases, such as P450 oxidoreductase deficiency were identified. The phenotype was widened for a number of conditions, such as SF-1 deficiency, lipoid adrenal hyperplasia and WNT4 deficiency. The pathogenesis of germ cell cancer was better understood. New technologies, such as micro-arrays and exome sequencing, were introduced with considerable early success in gene discovery. The anatomy of the clitoris was discovered in 2005 to be very different from what had been thought. Secondly, some radical thinking about treatment led to new ideas about the future roles of stem cell therapy and transplantation in the restoration of fertility. Some patients with CAH were treated using bilateral adrenalectomy. The third area of endeavour in the past decade resulted in many long-term outcome studies and a serious attempt to review diagnostic and treatment strategies in the light of the findings from these studies. There was a strong focus on sex, gender, sexuality and quality of life, with psychologists and biomedical ethicists coming to centre stage. Consensus guideline workshops were held on DSD, CAH and GID, bringing about major changes in clinical practice. Fourthly, trans-cultural outcome studies were carried out in countries such as India, Brazil, Vietnam, Egypt and the Middle East, highlighting the great suffering and much shorter survival of children with CAH and DSD who live in resource-poor areas of the world and the gross inequities that need to be corrected. Looking ahead to the next decade, it is clear there is much to be done. Between 50 and 60% of patients with 46,XY DSD never receive a definitive diagnosis. Decision making in relation to Y chromosome positive DSD is as difficult as it ever was and in some parts of the world, the threat of legal action is driving the referral of difficult medical decisions into the courts, which is highly undesirable. Ways of improving long term outcomes for common conditions are urgently needed, as are better ways of funding, organizing and delivering multidisciplinary services. I hope there will also be a big effort on the part of wealthy and developed countries to support the development of services for DSD patients in less fortunate parts of the world.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
16 3rd International Symposium on Disorders of Sex Development
K-02 NEW GENES FOR DSD - CGH MICROARRAY
# P. Wieacker1 1Westfälische Wilhelms-Universität Münster, Institut für Humangenetik, Münster, Germany
Objectives: Disorders of gonadal development are very heterogeneous disorders. XY gonadal dysgenesis (XY-GD) is characterized by a failure of testis differentiation leading to streak gonads and female genitalia. Premature ovarian failure (POF) is characterized by the onset of amenorrhea before age 40 years associated with increased concentrations of gonadotropins. XX ovarian dysgenesis (OD) can be seen as the most severe manifestation of POF with complete depletion of follicles before puberty. Although an increasing number of causative genes could be detected in the past, currently the majority of cases of XY-GD, OD or POF remain unexplained. Mayer-Rokitansky-Küster-Hauser syndrome (MRKH) is a congenital anomaly of the Müllerian ducts characterized by rudimentary or absent uterus and agenesis of the upper vagina. Methods: We performed molecular karyotyping based on array-CGH (Comparative Genomic Hybridization) in large cohorts of patients affected by XY-GD, OD or POF and MRKH. Results: In 25% of syndromic and 10% of non-syndromic cases with XY-GD causative microdeletions or - duplications involving DMRT1, DAX1 and SOX9 could be detected. Array-CGH in POF- and OD-patients led to the identification of microdeletions or -duplications including genes involved in i) meiosis, ii) DNA repair and iii) folliculogenesis or male infertility in homologs of model organisms. In MRKH-patients 3 recurrent microdeletions including candidate genes could be detected. Conclusion: Molecular karyotyping is a powerful method to identify microdeletions or microduplications affecting genes involved in sexual development.
20 – 22 May 2011, Lübeck, Germany 17
K-03 OLD GENES, MUCH FASTER EXPLORED-DSDCHIP
# K. McElreavy1 1Institut Pasteur, Paris, France
The diagnostic sequencing of genes known to be responsible for DSD using conventional Sanger sequencing is useful to identify novel mutations associated with gonad development and it is the mainstay of most diagnostic laboratories. However, this approach is both time-consuming and laborious. An accurate diagnosis is still lacking in approximately 40% of all patients with DSD. This frequency is even higher in patients with 46,XY DSD. High-throughput, high-density sequencing offers the possibility of rapidly and accurately sequencing portions of the genome at reduced costs. Several next-generation sequencing (NGS) technologies are available for whole genome or exome analyses but at present the costs and considerable data handling that are involved means that these approaches will be limited to the research environment for the foreseeable future. In contrast, sequencing by hybridisation is suitable for situations in which a moderate amount of sequence is being analysed (10-300 kb, representing several to tens of genes) in a repetitive manner. Here, I will describe a DSDChip that is being developed for the complete analyses of 36 genes corresponding to three functional groups (steroid hormone synthesis, hypothalamic-pituitary and gonadal) as well as the SOX9 TESCO enhancer sequence. The array also includes 50 bases of the intronic sequence bordering exons (potential splice sites), and 500 bases upstream from the first exon (potential regulatory elements) for each of the DSD gene. Protocols have been developed for targeted enrichment of patient DNA using locus-specific long-range PCR amplification of regions of interest. Long-range PCR minimises the number of reactions required. Although these PCR products can then pooled, fragmented, labeled and subsequently hybridised with the microarray and then scanned, these protocols can also be modified for targeted NGS approaches. The generation of vastly improved data sets of DNA sequences for each patient sample, both rapidly and at a relatively limited cost will transform the field of DSD by offering an accurate diagnosis, and by providing novel insights into the genetics and physiology of DSD.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
18 3rd International Symposium on Disorders of Sex Development
K-04 BEYOND THE GENES: EPIGENETIC CONTROL
# P.-M. Holterhus1 1Christian-Albrechts-Universität Kiel, UK S-H, Campus Kiel, Klinik für Allgemeine Pädiatrie, Kiel, Germany
Sex-specific development of the human comprises irreversible sexual differentiation of the external genitalia during embryogenesis, extra-genital sexual dimorphism during puberty (e.g., body proportions, pubertal voice change) as well as sex-specific development of the brain. The presence or the absence of normal androgen biosynthesis and normal androgen action via the androgen receptor play key roles in these processes. At the single cell level, androgens induce specific reversible changes of the cellular transcriptional program responsible for the biological short-term effects eventually leading to long-term changes of morphology and function. Our genome-wide gene expression analyses on cultured genital fibroblasts as well as on mononuclear cells of the blood derived from normal males, females and individuals with defined DSD- conditions have shown that androgens have imprinted comprehensive long-term changes of the basal transcriptome. We have called this phenomenon a transcriptional androgen memory. Work package 4 of the EuroDSD project has investigated the epigenomic background of androgen memory. Indeed, our large scale methylome analyses on labioscrotal fibroblasts using Illumina microarrays have revealed the existence of significantly (q=0.05) different epigenotypes comparing normal males with 46XY patients having molecular proven AIS. While the presence or absence of an androgen receptor gene mutation is a significant determinant for differential gene methylation, there is only a poor epigenotype - external genital phenotype - correlation. This implies that the role of the androgen receptor and of androgen receptor gene mutations in altering epigenomic programming is not restricted to the degree of genital virilization but must also include extragenital aspects of sex-specific development and function.
20 – 22 May 2011, Lübeck, Germany 19
K-05 COFACTORS OF ANDROGEN ACTION
# R. Werner1, H. Grötsch1, M. Kunert1, Z. Gao2, O. Hiort1 1Universität zu Lübeck, Department of Paediatrics, Lübeck, Germany 2The Children’s Hospital Zhejiang, University School of Medicine, Department of Pediatric Surgery, Zhejiang, China
Male genital development is strongly dependent on the presence of androgens during a small time window ranging from the 7th-12th week in humans or embryonic stages E15-E17 in mice. Androgen action is mediated by the androgen receptor (AR), a nuclear receptor and transcription factor that binds to its cognate hormone responsive elements and elicits transcriptional regulation of androgen responsive target genes. Transcriptional regulation by steroid receptors is a complex dynamic process involving a large set of coregulators. So far more than 200 androgen receptor coregulators are known possessing coactivating or corepressing functions. We presumed that specific AR coregulators might be expressed during the androgen-sensitive time window of genital development and could contribute to so far unknown causes of AIS. To identify androgen receptor coregulators expressed in the male genital tubercle during the critical phase of androgen-dependent development, we constructed three independent cDNA libraries of male genital tubercles from male mouse embryos at stage E15, E16 and E17 and searched for AR interacting proteins by independent yeast two-hybrid screenings. We identified 15 known androgen receptor coregulators in 37 independent clones, half of them in at least two independent screenings. Beside the known AR coregulators we identified 25 different AR-binding proteins with functions in transcriptional and cell cycle control.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
20 3rd International Symposium on Disorders of Sex Development
K-06 NEW APPROACHES TO MEASURING HORMONES
# F. Riepe1, A. Kulle1 1Christian-Albrechts-Universität Kiel, UK S-H, Campus Kiel, Klinik für Allgemeine Pädiatrie, Kiel, Germany
Liquid-chromatography - tandem mass spectrometry (LC-MS/MS) is becoming the method of choice for clinical steroid analysis. In most instances it has the advantage of higher sensitivity, better reproducibility and greater specificity than commercial immunoassay techniques. The method requires only minimal sample preparation and a small sample volume. Furthermore it has the potential to analyze multiple steroids simultaneously. Modern instruments guarantee high throughput, allowing an affordable price for the individual assay. All this makes LC-MS/MS an attractive method for use in a research or clinical setting. Reliable reference ranges for the detected analytes are the prerequisite for their clinical use. By this, LC- MS/MS can find its application for diagnostics and follow up of patients with disorders of sex development (DSD). In contrast to GCMS methods LC-MS/MS uses an analyte focussed approach rather than a global description of the steroid metabolome. However due to high throughput with minimal sample volumes as well as low costs LC-MS/MS is the ideal tool for initial diagnostic as well as for repetitive steroid determinations e.g. for treatment monitoring. GCMS has its advantage in equivocal diagnostic settings. Within the EURO-DSD network we applied the technique in patients with DSD and were able to reassure or make the diagnosis of various forms of congenital adrenal hyperplasia and other disorders of sex development due to disturbances of steroid biosynthesis. An overview on the application of LC-MS/MS in clinical routine and in research settings is given and results from the network study are presented.
20 – 22 May 2011, Lübeck, Germany 21
K-07 URINARY STEROIDS: SENSE & SENSITIVITY
# W. Arlt1, N. Krone1, A.E. Taylor1, P. Schneider1, C.H.L. Shackleton1 1University of Birmingham, Institute of Biomedical Research, Birmingham, United Kingdom
Mass spectrometry based steroid detection represents the state-of-the-art in steroid analysis, a most important approach for the rapid and specific diagnosis of adrenal and gonal disorders including 46,XX and 46,XY disorder of sex development (DSD). Since the 1930s almost all inborn errors of steroidogenesis have been first defined through the detection of their urine steroidobolome. In the last 30 years this has been exclusively carried out by gas chromatography/mass spectrometry (GC/MS), a unique and comprehensive discovery tool that has helped to define novel disorders including P450 oxidoreductase deficiency and apparent cortisone reductase deficiency. Moreover, analysis of maternal urine is an invaluable tool for prenatal detection of steroidogenic disorders such as Smith-Lemli-Opitz syndrome. A key advantage of GC/MS is its non-selective nature, allowing for concurrent analysis of the entire steroidobolome of an individual, with current scanning runs automatically identifying and quantifying more than 40 steroids. The use of diagnostic ratios consisting of substrate metabolite over product metabolite of a distinct enzymatic reaction allows for specific diagnosis of DSD and other steroidogenic disorders often even when employing spot urine only, e.g. 17,20 lyase deficiency, 5α-reductase deficiency, 17b-hydroxysteroid type 3 deficiency. GC/MS data become more accessible if presented in simplified graphical visualizations, which have been recently developed and tested for its usefulness in diagnosing DSD. Importantly, novel machine learning approaches allow for the computational analysis of steroid data and guide the way for the discovery of hitherto unknown disorders.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
22 3rd International Symposium on Disorders of Sex Development
K-08 DISORDERS OF SEX DEVELOPMENT: Pubertal outcome: predicting changes
# I. Hughes1 1Department of Paediatrics, University of Cambridge, UK
Disorders of sex development (DSD) present primarily at birth with an overriding urgency for the family and medical attendants to reach a decision on sex assignment. Coupled to that decision is attempting to predict the outcome for puberty and later reproductive function. Causes under the heading XY DSD pose additional difficulties in predicting changes at puberty when the affected infant is assigned male. Outcome data are sparse, particularly in partial androgen insensitivity syndrome (PAIS) and androgen biosynthetic disorders such as 5alpha-reductase (5RD) and 17beta-hydroxysteroid dehydrogenase (17HSD) enzyme deficiencies. The latter two conditions may present de novo at puberty and prompt a decision on sex re-assignment. The development of an external masculinisation score (EMS) based on examination of external genital development in males (normal score, 12) allows an objective and reproducible assessment of under- masculinisation in early infancy which can be tracked to subsequent masculinisation at puberty. Thus, using an arbitrary EMS division at 5 in 16 PAIS individuals with a proven dysfunctional mutant androgen receptor, puberty developed spontaneously with an EMS >5 whereas high-dose androgens were needed to induce puberty with an EMS <5. There was also associated information relating to adult height, presence of gynaecomastia and evidence of spermatogenesis. There was a reasonable correlation with responsiveness to androgens in an in vitro transactivation assay, particularly at the extremes of the EMS range. Individuals with the commoner XY DSD condition comprising a PAIS-like phenotype but a normal androgen receptor displayed EMS values of a similar range and a tendency to more spontaneous onset of puberty. A system is now available as a prediction model of puberty outcome in XY DSD individuals assigned male that will require multi-centre participation for its validation.
20 – 22 May 2011, Lübeck, Germany 23
K-09 ESTROGEN AND SEXUAL DIFFERENTIATION: LESSONS LEARNED FROM THE SPOTTED HYENA, NORMAL AND MUTANT MICE
# L.S. Baskin1 1University of California, San Francisco, Department of Urology, San Francisco, United States
According to Jost, masculinization of the external genitalia involves action of androgens. Feminine differentiation has been characterized as a “default condition” due to absence of androgen action, thus discouraging the search for active mechanisms involved in feminine differentiation. Although Jost is undoubtedly correct in regard to androgens and has provided a remarkably successful foundation for research in this area, acceptance of the Jost model has led to neglect of other potential mechanisms such as direct genetic effects, or a role for non-androgenic hormones. We propose to extend the classic Jost hypothesis of sexual differentiation by showing that female sexual differentiation is not simply a default pathway secondary to a lack of androgens but an active process dependent upon estrogen. Furthermore, male external genitalia (ExG) differentiation may not be solely a function of androgens but also influenced by estrogens. The overall hypothesis of our work is that normal male and female development of the external genitalia is dependent upon the correct balance of androgen and estrogen that signal through their respective receptors. Lessons Learned from the Spotted Hyena along with normal and mutant mice will be presented that support our hypothesis.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
24 3rd International Symposium on Disorders of Sex Development
K-10 GONADAL DEVELOPMENT AND TUMOUR RISK
# M. Cools1,2, L. Looijenga3,4 1University Hospital Ghent, Ghent, Belgium 2University of Ghent, Ghent, Belgium 3Erasmus Medical Center, Department of Pathology, Rotterdam, Netherlands 4Josephine Nefkens Institute, Rotterdam, Netherlands
In recent years, knowledge on genes implicated in gonadal determination and differentiation has dramatically increased. The human process of gonadal development however remains somewhat enigmatic, and almost totally relies on extrapolation of data obtained from the functional characterization of these genes in animal and in vitro models. Morphologic and immunohistochemical studies in gonads from DSD patients with identified genetic anomalies represent a unique and highly informative source of information in this context. Comparison of data obtained in over 200 gonadal samples from gonadal dysgenesis patients with literature data shed a new light on parallels and differences between the processes of sex determination and differentiation in humans and mice, and describe the role of proteins such as SRY, SOX9 and FOXL2 in human gonadal development. The degree of gonadal differentiation / testicularization is related to the risk for its malignant transformation, in particular the formation of malignant germ cell tumors (GCTs). Incompletely matured Sertoli/granulosa cells are insufficiently capable of directing the normal mitotic block / meiotic induction germ cell program, and germ cells in such a disturbed micro-environment are delayed or blocked in their normal maturation process. Thereby, they remain pluripotent and gain increased mitotic and survival characteristics, being the first step in the pathogenesis of GCTs. The patient's phenotype might be informative in assessing the degree of gonadal testicularization on a clinical basis. This contribution integrates genetic and immunohistochemical data on gonadal (mal)development in humans, actual knowledge on the pathophysiology of tumor development, and possible correlations between the patient's phenotype and his / her risk for GCT development. Current knowledge allows development of an informative cancer risk assessment of DSD patients.
20 – 22 May 2011, Lübeck, Germany 25
K-11 EVALUATION OF DSDS WITH ABNORMAL GENITALIA: A SURGICAL APPROACH
# P. Mouriquand1 1HCL-Lyon, Bron, France
The complexity and heterogeneity of presentation along with the variable management of DSDs with abnormal genitalia demands a consensual initial approach and post-treatment evaluation. Data currently available is too scattered and flawed to select the optimal treatments for these patients. Clinicians face 3 main types of abnormal genitalia: 1) The 46, XY DSDs essentially represented by Hypospadias; 2) The 46,XX mainly represented by Congenital Adrenal Hyperplasia; 3) The heterogenous group of gonosomal anomalies including Y deficient patients essentially represented by the Mixed Gonadal Dysgenesis . The first group mostly requests “masculinization” procedures i.e. hypospadias repair with variable techniques all bearing a significant number of complications. The second group commonly requests “feminization” procedures i.e. a connection of the vagina to the perineum, a possible reduction of the clitoris and a perineoplasty. Timing and nature of this surgery remain highly debatable. The third group raises the most difficult discussions for the choice of gender and interacts with several concepts which are not clearly defined: 1) Individual Sexual Identity which is what the individual considers himself or herself to be, 2) Social Identity (= gender) which is the way the “society” looks at the individual and 3) The Behavioral Identity which is the erotic inclination of each individual. The first and the third ones are not visible in the neonatal period which leads to the paradoxical situation where the “society” i.e. doctors and parents choose a gender (= social concept) for an individual without knowing his (her) Individual and Behavioral Identities. This critical situation has led some teams to delay any irreversible decision until a later stage of life when the individual will be able to express his (her) “subtle” identities. This position is however not shared by the entire scientific community and most DSD teams will select a gender soon after birth to avoid or diminish the psychological impact of a non- decision policy which implies “non-visibility” in the society. The gender decision made at an early stage of life is currently based using various tools which can be separated into 4 groups: 1) The Internal Sex represented by the genetic material, gonads and hormones; 2) The External Sex represented by the underdeveloped genital tubercle and the potential presence of a mullerian retro-urethral cavity; 3) The Functional Sex which is the potentiality for the individual to have intercourse and children (as a male or a female); 4) The Social Sex which is the social and cultural medium in which is the individual is going to be raised. The objective of this surgical evaluation is to define a common ground between international DSD teams to compare and improve our results. Surgical evaluation is the most reliable Ariadne’s thread to identify DSDs and their severity.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
26 3rd International Symposium on Disorders of Sex Development
K-12 CHECKLIST FOR THE STRUCTURAL DESCRIPTION
# L. Wünsch1 1Universität zu Lübeck, UK S-H, Campus Lübeck, Klinik für Kinder- und Jugendmedizin, Lübeck, Germany
Background: Disorders of sex development are characterized by variable spectrum of urogenital malformations, some of which require surgical intervention. Genital surgery and the control of tumour risk are particularly controversial issues that require further research. Outcomes depend on the quality of interventions but also on the underlying condition and the individual anatomy. We found that little attention has been given to the quality of the initial anatomic evaluation. Our own experience and a review of the literature showed a wide variability of diagnostic procedures and the terminology that was used to report the findings. From these observations we concluded that a standardizing the elements of the surgical evaluation would be useful. Based on the reevaluation of 66 patients with various DSD treated at our institution we propose a structured examination schedule. It should be discussed as a base for a future standard operating procedure. This may facilitate clinical decision making, create a comparable data base for future investigations and improve quality by eliminating ambivalent terminology. Patients and Methods: During 2022 to 2011, 66 patients underwent diagnostic evaluation at our institution. All patients who had either laparoscopy, endoscopy or both were included. Diagnosis were complete androgene insensivity (7), 46 XY complete gonadal dygenesis (11), partial or mixed gonadal dysgenesis (11), defects of androgene synthesis (7), ovotesticular dsd (3), congenital adrenal hyperplasia (15), 46 XX gonadal dysgenesis (1) Turner Syndrom (2), persistent Mullerian duct syndrome (2), partial androgene insensivity 1, Frazier Syndrom, 1 unclear 5. Patients with congenital adrenal hyperplasia were examined by only by genitoscopy/urethroscopy, most other patients had both procedures for diagnosis, gonadal biopsy or gonadectomy. Results: Within all diagnostic subgroups, notable variations in urogenital anatomy were found. These findings were reported with variable terminology and documented with a variable number of digital images. From this analysis emerged that most anatomic details can be efficiently recorded within few images. A 5 item-checklist to define the anatomy of gonads and sex ducts on both sides and imaging of the retrovesical space is proposed. For cysto-genitoscopy, also a 5-item- list is suggested. It contains the two ureteral orifices, the bladder neck, the are of the confluens/verum montanum and an image of the vagina/cervix or an utricular cyst. Discussion: We propose a structured examination schedule and a reporting system based on digital images during laparoscopy and cysto/genitoscopy. Our check-list items are clearly defined, based on a rationale, and easily applicable. Laparoscopy and cystogenitoscopy provide the most detailed anatomic information. Documenting anatomical findings in detail and high quality may improve future outcome studies. Size. shape and surface anatomy of the gonads are highly variable, an these factors may be relevant for the individual tumour risk. Sex duct anatomy also varied individually. Written reports capture only a limited amount of this variability. The endoscopy items are relevant for genital reconstruction and function. The distance between bladder neck and vaginal confluens is important for vaginal reconstruction. Anomalies of the verum montanum are common and contribute to the influx of urine, vaginal length is relevant for sexual function and treatment planning. We propose our two 5 item check-lists for discussion as standard operating procedures.
