Etiological Diagnosis of Undervirilized Male / XY Disorder of Sex Development

Transcription

1 ORIGINAL ARTICLE Etiological Diagnosis of Undervirilized Male / XY Disorder of Sex Development Irum Atta, Mohsina Ibrahim, Arit Parkash, Saira Waqar Lone, Yasir Naqi Khan and Jamal Raza ABSTRACT Objective: To do clinical, hormonal and chromosomal analysis in undervirilized male / XY disorder of sex development and to make presumptive etiological diagnosis according to the new Disorder of Sex Development (DSD) classification system. Study Design: Case series. Place and Duration of Study: Endocrine Unit at National Institute of Child Health, Karachi, Pakistan, from January 2007 to December Methodology: Patients of suspected XY DSD / undervirilized male visiting endocrine clinic were enrolled in the study. Criteria suggested XY DSD include overt genital ambiguity, apparent female/male genitalia with inguinal/labial mass, apparent male genitalia with unilateral or bilateral non-palpable testes, micropenis and isolated hypospadias or with undescended testis. The older children who had delayed puberty were also evaluated with respect to DSD. As a part of evaluation of XY DSD, abdominopelvic ultrasound, karyotype, hormone measurement (testosterone, FSH, LH), FISH analysis with SRY probing, genitogram, laparoscopy, gonadal biopsy and HCG stimulation test were performed. Frequencies and percentages applied on categorical data whereas mean, median, standard deviation were calculated for continuous data. Results: A total of 187 patients met the criteria of XY DSD. Age