20 – 22 May 2011, Lübeck, Germany 27
K-13 OUTCOME STUDIES
# U. Thyen1, M. Jürgensen1, E. Kleinemeier1 1Universität zu Lübeck, UK S-H, Campus Lübeck, Klinik für Kinder- und Jugendmedizin, Lübeck, Germany
The German Network of Disorders of Sex Development (DSD)/Intersexuality conducted a multicenter clinical evaluation study on psychological adaption, psychosexual development, and coping with diagnoses and therapies in individuals with disorders of sex development (DSD). We used a broad generic definition of DSD following the classification of the expert consensus statement ESPE/LWSPE 2006. Given the lack of large datasets in rare conditions such as DSD and often biased results from small scale clinical case series, the study aims to describe the variation in outcomes and to compare age groups in order to generate concrete hypotheses to develop evidence-based guidelines. This presentation focuses on the results of children and adolescents with disorders of sex development (DSD), who often are exposed to specific stressors like surgery, lack of virilization in boys and over-virilization in girls. For children and adolescents with DSD symptoms interfere in particular with developmental tasks, especially entering puberty. Methods: We interviewed children and adolescents between 4 and 16 years between 2005 and 2007 in Germany, Austria and Switzerland. Health-related quality of life (HrQoL), mental health, gender identity and gender role behaviour were measured with generic instruments. We asked for disease-specific aspects with self-constructed parent’s and adolescent’s questionnaires. Further, we collected medical data by the attending physician. Results: 166 children (4-12 years) and 66 adolescents (13-16 years) with DSD and their parents participated, 170 raised as girls, 62 raised as boys. For statistical group comparisons we used four diagnostic subgroups: girls with 46,XX & overvirilization/CAH (N=95), girls with 46,XY & partial androgen effects (N= 55), girls with 46,XY without androgen effects (N= 20), boys with lack of virilization (N= 62). Results from the HrQoL and the mental health did not reveal differences between the four diagnostic groups. Children’s HrQoL was impaired concerning emotional well-being in comparison with norm data. Adolescent’s HrQoL was not impaired in comparison with the norm group. We did not identify high risk for mental health problems. Children without androgen effects demonstrated a tendency for female-typical behaviors, while children with androgen effects showed more male-typical behaviors. Boys with DSD did not differ from control boys. Although girls with androgen effects showed more cross-gender behaviors than girls without androgen effects and boys with androgen effects, we found a high congruence between gender identity and gender of rearing in children and adolescents with DSD. However, adolescent girls with DSD show fewer activities like their female peers. Furthermore adolescents with DSD try to hide physical differences. Discussion: The impairments concerning HrQoL in children demonstrate a quite significant impact of DSD on the emotional well-being. The fact that mental health is not affected in pre-pubertal children can be explained with social support and personal resilience. The result that children with androgen effects more male typical behaviors compared to peers indicates prenatal androgen effects on psychosexual development. Children and adolescents with DSD did not show an increased risk for cross-gender behavior or gender identity confusion in general. Adolescents did not report obvious psychological impairments, however, being different is a source of fear, insecurity and the need to conform. Treatment teams should aim to address the age specific needs of children and adolescents independent of their parents needs for psychosocial support. Literature: Hughes IA, Houk C, Ahmed SF, Lee PA, and LWPES/ESPE Consensus Group. Consensus statement on management of intersex disorders. Arch Dis Child 2006; 91: 554-63. Jürgensen, M., Kleinemeier, E., Lux, A., Steensma, T. D., Cohen-Kettenis, P. T., Hiort, O., et al. (2010). Psychosexual development in children with disorder of sex development (DSD)--results from the German Clinical Evaluation Study. J Pediatr Endocrinol Metab, 23, 565-578. Kleinemeier, E., Jürgensen, M., Lux, A., Widenka, P. M., & Thyen, U (2010). Psychological adjustment and sexual development of adolescents with disorders of sex development. J Adolesc Health, 47(5), 463-471. Lux, A.; Kropf, S.; Kleinemeier, E.; Jürgensen, M.; Thyen, U. (2009). Clinical evaluation study of the German network of disorders of sex development (DSD)/intersexuality: study design, description of the study population, and data quality. BMC Public Health, 9, 110.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
28 3rd International Symposium on Disorders of Sex Development
K-14 THE LOGISTICS AND ETHICS OF DATA BASES
# F. Ahmed1 1University of Glasgow, Glasgow, United Kingdom
Disorders of sex development (DSD) are a rare group of conditions which require further research. Effective research into understanding the aetiology, as well as long-term outcome of these rare conditions, requires multicentre collaboration often across national boundaries. The EU-funded EuroDSD programme (www.eurodsd.eu) is one such collaboration involving clinical centres and clinical and genetic experts across Europe. At the heart of the EuroDSD collaboration is a European DSD registry and a targeted virtual research environment (VRE) that supports the sharing of DSD data. Security, ethics and information governance are cornerstones of this infrastructure. This paper describes the infrastructure that has been developed, the rules and responsibilities of users, the inherent challenges in security, availability and dependability that must be overcome for the enterprise to succeed and provides a sample of the data that are stored in the registry.
20 – 22 May 2011, Lübeck, Germany 29
K-15 APPLICATION OF AN ESPE DSD E-LEARNING WEBPORTAL
# S. Drop1, L. Kranenburg1, K. Grijpink1, C. de Vugt1, M. Belder1, R. Nanninga1 1Erasmus University of Rotterdam, Sophia Children Hospital, Rotterdam, Netherlands
The aim of the webportal is to provide entrance to an interactive learning environment for an up-to-date program on DSD including normal development, patho-physiological mechanisms, diagnostic and therapeutic interventions, psychological counselling and outcome. Target groups are medical students, residents, fellows, specialists, consultants and teachers around the world. The portal is developed in the English language with two levels of learning: a. basal (medical student): the focus is the understanding of the normal development and its patho-physiology with clinical and social implications. b. advanced (post-doc, etc): the fellow is additionally invited to analyze and diagnose disorders, to solve problems, to appraise scientific evidence and to communicate with professionals, patients and parents. Following an intensive phase of construction the webportal has gone live: www.espe-elearning.org. Presently, the main emphasis is on expanding data entry, i.e text for chapters and case descriptions. Authors have been invited to contribute content. An editorial working group has been formed and a technical staff is involved in the construction, lay out and function of the web portal. Results will be presented of a pilot study evaluating content and user experience of a set of chapters and cases in the current portal in order to obtain recommendations for further improvement. Furthermore, it is the intention to develop functionalities for scoring of questions and to develop formative assessment of competencies of students and portal users making use of a forum functionality. The educational background and objectives have been summarised recently 1. (1) Grijpink-van de Biggelaar K, Drop SL, Schuwirth L. Development of an ESPE e-learning portal: educational considerations. Horm Res Paediatr 2010; 73:223-30
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
30 3rd International Symposium on Disorders of Sex Development
K-16 DSD RESEARCH - BEYOND DESCRIPTIVE STUDIES
# O. Hiort1 1Universität zu Lübeck, UK S-H, Campus Lübeck, Klinik für Kinder- und Jugendmedizin, Lübeck, Germany
The last decade has seen major advances in the professional and public views on disorders of sex development (DSD). An important focus was put on the nomenclature and classification, as well as the overall management of DSD. The consensus statement of the major Western paediatric endocrine societies in 2005 has been widely accepted and incorporated into text books and scientific presentations1. Moreover, a widespread view has been published that both the diagnosis and the management of DSD needs a very high degree of professionalism and the establishment of Multidisciplinary Teams that will give the patient and the family a personalized approach to the individual diagnosis and prognosis. However, a survey in Germany demonstrated that the definitive diagnosis of the underlying condition is still lacking in many cases, and expert institutions are not validated 2. DSD have been seen as a complex conglomerate of rare to very rare conditions affecting the urogenital tract and often the endocrine reproductive system. In this context, research on DSD was promoted by the European Union for the benefit of its members. The approach undertaken in the EuroDSD project aimed to establish a European consortium on the basis of genuine and equal partnership between leading clinical centres, academic groups, cell biologists, and geneticists to provide further knowledge about the physiology and pathophysiology of sex development. The results are presented at this meeting. In the future, EU research in rare diseases will centre on observational trials utilizing an existing pool of clinical data and biomaterials to develop new technologies for diagnosis and treatment and furthermore to put an emphasis on drug development. A major effort is envisaged, both in using -omics technologies as well as other modern methods to achieve better diagnosis and treatment. There will also be a major push for pre- clinical or clinical development of orphan drugs, as well as observational trials, aiming to improve clinical practices. DSD research in this context needs to strengthen observational and clinical trials to develop evidence-based guide lines for diagnostic steps as well as for measurement of outcome data to achieve a personalized yet standardized care for our patients. This can best be achieved through a networking platform supporting the collection of standardised and validated data and the exchange of information providing evidence for best clinical management. This will essentially lead to the definition of requirements for the implementation of specialized reference centres for DSD. 1Paterski V, Prentice P, Hughes IA (2010). Consequences of the Chicago consensus on disorders of sex development (DSD): current practices in Europe. Arch Dis Child 95: 618-623 2Thyen U, Lanz K, Holterhus P-M, Hiort O (2006): Epidemiology and initial management of ambiguous genitalia at birth in Germany. Horm Res 66:195-203
20 – 22 May 2011, Lübeck, Germany 31
Abstracts Oral Presentations
O-01 DISCLOSURE IN DISORDERS OF SEX DEVELOPMENT: PRELIMINARY ANALYSIS USING PARENT REPORT
# V. Pasterski1, K. Mastroyanopoulou1, I. Hughes1 1Addenbrookes Hospital, University of Cambridge, Paediatrics, Cambridge, United Kingdom
Objective: The objectives of the current study were two-fold. Firstly, we aimed to chart the disclosure process by interviewing parents about their experiences of learning about their child’s DSD. Secondly, we assessed psychological and emotional well-being in parents and their children so that we might relate outcome to disclosure practices. Method: Semi-structure interviews were conducted either in person or over the phone. Participants also completed standardised questionnaires assessing psychological functioning, emotional well-being, and self- esteem in themselves as well as psychosocial/psychosexual functioning in their children diagnosed with DSD. Results: Trends in content analysis of the 45 interviews assessing parents’ experiences suggested that key issues for them included: considerations of privacy in such delicate matters as sex of the child in cases of ambiguity; misunderstandings of medical jargon; lack of digestible information; and lack of social support present in the first moments of disclosure. Cases where gender assignment was a concern presented the most difficulty in that parents left in a state of uncertainty had few or no coping strategies. Conclusions: Disclosure in the case of DSD is a matter which should be handled in particularly sensitive manner. The experience of learning of DSD for parents can have lasting effects on their own and their child’s mental health as well having an affect on the bond between parent and child. Recommendations for case management include: early disclosure in a private setting with both parents present, where possible; Health care professionals (HCPs) should avoid the use of confusing medical jargon or pejorative terminology such as “hermaphrodite”; and assessment of individual levels of understanding should be taken into consideration where technical information is given. In this case specialised information leaflets have been identified as particularly helpful. Note: Outcome data will also be presented as it relates to various disclosure practices. These data are currently being analysed.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
32 3rd International Symposium on Disorders of Sex Development
O-02 FROM ETHICAL GUIDELINES TO CLINICAL PRACTICE - ASSESSING THE IMPACT OF VALUE CHANGES IN CLINICAL PRACTICE
# J. Streuli1,2, V. Schönbucher3, S. Morichetti2 1University of Zurich, Institute of Biomedical Ethics, Zürich, Switzerland 2Swiss Network of Intersex and Variations of Sex Development, Zurich, Switzerland 3University Children’s Hospital, Zurich, Switzerland
Objectives: Ethical guidelines and recommendations for clinical management of intersex are showing clear evidence for a significant change in underlying values. However, there has been only few attention on the related change in clinical practice. Likewise there is little data showing how clinical procedures could be supported to integrate current recommendations, e.g. multidisciplinary approaches, upholding children’s rights to participate in decision-making , protecting the parent-child relationship. Methods: To fulfill these demands, an interdisciplinary network of health care professionals for ethics and professionalism in clinical practice have recently been formed in Switzerland. It comprises a study group evaluating clinical practice and a center of clinical competence. Thereby, it focuses on close collaboration with self-help groups and specialists from endocrinology, surgery, genetics and other disciplines. The study group developed a register, which collects a selection of epidemiological and psychosocial data. In addition a nation-wide qualitative follow-up study regarding certain vulnerable patient groups is offered. Results: Preliminary results of our group indicate that a change of underlying values has happened not only in ethical guidelines but also in clinician’s attitudes. However, subsequent change in clinical practice and its impact on psychosocial parameters are still based on anecdotal evidence. Our first experiences with the register combined with a qualitative follow-up showed an increase of the awareness of clinical problems concerning the value changes with high willingness to solve them. Conclusion: The presented measures should improve the correspondence between ethical guidelines and clinical practice. Combined with a follow-up study the impact on children and families can be used as an early-warning system for positive and negative influences due to changes in clinical practice. For further research we suggest an integration of the presented assessment tools into a workpackage of EuroDSD.
20 – 22 May 2011, Lübeck, Germany 33
O-03 MANAGEMENT OF VAGINAL HYPOPLASIA IN DISORDERS OF SEX DEVELOPMENT: A RETROSPECTIVE STUDY COMPARING SURGICAL AND NON-SURGICAL OPTIONS
# N. Callens1,2, A.B. Dessens1, G. De Cuypere3, K.P. Wolffenbuttel4, Y.G. Van der Zwan1, S.L.S. Drop1, P. Hoebeke5, E. Van Hoecke6, P. De Sutter7, C. Beerendonk8, D. Study Group on DSD1,8,9,10,11, M. Cools2 1Erasmus Medical Centre- Sophia, Pediatric Endocrinology, Rotterdam, Netherlands 2Ghent University and University Hospital Ghent, Pediatric Endocrinology, Ghent, Belgium 3Ghent University and University Hospital Ghent, Sexology and Gender problems, Ghent, Belgium 4Erasmus Medical Centre - Sophia, Urology, Rotterdam, Netherlands 5Ghent University and University Hospital Ghent, Urology, Ghent, Belgium 6Ghent University and University Hospital Ghent, Child Psychology, Ghent, Belgium 7Ghent University and University Hospital Ghent, Gynecology, Ghent, Belgium 8University Medical Centre St Radboud, Gynecology, Nijmegen, Netherlands 9VUMC, Amsterdam, Netherlands 10UMCU, Utrecht, Netherlands 11UMCG, Groningen, Netherlands
Objectives: Females with DSD requiring vaginal reconstruction, because of vaginal hypo- or aplasia, are complex and varied in their presentation. Although health care providers agree on the importance of the construction of a good functional vaginal substitute that is durable and provides long-term sexual satisfaction, there is substantial disagreement as to which technique best accomplishes these goals. Operative vaginoplasty and vaginal dilation are both valid vaginal substitution treatments. The American College of Obstetricians and Gynecologists recommends vaginal dilation as a first choice treatment, because of its less invasive character and high success rate. To our knowledge, however, no studies compare the psychosexual functioning of women after vaginoplasty to that after vaginal dilation in a standardized manner and followed in the same clinical setting. Methods: Psychosexual functioning of 32 women at least two years after vaginoplasty (n=15) or vaginal dilation (n=17) was investigated. Questionnaire (Female Sexual Function Index, Female Sexual Distress Scale- Revised) scores of study participants were compared against the scores of the test validation population. Additionally, a semi-structured interview and a gynecological examination were performed. Results: A shortened vaginal length of less than 6.6 cm, more than 2 standard deviations below the published mean value, was found in 26% (7 out of 26) of the women, after either treatment. Irrespective of the treatment, 44 % of the patients had (a) sexual dysfunction(s) and experienced sexual distress. However, after vaginoplasty, patients reported significantly more problems with lubrication (p=0.025, p<0.05) than after vaginal dilation. Conclusions: Vaginal dilation therapy is a non-invasive, valuable alternative to vaginoplasty, possibly with less side effects and post treatment problems, and leads to an equal or superior outcome with regard to long- term psychosexual functioning. However, if this method wants to reach a high success rate, gradual self-dilation has to be supported by an expert multidisciplinary team that integrates endocrinology, gynecology, sexology and clinical psychology expertise. Equal priority should be given to quality of life outcomes, including adequate psychological and sexual treatments, as is currently given to the traditional clinical concerns, such as anatomical outcome. Further longitudinal studies, incorporating prospective evaluation of vaginal reconstruction interventions, are highly needed.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
34 3rd International Symposium on Disorders of Sex Development
O-04 LONG TERM OUTCOMES IN XY DSD RAISED AS FEMALES TREATED IN A SINGLE INSTITUTION
# K. Gueniche1,2, E. Thibaud1, M. Jacquot2, C. Duflos1,3, P. Touraine3, S. Lortat-Jacob4, M. Polak1 1Hôpital Necker Enfants Malades, Pediatric Endocrinology and Gynecologie, Centre des maladies rares gynécologiques et endocriniennes de la croissance, Paris, France 2Université Paris Descartes, Institut de Psychologie, LPCP EA 4056, Boulogne, France 3Hôpital Pitié-Salpétrière, Endocrinology and Reproductive Medicine – Centre des maladies rares gynécologiques et endocriniennes, Paris, France 4Hôpital Necker Enfants Malades, Pediatric Visceral Surgery – Centre des maladies rares gynécologiques, Paris, France
Few studies have addressed the question of the long term outcomes including well-being, emotions and feelings in DSD patients. Our study aimed to assess cognitive, psychic functioning, affective life, sexual behavior and quality of life in a cohort of XY DSD raised as girls, treated at a single institution. Subjects and methods: 18 patients aged 18 to 41 with a 46 XY DSD, whose gender was assigned as female, whose treatment included genital surgery and raised as girls, were recruited and studied. The protocol included a gynecological examination, a semi-directive interview and a WAIS III-R intelligence test, two projective tests (Rorschach, TAT), the WHO quality of life 26 and a questionnaire of sexual behavior to be compared to normative data from the same french population. Results: The global cognitive abilities of the cohort are in the lower half of the normal (average IQ = 90). The assessment given by the women to their quality of life as concerns social relationships (56.4%), was significantly lower than that of the reference population (RP) (p = <0.05). Six of the fifteen women had never had sex; a proportion much higher than average among women of their age (40% versus 5.2%) (p= <0.05). Homosexual attraction (38.3%) and genital sexuality (30%) are much more frequent than in the RP (respectively 6.5% and 1.9%) (p = <0.05). The question of sexuality was addressed as a medical issue (27.7%, against 5.2% for the RP) (p= <0.05). Despite medically satisfactory external genitalia, interviews underline 6 recurrent themes in the narratives: 1) a great deal of confusion between the perceived anomalous genitalia and sterility 2) a vagueness in the ability to name the disorder, to recall the information that was indeed given 3) fears for the sexual and love life 4) high degree of difficulty with identification to a feminine maternal image 5) a feeling of secrecy, guilt and shame 6) a social and family, mostly on the maternal side, isolation. Besides, for 15, psychic functioning is organized on a borderline-narcissistic mode; for 3 diagnosed during adolescence, it is fluid with appropriate defense mechanisms on a normal-neurotic mode. Conclusions: The analysis revealed these women's dissatisfaction with their emotional and social life. They are in great psychic pain. Even if they do not always express it verbally, they clearly suffer from depression, along with more or less severe mood disorders (despair, anxiety, anguish and suicidal thoughts). Their quality of life is greatly altered, their sexual and love life inhibited. They are not subjectively satisfied with the surgery (despite satisfactory gynecological examinations) and with the knowledge of their disorder. The mental process that enables girl to recognize themselves (physically and psychically) as women is impaired. Their doubts concerning their identity as women were stronger when the relationship with their mother was either non-existent or impossible. Supported by the WYETH foundation
20 – 22 May 2011, Lübeck, Germany 35
O-05 SATISFACTION WITH TREATMENT, GENITAL SURGERY AND SEXUAL LIFE IN ADULTS WITH XY DISORDERS OF SEX DEVELOPMENT - RESULTS FROM THE GERMAN CLINICAL EVALUATION STUDY
# B. Köhler1, E. Kleinemeier2, A. Lux3, A. Grüters1, O. Hiort4, U. Thyen2 1Charité, CVK, Pediatric Endocrinology, Berlin, Germany 2University of Lübeck, Pediatric and Adolescent Medicine , Lübeck, Germany 3Otto von Guericke University, Institute of Medical Statisitics, Magdeburg, Germany 4University of Lübeck, Pediatric Endocrinology, Lübeck, Germany
Background: Prenatal deficit of androgens or androgen action results in atypical genitalia in individuals with XY disorders of sex development (XY,DSD). XY,DSD include mainly disorders of gonadal development, testosterone synthesis and action. Previously most XY,DSD individuals were assigned to the female sex. Genital surgery allowing heterosexual intercourse, gonadectomy and hormone therapy for feminisation were often performed. However, outcome studies are scarce. Objective: Evaluation of satisfaction with treatment, genital surgery and sexual quality of life (SexQoL) in adults with XY,DSD. Design and methods: Fiftyseven individuals with XY,DSD from the German multicenter clinical evaluation study, which were divided in three subgroups reflecting the absence/presence of partial androgen effect, were evaluated by a questionnaire. Results: Homo- or bisexual orientation was more common in 46,XY females with partial androgen effects (24.9%). Many participants were dissatisfied with treatment and genital surgery (35.1%). Dissatisfaction with overall sex-life was significantly increased after clitorisreduction (56.2% vs. 27.3%). Sexual anxieties were considerably higher in XY,DSD individuals (55.4%). Problems with arousal (44.4%), dyspareunia (56.4%) and vaginism (20.5%) were about twice as common in XY females compared to controls (28.5%, 30.2% and 9.5% respectively). Problems with arousal (60%) and vaginism (33.3%) were increased after multiple surgery. The majority of individuals with complete androgen insensitivity (CAIS) reported decreased sexual desire (81.8%) and problems with arousal (63.6%). Conclusions: Genital surgery should be minimized and care has to be improved in XY,DSD. Feminization surgery should be performed mainly in adolescence or adulthood with the patients' consent. Individuals with DSD and their families should be informed with openness and sensitivity about the condition. Sexuality should be addressed and worked on in puberty together with a psychologist/sex therapist.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
36 3rd International Symposium on Disorders of Sex Development
O-06 VOICE CHARACTERISTICS IN ADULT CAH-WOMEN
# A. Nordenskjold1, K. Hagenfeldt1, U. Nygren2, M. Södersten2, H. Falhammar3, M. Thorén3 1Karolinska Institutet, Women and Children Health, Stockholm, Sweden 2Karolinska Institutet, Clinical Sciences, Stockholm, Sweden 3Karolinska Institutet, Molecular Medicine and Surgery, Stockholm, Sweden
Objectives: Congenital adrenal hyperplasia (CAH) is an autosomal recessive inherited disorder resulting in lack of cortisol and often aldosterone and overproduction of androgens. Raised levels of androgens can lead to increased mass in the laryngeal tissues, which may lower the fundamental frequency (F0) of the voice. Methods: A follow-up study of voice characteristics in 38 women with CAH and 24 age-matched controls between 18 and 63 years of age using subjective self-ratings of voice symptoms and acoustic and perceptual analyses. The results were correlated to mutation and treatment. The subjects rated degree of hoarseness, dark voice, voice problems and vocal fatigue on visual analogue scales. Audio recordings were made of a standard reading passage in a sound treated booth. Acoustic analyses were made of the subjects’ mean, minimum and maximum F0 in the habitual speech range. Three voice clinicians made perceptual evaluations of dark voice in terms of timbre. Results: Women with CAH rated higher values than controls with regard to the statement ‘my voice is a problem in my daily life’. They spoke with significantly lower mean, lower minimum and lower maximum F0, as well as darker voice quality as compared with the controls. Affected voice symptoms were associated with a late diagnosis or problems with medication. Conclusions: The voices in women with CAH can be virilized because of late diagnosis or suboptimal treatment with glucocorticoids. These voice problems may need referral for voice assessment and should be considered when treating CAH.
20 – 22 May 2011, Lübeck, Germany 37
O-07 BONE MINERAL DENSITY IN WOMEN WITH COMPLETE ANDROGEN INSENSITIVITY SYNDROME
# E. Dati1, # S. Bertelloni1, S. Ghione1, A. Petracchi1, G.I. Baroncelli1 1University of Pisa, Dept. of Obstetrics, Gynecology and Pediatrics, Pisa, Italy
Background: Bone health represents a clinical concern in women with complete androgen insensitivity syndrome (AIS). Reduced bone mineral density (BMD) has been reported, but methodological issues may give some inferences on data interpretation, for examples methods of BMD assessment or AIS diagnosis, because some reported patients have not molecular assessment. Another problem can be the use of female (the phenotypic sex) or male (the genetic/gonadal sex) references values to evaluate BMD status. Objective: To evaluate BMD in women with molecular diagnosis of complete AIS. Patients and methods: Twenty-eight women (age 15-47 years) with complete AIS (diagnosis based on phenotype, 46,XY karyotype, hormonal pattern and androgen receptor gene mutation); 7 patients were not gonadectomised at BMD assessment, while the others were. The women with removed gonads were on hormone replacement therapy (HRT). BMD [lumbar spine (L2-L4), femoral neck (FN), total body (TB)] were assessed by DXA. The values are expressed as T-score in comparison with normative values for females. Results: Mean BMD values (SDS) in the whole group are: L2-L4 -1.57 ± 1.17 (Mean ± SD) (p < 0.001 vs 0); FN - 0.62 ± 1.29 (p < 0.002 vs 0); TB 0.68 ± 1.37 (p = NS vs 0). BMD (SDS) was significantly reduced in women with removed gonads in comparison with women with intact gonads [FN, SDS: gonadectomized -0.98±1.04 vs not gonadectomized 0.63±1.38 (p = 0.003); L2-L4: gonadectomized -1.86±0.90 vs not goadectomized -0.55±1.51 (p = 0.0095); TB: gonadectomized -0.75±1.07vs not gonadectomized 1.35±1.65 (p = 0.0006)]. Good compliance with HRT was associated with better BMD values. Conclusion: Gonad removal is associated with poorer bone health in women with proven complete AIS. HRT may improve BMD status in complete AIS women who had gonads removed. Testicular protein hormones (f.e. INSL3) may have a role on bone health in women with complete AIS. Larger series of homogeneous patients will be investigated to obtain conclusive data and clear indications for the optimal management of people with complete AIS.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
38 3rd International Symposium on Disorders of Sex Development
O-08 DEVELOPMENT OF A NOVEL UPLC-MS/MS ASSAY FOR EIGHT C21 STEROIDS, REFERENCE DATA FOR CHILDREN, AND ITS APPLICATION ON CONGENITAL ADRENAL HYPERPLASIA
# A. Kulle1, P.-M. Holterhus1, F. Riepe1 1University Hospital Kiel, Endocrinology and Diabetes, Kiel, Germany
Introduction: Congenital Adrenal Hyperplasia (CAH) is composed of several inherited defects of steroid biosynthesis that impair cortisol and aldosterone biosynthesis. The incidence of CAH worldwide is approximately one in 15,000 live births. Due to ACTH-driven elevation of adrenal androgens, CAH is also a common cause for 46,XX DSD. The most frequent subtype is 21-hydroxylase deficiency (21-OHD) followed by 11ß-hydroxylase deficiency (11-OHD), which is responsible for 95 % and 5 % of all cases, respectively [1]. 17- hydroxyprogesterone is elevated in 21-OHD and stands as key marker steroid for diagnosis of CAH. But to discriminate exactly cases of 11-OHD from case of 21-OHD at least the following steroids are needed: 17- hydroxyprogesterone, 21-deoxycortisol and 11-deoxycortisol [2]. Method: A sensitive and reliable method for the simultaneous detection and quantification of prog-esterone (P), 17-hydroxyprogesterone (17OHP), deoxycorticosterone (DOC), 11-deoxycortiol (11S), 21-deoxycortisol (21S), corticosterone (B), cortisol (F) and cortisone (E) by UPLC-MS/MS was developed. Samples of 359 children were measured and grouped in to ten age groups. 16 cases with different forms of congenital adrenal hyperplasia (CAH) were analyzed. Results: The separation of the seven C21 adrenal steroids was achieved in 5.3 min. The assay was linear up to 200 nmol/L (r2>0.992) for each steroid. The lower limit of detection was 0.03 to 0.2 nmol/L and the lower limit of quantification was 0.1 to 0.8 nmol/L. All patients with suspected CAH could be diagnosed because of pathologic concentrations of the required adrenal steroid hormones. Conclusion: We developed a sensitive and reliable UPLC-MS/MS method for the simultaneously determination of adrenal hormones. With the required samples volume of 0.1 mL this method is highly suitable for the peadiatric clinical routine and for cases of research as well. The data set allows the diagnosis of different adrenal diseases. 1. Speiser, P.W., et al., Congenital adrenal hyperplasia due to steroid 21-hydroxylase deficiency: an Endocrine Society clinical practice guideline. J Clin Endocrinol Metab. 95(9): p. 4133-60. 2. Tonetto-Fernandes, V., et al., Serum 21-Deoxycortisol, 17-Hydroxyprogesterone, and 11-deoxycortisol in classic congenital adrenal hyperplasia: clinical and hormonal correlations and identification of patients with 11beta-hydroxylase deficiency among a large group with alleged 21-hydroxylase deficiency. J Clin Endocrinol Metab, 2006. 91(6): p. 2179-84.
20 – 22 May 2011, Lübeck, Germany 39
O-09 RECURRENT ABERRATIONS IDENTIFIED BY ARRAY-CGH IN PATIENTS WITH MAYER-ROKITANSKY- KÜSTER-HAUSER SYNDROME
# S. Ledig1, C. Schippert2, R. Strick3, M. Beckmann3, P. Oppelt3, M. Cools4, K. Wolffenbuttel5, S. Drop5, P. Wieacker1 1University Münster, Institute of Human Genetics, Münster, Germany 2Medizinische Hochschule Hannover, Klinik für Frauenheilkunde und Geburtshilfe, Hannover, Germany 3Universitätsklinikum, Frauenklinik, Erlangen, Germany 4University Hospital Ghent and Ghent University, Department of Pediatric Endocrinology, Ghent, Belgium 5Erasmus MC, Sophia Children’s Hospital, Rotterdam, Netherlands
Objectives: MRKH (Mayer-Rokitansky-Küster-Hauser) is a congenital anomaly of the female genital tract characterized by agenesis of the upper vagina and rudimentary or absent uterus. Typically, the external genitalia as well as the ovaries are normal and the karyotype is 46,XX. A variety of different defects, especially renal anomalies, can be associated. To date, in most of the cases the cause remains unclear. Methods: We performed array-CGH (comparative genomic hybridization) analysis in a group of 53 MRKH patients (2 patients from the Euro-DSD-database) by using high-resolution Agilent oligonucleotide arrays. Results: We detected 30 losses and gains being potentially causative. Of these regions we could delineate three definitively relevant regions (1q21.1, 17q12, and 22q11.21), which have been repeatedly observed in our and other MRKH patients. Furthermore, sequential analysis of LHX1 and HNF1B, being located in candidate region 17q12, revealed in one MRKH patient a missense mutation in LHX1 gene, making LHX1 a strong candidate gene for MRKH. However, HNF1B is another candidate gene in the 17q12 region because mutations of this gene have been observed in patients with renal cysts, diabetes, and Müllerian aplasia. Conclusion: Array-CGH is a very useful tool for delineating genetic causes of MRKH, revealing in this study three relevant regions and the LHX1 gene as important causative candidate.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
40 3rd International Symposium on Disorders of Sex Development
O-10 EXOME SEQUENCING REVEALS NOVEL GENETIC FACTORS INVOLVED IN DSD
# A. Bashamboo1, R. Brauner2, D. Lourenço1, J. Bignon-Topalovic1, V. Tillman3, K. McElreavey1 1Institut Pasteur, Paris, France 2Université Paris Descartes, Faculté de médecine and AP-HP, Hôpital Bicêtre, Le Kremlin Bicêtre, France 3Tartu University Hosptial, Pediatrics, Tartu, Estonia
Objectives: It has been estimated that a molecular diagnosis is made in only around 20% of DSD, except in cases where the biochemical profile indicates a specific steroidogenic block. These conditions are difficult to analyse using classic genetic approaches since familial cases are rare and sex-determination, as a developmental process, is not evolutionary well-conserved. Here, we have used the exome sequencing approach on 10 cases of either familial or sporadic 46,XY DSD and 46,XX DSD in order to identify new genetic causes of these pathologies. In 7 of these cases associated somatic anomalies were also present, which included autism, learning difficulties and congenital heart disease. Methods: Exons were enriched, from genomic DNA, using the Agilent SureSelect Exome Kit (50 Mb) and sequencing was performed using SOliD sequencing protocols (Life Sciences). Accuracy enhancement was carried out using the SAET program. Reads were mapped using rmapper (ABI) to the reference Human genome (GRCh37/hg19) with inclusion of -/+ 500 nt at 5’ or 3’ from the set of sequences corresponding to the total of 171,744 exons designed from the CCDS dataset. Alignments of the mapped reads were subjected to two different computational pipelines: smallIndel and diBayes. This procedure was repeated using data from the 1000 genomes project. An in-house script classified variants as « known » or « novel ». These variants were analysed using the EnsEMBL SNP Effect Predictor, and with the FANS site (http://fans.ngc.sinica.edu.tw). All potentially pathogenic variants were confirmed by Sanger sequencing in the affected individuals and inheritance investigated in other family members. Functional studies were also performed for some variants. Results: We performed exome sequencing with an average coverage of x200. With this average coverage we accurately analysed 96% of the Agilent High Quality Exome with a coverage of >x20. We identified several pathogenic mutations in the 10 cases of DSD analysed. For example, an individual with 46,XY DSD and learning difficulties, who was born to consanguinous parents carried two missense mutations in the FOG2 gene, one homozygous the other de novo heterozygous. Murine Fog2 mutations are associated with a complex phenotype that includes anomalies of testis development. Functional studies indicated that the FOG2 protein carrying both the mutants exhibited aberrant biological activity. The analyses of other DSD patients revealed pathogenic mutations including a missense mutation in a novel MAP kinase gene and a nonsense mutation in the TITIN (TTN) gene that is associated with congenital heart disease. Conclusions: These data highlight the power of an exome sequencing approach to understand the genetic basis of complex disorders that are difficult to study by classical approaches.
20 – 22 May 2011, Lübeck, Germany 41
O-11 FUNCTIONAL ANALYSES OF NOVEL ANDROGEN RECEPTOR VARIANTS PREDICT PHENOTYPES IN PATIENTS WITH ANDROGEN INSENSITIVITY SYNDROME
# A. Brinkmann1, P. Elfferich2, M. Van Royen3, D. Van de Wijngaart3, S. Lusher4, R. Bosch4, J. Trapman3, T. Bunch5, S. Drop6, M. Cools7, H. Bruggenwirth2 1ErasmusMC, Reproduction and Development, Rotterdam, Netherlands 2ErasmusMC, Clinical Genetics, Rotterdam, Netherlands 3ErasmusMC, Pathology, Rotterdam, Netherlands 4MerckResearch Laboratories, Molecular Design & Informatics, Oss, Netherlands 5University Cambridge, Paediatrics, Cambridge, United Kingdom 6ErasmusMC, Pediatric Endocrinology, Rotterdam, Netherlands 7University Ghent, Pediatric Endocrinology, Ghent, Belgium
Objectives: Androgen Receptor (AR) gene mutations in the X-linked androgen insensitivity syndrome (AIS) are associated with a variety of clinical phenotypes. The aim of the present study was to determine the molecular properties and potential pathogenic nature of 8 novel and 3 known AR variants of unknown clinical significance. These extensive studies may provide treatment strategies and optimal counseling. Methods: Based on AR sequencing (exons 1 to 8 and flanking intronic sequences), the found mutations were introduced in AR cDNA expression vectors by site directed mutagenesis. Next transcriptional activation assays on an androgen-regulated LUC-reporter, western blot analysis, as well as interactions with several coactivator motif (FxxLF-like) containing peptides were performed. Using green fluorescent protein (GFP) tagged mutants nuclear dynamics (Fluorescence recovery after photobleaching (FRAP)) and intracellular distribution were studied. In addition Scatchard plot analysis was done for equilibrium dissociation constant (Kd) determination. Furthermore, molecular modelling was performed in PyMOL for predicting structural consequences of the mutations. Results: The following 8 novel mutations were found: R608M, L700F, S740Y, R774L and I914T in complete AIS (CAIS) patients and C175G and F813C in partial AIS (PAIS) patients. In one case (K912X) the exact phenotype description was not available, but the patient had a female gender identity. In addition D690del and L722F (both CAIS) and A765S (PAIS) were included in the functional studies. Data obtained in the transcriptional activation assays correlated well with the data from the FxxLF-like peptide binding assays, with exception for the R608M, L722F and R774L mutations, which displayed a relatively higher peptide binding activity. In the FRAP experiments all transcriptional inactive mutants lacked DNA binding. For some mutants (C175G, D690del, L722F, A765S and I914T) intermediate or wild-type values were found in both assays (transcription activation + FRAP). All novel mutations in the ligand binding domain (LBD) with a low to intermediate transcriptional activity displayed an aberrant Kd value, also reflected in the decreased nuclear translocation of GFP-tagged mutants at lower hormone concentrations. Molecular modelling programs applied to mutations in the LBD predicted for 6 mutations (S740Y, L700F, R774L, L722F, I914T, F813C) distorted structural consequences of the LBD. Remarkably some mutants (L722F, I914T and D690del) displayed significant functional activities in 3 different assays, while the clinical phenotype was CAIS. Conclusions: The molecular and biological phenotypes of 8 novel and 3 known AR mutations in AIS patients based on 6 different functional assays can predict possible activities of the mutant receptors in vivo and match to a large extent (in approx. 75% of the cases) the clinical phenotypes.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
42 3rd International Symposium on Disorders of Sex Development
Abstracts Poster Presentations
P-01 SEVEN NOVEL NR5A1 GENE MUTATIONS IN A COHORT OF 16 ITALIAN PATIENTS WITH 46,XY DISORDERS OF SEX DEVELOPMENT (DSD) WITHOUT ADRENAL INSUFFICIENCY
# L. Baldazzi1, A. Balsamo1, A. Nicoletti1, S. Menabò1, D. Rinaldini1, G. Cangemi1, P. Pirazzoli1, A. Cicognani1 1S.Orsola-Malpighi University Hospital Pediatric Unit, Gynecology, Obstetric and Pediatric Sciences, Bologna, Italy
Steroidogenic factor 1 (SF1, NR5A1) is a nuclear receptor that regulates multiple genes involved in adrenal and gonadal development. Recent papers reported that heterozygous mutations in SF1 are a relatively frequent cause of 46,XY DSD in humans, in particular among patients with severe underandrogenization and normal adrenal function and with hormonal profiling in the neonatal period that may lead to a presumptive diagnosis of partial androgen insensitivity syndrome (PAIS). We analyzed the NR5A1 gene by automated sequencing in 16 of the patients, among a cohort of 24 Italian patients with 46,XY DSD (mostly referred for a suspected PAIS or 5α reductase deficiency) that did not show any AR/SRD5A2 gene mutation. Heterozygous SF1 mutations were found in 7 cases. Of these, two present a mutation predicted to disrupt RNA stability: the nonsense mutation (p.E51X) in the DBD and the frameshift mutation (c.1016 _1025delGGCGGGCT) starting from helix 6 in the LBD; two are missense mutations that affect a perfectly conserved residue in the DNA binding domain (p.T29K, p.Y47C) and are predicted to compromise this function of the protein; three are missense mutations that map in the LBD and affect perfectly conserved residues (p. R313H in helix 5, p.A351E in helix 7 and p.Q460R at the last but one residue). As far as we know, all the identified mutations are novel. All patients present with severe underandrogenization at birth: female external genitalia with enlarged clitoris, inguinal retained testis and, in a few cases, a very mild gonadal dysgenesis. With the exception of the 2 most recently observed, all were submitted to gonadectomy and female correction of external genitalia in the first years of life. To date, adrenal insufficiency has not occurred in any of the patients. DNA analysis of the parents and clinical evaluation of the identified carriers are running. The degree of impairment of transcriptional activation of SF1-responsive target genes caused by the novel missense mutations will be tested by functional studies, but the degree of conservation of the affected residues, as well as the type of substitution (not conservative in most of the cases), strongly suggest an implication in the phenotypes observed. This study has enlarged the spectrum of NR5A1 gene mutation and clearly confirmed the high frequency of SF1 heterozygous mutations among 46,XY DSD patients without AR/SRD5A2 gene mutations, thus underlining the role of SF1 haploinsufficiency in humans. It also confirms the importance of performing NR5A1 gene analysis in this group of patients for the genetic counselling and in order to take into consideration alternative strategies for the treatment and follow up for both the patients and the 46,XX carriers.
20 – 22 May 2011, Lübeck, Germany 43
P-02 ARRAY-CGH TO IDENTIFY NOVEL GENES INVOLVED IN 46,XY AND 46,XX GONADAL DYSGENESIS: HOW MANY ARE STILL TO BE FOUND?
# A. Norling1, A. Hirschberg Linden2, E. Iwarsson1, A. Wedell1,3, M. Barbaro1,3 1Karolinska Institutet, Molecular Medicine and Surgery, Stockholm, Sweden 2Karolinska Institutet, Department of Woman and Child Health, Stockholm, Sweden 3Karolinska University Hospital, Center for Inherited Metabolic Diseases, Stockholm, Sweden
Objective: Gonad development has been shown to be sensitive to gene dosage. Thus losses or gains of functional gene copies involved in normal gonadal development can lead to gonadal dysgenesis (GD). Currently only approximately 20% of cases of GD have a genetic diagnosis. We applied array comparative genomic hybridization (array-CGH) to identify submicroscopic deletions or duplications in a cohort of non- syndromic patients with GD and a 46,XY or 46,XX karyotype. Genes within these regions would represent candidate genes for GD. Methods: A customized high-resolution 1M arrayCGH platform has been developed at OGT. The platform has a whole genome coverage leading to a 2.1 kb theoretical average resolution, and has been enriched in probes targeting 77 known genes involved in GD or other disorders of sex development. Results: Experimental conditions have been optimized for a 1M platform with some modification from the manufactured protocol. Different DNA sources have also been compared for two cases. A total of 20 patients have been successfully analysed, 8 patients with 46,XY GD, 9 with 46,XX GD and three with premature ovarian failure in younger teens. Several copy number changes have been identified in each patient. The size ranges from 300 kb to 10 Kb. After exclusion of reported copy number variants (CNVs) present in several patients, several regions of interest still remain that represent novel changes or are only partially overlapping with known CNVs, thus representing potential genetic explanations for the dysgenetic gonadal phenotype. Conclusions: Array-CGH is a valuable tool to complement genetic investigations in patients with XY as well XX GD. DNA source and array platform can affect the final results. CNVs need to be carefully evaluated. Genes within the novel candidate regions need to be further characterized to determine their function in normal and abnormal gonadal development.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
44 3rd International Symposium on Disorders of Sex Development
P-03 IDENTICAL TWINS OF OPPOSITE SEX
# H.L. Miles1, J.J. Buck1,2, I. Simonic1,2,3, J. Whittaker1,2,3, R.N. Sandford1,2,3, I.A. Hughes1,2,3 1University of Cambridge, Department of Paediatrics, Cambridge, United Kingdom 2Ipswich Hospital, Department of Peadiatrics, Ipswich, United Kingdom 3Addenbrookes Hospital, Department of Cytogenetics, Cambridge, United Kingdom
Objectives: A female twin presented with an inguinal hernia shortly after birth. Karyotype analysis revealed 46XY(58)/46XX(2).Genetic investigations were instituted to investigate the origin of this unusual karyotype. Methods: History: Twins were born at 28 weeks gestation following ART. The phenotype of twin 1 was male and twin 2 female. Eggs were fertilised using ICSI with fresh embryo transfer of two eggs at the 4 cell stage. Early USS showed a triplet pregnancy. However one fetus was lost at 10 weeks gestational age. Further scans showed a single amniotic sac and placenta suggesting identical twins. Twin 2 with normal female genitalia developed a right inguinal hernia with no gonad palpable. Initial investigations revealed a karyotype of 46XY(58)/ 46XX(2), LH >50IU/l and FSH>50 IU/l. Baseline testosterone was 2.3 nmol/l and there was no increase following a 3 day HCG stimulation test. Pelvic USS demonstrated a normal female genital tract, confirmed by laparoscopy at the time of inguinal hernia repair. Further karyotype analysis revealed varying proportions of 46XX and 46XY cell lines in skin and buccal mucosa (skin 46XY(27)/46XX(33), buccal smear 46XY(19)/46XX(81). The karyotype of twin 1, with a male phenotype, was 46XY(95)/46 XX(5) and buccal smear revealed 46 XY in all cells. A panel of microsatellite markers across the sex chromosomes and all autoosmes was used to identify the origin of te hdifferent cell lines. Gonadal biopsy of twin 2 was also performed to investigate gonadal dysgenesis. Results: There were 4 informative X chromosome markers confirming the presence of three X chromosomes in the 46 XX/46XY female twin ( 2 maternal and one paternal). There were 5 informative autosomal markers confirming the presence of 3 cell lines within venous blood, buccal mucosa and cultured cells of twin 2. Gonadal biopsy revealed ovarian stroma with abundant primordial follicles. Genetic testing of ovarian biopsies showed the Y line present bilaterally in 8 of 100 (left) and 11 of 100 (right) cells examined. Conclusions: The karyotypic findings in this case may be due to chimerism which has been previously described in ART conceptions. Alternative explanations are 47XXY with loss of lines or bone marrow colonisation of a co twin. This case illustrates the importance of karyotyping a female infant with an inguinal hernia.
20 – 22 May 2011, Lübeck, Germany 45
P-04 MUTATION ANALYSIS OF NR5A1 ENCODING STEROIDOGENIC FACTOR 1 IN 81 PATIENTS WITH 46,XY DISORDERS OF SEX DEVELOPMENT (DSD) INCLUDING HYPOSPADIAS
# K. McElreavey1, S. Allali2, J.-B. Muller2, R. Brauner2, D. Lourenço1, C. Trivin3, H. Lottmann4, S. Lortat-Jacob4, V. Karageorgou1, R. Boudjenah1, C. Nihoul-Fékéte5, O. De Dreuzy6, A. Bashamboo1 1Institut Pasteur, Paris, France 2Université Paris Descartes, Faculté de médecine and AP-HP, Hôpital Bicêtre, Le Kremlin Bicêtre, France 3Hôpital Necker-Enfants Malades, Service d’explorations fonctionnelles, Paris, France 4Hôpital Necker-Enfants Malades, Service de chirurgie viscérale pédiatrique, Paris, France 5Université Paris Descartes, Paris, France 6Hôpital Bicêtre, Le Kremlin Bicêtre, France
Objectives: Mutations of the NR5A1 gene encoding steroidogenic factor-1 have been reported in association with a wide spectrum of 46,XY DSD phenotypes including severe forms of hypospadias. We evaluated the frequency of NR5A1 gene mutations in a large series of patients presenting with 46,XY DSD and hypospadias. Methods: Based on their clinical presentation 81 patients were classified either as complete and partial gonadal dysgenesis (uterus seen at genitography and/or surgery, n=15), ambiguous external genitalia without uterus (n=33) or hypospadias (n=33).All cases were screened for mutations in the NR5A1 gene. Results: We identified heterozygous NR5A1 mutations in 4 cases of ambiguous external genitalia without uterus (12.1%; p.Trp279Arg, pArg39Pro, c.390delG, c140insCACG) and a de novo missense mutation in one case with distal hypospadias (3%; p.Arg313Cys). Mutant proteins showed reduced transactivation activity and mutants p.Arg39Pro and p.Arg313Cys did not synergize with the GATA4 cofactor to stimulate reporter gene activity, although they retained their ability to physically interact with the GATA4 protein. Conclusions: Excluding individuals with karyotype anomalies or carrying mutations in the SRY gene, mutations in NR5A1 were observed in 5/77 (6.5%) cases of DSD including hypospadias. Excluding the cases of 46,XY gonadal dysgenesis the incidence of NR5A1 mutations was 5/66 (7.6%). An individual with isolated distal hypopadias carried a de novo heterozygote missense mutation, thus extending the range of phenotypes associated with NR5A1 mutations and suggesting that this group of patients should be screened for NR5A1 mutations.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
46 3rd International Symposium on Disorders of Sex Development
P-05 A NOVEL SRY MISSENSE MUTATION IN A 46,XY FEMALE PATIENT WITH BILATERAL GONADOBLASTOMA AND DYSGERMINOMA
# Y. Van der Zwan1, R. Hersmus2, H. Stoop2, J.W. Oosterhuis2, K.P. Wolffenbuttel3, S.L.S. Drop1, L.H.J. Looijenga2 1ErasmusMC, Pediatric Endocrinology, Rotterdam, Netherlands 2ErasmusMC, Josephine Nefkens Institute, Pathology, Rotterdam, Netherlands 3ErasmusMC, Pediatric Urology, Rotterdam, Netherlands
Introduction: Disorders of Sex development (DSD) are congenital conditions of incomplete or disordered gonadal development leading to discordance between genetic sex, gonadal sex, and phenotypic sex. Especially patients with gonadal dysgenesis and hypovirilization are at risk of developing malignant type II germ cell tumors (GCTs) (seminoma/dysgerminoma and nonseminoma), preceded by either carcinoma in situ (CIS) or gonadoblastoma (GB). SRY mutations residing in the HMG (High Mobility Group) domains are found in 10 -15% of the 46,XY gonadal dysgenesis cases and affect binding to and bending of DNA. This domain contains two nuclear localization signals (NLSs). Materials & Methods: Immunohistochemical staining was performed on formaline - fixed paraffin - embedded slides of 3 μm thickness. The antibodies directed against OCT3/4, c-KIT, Stem Cell Factor (SCF), Testis Specific Protein on the Y chromosome (TSPY), SOX9 and FOXL2 have been described before In addition direct sequencing of the SRY gene on peripheral blood DNA of the patient was performed. Results: A unique case is here reported of a phenotypical normal woman (age 22), who had primairy amenorrhoea at presentation, later diagnosed as hypergonadotropic hypogonadism on the basis of 46,XY gonadal dygenesis with a missense mutation on the SRY gene just before the second NLS (K128R). Laparoscopic examination revealed streak ovaries and a normal, but small, uterus. Pathological examination showed the presence of bilateral GB and dysgerminoma, confirmed by immunohistochemistry including OCT3/4, c-KIT, Stem Cell Factor (SCF), Testis Specific Protein on the Y chromosome (TSPY), SOX9 and FOXL2. The patient had a severe kidney impairment and (treatment-resistant) osteoporosis. Discussion & Conclusion: Most likely the identified mutation results in reduced transcription activation or binding and bending of the DNA, currently under investigation. This will result in less SOX9, i.e., absence of Sertoli cell development, and a maturation block of embryonic germ cells, increasing the risk for malignant transformation. In addition, it might be linked to the osteoporosis and kindney anomalies found. This further supports the importance of proper diagnosis of DSD patients, especially those with an increased risk for GCTs, allowing early diagnosis and treatment, thus preventing the development of invasive cancer.
20 – 22 May 2011, Lübeck, Germany 47
P-06 ARRAY-CGH ANALYSIS IN THE GHENT UNIVERSITY HOSPITAL DISORDERS OF SEXUAL DEVELOPMENT POPULATION
# B. Delle Chiaie1, B. Menten1, T. Acke1, E. De Baere1, M. Cools2 1University Hospital Ghent, Center for Medical Genetics, Ghent, Belgium 2University Hospital Ghent, Department of Pediatric Endocrinology, Ghent, Belgium
Objectives: Disorders of sexual development (DSD) are one of the most common human birth defects, occuring in almost 3% of newborns. Despite the high incidence, the underlying molecular mechanisms for DSD are only partially understood. Conventional karyotyping and molecular analysis of specific genes lead to a diagnosis in only a minority of patients. Since its introduction, array comparative genomic hybridization (array-CGH) has proven to be a helpful tool to find a molecular diagnosis in DSD patients. In addition, array- CGH leads to the identification of novel DSD candidate genes. Methods: We report 21 DSD patients, 10 syndromic and 11 non-syndromic cases, in which high resolution Agilent oligonucleotide array has been perfomed. Results: In 14 patients we identified a copy number variation (CNV) which has not been described as polymorphism. Six of them, all in syndromic DSD patients, were found to be causal. They had either previously been described to be involved in DSD or included known DSD genes (SF1, DMRT1). In eight patients (syndromic and non-syndromic) we found a CNV of unknown significance, mostly inherited from a healthy parent. Conclusion: By array-CGH, the cause of DSD has been elucidated in 60% (6/10) of syndromic DSD cases. None of the non-syndromic DSD patients carried a known causal aberration. In our experience, array-CGH appears to be an indispensable tool in the diagnostic work-up of syndromic forms of DSD, whereas its place in the diagnosis of isolated DSD remains to be determined.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
48 3rd International Symposium on Disorders of Sex Development
P-07 SYNERGISTIC EFFECT FOR P.Q798E AND P.C806F MUTATIONS IN A PATIENT WITH COMPLETE ANDROGEN INSENSITIVITY SYNDROME
# R.J. Petroli1, A. Maciel-Guerra2, F.C. Soardi2, F. Leme de Calais1, G. Guerra-Junior3, M. Palandi de Mello1 1Universidade Estadual de Campinas, Centro de Biologia Molecular e Engenharia Genética, São Paulo, Brazil 2Universidade Estadual de Campinas, Faculdade de Ciências Médicas, Departmento de Genética Médica, São Paulo, Brazil 3Universidade Estadual de Campinas, Faculdade de Ciências Médicas, Departmento de Pediatria, São Paulo, Brazil
The androgen insensitivity syndrome is the most frequent cause of sex development disorders in 46,XY patients. The molecular analysis the androgen receptor (AR) gene can confirm the clinical diagnosis, thus the aim of the study was to search for mutations in the AR gene in a patient with complete androgen receptor insensitivity syndrome (CAIS). A 13 years old female was referred to us with typical female external genitalia and B2P2 pubertal stage. She was born at term by normal delivery after an uneventful pregnancy; her birth weight was 3,950 g. Inguinal hernias were detected in the neonatal period therefore she was submitted to surgical repair at 2 months old. Bilateral testes were found upon inguinal herniorrhaphy and both were removed when the child was 16 months old. Estrogens had been prescribed five months before her first visit to our service. She had two normal sisters aged 19 and 17 years and a younger 2.5-year-old brother. Her hormonal evaluation revealed high FSH (92.6 mUI/mL; normal range (NR): 1.5-12.4 mUI/mL) and LH (20.6 mUI/mL; NR: 1.7-8.6 mUI/mL) levels and prepubertal levels of total testosterone (4.41 ng/mL; NR: 2.86-8.1 ng/mL) levels; her karyotype was 46,XY. Genomic DNA was obtained from peripheral blood and was extracted by proteinase K protocol. The eight exons of AR gene were amplified by PCR followed by sequencing. Mutant sequence was compared with the normal sequence of this gene. The model of mutant AR protein was built using the resolved 3D structure of human AR (PDB-ID 2AM9) as template. Molecular modeling was performed using MODELLER webserver program. The model images were examined and edited by PyMOL® program and Millennium STING (CNPTIA-Embrapa, Brazil). The human AR sequence was compared with the corresponding mammalian protein sequences in the ClustalW. Two nucleotide changes were found in the exon 6 for this patient. The first leads to the p.Q798E mutation that has already been described as causing the partial androgen insensitivity syndrome (PAIS) and the second causes the novel mutation p.C806F. The structural analysis showed internal contacts alterations. The diagnosis of the patient was CAIS, which did not correspond to the phenotype associated to the p.Q798E mutation. Therefore, the patient’s phenotype should be a direct result of the mutation p.C806F or due to a synergistic effect of both mutations, p.Q798E and p.C806F. Considering the structural analysis, those alterations modify internal contacts for the two mutated amino acids. The cysteine to phenilalanine change is under investigation, however, preliminary data suggest an important protein modification considering that they may cause the disruption of naturally occurring disulfide interactions, which will result in overall structure destabilization. Therefore, the molecular analysis of AR gene reveled that the combination of both mutations is probably responsible for the CAIS phenotype of the patient. Financial Support: Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP); Conselho Nacional de Pesquisa e Desenvolvimento (CNPq); Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES).
20 – 22 May 2011, Lübeck, Germany 49
P-08 ANALYSIS OF DSD PATIENT MUTATIONS LOCATED IN THE N-TERMINAL DOMAIN OF THE ANDROGEN RECEPTOR: HELPING TO UNDERSTAND DOMAINS WITH MOLTEN GLOBULAR STRUCTURE
# I. Hughes1, R. Tadokoro Cuccaro1, K. Watt2, J. Davies1, T. Bunch1, I.J. McEwan2 1University of Cambridge, Dept. Paediatrics, Addenbrookes Hospital, Cambridge, United Kingdom 2University of Aberdeen, Institute of Medical Sciences, Aberdeen, United Kingdom
World-wide screening of DSD patients for AR gene mutations has resulted in the identification of a large number which in many cases is proven to be the cause of AIS. The frequency distribution is receptor domain dependent, with few missense mutations described in the large N-terminal domain (NTD). The mutation rate is likely to be constant throughout the gene, suggesting a reduced impact of mutations on NTD function compared to those located in the ligand or DNA binding domains. The aim of this study was to establish a pathogenic role for NTD mutants in disease, and to use them as tools for defining the structure-function relationships of this poorly understood domain. Ten mutations were investigated: Q118E, five in the Tau1 region (A157T, G214R, N233K, G246V, L270F) and two in Tau5 (P378R, P390S). Two were trinucleotide slippage mutations adjacent to the glutamine repeat (insL57 and delL57). All mutations were recreated and activity measured in two reporter gene assays (GRE and PEM promoters). Loss of transactivation was observed for A157T, N233K and G246V. Protein levels of InsL57 were less than wild-type AR in stable isogenic expressing cell lines. Transactivation activity was not reduced when expressed at the same level as wild-type in transient transfection assays. Other mutants exhibited wild-type transactivation activity and protein expression levels. Kindred analysis revealed inheritance via the paternal grandfather for two probands (Q118E, Del57L). We were unable to define a pathological role for 7 NTD mutations, of particular concern when associated with more severe clinical phenotypes (PAIS). Nevertheless, the study supports a pathogenic role in AIS for patients harbouring mutants A157T, N233K and G246V. These three mutants are potential tools to define the molecular mechanism of AF1 function. Residue A157 maps close to the RAP74 (TFIIF) binding motif PSTLSL. Furthermore, N233 and G246 are located in a conserved alpha-helical region of AF1 thought to be involved in Tau1 transactivation activity / RAP74 binding. Intrinsic fluorescence emission spectroscopy and partial proteolysis assays were conducted to probe for structural changes of mutant AF1. No change was observed for the three mutants, possibly due to low resolution of the assays. CONCLUSION: The study highlights uncertainties about a pathological role for a number of NTD mutations. However, mutants shown to cause AIS will contribute to understanding the structure and function of this disordered “molten-globular” domain.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
50 3rd International Symposium on Disorders of Sex Development
P-09 DEVELOPING CELL ASSAYS SUITABLE FOR CHARACTERIZING AR MUTATIONS ASSOCIATED WITH ANDROGEN INSENSITIVITY SYNDROME
# K. Mooslehner1, J. Davies1, I. Hughes1 1Addenbrookes Hospital, Paediatrics, Cambridge, United Kingdom
Objective: Partial Androgen Insensitivity Syndrome (PAIS) is associated with impaired male genital development and often mutations in the Androgen Receptor (AR) are involved. The aim of this study is to develop an in vitro model for PAIS. For this purpose male genital development relevant murine cell lines are genetically modified to express mutant and wild-type AR. The modifications allow purification of multi- protein complexes of mutant and wild type AR under native conditions and analysis by mass spectrometry. Methods: In the ongoing work we are optimizing His and FLAG -tag purification conditions to allow tandem purification of AR co-regulator complexes. We aim to replace the endogenous AR with N-terminal Tandem Affinity Purification Tag (NTAP) AR. For that purpose a “flip in” vector has been developed to target exon 1 of the endogenous murine (m) AR. We are currently screening pools of clones for homologous recombinants. Once identified, Flip recombinase will be used to “flip in” NTAP-AR mutant or WT cDNA replacing the endogenous AR. A Recombination Mediated Cassette Exchange (RMCE) has been developed and is going to introduce LoxP recombination sites into the 4th intron and the 3’flanking region of the endogenous mouse (m) AR, which then enables us to exchange the wild-type Ligand Binding Domain (LBD) with a mutant LBD. Results: We have developed the epithelial and mesenchymal mouse cell lines P17 and M7, which stably express a tandem NH2-terminal affinity tag (3xFlag-6xHis=NTAP) fused to the mAR cDNA. This tag allows us to co-purify AR associated protein complexes under native conditions. The two cell lineages are derived from the proximal caput epididymus (P17) and the mesenchyme of the fetal vas deferens (M7) of the mouse, both express equal levels of NTAP-mAR and endogenous untagged mAR, and respond well to androgens. We were able to show that NTAP-AR translocates to the nucleus in P17 and M7, and that it activates an androgen responsive promoter in COS-1 cells. Using FLAG antibody coupled magnetic beads we optimized conditions for pulling out the co-regulators β-Catenin and SRC-1 bound to NTAP-mAR in P17. Conclusions: By expressing the tagged AR mutants in our system we aim to be able to identify mutant co- regulator binding patterns. The RMCE can now be tested in an ES cell culture system, and if successful would be suitable for generating AIS mouse models.
20 – 22 May 2011, Lübeck, Germany 51
P-10 RWDD1: AN ANDROGEN RECEPTOR COREGULATOR WITH A PUTATIVE ROLE IN MALE GENITAL DEVELOPMENT
# H. Grötsch1, M. Kunert1, K. Mooslehner2, D. Struve1, Z. Gao3, I. Hughes2, O. Hiort1, R. Werner1 1University of Lübeck, Department of Paediatrics, Lübeck, Germany 2University of Cambridge, Department of Paediatrics, Cambridge, United Kingdom 3Children’s Hospital Zhejiang, Department of Paediatric Surgery, Zhejiang, China Objective: The androgen receptor promotes hormone-dependent transcription as part of a multi-protein complex, including other transcription factors and so-called androgen receptor coregulators (coactivators and corepressors). Although it is known that androgen receptor-dependent transcription is required for the development of the male phenotype during embryonic development, the exact molecular interactions involved in this process have not been analyzed so far. The aim of the project is to identify androgen receptor coregulators required for the genital development of male mouse embryos. Methods: We have constructed cDNA libraries of genital tubercle tissue from male mouse embryos of the developmental stages E15, E16 and E17. Using a yeast two-hybrid system, the libraries were screened for proteins interacting with the AR in the presence of dihydrotestosterone. Candidate genes that play a role in proliferation, cell cycle control and differentiation were further analyzed by cellular reporter assays. The binding of the corresponding proteins to the AR was confirmed by in vitro protein binding assays and by coimmunoprecipitation experiments using an epithelial Wolffian duct cell line. Results: One newly identified binding partner of the AR was found in 9 independent clones from all three cDNA libraries. The cellular function of this protein, Rwdd1, is unknown so far. In cotransactivation experiments Rwdd1 caused an enhancement of AR transcriptional activity at three different promoters in different cell types. Moreover, Rwdd1 had a positive effect on the activity of the glucocorticoid receptor and the mineralocorticoid receptor, indicating that this protein might be a general nuclear receptor coactivator. Protein binding assays demonstrated a physical DHT-independent interaction between Rwdd1 and the AR. Immunohistochemical staining of Rwdd1 in genital tubercles of male mice at E16 and E17 showed a mainly cytoplasmic localization with a more pronounced expression in the urethral plate epithelium of the developing urethra. Conclusions: These results suggest that Rwdd1 might be an androgen receptor coregulator involved in genital development.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
52 3rd International Symposium on Disorders of Sex Development
P-11 INVESTIGATING THE PATHOGENICITY OF ANDROGEN RECEPTOR (AR) MIS-SENSE MUTATIONS IN ANDROGEN INSENSITIVITY SYNDROME (AIS)
# T. Bunch1, K. Zieliñska1, I. Hughes1 1University of Cambridge, Paediatrics, Cambridge, United Kingdom
Objectives: Patients with suspected Androgen Insensitivity Syndrome (AIS) are routinely screened for mutations in the androgen receptor (AR) gene. While non-sense mutations are usually assumed to be pathogenic, the role of mis-sense mutations has to be established. We investigated mutations from the Cambridge DSD database using transactivation assays, N-Terminal/C-Terminal interaction assays (N/C) and ligand binding assays. Methods: Transactivation asssays were carried out in COS1 cells which were transfected with AR and either GRE, PEM or probasin reporter plasmids. AR's were induced with Mibolerone, and luciferase measured after 24hours. N/C assays were performed by co-transfecting COS1 cells with pM-LBD, pVP16-hAR (2-656) and luciferase reporter plasmid pCMX-UAS-TK-Luc. Ligand binding assays (Kd) were performed in COS1 cells in the presence of increasing concentrations of 3H Mibolerone at 37˚C for 1hr. Results: DNA Binding Domain mutations V581G, I603S and R615P all showed loss of transactivation ability in-keeping with the patient phenotypes. The results are supported by crystal structure and published data showing that these residues lie within regions known to be required for normal DNA binding. The McGill AR mutation database also shows a number of other mutations at the same locations, further suggesting a pathogenic role for these mutations. Hinge region mutation I664T and Ligand Binding Domain mutations F673C, E678G, D690E, G708R, A721T, F827S, T860I, L881I and H917R were all found to have reduced activity in the GRE transactivation assay. Most of the mutations tested demonstrated reduced ligand binding affinity in-keeping with the loss of transactivation activity. Two exceptions were D690E, which had a wild-type Kd, and H917R where the increased Kd was insufficient to explain the CAIS phenotype. Mutations A870V and A896E both showed normal transactivation in the GRE and probasin reporter assays, and both had wild-type Kd's. The only abnormality identified thus far is an increased N/C interaction. Analysis of the AR crystal structure shows that these two mutations are located adjacent to the AF-2 region, known to be important for N/C interaction and co-regulator binding. The McGill database lists several mutations at residue A870 including A870V, supporting the hypothesis that this mutation is pathogenic to normal sex development. Conclusions: Functional tests such as the transactivation assay provide useful information about the effects of mutations on the structure and function of the AR, but the data should be interpreted in the context of available clinical and research information. Mutations such as A870V and A896E highlight the need to develop new assay systems that are more relevant to sex development.
20 – 22 May 2011, Lübeck, Germany 53
P-12 IDENTIFICATION OF THE FKBP4 MUTATION C.T1575C (PX460Q) IN A DSD PATIENT WITH ABERRANT GENITAL SKIN FIBROBLAST LIGAND BINDING, BUT A NORMAL AR
# I. Hughes1, J. Davies1, V. Pilfold-Wilkie1 1University of Cambridge, Department of Paediatrics, Addenbrookes Hosp., Cambridge, United Kingdom
In contrast to the numerous genes implicated in gonadal dysgenesis or androgen deficiency, the Androgen Receptor gene (AR) is the only known cause of Androgen Insensitivity Syndrome (AIS) in humans. Loss of AR function due to the disruption of co-regulators is an alternative possible mechanism for AIS. One such candidate is the co-chaperone FKBP52 encoded by the gene FKBP4. FKBP52 interacts with the AR regulating ligand binding and transactivation. Furthermore, the FKBP4 knock out male mouse presents with a PAIS phenotype. We hypothesised that humans with disorders of sex development (DSD) could have mutations of FKBP4 thereby perturbing androgen binding to wild-type AR. To test this hypothesis we selected 12 idiopathic DSD patients on the basis of aberrant ligand binding kinetics measured in genital skin fibroblasts; all subjects had a normal AR. Genomic DNA sequence analysis of the FKBP4 gene was performed for the 12 probands; a heterozygous DNA sequence variation c.T1575C (pX460Q) was identified in one subject (accession ref: NM_002014.3). The change of the stop codon to a glutamine resulted in a 30 amino-acid extension of the C-terminal domain of FKBP52. The same mutation was identified in two 46XY siblings; one had DSD and the other a normal male genital phenotype. Additional variants were identified in the study, including the common intronic variant IVS3+50 g>a found in normal and DSD patients. No compounding FKBP4 mutation was identified, so that a pathogenic role for mutant FKBP52 in DSD was not established. However, the function of this unique FKBP52 mutant remains to be determined and we have not excluded that it may contribute to an AIS-like phenotype.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
54 3rd International Symposium on Disorders of Sex Development
P-13 CLINICAL OUTCOME STUDIES FOR DISORDERS OF ANDROGEN PRODUCTION: SRD5A2 AND 17&BETA;HSD-3
# H.L. Miles1, T.I. Bunch1, V. Pilford-Wilkie1, I.A. Hughes1 1University of Cambridge, Department of Paediatrics, Cambridge, United Kingdom
Objectives: SRD5A2 and 17βHSD-3 enzymes are essential to produce sufficient androgens for male development, mutations result in XY DSD. There is limited outcome data for these disorders. Methods: Patients were identified with SRD5A2 and 17βHSD-3 mutation from our DSD database. Presentation, biochemistry and pubertal outcome were documented. Results: Information was available for 34 patients with SRD5A2 and 25 patients with 17βHSD3. Parental consanguinity was high for both disorders and high rates of homozygote missense mutations documented. Homozygous splice site mutations occurred in one third of affected males with 17βHSD-3. SRD5A2 affected males presented at birth with genital ambiguity. Sex of rearing was equally split between male and female. 17βHSD-3 affected males mainly presented with a female phenotype at birth, sex of rearing was female with the majority of cases presenting in childhood with inguinal masses. A lack of genotype- phenotype correlation was observed for both enzyme deficiencies. Typical biochemistry was specific but not universal. Low testosterone: androstendione ratio post HCG stimulation (<0.8) was 89% sensitive for 17βHSD3. Diagnostic sensitivity for SRD5A2 using post HCG testosterone: DHT ratio was poor for traditional values of a ratio greater than 30 but increased to 74% by using a ratio of greater than 10. A minority of patients in our cohort presented at pubertal age. One affected male with SRD5A2 presenting with virilisation changed sex to male whereas a second elected to remain female, a 50% change of sex observed in keeping with previous data. None of the 6 patients in our cohort with 17βHSD-3 who presented at puberty chose to change sex. We present pubertal follow up data for 18 patients with SRD5A2 (9 assigned male at birth who remained male, 7 assigned female at birth who remained female, one female reassigned male at the age of 2 years, one female reassigned male following virilisation at puberty) and 14 affected males with 17βHSD-3 deficiency who were raised female and remained female. Conclusions: The presentation of these two enzyme disorders overlaps both with each other and the phenotype of CAIS/PAIS. Biochemistry is specific but sensitivity is low for SRD5A2. Fewer cases presenting with virilisation at puberty in our cohort chose to change sex than published data. Pubertal outcome as a group was good with genital staging between G3 and G5 for those raised male and breast development B3 to B5 for those raised female receiving ethinyloestradiol replacement.
20 – 22 May 2011, Lübeck, Germany 55
P-14 DISORDERS OF SEXUAL DEVELOPMENT IN A GROUP OF SUDANESE: HARMONICAL DIAGNOSIS
# M. Ellaithi1,2, R. Werner3, F. G. Riepe4, N. Krone5, A. E. Kulle4, A. Kamil6, D. Gissselsson7,8, W. Alt5, P.-M. Holterhus4, O. Sabir1, O. Hiort3 1Al-Neelain University, Faculty of Medicine and Health Sciences, Khartoum, Sudan 2Ahfad University for Women, Faculty of Pharmacy, Omdurman, Sudan 3University of Lübeck, Division of Pediatric Endocrinology and Diabetes, Department of Pediatrics, Lübeck, Germany 4University of Kiel, Division of Pediatric Endocrinology and Diabetes, Department of Pediatrics, Kiel, Germany 5University of Birmingham, Centre for Endocrinology, Diabetes and Metabolism (CEDAM), School of Clinical and Experimental Medicine, Birmingham, United Kingdom 6National Health Centre, Human Genetics, Cairo, Egypt 7Lund University, Department of Clinical Genetics, Lund, Sweden 8Lund University, Department of Pathology, Regional and University Laboratories, Lund, Sweden
Introduction: Given the high consanguineous marriage rate and the autosomal recessive inheritance of a large number of disorders of sexual development (DSDs), we can assume a significant problem of DSDs in the Sudanese society. Here we show systematic clinical, hormonal, and genetic diagnosis in some Sudanese cases. Materials: Twelve index cases, aged between 6 months - 33 years (median 13. 4 years old). Ten were raised as females, two as males; all were clinically diagnosed with DSD. Among the cases there were three families, one case with an extended family history of DSD and only one case with no family history. Consanguinity is recorded among all cases. Steroids were analyzed in urine and plasma; accordingly genes correlated were analyzed. Results: Seven cases had a 46, XY and five a 46, XX karyotype. In three cases, results from urine analysis and plasma steroid analysis were discrepant, later gene mutation analysis correlated with urinary steroids more than with plasma steroids. Both urine and plasma steroids were congruent with molecular genetic diagnosis in 6 cases. In three cases, no samples for steroid analyses were available. In the group of 46,XY, three patients had a pathologic HSD17B3 mutation, 4 cases a SRD5A2 mutation, in the group of 46,XX patients, 4 had a CYP21A2, and one case with a CYP11B1 mutation. All mutations are homozygous mutations except for two siblings who have compound heterozygous mutations in HSD17B3 gene. Conclusions: The abnormal steroid profiling was followed by detection of the disease causing genes mutation in all cases with suspected recessive inheritance. Therefore, a stringent and stepwise analysis will give a correct diagnosis. This is of high importance also for the Sudanese society, as most of the patients were quite old at medical investigations and diagnosis was lacking.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
56 3rd International Symposium on Disorders of Sex Development
P-15 SEVERE VIRILIZATION OF A 46,XX NEWBORN AND HER MOTHER DURING PREGNANCY: A NOVEL CASE OF AROMATASE DEFICIENCY
# S. Riedl1, A. Springer2, J. Huber3 1Medical University, St Anna Children´s Hospital, Vienna, Austria 2Medical University, Paediatric Surgery, Vienna, Austria 3Medical University, Obstetrics and Gynaecology, Vienna, Austria
Case report: A female newborn with Prader stage 4-virilized external genitalia, 3rd child from Turkish consanguineous parents, presented at the paediatric endocrinology outpatient clinic of the Medical University of Vienna. Starting from the second trimester, the mother had developed progressive hirsutism, acne, and deepening of her voice. The preceding pregnancies, one girl, one boy, had been uneventful. At presentation on day 12 post-partum, her LH and FSH were suppressed (<0.1 mU/ml) in the presence of extremely high testosterone (T; 10.4 ng/ml [0.08-0.48]), androstendione (A; 69.4 ng/ml [0.3-3.3]), 17-hydroxy progesterone (17-OHP; 11.4 ng/ml [0.9-2.5]), and estrogen (E2; 2.787 pg/ml [22-232]). T, A, 17-OHP, and E2 declined rapidly to appr. 1/10 of their initial level on day 20. Tumour markers were negative. Sonography revealed markedly enlarged ovaries with PCOS morphology. An adrenal/ovarian virilizing tumour was ruled out by MRI. The baby had a 46,XX karyotype and was negative for SRY (FiSH). A normal uterus could be visualized on sonography. Biochemically, the girl showed constantly elevated LH (65, 67, and 59 mU/ml, respectively) and FSH (112, 123, and 142 mU/ml, respectively) without measureable E2 (<10 pg/ml) on 3 occasions during the first 2 months of life. Initial T was 0.37 ng/ml and anti mullerian hormone undetectable. Cortisol rise following 125 μg of synacthen was normal (43 μg/dl at 60 min). Urinary steroid profile showed normal excretion of CYP21 and CYP17 metabolites. Genetic testing for a CYP19 defect is currently under way. The baby is now 2 months old and has been raised as a boy so far. Conclusions: Clinically and biochemically, our case illustrates several features, that are distinctive of aromatase deficiency. Accordingly, the mother showed signs of severe virilization due to a lack of placental aromatisation of androgens originating from the fetal adrenal gland. Other disorders that may cause virilization of both mother and a female fetus like CYP21 defect, P450 oxidoreductase deficiency, a virilizing tumour or luteoma could be excluded, a maternal cause being unlikely since abundance of placental aromatase would normally protect a female fetus from severe virilization. Interestingly, the mother´s ovaries showed PCOS morphology, which was probably caused by high LH originating from the fetus, leading to marked ovarian hyperandrogenism. So far, 19 cases of aromatase deficiency have been published. To our knowledge, the changes observed in the mother during the first 2 months after birth have not been documented yet.
20 – 22 May 2011, Lübeck, Germany 57
P-16 HORMONE PROFILES IN ADOLESCENTS AND ADULTS WITH COMPLETE ANDROGEN INSENSITIVITY SYNDROME (CAIS)
# U. Doehnert1, S. Bertelloni2, A. Richter-Unruh3, R. Werner1, O. Hiort1 1University of Lübeck, Department of Pediatric Endocrinology and Diabetes, Lübeck, Germany 2University of Pisa, Department of Reproductive Medicine and Pediatrics, Pisa, Italy 3Endokrinologikum Ruhr, Wattenscheid, Germany
Background: Due to an increased risk of malignant degeneration of dysgenetic gonads early gonadectomy has often been recommended in individuals with disorders of sex development. Therefore data about reference values of gonadotropins and sex steroids in postpubertal women with CAIS are rare. Patients: Hormone profiles (testosterone, estradiol, sex-hormone-binding globulin SHBG, luteinizing hormone LH, follicle-stimulating hormone FSH) were studied in 36 women with proven mutations in the androgen receptor gene. At that time the women were aged 14 to 40 years (median = 17) and showed a CAIS phenotype with a breast development corresponding Tanner stadium 5 and absent or sparse pubic hair. Gonadopexy had been performed in three women, in the others the gonads had remained in situ. Results: Basal testosterone levels (1.8-15 μg/l, mean 6.4 μg/l) were within the male reference range for Tanner 5. Estradiol levels (17-70 pg/ml, mean 33 pg/ml) were measured within the male reference range for Tanner 5 or female reference range for Tanner 3. At the same time LH levels (5.5-51 IU/l) were elevated with normal FSH levels (0.4-14.6 IU/l). SHBG levels (16-180 nmol/l, mean 58 nmol/l) were in the upper male or lower female reference ranges respectively. Conclusions: The 36 women show a specific hormone profile with testosterone, estradiol and FSH levels in the male reference ranges for Tanner 5 and elevated LH levels. Further studies might reveal statistically significant CAIS-specific hormone profiles. Since the prevalence of germ cell tumors in CAIS has been estimated <1% until puberty in recent publications, we tend to recommend to leave the gonads in situ in order to preserve endogenous hormone production. In case of gonadectomy hormone replacement therapy could be steered individually according to CAIS-specific hormone profiles.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
58 3rd International Symposium on Disorders of Sex Development
P-17 EXPLORING THE UTILITY OF URINE STEROID METABOLITE RATIOS IN EVALUATING OVER 500 SUSPECTED CASES OF DISORDERS OF SEX HORMONE SYNTHESIS
# M. Rodie1, K. Rankin2, R. Howarth1, N. Liu1, D. Shapiro2, F. Ahmed1 1Royal Hospital for Sick Children, Department of Child Health, Glasgow, United Kingdom 2Glasgow Royal Infirmary, Clinical Biochemistry, Glasgow, United Kingdom
Background: Calculation of a urinary steroid metabolite ratio (uSMR) may be a useful method of improving diagnostic yield when investigating disorders of steroid hormone synthesis. However, the reliability of this method needs further exploration. Objective & Hypothesis: To determine the range of uSMR and the relationship between the uSMR and the diagnostic outcome in cases investigated for disorders of steroid hormone synthesis. Population / Methods: Ten ratios were calculated on steroid metabolite data previously analysed by GC-MS in urine samples from 544 patients less than 18 years of age collected between 2008 and 2010. In addition to comparing to the published data, information on clinical outcome on those beyond the 97th centile was collected. Results: Out of 544 patients, 334 (62%) were female, 192 (35%) were male and in 18 (3%) the sex was unknown or not documented when the test was requested. Indications for performing the test included adrenarche in 239 (44%) cases and XY DSD in 68 (13%) cases. Median age at test was 7.4 yrs (range 1 day to 18 years) with 11%, 4%, 3%, 3%, 7%, 16%, 40%, 6%, 6% and 4% of tests performed in the following age bands, <1wk, 2wks-2mths, 3-5mths, 6-11mths, 1-3yrs, 4-6yrs, 7-9yrs, 10-12yrs, 13-15yrs and 16-18yrs, respectively. In 97 (18%) patients, one or more uSMR was above the 97th centile. The proportion of uSMR which were above the 97th centile according to the above age bands, was 0%, 0%, 2%, 11%, 33%, 6%, 16%, 8%, 7% and 17%, respectively. Conclusions: Applying simplified diagnostic ratios to urinary steroid profile results identified 97 patients with abnormal results who may not have been picked up through conventional interpretation of the data. There is a need to assess the sensitivity and specificity of the uSMR and reference ranges for these ratios need to be age and sex matched.
20 – 22 May 2011, Lübeck, Germany 59
P-18 FEMINIZING SURGERY IN WOMEN WITH CONGENITAL ADRENAL HYPERPLASIA: LONG TERM OUTCOME STUDY ON ANATOMY, COSMETICS, SEXUAL- AND PSYCHOLOGICAL FUNCTIONING
# Y. Van der Zwan1, E. Janssen2, N. Callens3, K. Wolffenbuttel4, M. Van den Berg5, A. Dessens1, C. Beerendonk2, study group on DSD6 1ErasmusMC, pediatric endocrinology, Rotterdam, Netherlands 2Radboud UMC , Gynaecology, Nijmegen, Netherlands 3University Hospital Ghent, pediatric endocrinology, Ghent, Netherlands 4ErasmusMC, Pediatric Urology, Rotterdam, Netherlands 5UMC Groningen, Gynaecology, Groningen, Netherlands 6EMC, UMCN, VUMC, UMCG, the Netherlands, Netherlands
Objective: To investigate long term cosmetic, anatomical and psychosexual outcomes in patients with Congenital Adrenal Hyperplasia (CAH). Methods: Forty women with CAH, aged 15 and older from two Dutch medical centres were included. We assessed the physical and functional status by a gynecological interview and examination. Sexual development and functioning was assessed by validated rating scales and a structured interview. Results: 36 of 40 CAH patients underwent feminizing surgery; 25 patients underwent more than one operation. Seven out of 13 females needed re-surgery after a single stage procedure. Multiple regression showed that only the level of confluence had a significant effect on cosmetic outcome. CAH patients had a decreased sexual functioning and increased sexual distress compared to control women. Only 20 women (50%) had intercourse experience. The median age of sexarche was 19 years. Eight patients reported dyspareunia. Seven patients reported urinary incontinence. Conclusions: Our data show that 54% of the females needed re - surgery after a single stage procedure. The most important determinant of cosmetic outcome is the level of confluence. Although the overall cosmetic result had been evaluated as satisfactory by the patient and gynaecologist, the functional outcome in terms of being able to enjoy intercourse is poor.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
60 3rd International Symposium on Disorders of Sex Development
P-19 A FATE OF DYSGENETIC GONADS
# J. Slowikowska-Hilczer1, M. Szarras-Czapnik2, J.K. Wolski3, L. Jakubowski4, E. Oszukowska1, K. Marchlewska1, R. Walczak-Jedrzejowska1, E. Filipiak1, K. Kula1 1Medical University of Lodz, Dpt. of Andrology and Reproductive Endocrinology, Lodz, Poland 2Children’s Memorial Health Institute, Dpt.of Endocrinology and Diabetology, Warsaw, Poland 3Maria Sklodowska-Curie Memorial Cancer Centre, Urology-Oncology Dpt., Warsaw, Poland 4Institute CZMP, Dpt.of Genetics, Lodz, Poland
Objectives: Patients with gonadal dysgenesis (GD) and Y chromosome are usually submitted to the bilateral gonadectomy in early childhood to prevent germ cell neoplasia. Because of the paucity of older patients with GD and preserved gonads, a natural history of dysgenetic gonads is not exactly known. Methods: 22 patients with GD, aged 15-32, with 46,XY, 45,X/46,XY or 45,X/47,XXY, were investigated. In all ambiguous or female genitalia were found. In 8 cases single hypotrophic gonads were removed before the 10th year of age. The remaining gonads were preserved because of their relatively good development and intact testosterone secretion or a diagnostic process started later, around the 15th year of age, because of delayed puberty. Preserved gonads were biopsied or removed (suspicion of germ cell tumour). Histological evaluation, including immunohistochemical reaction with monoclonal antibodies against placental like alkaline phosphatase (PLAP), a marker of neoplastic germ cells, was performed. Results: In 9 cases streak gonads on both sides (pure GD) were found and in the next 7 a streak gonad on one side and an underdeveloped testis on the other (mixed GD). In the remaining 6 cases underdeveloped testicular structure on both sides (partial GD) was recognized. Seventeen out of 36 gonads (47%) revealed germ cell neoplasia: 6 overt germ cell tumours (seminoma, immature teratoma, embryonal carcinoma), 6 intratubular germ cell neoplasia (testicular carcinoma in situ - CIS) and 6 gonadoblastoma. In a 25-year-old patient GDB revealed invasive transformation and was connected with seminoma. In gonads with testicular structure dominated CIS (40%), while in the streak gonads gonadoblastoma (27%). Spermatogenesis appeared in 4 gonads (29% among cases with testicular structure and 0.9% among all GD) beside features of testicular underdevelopment (diminished tubular diameter, increased thickness of tubular membrane and increased intertubular spaces). In one of these cases number of germ cells was diminished and single elongated spermatids were found, while in others spermatogenesis was arrested at the level of spermatogonia or pachytene spermatocytes. Sertoli cell only syndrome was found in 5 gonads with testicular structure (36%) and in one gonad seminiferous tubules totally degenerated (0.7%). Conclusions: Dysgenetic gonads preserved to the age above 15 are at high risk of germ cell neoplasia, independently of the severity of aberrant organogenesis. In dysgenetic gonads with the presence of testicular structure seminiferous epithelium may be totally deprived of germ cells or degenerated and spermatogenesis, if maintained, is disturbed.
20 – 22 May 2011, Lübeck, Germany 61
P-20 SENSORIAL ROLE OF THE EPITHELIUM OF THE LABIA MINORA IN PREPUBERTAL GIRLS
# J. Schober1,2, N. Martin Aguacil2, D. Pfaff2 1UPMC Hamot, Urology, Erie, United States 2Rockefeller University, Neurobiology and Behavior, New York, United States
A variety of stimuli like friction, pressure and temperature on the labia minora skin activate the profuse and often rich distribution of endings of sensory neurons called receptors that are then transmitted to the spinal cord and to the brain for interpretation and response. A single sensory receptor can encode stimulus features such as intensity, duration, and direction. It usually activates many receptors, and is the central nervous system (CNS) the one that interpret the activity of the different receptors involve in the sensation and use it to generate coherent perceptions. The process by which the receptive portion of an afferent fiber converts natural stimulus energy into neural activity is called stimulus transduction. In order to activate a receptor, a stimulus must be of suitable intensity as well as of suitable quality. Physiologically identified responses have been linked to the morphologically distinct cutaneous receptors. In the majority of receptors in the skin, mechanical energy pressed on, displaces, or deforms the receptor (mechanoreceptor), which convert the energy into an electrochemical event, resulting in a nerve impulse. Free nerve ending and similar corpuscular receptors including Pacinian, Ruffini, and Merkel´s corpuscles, were found in the clitoris and on the the labia minora. The axonal endings respond to displacement of the collagen fibers induced by sustained or continuous mechanical stimuli. Free nerve endings are the simplest type of receptors and are receptive to touch, temperature and pain. Little is known about how signals from the numerous specialized nerve endings in the genital sites are converted into sexual pleasure, ultimately leading to orgasm. Their afferent impulses not only create spinal reflexes that influence genital motility and blood flow but also ascend the cord's spinothalamic and spinoreticular tracts to the brain. Here they are decoded and interpreted as sexual arousal/pleasure, and efferent outputs to the genitals and various organs become activated. Eventhough the labia minora skin can detect patterns at a very fine and smaller scale, nerve fibers are not always so numerous, which suggests that nerve terminals are helped by some other system. One possibility is that the stimulus energy may be transmitted through different cells within the skin, thus cells of the epidermis, not only Merkel's cells, may play an important role for sensation. In order to understand the role of the labia minora skin in sensing during sexual arousal it is important to study the morphology of the skin, to identified the different mechanoreceptors and to understand the role that epidermal cells may play. In other words we will study how the epidermal cells are arranged and how they could intercommunicate.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
62 3rd International Symposium on Disorders of Sex Development
P-21 IMMUNOCYTOCHEMICAL CHARACTERIZATION OF PACINIAN-LIKE CORPUSCLES IN THE LABIA MINORA OF PREPUBERTAL GIRLS
# J. Schober1, N. Martin Alguacil2, D. Pfaff2, Y. Litvin2, L. Mayaglou1,2 1UPMC Hamot, Urology, Erie, United States 2Rockefeller University, Neurobiology and Behavior, New York, United States
Study Objective: To better understand the precise role of sensory corpuscles within the female external genitalia. Design: After IRB approval, waste tissue samples were obtained from 10 normal girls (aged 2-9 years) who underwent surgery for labial fusion. Immunocytochemistry against protein gene product 9.5 (PGP 9.5), neuron-specific enolase (NSE), vasoactive intestinal peptide (VIP), 5-hydroxytryptamine transporter (5HTT), 5- hydroxytryptamine receptor 1A (5HT1A), Neuronal Peptide Y (NPY), neuronal nitric oxide synthase (nNOS) and estrogen receptors (ER) α and β was performed. Results: Pacinian-like corpuscles (PC) were identified in epithelium of labia minora of prepubertal girls. A central structure composed of an axon surrounded by a central core, outer core, external capsule, surrounded by encapsulated stroma, and a subsidiary innervation in the outer aspect of the corpuscle stroma stained for PGP 9.5 in the outer core and layers of the external capsule, NSE positive cells in layers of the outer core, 5HTT in stroma of the corpuscle and cells located in layers of the outer core, 5HT1A in cells of outer core, NPY in stroma of the corpuscle and nNOS in external core and external capsule of the central structure. ERα was present in stroma, external core and external capsule, ERβ in stroma of the corpuscle with subsidiary innervation in the stroma positive to PGP 9.5, VIP, and NPY. Conclusions: PGP 9.5, NSE, ERα, nNOS, and 5HTT immunoreaction detected in the outer core and external capsule could indicate these areas may play an important role in the functional aspects of the Pacinian-like corpuscle.
20 – 22 May 2011, Lübeck, Germany 63
P-22 SENSORY NEURAL CHARACTERISTICS OF MERKLE CELLS IN THE LABIA MINORA OF PREPUBERTAL GIRLS
# J. Schober1, N. Martin Aguacil1, D. Pfaff1, Y. Litvin1,2, L. Mayaglou1,2 1UPMC Hamot, Urology, Erie, United States 2Rockefeller University, Neurobiology and Behavior, New York, United States
Study Objective: Merkel Cells (MC) are characterized by their large oval shaped cells containing dense core vesicles near the nerve fiber junction and microvilli-like projections which contact surrounding keriatinocytes. Merkel cells have been located in a diverse group of mammalian tissues such as; glabrous skin, hairy skin, sinus hairs, ectodermal derived mucosa (oral, anal, and nasal), and taste buds. MC express sensor proteins and neuropeptides regulating the neuro-immuno-cutaneous system. The goal of this study is to better understand the location and neural characteristics of Merkle cells within the female external genital epithelium of the labia minora. Design: After IRB approval, waste tissue samples were obtained from 10 normal girls (aged 2-9 years) who underwent surgery for labial fusion. 3,3'-Diaminobenzidine (DAB) immunocytochemistry against protein gene product 9.5 (Abcam), and CK20 (Santa Cruz )was conducted in order to assess protein expression. In order to assess colocalization of these proteins (with DAPI as well; 1:10000, Sigma), we conducted double immunofluorescent histochemistry. . Images were taken using an Inverted LSM 510 laser scanning confocal microscope (Zeiss). Results: Merkel cells appear as epithelial cells associated to a nerve terminal. In the labia minora Merkel's cells were detected as a single cell isolated near the basal lamina or grouped, either forming clusters of cells in the granulous layer, or around the protrusion of a dermal papilla, when it protrudes through the epidermis, or sometimes they were located in more superficial layers of the epidermis. Merkels cells were never located along the whole layer, but in some spots of the layer. Conclusions: Merkle Cells are identified in the labial minora of prepubertal girls by colocalization with double immunoflourescent staining for PGP 9.5, and CK20.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
64 3rd International Symposium on Disorders of Sex Development
P-23 HYPOSPADIAS REPAIR WITH PRESERVATION OF THE VAGINA IN BOYS WITH DSD
# K. Wolffenbuttel1, A. Dessens1, J. Van den Hoek1 1Erasmus MC, Pediatric Urology, Rotterdam, Netherlands
Objectives: A large utricle is a common feature in boys with DSD. In our hospital utricles are only removed when they cause symptoms. In rare cases there is a separate perineal opening of the utricle. This so-called male vagina interferes with hypospadias correction and used to be removed. As gender dissatisfaction after puberty in this group of patients with DSD is common, genital tissues should be preserved whenever possible. We report on our modified technique for hypospadias repair with intentional preservation of the vagina. Methods: Between 2000 to 2003 three neonates with ambiguous genitalia and a male vagina presented to our multidisciplinary team. Karyotype was 46,XY in 2 and 45,X/46,XY in 1 patient. Only in the patient with mosaic karyotype the vagina was visualized on ultrasonography; in the other two the vagina was detected during the first hypospadias repair. After genetic and hormonal analysis all patients were assigned the male gender, and scheduled for 2-stage hypospadias repair in early childhood. Two boys with a small penis were pre-treated with topical dihydrotestosterone (DHT). In the second stage the vaginal opening was incorporated in the neo-urethra, and in this way converted into a utricle. Results: On follow-up all patients, at age 7, 8 and 11 years respectively, are asymptomatic after a mean period of 5.5 years since last surgery. They are able to void in a standing position with a satisfactory urinary stream (Qmax 11-15 ml/s). Utricle size remained stable on ultrasonography. There are no signs of gender dissatisfaction; psychological evaluation of the oldest boy shows a male gender role and behaviour. Conclusions: As future gender identity in DSD-patients with ambiguous phenotype and (mosaic)Y genotype is difficult to predict, genital tissues should be preserved whenever possible. Raising these children as males will allow early genital reconstruction without loss of tissue. Our surgical approach of preservation of the vagina by conversion into a utricle during hypospadias repair, combines the advantages of reversible genital surgery and prevention of adverse psychological effects of growing up with ambiguous genitalia.
20 – 22 May 2011, Lübeck, Germany 65
P-24 CLINICAL AND MOLECULAR GENETIC INVESTIGATION OF CONGENITAL MÜLLERIAN ABNORMALITIES
# R. McGowan1, R. Crawford2, D. Shapiro3, N. Morrison4, S. Logan5, A. Balen6, F. Ahmed7, M. Deeney8, J. Tolmie1, E. Tobias9 1Yorkhill Hospital, Department of Clinical Genetics, Glasgow, United Kingdom 2Glasgow Royal Infirmary, Department of Clinical Psychology, Glasgow, United Kingdom 3Glasgow Royal Infirmary, Department of Clinical Biochemistry, Glasgow, United Kingdom 4Yorkhill Hospital, Department of Molecular Cytogenetics, Glagsow, United Kingdom 5NHS Grampian, Department of Sexual and Reproductive Healthcare, Aberdeen, United Kingdom 6Seacroft Hospital, Leeds Centre for Reproductive Medicine , Leeds, United Kingdom 7University of Glasgow, Department of Paediatric Endocrinology, Glasgow, United Kingdom 8Glasgow Royal Infirmary, Department of Gynaecology, Glasgow, United Kingdom 9University of Glasgow, Duncan-Guthrie Institute of Molecular Genetics, Glasgow, United Kingdom
The prevalence of congenital Müllerian duct abnormalities may be as high as 4% but there is little information on the underlying aetiology. To understand the aetiology, there is a need for careful delineation of the phenotypic spectrum associated with these conditions.
Objectives: The objectives are: to clinically phenotype women over 16 years who have congenital Müllerian abnormalities, to determine the prevalence of associated congenital malformations and associated psychological effects and to improve our understanding of the underlying genetics. Methods: Women are recruited from gynaecology clinics or identified from the SGAN register that contains a dataset on patients and provides a unique resource for recruitment. Results: To date, 20 women with a karyotype of 46XX and Müllerian abnormalities have been recruited: 7 with MRKH (Mayer-Rokitanksy-Küster-Hauser syndrome), 3 with uterus didelphus, duplex cervix and bifid upper vagina, 3 with bicornuate uterus (1 with partial vaginal atresia and one with endocervical atresia), 2 with septate uterus (1 with transverse vaginal septum and absent lower third of vagina), 3 with unicornuate uterus (2 associated with Müllerian aplasia, renal aplasia and cervicothoracic somite dyslasia, MURCS) and 2 with vaginal agenesis. The mean age is 31.6 years ± 9.9. None of these women have consanguineous parents, there is a family history of Müllerian abnormality in one case only. All seven women with MRKH and one woman with bicornuate uterus presented with primary amenorrhea. Other presenting features were abdominal pain, dyspareunia, weight loss, infertility or abnormality identified on routine health check. The diagnosis was made by one or a combination of the following: US, EUA, vaginogram, laparoscopy and laparotomy, with MRI used in 7/20 cases. Abnormalities of the renal tract were present in 50% (e.g. agenesis, hypoplasia), skeletal anomalies in 25% and cloacal anomalies in 25% of women. Other congenital anomalies were present in 13/20 women (e.g. absent thumb, hearing impairment or duplicated IVC). The mean BMI of affected women was 23.6 ± 5.3 and acne and hirsuitism were present in 6 and 1 out of 20 cases but there was no biochemical hyperandrogenism. Serum gonadotrophins and oestrogen were normal apart from one case suggestive of premature ovarian failure at the age of 39 years. Urinary steroid metabolite profiles were normal. All women are independent; all but one attended a mainstream school. Only three are unemployed. Genetic analyses are underway. Discussion: Our preliminary studies suggest that congenital Müllerian abnormalities are often associated with renal and skeletal abnormalities and all affected women should be thoroughly assessed. Biochemical evidence of androgen excess is not common in these cases previously identified in some women with MRKH. Given the range of problems encountered by these women, there is a need to study long-term outcome as well as the genetic basis to these conditions.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
66 3rd International Symposium on Disorders of Sex Development
P-25 DELAYED RECOGNITION OF TWO DISORDER OF SEX DEVELOPMENT (DSD) CASES: A MISSED POSSIBILITY FOR EARLY DIAGNOSIS OF MALIGNANT GERM CELL TUMORS
# R. Hersmus1, H. Stoop1, S. White2, S. Drop3, L. Looijenga1 1ErasmusMC, Pathology, Rotterdam, Netherlands 2University of Melbourne, Molecular Development Research Group, Melbourne, Australia 3ErasmusMC, Pediatric Endocrinology, Rotterdam, Netherlands
Summary: Disorders of sex development (DSD) are defined as a congenital condition in which development of chromosomal, gonadal or anatomical sex is atypical. DSD occurs with an incidence of about 1:4,500 to 1:5,000 live births. Specific patients with this disorder, i.e. those with gonadal dysgenesis or hypovirilization, with part of the Y-chromosome (GBY), are known to have an increased risk to develop malignant type II germ cell tumors (GCTs: seminomatous and non-seminomatous tumors). DSD may be diagnosed in a newborn (e.g., presence of ambiguous genitalia), or later in life, even at or after puberty, when the anomalies are more subtle. Here two independent cases are presented in which DSD was retrospectively recognized, initiated by the diagnosis of malignant GCTs after puberty. Materials and Methods: Immunohistochemistry and fluorescent in situ hybridisation (FISH) was performed as described in R. Hersmus et al. EJHG. 2009; 17:1642-1649 Results: Patient no. 1 had multiple hypospadias corrections at young age, and the left testis was positioned from inguinal to scrotal, while no right gonad was found. At 26 years of age the patient underwent surgery for an inguinal hernia during which the right (inguinal) testis was identified and removed. Detailed pathological examination revealed a dysgenetic gonad with gonadoblastoma (GB), dysgerminoma (DG), carcinoma in situ (CIS) and seminoma (SE), supported by immunohistochemistry (OCT3/4, TSPY, SOX9 and FOXL2; Figure 1). Irradiation was given because of metastasized disease. Patient no. 2 was initially diagnosed with a right intra-abdominal testicular SE at the age of 20 years. Half a year later, a biopsy from the left inguinal gonad showed malignant changes, confirmed as GB, DG, CIS and SE in the gonadectomy specimen (Figure 2). Hypospadias and a uterus was found. Chemotherapy was given because of metastasized disease. Both patients showed a normal male karyotype, based on FISH analysis (Figure 1 and data not shown) and karyotyping (data not shown). Currently high throughput array-based studies are performed to screen for genomic changes, possibly explaining the molecular basis of the 46, XY DSD. Conclusions: These two cases demonstrate the overlap between the so-called Testicular Dysgenesis Syndrome (TDS) and DSD. In fact, identification of common characteristics of TDS and DSD, e.g., hypospadias and (bilateral) cryptorchidism, is informative to identify patients at high risk for malignant GCTs. In addition, the presence of GB must trigger further investigation of possible DSD. This knowledge will allow early diagnosis of the disease, resulting in optimal treatment of the patients at early stage, likely excluding the need for irradiation and chemotherapeutic intervention.
20 – 22 May 2011, Lübeck, Germany 67
P-26 SURGICAL CORRECTION OF WEBBED PENIS ASSOCIATED WITH PENOSCROTAL TRANSPOSITION
# H. Özbey1, C. Münevveroglu1 1Istanbul University, Pediatric Surgery & Pediatric Urology, Istanbul, Turkey
Concealed penis (CP) is an uncommon congenital anomaly, that is an inconspicious or hidden appearance of the penile shaft. Primary concealment may be due to buried penis, webbed penis or penoscrotal transposition. Secondary concealment is usually iatrogenic, such as trapped penis seen after circumcision. In this report, the surgical correction of a webbed penis associated with penoscrotal transposition is described. The surgical treatment of concealed penis requires an awareness of the condition and has to be tailored to its aetiology. The described approach to webbed penis with penoscrotal transposition is a simple technique, in which both penile and scrotal relocations were provided with successful lengthening of the penile body.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
68 3rd International Symposium on Disorders of Sex Development
P-27 SEXUAL QUALITY OF LIFE IN INDIVIDUALS WITH COMPLETE ANDROGEN INSENSITIVITY (CAIS) AND MAYER-ROKITANSKY-KÜSTER-HAUSER SYNDROME (MRKHS)
# K. Huber1, M. Fliegner1, S. Brucker2, H. Richter-Appelt1 1Universitätsklinikum Hamburg-Eppendorf, Institut für Sexualforschung, Hamburg, Germany 2Universitäts-Frauenklinik, Tübingen, Germany
Objective: Individuals with a Complete Androgen Insensitivity Syndrome (CAIS; 46,XY-karyotype) are born with an unremarkable female body appearance. Androgens are produced, but the target organs show an insensitivity to these hormones. External genitalia develop along a female pattern, but due to the XY- karyotype no uterus or vagina develop. Testosterone is produced by undescended testes. In the Mayer-Rokitansky-Küster-Hauser Syndrome (MRKHS), some similar characteristics are found insofar as these individuals are also born without uterus and vagina. In contrast to CAIS, MRKHS occurs in individuals with 46,XX-karyotype and ovaries. In the literature, individuals with CAIS and MRKHS have sometimes even been seen as one homogenous group; the differences between them remain unexamined. The current study takes a closer look at the sexual experiences and sexual quality of life of these two groups separately. Methods: Data collection took place as part of the larger study “Androgens, Quality of Life and Femininity”*. Sexual quality of life is measured as part of a comprehensive questionnaire using the Female Sexual Function Index (FSFI). Additional questions concerning partnership, sexual activity, sexual quality of life and medical treatment, especially surgical procedures (e.g. the creation of a so-called “neovagina”), are asked. Results for individuals with CAIS (n = 10) and MRKHS (n = 44) are described and compared to each other as well as to data from the literature concerning non-clinical samples. Results: Descriptive data concerning partnership, sexual activities, experience and satisfaction are reported. The majority of individuals in this sample have undergone a surgical operation to create an artificial vagina (neovagina). This has been reported more often in the group with MRKHS than in the group with CAIS. Some individuals describe a sexual quality of life that is impaired. The results are compared to data from the literature. Conclusions: Although there are some similarities between individuals with CAIS and MRKHS, differences in sexual quality of life and medical treatment suggest that these two groups should be looked at separately. Sexuality and sexual quality of life are important topics that should be considered and addressed when dealing with individuals with CAIS and MRKHS. * supported by the Else Kröner-Fresenius-Stiftung
20 – 22 May 2011, Lübeck, Germany 69
P-28 INFERTILITY AND THE WISH FOR A CHILD: A STUDY ON ATTITUDES TOWARDS PARENTHOOD IN PEOPLE WITH COMPLETE ANDROGEN INSENSITIVITY SYNDROME (CAIS) AND MAYER-ROKITANSKY- KÜSTER-HAUSER SYNDROME (MRKHS)
# M. Fliegner1, K. Huber1, S. Brucker2, H. Richter-Appelt1 1Universitätsklinikum Hamburg-Eppendorf, Institut für Sexualforschung, Hamburg, Germany 2Universitäts-Frauenklinik, Tübingen, Germany
Objective: Research regarding the wish for a child has mainly been conducted in the context of government policies and population issues, the use of contraception, and Assisted Reproductive Technology (ART). Little is known about family planning in individuals with permanent infertility conditions. The current study was designed to investigate the wish for a child in individuals with Complete Androgen Insensitivity Syndrome (CAIS) and in individuals with Mayer-Rokitansky-Küster-Hauser Syndrome (MRKHS). Individuals with (CAIS) can neither procreate nor carry a child. Their appearance is inconspicuously female, however, they present with uterine as well as partial vaginal aplasia. Their gonads consist of testicular tissue and are situated somewhere in the pelvic area. They do not show regular functioning as to the production of semen. There are no medical procedures available to assist them in reproduction. Individuals with MRKHS likewise face such situations regarding their uterine and vaginal constitution. In contrast to people with CAIS, these individuals possess fully-functioning ovaries. By means of medical assistance and with the help of surrogate mothers it is possible for them to have genetically-related offspring. In Germany, however, surrogate child-bearing is illegal. Thus within the confines of German law individuals with CAIS and MRKHS face comparable situations. The objective of the study was to gain more insight into individuals’ attitudes and their hopes and concerns regarding parenthood. Methods: The data were collected within the scope of the study “Androgens, Quality of Life and Femininity*”. Sample sizes amounted to n = 10 for CAIS, n = 44 for MRKHS, and n = 932 for the non-clinical group. After signing up for the study, individuals were sent extensive questionnaires. They were asked about their current wish for a child and were to rate its intensity. The questionnaires contained a self-developed scale on “Attitudes towards Motherhood” and questions about alternative ways to have children. The complete version of the questionnaire comprised many other scales and questions, only the respective parts of the data were chosen for the present analyses. Non-clinical comparison data were at hand from a preliminary study. Results: Results concerning the presence of a wish for children and its strength will be presented for each sample respectively (CAIS, MRKHS, non-clinical). Furthermore, their attitudes towards parenthood will be analyzed and compared. Data concerning the starting of a family through other means (adoption, foster care, surrogate mothers) will be presented. Conclusions: The wish for a child has not been a subject of intensive research regarding individuals with permanent infertility conditions. Just the same, our data suggest that starting a family is of vital importance to many and may be an important matter to explore when counselling infertile individuals. * supported by the Else Kröner-Fresenius-Stiftung
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
70 3rd International Symposium on Disorders of Sex Development
P-29 BODY EXPERIENCE IN INDIVIDUALS WITH DIFFERENT CONDITIONS OF INTERSEXUALITY
# C. Prochnow1, K. Schweizer1, H. Richter-Appelt1 1Universitätsklinikum Hamburg Eppendorf, Institut für Sexualforschung und Forensiche Psychiatrie, Hamburg, Germany
In recent years the topic of intersexuality/ Disorders of sex development (DSD) has gained more attention in medical and psychological research. Intersexuality refers to a variety of conditions in which an individual is born with a reproductive or sexual anatomy that does not seem to fit typical definitions of male and female individuals. Although the experience of one´s own body is closely connected to the experience of one´s self there has been little research on how individuals with intersexuality experience their own body. Previous findings have shown disturbances of body image in individuals with disorders of sex development. The Hamburg research project on intersexuality focused on quality of life and treatment experience of adults with different forms of intersexuality. The results presented here are a part of the Hamburg project and deal with the question whether individuals with different forms of intersexuality differ from a non intersexual control group in respect to their general assessment of their body and their satisfaction with their own body and different body parts.
20 – 22 May 2011, Lübeck, Germany 71
P-30 COMPARISON OF CLINICAL AND METABOLIC EFFECTS OF TESTOSTERONE AND ESTRADIOL IN ADULT GONADECTOMIZED PATIENTS WITH 46, XY DSD DUE TO COMPLETE ANDROGEN INSENSITIVITY SYNDROME (CAIS)
# W. Birnbaum1, L. Marshall1, D. Schnabel2, M. Bals-Pratsch3, A. Richter-Unruh4, R. Wagner5, S. Kropf6, O. Hiort1 1Universität zu Lübeck, UK S-H, Campus Lübeck, Klinik für Kinder- und Jugendmedizin, Lübeck, Germany 2Charité Berlin, Department for Experimental Pediatric Endocrinology, Berlin, Germany 3Fertility Center Regensburg, Regensburg, Germany 4Endokrinologikum Ruhr, Bochum, Germany 5University of Münster, Trial Supporting Facility, Münster, Germany 6Otto-von-Guericke-University Magdeburg, UKL Magdeburg, Institute for Biometry and Medical Informatics, Magdeburg, Germany
Objectives: Based on sporadic experiences in self-trials, it is expected that an androgen replacement instead of the usual estradiol substitution might improve the quality of life in patients with complete androgen insensitivity (CAIS) after gonadectomy. The biological pathways are yet unknown. Until now, no clinical trials have been conducted to elucidate any specific endocrine substitution scheme that will take into account the special medical needs of patients with CAIS. The Primary Objective is to detect differences in the effects of testosterone versus estradiol treatment on quality of life and well being in patients with CAIS due to mutation of the androgen receptor. The Secondary Objectives are to compare the levels of testosterone and estradiol, as well as other sex steroid metabolites between both treatments. We intend to investigate quality of life including sexual satisfaction and psychological well-being. Furthermore correlations between these findings and hormone levels will be analysed. Study design: We initiated a phase III-drug-study designed as a double-dummy, double-blind, cross-over, randomized, multicenter clinical trial. Subjects: 25-30 adult CAIS patients after gonadectomy. Experimental intervention: One month will be without any sex steroid substitution prior to treatment (wash-out-period). Thereafter patients will take either testosterone 50 mg or estradiol 2 mg and matching placebo respectively. Study medication is administered daily in a single dose on the skin in a blinded fashion for a duration of six months. Following a second wash-out-period the change to the reverse medication (either estradiol or testosterone and matching placebo) is conducted. Both interventions are successively applied to all patients in a cross-over design. Methods: Measures of metabolic and clinical parameters take place after three and six months on study medication. A follow-up is planned three months past treatment. General quality of life including sexual satisfaction and psychological well-being will be measured via questionnaires (SF-36, BSI, FSFI-d, questionnaire for adult DSD patients from the DSD-network). Steroid extraction profiles and other laboratory findings related to androgen and estrogen action respectively will be determined as control parameters. Further molecular genetic analysis is conducted depending on prior documented mutations and clinical findings. Psychological support is provided throughout the study course. Conclusions: The lack of previous studies or clinical trials on the special endocrine profiles in CAIS warrants this study. The results could lead to a complete new recommendation for hormone replacement encompassing the enhancement of quality of life in patients with CAIS.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
72 3rd International Symposium on Disorders of Sex Development
P-31 EVALUATION OF GONADECTOMY IN ADULTS WITH COMPLETE ANDROGEN INSENSITIVITY SYNDROME (CAIS): A PATIENTS&ACUTE; PERSPECTIVE
# F. Brunner1, K. Schweizer1, H. Richter-Appelt1 1Institute for Sex Research and Forensic Psychiatry, University Hospital Hamburg-Eppendorf, Hamburg, Germany
Background and Objectives: Complete Androgen Insensitivity Syndrome (CAIS) is one of the most common forms of 46,XY DSD. Minimal incidence is estimated at 1:99,000 (Boehmer et al., 2001). An androgen receptor defect results in insensitivity to androgens, leading to a phenotypic female appearance. There are, however, no uterus, a blind-ended vagina, and scant or absent pubic and axillary hair. Clinical management of all intersex conditions is controversial because available evidence is limited and conflicting. Regarding gonadectomy in people with CAIS condition different expert opinions exist. In particular, researchers disagree on the timing (for delayed gonadectomy see: Allen, 2009; for early gonadectomy see: Kiddo, 2009). Furthermore, recent studies reveal only a low incidence rate of germ cell tumors in the CAIS population (0.8%: Oakes et al, 2008). Predominantly support groups, therefore, call the necessity for gonadectomy in CAIS population into question. The aim of this study is to present the patients´ retrospective experiences and to gain insight on the psychological impact of this intervention. Furthermore, implications for an improved management with CAIS population will be discussed. Methods: The Hamburg Intersex Study investigates treatment experiences and quality of life in adults with different DSD conditions. Medical treatment experiences were assessed by a questionnaire and patients’ medical records. To gain insight on the retrospective experience a self-constructed questionnaire was developed: Satisfaction with the surgery result was assessed by a 5-point Likert scale. Further, participants described their feelings and cognitions about the gonadectomy. 13 adults with CAIS condition aged 24 to 47 were investigated. All participants live in the female gender role. Results: 12 of 13 participants with CAIS were gonadectomized. Gonadectomy was conducted at the age of 3 to 32. Only one person had additional surgical interventions (vaginal corrections). 6 of 12 persons (50%) were very dissatisfied with the surgery result, 4 (33%) dissatisfied and 2 (17%) satisfied. 9 participants had negative feelings and cognitions about the gonadectomy (fear: n=3; sadness because of infertility: n=2; feeling of “castration”: n=2; uncertainty: n=2; bad management: n=1). In one person, positive feelings referred to relieve of the tumor risk. Conclusions: The findings illustrate the psychological impact of gonadectomy and dissatisfaction with the surgery result. Patients should be prepared for gonadectomy and offered a counseling service. Professionals should be sensitive to the meaningfulness of a gonadectomy for the patient. Further research is necessary to investigate the reasons for dissatisfaction with gonadectomy in CAIS population. References: Allen, L. (2009). Opinion One: A Case for Delayed Gonadectomy. Journal of Pediatric and Adolescent Gynecology, 22(6), 381-384. Boehmer, A. L. M., Brüggenwirth, H., van Assendelft, C., Otten, B. J., Verleun-Mooijman, M. C., Niermeijer, M. F., et al. (2001). Genotype versus phenotype in families with androgen insensitivity syndrome. Journal of Clinical Endocrinology & Metabolism, 86(9), 4151-4160. Kiddo, D. A. (2009). Opinion Two: A Case For Early Gonadectomy. Journal of Pediatric and Adolescent Gynecology, 22(6), 384-386. Oakes, M. B., Eyvazzadeh, A. D., Quint, E., & Smith, Y. R. (2008). Complete Androgen Insensitivity Syndrome-A Review. Journal of Pediatric and Adolescent Gynecology, 21(6), 305-310.
20 – 22 May 2011, Lübeck, Germany 73
P-32 BODY IMAGE AND SEXUALITY IN INDONESIAN TREATED AND UNTREATED SUBJECTS WITH DSD
# A. Ediati1,2, A.Z. Juniarto2,3, S.L. Drop4, S.M. Faradz2,3, A.B. Dessens4 1Diponegoro University, Faculty of Psychology, Semarang, Indonesia 2Diponegoro University, Center for Biomedical Research, Faculty of Medicine, Semarang, Indonesia 3Dr. Kariadi Hospital, Semarang, Indonesia 4ErasmusMC-Sophia, Rotterdam, Netherlands
Objective: Since 2004, the Sexual Adjustment Team of the Faculty of Medicine Diponegoro University/Dr. Kariadi Hospital became more accessible for large group of patients with disorders of sex development (DSD). Many of our patients had been identified lately and had not received treatment before they entered our clinic. Consequently, all patients suffered from genital ambiguity and in many patients ambiguous secondary sex characteristics had developed. This study has been carried out in order to investigate body image and sexuality in these patients with DSD. Methods: Design: Comparison of treated and untreated patients with DSD and healthy controls on body image and sexual functioning. Subjects: 15 treated and 10 untreated male and 8 treated and 10 untreated female patients (aged 12-41y) and 43 healthy controls matched for gender, age and living area (rural/urban). Measurements: Degree of satisfaction with different body parts was measured with an Indonesian version of a Body Image Scale (BIS). Sexuality was investigated by application of the Indonesian translations of the Female Sexual Distress Scale-Revised (FSDS-R), the Female Sexual Functioning Index (FSFI), the Male Sexual Health Questionnaire (MSHQ), and the Klein Sexual Orientation Grid (KSOG). The scales had been applied verbally, due to the subjects’ unfamiliarity with self-report measurements and illiteracy. Statistics: Mann-Whitney test and Fisher Exact significance were applied. Results: Body Image: Results on the BIS revealed that female patients felt dissatisfied with their primary sex characteristics but not with other body parts. The treated males felt less satisfied than the untreated one with respect to secondary sex and non-sex characteristics. Sexuality: Data on the FSDS-R demonstrated that females with DSD experienced more sexual distress. Findings on the FSFI indicated that none of the female patients had a partner and never had been engaged in sexual relationships. Males did not report more sexual problems compared to controls. With respect to sexual orientation, control subjects considered themselves exclusively heterosexual whereas male and female patients with DSD considered themselves mainly heterosexual. A similar study has been carried out in Dutch treated patients. Findings in Indonesian and Dutch patients will be compared. The data will be placed in a cultural context and will be presented at the conference. Keywords: body image, sexuality, disorders of sex development, untreated, Indonesia
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
74 3rd International Symposium on Disorders of Sex Development
P-33 COGNITIVE AND EDUCATIONAL OUTCOME OF OFFSPRING FROM DEXAMETHASONE-TREATED PREGNANCIES AT RISK FOR CONGENITAL ADRENAL HYPERPLASIA DUE TO 21-HYDROXYLASE DEFICIENCY
# H.F. Meyer-Bahlburg1, C. Dolezal1, R. Haggerty2, M. Silverman3, B. Kassai4, P. Chatelain5, M.G. Forest5, M.I. New6 1NYS Psychiatric Institute / Columbia University, Psychiatry, New York, United States 2Columbia University, Psychiatry, New York, United States 3Mount Sinai School of Medicine, Psychiatry, New York, United States 4Inserm, CIC201, EPICIME, and Universite de Lyon, Bron, France 5Hopital Femme Mere Enfant de Lyon, Endocrinologie & Diabetologie Infantiles, Bron, France 6Mount Sinai School of Medicine, Pediatrics, New York, United States
Objectives: Pregnancy treatment with dexamethasone is used to prevent genital masculinization in 46,XX fetuses with classical congenital adrenal hyperplasia (CAH), but has become increasingly controversial due to animal studies suggesting potentially adverse side effects. This report presents data from studies designed to test whether prenatal DEX exposure impairs cognitive functioning and educational outcome in offspring from CAH-risk pregnancies. Methods: We present data from two observational follow-up studies of DEX-exposed offspring and controls in the U.S. using neuropsychological testing and one questionnaire study that combined data from the U.S. and France, together covering ages from childhood to young adulthood. Study 1 included 42 patients with CAH and 98 participants without CAH, age 5-12 years; Study 2 included 13 patients with CAH and 7 participants without CAH, age 11-24 years; and Study 3 included 68 patients with CAH and 65 participants without CAH, age 11-39 years. The first two studies employed several neuropsychological tests (blindly administered): In Study 1, the Kaufman Assessment Battery for Children (K-ABC; Kaufman & Kaufman, 1983) and the Wide Range Assessment of Memory and Learning (WRAML; Sheslow & Adams, 1990); in Study 2, the Wechsler Abbreviated Scale of Intelligence (WASI; Psychological Corporation, 1999; two-test format) and the Wide Range Assessment of Memory and Learning, 2nd ed. (WRAML-2; Sheslow & Adams, 2003). Study 3 assessed educational attainment by way of a written questionnaire. Neuropsychological testing and questionnaire completion was done in hospital settings or patients’ homes. Results: None of the findings indicate an adverse effect of short-term prenatal DEX exposure. However, a small sample of DEX-exposed girls with CAH and long-term DEX exposure showed lower mean scores than controls on two of eight neuropsychological subtests, K-ABC Sequential Processing and K-ABC Mental Processing Composite (which includes the former) in contrast to the short-term exposure group (DEX-by- CAH interaction p≤.05). Yet, partial correlations of DEX-exposure duration and cognitive outcome did not corroborate this finding. Conclusions: The finding for DEX-exposed girls with CAH cannot be attributed with certainty to the prenatal DEX exposure, but needs to be followed up by examination of additional samples. Acknowledgments: This work was supported in part by USPHS grants R01 HD00072-33A, U54 RR019484, and P30 MH43520
20 – 22 May 2011, Lübeck, Germany 75
P-34 THE PROS AND CONS OF A THIRD GENDER: ATTITUDES AMONGST INTERSEX ADULTS WITH COMPLETE (CAIS) OR PARTIAL (PAIS) ANDROGEN INSENSITIVITY SYNDROME
# C. Handford1, K. Schweizer1, F. Brunner1, H. Richter-Appelt1 1Universitätsklinikum Hamburg-Eppendorf, Institut für Sexualforschung und Forensische Psychiatrie, Hamburg, Germany
Introduction: People with Complete (CAIS) or Partial (PAIS) Androgen Insensitivity Syndrome have a chromosomal karyotype of 46,XY. Because of an androgen receptor defect the outer appearance of the genitals in CAIS is female whereas female internal organs are absent and undifferentiated gonads are present. In the case of PAIS, appearance varies according to the level of virilization. Objectives: Standard treatment at present consists of assigning a male or female sex as early as possible and bringing the appearance of the body as far into line with this sex as is feasible by means of surgery and hormone treatment. However, in the light of the inadequacy of the binary perception of sex for describing people with intersexuality, the question is considered as to whether the current treatment approach and 2-gender option are appropriate or sufficent. The attitudes of 25 people with CAIS or PAIS were investigated with regard to whether the introduction of a third gender would make sense to them. In addition, the participants’ level of satisfaction with their assigned gender was examined in relation to their attitudes towards the issue of a third gender. Methods: Data were collected from 13 people with CAIS and 12 people with PAIS as part of the Hamburg Intersex Study by means of the following two items: Open question with binomial answer category: We would like to ask your opinion on whether introducing a third gender would make sense to you (Yes/no; space for comment) Item with 5-point scale and space for comments: How satisfied are you today with your assigned gender? (very, fairly, more or less, not very, not at all) A qualitative content analysis approach was used to analyse the open question responses. Thematic categories were formulated and illustrated by means of quotes. Results: The following attitude categories could be identified: Supporting a Gender Definition Beyond the Sexual Binary Social Acceptance: Positive Effect Social Acceptance: Negative Effect Conclusions: CAIS: Despite personal satisfaction with assigned gender, the majority of participants nevertheless criticise the sexual binary. PAIS: Overall responses were more heterogeneous, which was consistent with the more heterogeneous nature of the group.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
76 3rd International Symposium on Disorders of Sex Development
P-35 MULTIDIMENSIONAL BODY IMAGE AND SEXUAL FUNCTIONING COMPARISONS AMONG PATIENTS WITH DISORDERS OF SEX DEVELOPMENT (DSD) AND CONTROLS: A MULTISITE STUDY
# N. Callens1,2, K.P. Wolffenbuttel3, Y.G. Van der Zwan1, S.L.S. Drop1, M. Cools2, C. Beerendonk4, M. Van Kuyk5, M. Van den Berg6, D. Study Group on DSD7,8,9,6,5, A.B. Dessens1 1Erasmus Medical Centre- Sophia, Pediatric Endocrinology, Rotterdam, Netherlands 2Ghent University and University Hospital Ghent, Pediatric Endocrinology, Ghent, Belgium 3Erasmus Medical Centre - Sophia, Urology, Rotterdam, Netherlands 4University Medical Centre St Radboud, Gynecology, Nijmegen, Netherlands 5University Medical Centre St Radboud, Nijmegen, Netherlands 6Erasmus Medical Centre- Sophia, Rotterdam, Netherlands 7VUMC, Amsterdam, Netherlands 8UMCU, Utrecht, Netherlands 9UMCG, Groningen, Netherlands
Objectives: Having a DSD has extensive clinical and psychosocial implications. Clinical management often implies early gender assignment followed by surgical and endocrine treatment in order to correct or to prevent an ambiguous physical appearance and/or to facilitate sexual intercourse. The pressure to conform to a narrowly defined body ideal, and in some cases, the failure to attain this ideal, might be related to a negative body image and associated emotional problems (such as depression). Although body image dissatisfaction is not considered a core characteristic of DSD, it might be of overwhelming importance in its impact on quality of life, sexual well-being and adaptation to living with this condition. This study examined the multidimensional nature of body image functioning and its effect on sexuality among individuals with DSD, relative to female and male controls. Methods: Participants (n = 91 females, 9 males), aged 14- 48, were recruited from two treatment centers in the Netherlands and completed psychometrically validated and standardized self-report measures of body image (BIS) and sexual functioning (FSFI, FSDS-R, MSHQ, KSOG). A semi-structured interview with additional questions related to sexuality and coping with DSD and a gynecological/urological examination were also performed. Reference and control data were available from a student survey. Results: Overall, the DSD groups were characterized by elevated disturbances in most sex-related body image dimensions relative to their gender-matched controls. Male DSD patients reported more body image impairment than female patients with reference to primary sex characteristics (such as penis and vagina) but not secondary sex characteristics (such as facial hair or voice). Male patients with DSD were more likely to be exclusively heterosexual than their female counterparts. However, no overall difference on the global measure of body dissatisfaction between heterosexual and non-heterosexual individuals with DSD was found. Data on the FSDS-R demonstrated that females with DSD experienced more sexual distress, whereas this was not the case in males. Findings on the FSFI and MSHQ indicated that sexual function was impaired, but no association between body dissatisfaction and prevalence of sexual difficulties was apparent. Conclusions: DSD is characterized by body image dissatisfaction, especially with respect to sex-related body dimensions. However, it is not clear how cognitive, behavioral, and emotional elements of body image functioning in this specific population interact. Interdisciplinary health care teams should focus on the pressure of conformity and openly discuss it with the patients in context of treatment decisions. Furthermore, individually tailored counseling concerning sexuality and coping with the condition in daily life is needed to optimize psychosexual and social comfort.
20 – 22 May 2011, Lübeck, Germany 77
P-36 USE OF GNRH ANALOGUES IN THE MANAGEMENT OF DISORDERS OF SEX DEVELOPMENT DIAGNOSED AT PUBERTY
# S.L.C. Meroni1, A. Di Lascio1, M. Ferrario2, G. Russo1, G. Chiumello1 1San Raffaele Hospital, Pediatrics, Milan, Italy 2Sant'Anna Hospital, Pediatrics, Como, Italy
Introduction: The management of a newly presenting adolescent with DSD requires a holistic management from the outset and an appropriately trained multidisciplinary team. Decision on gender assignment should only be made after a thoughtful balancing of medical and psychosocial pros and cons and is based on several factors, such as diagnosis, genital appearance, surgical options, need for life-long replacement therapy, potential for fertility, views of the family and, sometimes, circumstances relating to cultural practices. Each adolescent should receive counseling by an experienced psychologist, specialized in gender identity, who must act as soon as the diagnosis is suspected and then follow the adolescent periodically. The knowledge of the disease and its consequences should be given incrementally in an age-appropriate manner. Case report: We describe an XY individuals affected by 17β-HSD3 deficiency and raised as female. She had no significant medical or surgical history, with the exception of unilateral inguinal hernia surgery in infancy. She was evaluated for the first time at puberty because of progressive virilization. The diagnosis was substantiated by clinical presentation, hormonal profile and karyotype and was confirmed by molecular analysis of 17β-HSD3 gene. The dilemma of sex reassignment was seriously considered. We proposed to block temporarily gonadic hormonal secretion in order to prevent gender identity disorder and to provide time to confirm or change the birth sex assignment. The suppression of sex hormone secretion is achieved by using a long-acting GnRH analogue. The final decision and the resulting future therapeutic management were taken only after an adequate and extensive psychological evaluation with numerous interviews with the adolescent and her parents and in consultation among the paediatric endocrinologist, the surgeon and the psychologist. The overall conclusion was that the patient had a clear female identity and she should be helped to retain female gender. Discussion: Adolescents with DSD, who were misdiagnosed or late diagnosed, create special challenges to their healthcare providers. The choice of sex reassignment should be done with extreme caution, after a thoughtful balancing of pros and cons, and in consultation among the paediatric endocrinologist, the surgeon and the psychologist. Treatment with GnRH analogue can be recommended in individuals with DSD, who are diagnosed at puberty, in order to prevent gender identity disorder and to provide time to confirm or change the birth sex assignment. During this period of hormonal suppression, the adolescent and his/her family have to receive counselling by an experienced psychologist. The final decision about sex assignment is made after a complete medical and psychological evaluation of the adolescent, together with his/her family, in order to establish gender identity.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
78 3rd International Symposium on Disorders of Sex Development
P-37 CLINICAL HORMONAL AND CHROMOSOMAL ANALYSIS OF UNDERVIRILIZED MALE/46XYDSD - A 3YEARS EXPERIENCE OF NATIONAL INSTITUTE OF CHILD HEALTH
# I. Tufail1, S. Lone1, Y.N. Khan1, M. Ibrahim1, J. Raza1 1National Institute of Child Health, Pediatric Medicine, Karachi, Pakistan
Objective: To do the clinical, hormonal and chromosomal analysis in undervirilized male / 46XY DSD. To make a Presumptive diagnosis on the basis of clinical, chromosomal and hormonal assessment. Methodology: This study was conducted in National Institute of Child Health at Department of Pediatrics, Division of Endocrinology from January 2008 to December 2010. A Total of 127 Patient under age of 14 years with ambiguity, micropenis, hypospadias, cryptorchism and delayed puberty were selected and studied.USG Pelvis, HCG Stimulation test and Chromosomal analysis were carried out in all patients. Two types of HCG stimulation test were performed. Short HCG was done in children ten and less than ten years of age. Prolong HCG was performed in children more than ten years of age. Laproscopy and biopsy were carried out in patients who had mullerian duct structure on USG and also in patients with no gonads. FISH analysis was done in patients who were 46XX karyotype with testes. Result: Total no. of patients were 127. 43% presented with hypospadias, 17% with ambiguity, 20% with cryptorchism, 13% with micropenis and 5 % with delayed puberty.HCG stimulation showed high response (pre and post- testosterone) in 29%, flat in 28%, partial in 27% and normal in 16% of patients. On chromosomal analysis 123 (97%) patients were turned out to be 46XY, 3(2%) patients were 46XX and 1(1%) patient was 46XXY. FISH analysis performed in 46 XX patients showed Y translocation in one patient. 8(6%) 46 XY DSD patients had both wolffian and mullerian duct structure on ultrasonography. Laproscopy and biopsy performed in 4(3%) patients and proved ovotesticular DSD on histopathology. Laproscopy was also done in 2(1.5%) 46 XY patients with no gonads on ultrasonography and diagnosed as a case of testicular regression syndrome on per-operative findings .The diagnosis of Gonadal dysgenesis considered in patients who have partial testosterone response , Androgen Insensitivity in high testosterone response and testicular biosynthetic defect in flat pre and post testosterone response to HCG. Conclusion: Phenotypic presentation of 46XY DSD depends on the underlying defects. Defect in androgen action on the target tissues or production of active metabolite share common morphological features. Molecular study may help in differentiating these abnormalities and to make a final diagnosis.
20 – 22 May 2011, Lübeck, Germany 79
P-38 THE EUROPEAN DSD REGISTER - A PLATFORM FOR INTERNATIONAL COLLABORATIVE RESEARCH
# M. Rodie1, J. Jiang2, R. Sinnott2, F. Ahmed1 1Royal Hospital for Sick Children, Department of Child Health, Glasgow, United Kingdom 2University of Glasgow, National e-Science Centre, Glasgow, United Kingdom
Effective research into understanding the aetiology of DSD, as well as long-term outcome of these rare conditions, requires multicentre collaboration often across national boundaries. The EU-funded EuroDSD programme (www.eurodsd.eu) is one such collaboration involving clinical centres and clinical and genetic experts. At the heart of the EuroDSD collaboration is a DSD Register that supports the sharing of DSD data. Over the past 12 months the DSD Register has remained central to the EuroDSD project. In addition, the number of individual users has increased by approximately 30% with 23 new users from 14 new centres in 7 new countries and in 1 new continent. At last review (February, 2011) there were 847 cases on the Register from 12 countries. The United Kingdom has the largest number of cases on the Register currently (n=248), followed by the Netherlands (n=179). The age of first presentation ranges from <1month to 53years and the median year of birth was 1995 (range 1927-2010). 58% (n=489) cases were assigned female sex and 42% (n=358) were assigned male sex. There was a history of infertility or parental consanguinity in 8% (n=68) and 11% (n=89) cases respectively, and associated malformations were present in 27% (n=227) cases. The commonest disorder type on the Register continues to be disorders of androgen action (n=254), followed by disorders of gonadal development (n=194). The majority of cases had a 46XY karyotype (n=629), followed by 46XX (n=149). Mosaicism was present in 6% (n=51) cases. Samples are available in 41% (n=351) cases. The Register has attracted much interest internationally over the last year and is changing from a European initiative to an international activity. It provides a virtual research environment within which clinicians and investigators can interact and develop new DSD related studies.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
80 3rd International Symposium on Disorders of Sex Development
P-39 46XY AND FEMALE PHENOTYPE FETUS: A CHALLENGING DIAGNOSIS
# A. Di Lascio1, S.L.C. Meroni1, M. Ferrario2, G. Russo1, O. Hiort3, G. Chiumello1 1San Raffaele Hospital, Pediatrics, Milan, Italy 2Sant'Anna Hospital, Pediatrics, Como, Italy 3Campus Lübeck, Pediatrics, Lübeck, Germany
Introduction: Disorders of sex development (DSD) are congenital conditions in which the development of chromosomal, gonadal or anatomical sex is atypical. The 46,XY DSD may be caused by a defective synthesis or action of androgen hormones, leading to incomplete or absent virilization of the external genitalia. Clinical presentation in these cases is variable: some are diagnosed for ambiguous external genitalia at birth, others during evaluation of inguinal hernia in females during childhood, others for primary amenorrhea and/or virilization in girls at pubertal age. Case presentation: We present the case of a woman who underwent amniocentesis with fetal karyotype 46,XY, discordant with ultrasound appearance of female external genitalia. Prenatal analysis for 5α-reductase (SRD5A2) and androgen receptor (AR) gene resulted negative. At birth the infant presented normal female external genitalia, 46,XY karyotype and absence of uterus and ovaries at pelvic ultrasound. The newborn was assigned female sex. At four months of age, an HCG test was performed, which showed hormonal values suggestive of 17βHSD3 deficiency (testosterone/ Δ4-androstenedione ratio = 0.22 with normal values>0.8- 0.9), confirmed by HSD17B3 gene analysis. Discussion and conclusion: In the absence of anamnestic data, 46,XY patients with female phenotype are often unrecognized at birth and therefore female sex is assigned to them. In these cases diagnosis may be delayed until puberty for primary amenorrhea and/or spontaneous virilization. In some cases diagnosis can be made in the first months of life or during childhood for the occurrence of inguinal hernias. The increasing use of prenatal investigations makes more frequent the possibility to suspect disorders of sexual development during the fetal period. Complete androgen insensitivity syndrome (CAIS) represents the most common cause of female phenotype and 46,XY karyotype, but several other rarer conditions may have a similar presentation. In our case hormonal and genetic investigations showed a 17βHSD3 deficiency. Specific genetic investigations for these diseases should be performed in the prenatal period when possible. Subsequently, evaluation of clinical phenotype, abdominal ultrasound, karyotype and hormonal tests to evaluate gonadal and adrenal function should be performed after delivery. Early identification of DSD in the prenatal period may be a useful tool to detect diseases that might otherwise remain unidentified until puberty, and to eventually enable both the patients and their parents to be more conscious about the disease.
20 – 22 May 2011, Lübeck, Germany 81
P-40 CONCOMITANT GONADAL DYSGENESIS AND NON-CLASSIC CONGENITAL ADRENAL HYPERPLASIA IN A GIRL WITH 46,XY DSD
# M. Szarras-Czapnik1, A. Malinowska1, A. Lecka-Ambroziak1, E. Malunowicz2, M. Baka-Ostrowska3, J. Slowikowska-Hilczer4 1The Children's Memorial Health Institute, Dpt. of Endocrinology and Diabetology, Warsaw, Poland 2The Children's Memorial Health Institute, Dpt. of Biochemistry and Experimental Medicine, Warsaw, Poland 3The Children's Memorial Health Institute, Dpt. of Paediatric Urology, Warsaw, Poland 4Medical University of Lodz, Dpt. of Andrology and Reproductive Endocrinology, Lodz, Poland
Objective: Differential diagnosis of congenital clitoromegaly includes virilisation of female foetus or insufficient masculinisation of male foetus which result from disorders in steroidogenesis or testes dysfunction. The most common reason is congenital adrenal hyperplasia (CAH: 21-hydroxylase deficiency), other reasons are much less common. Case report: A preterm baby with clitoromegaly assessed as a sign of prematurity. The child's growth and development was normal. In 18 month of life the girl was hospitalized because of diarrhoea, without symptoms of salt losing syndrome. Diagnostic tests for clitoromegaly (Prader's I) in the girl with normal female phenotype were planned. Mild signs of 21-hydroxylase deficiency were present in steroid profile. Treatment with hydrocortisone (15 mg/m2) was started and stopped after 3 months due to a result of karyotype examination which was inconsistent with clinical picture (46, XY). According to increased level of 17-OHP in a Synacthen test (3450 → 11300 ng/dl) and results of molecular tests - compound heterozygous CYP21 mutations: g.655A/C>G, c.89C>T (p.P30L), c.842G>T (p.V281L) the diagnosis of CAH (non-classical form) was sustained. In spite of slightly decreased cortisol reserve (18.2 → 21.7 μg/dl) and due to normal levels of ACTH (46 pg/ml) and androstenedione (23.7 ng/dl) the hydrocortisone treatment was required only during stress. Further diagnostic tests showed: no acceleration of bone age, increased gonadotropin levels (LH: 6.7 IU/l, FSH: 53 IU/l), moderate rise of serum testosterone level in hCG test (0.052 → 0.423 ng/ml), presence of uterus and no visible gonads in an ultrasound examination. A diagnosis of 46,XY gonadal dysgenesis was suspected. Bilateral gonadectomy was performed at the age of 5 yr. Histological examination revealed a streak of connective tissue without testicular or ovarian structures on the right side and a streak gonad with two small nests of gonadoblastoma in the cortical part on the left side. Cross-sections of ovarian tubes und immature epididymal ducts were found. At the age of 6 yr growth velocity and bone age were advanced, glucocorticoid deficiency was detected by the Synacthen test (no reserve of cortisol 10.8 → 13.0 μg/dl), levels of 17-OHP (1680 ng/dl), androstendione (114 ng/dl) and DHEA-S (1785 ng/dl) were elevated. Hydrocortisone therapy (10 mg/m2) was introduced and 17-OHP (252 ng/dl) and androstedione (44.7 ng/dl) levels normalized. Conclusions: -clitoromegaly after birth in a phenotypic girl may be a symptom of not only CAH, but also gonadal dysgenesis; -presented child was diagnosed with both gonadal dysgenesis and non-classical form of CAH; -even mild genital virilisation in a girl should be an indication to perform a karyotype examination; -this observation highlights the importance to assess both gonadal and adrenal function in patients with 46,XY DSD.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
82 3rd International Symposium on Disorders of Sex Development
P-41 TRANSVERSE TESTICULAR ECTOPIA, PERSISTENT MÜLLERIAN DUCT AND BILATERAL DUPLICATION OF THE VAS DEFERENS. EMBRYOLOGY, PRESENTATION AND SURGICAL MANAGEMENT
# H. Özbey1, C. Münevveroglu1 1Istanbul University, Pediatric Surgery & Pediatric Urology, Istanbul, Turkey
Transverse testicular ectopia (TTE) is a rare congenital abnormality of testicular maldescent and is strongly associated with persistent Müllerian duct syndrome. A 6 month-old boy with transverse testicular ectopia, persistent Müllerian duct and bilateral duplication of the vas deferens, is reported. Treatment included partial excision of the Müllerian remnant with duplicated vas deferens through a right inguinal exploration, orchidopexy of the right and left (via transseptal route) testis. The embryology, presentation and surgical management of this rare malformation are discussed.
20 – 22 May 2011, Lübeck, Germany 83
P-42 BILATERAL TESTICULAR REGRESSION SYNDROME: REPORT OF A 2 YEAR-OLD BOY
# H. Özbey1, B. Erginel1, T. Günendi1, G. Kılıç2 1Istanbul University, Pediatric Surgery & Pediatric Urology, Istanbul, Turkey 2Ekin Pathology Laboratory, Istanbul, Turkey
Bilateral anorchia (vanishing testis syndrome or testicular regression syndrome) is defined as the absence of testicular tissue in 46, XY individuals with male phenotype. Accurate diagnosis requires surgical exploration, specific laboratory and histopathological findings. Extremely low or undetectable anti-Müllerian hormone (AMH) levels and complete absence of plasma testosterone in response to human chorionic gonadotropin (hCG) stimulation are needed. In addition, the finding of dystrophic calcification and haemosiderin deposition, with no evidence of viable testicular tissue in the presence of normal spermatic cord elements, supports the concept of testicular regression. Here we report a 2 year-old boy with normal penile development associated with bilateral testicular regression syndrome. The embryology, presentation and surgical management of this rare malformation are discussed.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
84 3rd International Symposium on Disorders of Sex Development
P-43 PATTERN OF DSD AND GENITAL ANOMALIES: EGYPTIAN EXPERIENCE
# I. Mazen1 1National Research Center, Clinical Genetic, Cairo, Egypt
Background: Disorders of sex development with birth of an infant with ambiguous genitalia require medical attention to elucidate the differential diagnosis. This group of disorders is not uncommon in Egypt (1:5000 live births). Aim: We want to provide an extensive review and pattern of the molecular study of patient collective with various types of disorders of sex development in Egypt. The DSD patients were recruited from the Department of Clinical Genetics at the National Research Center. Moreover we shall describe the first pilot study which was conducted in great Cairo and Kalioubia governerates in 2007 showing the relatively high incidence of disorder of sexual development (DSD) and genital abnormalities Subjects and Methods: 20,000 newborns and infants up to the age of 6 months coming for compulsory vaccination at primary health care units and centers in Great Cairo and Qalyubiyah governorates were examined in the years 2006-2007 for suspected genital anomalies. Results: There were 187 (93.5/10,000) cases with external genital anomalies among the screened 20,000 participants. Various abnormalities in the form of 46,XY DSD, undescended testis, hydrocele, hypospadias, micropenis, synechia of the labia and other genital anomalies were diagnosed and classified after thorough clinical examination, and hormonal, radiological, and laparoscopic investigations. Conclusion: This first pilot study in Great Cairo and Qalyubiyah governorates showed a relatively high incidence of genital anomalies and DSD. Therefore, we recommend more studies including larger population sizes to detect the actual incidence of genital anomalies and DSD in Egypt in order to serve those patients and their families. Furthermore, Disorders of sex development have a broad range of underlying causes in Egypt with some preference of rare monogenic disorders.
20 – 22 May 2011, Lübeck, Germany 85
P-44 FERTILITY AND PREGNANCY OUTCOME IN WOMEN WITH CONGENITAL ADRENAL HYPERPLASIA DUE TO 21-HYDROXYLASE DEFICIENCY
# A. Nordenskjold1, G. Holmdahl2, K. Hagenfeldt1, P.O. Janson3, H. Falhammar4, H. Filipsson5, L. Frisén6, M. Thorén4, 1Karolinska Institutet, Women and Children Health, Stockholm, Sweden 2Sahlgrenska Academy, Pediatric Surgery, Gothenburg, Sweden 3Sahlgrenska Academy, Department of Obstetrics and Gynaecology, Gothenburg, Sweden 4Karolinska Institutet, Molecular Medicine and Surgery, Stockholm, Sweden 5Sahlgrenska Academy, Endocrinology, Gothenburg, Sweden 6Karolinska Institutet, Department of Clinical Sciences, Division of Psychiatry, Stockholm, Sweden
Objectives: Low pregnancy rate has been reported in women with congenital adrenal hyperplasia (CAH) and little information on pregnancy and children is known. Methods: In a Swedish study, 62 adult women with CAH, aged 18-63 years, and 62 age-matched controls were followed-up. Medical records, including those concerning pregnancies and deliveries, were examined and the 21-hydroxylase genotype of patients was noted. All women answered a questionnaire concerning sexual and reproductive health including health of the children. Results: Pregnancy and delivery rates were significantly lower in women with CAH (P < 0.001, P < 0.0056, respectively), and the severity of the 21-hydroxylase-mutation correlated with the reduced number of children born. More women with salt-wasting CAH were single and had not attempted pregnancy. Pregnancies were normal except for a significantly increased incidence of gestational diabetes in CAH patients (P < 0.0024). The children had normal birthweight and no malformations were observed. A later follow-up of the children showed a normal intellectual and social development. The sex ratio of the offspring differed significantly, with 25% boys in the CAH group compared with 56% among controls (P < 0.016). CAH women had more gynaecological morbidity during menopause. Conclusions: Pregnancy and delivery rates are reduced in women with CAH mainly due to psychosocial reasons. The outcome of children did not differ from controls. The unexpected sex ratio in children born to mothers with CAH warrants further research.
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
86 3rd International Symposium on Disorders of Sex Development
Maps
Congress Venue: Universität zu Lübeck Ratzeburger Allee 160, 23562 Lübeck Audimax, Building 65
From Luebeck main station take bus 19 (Ö Hochschulstadt) or bus 9 (Ö Grillenweg), stop: Fachhochschule.
Universität zu Lübeck, Audimax Congress venue: Parking available along Mönkhofer Weg
g Parkin H
Bus 9 and 19 : Fachhochschule p Sto
20 – 22 May 2011, Lübeck, Germany 87
Scientific and technical exhibition – Audimax
Poster Presentation Audimax 3 Lilly Deutschland GmbH Lilly Deutschland GmbH Sandoz Pharmaceuticals 3 4
Ipsen Pharma GmbH Ipsen Pharma Novo Nordisk Pharma GmbH Pharma GmbH Novo Nordisk Audimax Foyer 1 2 Exhibition of companies
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu
88 3rd International Symposium on Disorders of Sex Development
Social Evening: Restaurant: Die Gemeinnützige, Restaurant Heinrichs Königstraße 5-7, 23552 Lübeck
How to get to Restaurant Heinrichs: From Audimax: Bus 9 or bus 19 (direction Bad Schwartau ZOB) from stop „Fachhochschule“ to stop „Königstraße“, go straight ahead for 400 meters, Restaurant Heinrichs is located on the right-hand side
Senator Hotel Senator Radisson Blue Gemeinnützige/ Restaurant Heinrichs Restaurant
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90 3rd International Symposium on Disorders of Sex Development
Imprint
Editor Universität zu Lübeck Ratzeburger Allee 160 D-23562 Lübeck Germany www.uni-luebeck.de Editorial Staff:
Coordinator: Olaf Hiort Klinik für Kinder- und Jugendmedizin Email: [email protected] Internet: www.kinderhormonzentrum-luebeck.uk-sh.de Lutz Wünsch Klinik für Kinderchirurgie Email: [email protected] Internet: www.kinderchirurgie.uni-luebeck.de
Administration: Renate Wagner Klinik für Kinder- und Jugendmedizin Email: [email protected] Internet: www.kinderhormonzentrum-luebeck.uk-sh.de
Conference Office: Kristin Bätzel event lab. GmbH Dufourstraße 15, 04107 Leipzig, Germany www.eventlab.org
Participating Institutions:
Universitätsklinikum Schleswig-Holstein
Klinik für Kinder- und Jugendmedizin
Klinik für Kinderchirurgie
20 – 22 May 2011, Lübeck, Germany 91
Sponsors
We are grateful to our major sponsors
EuroDSD – a European colloborative study on DSD· Funded by FP7 – www.eurodsd.eu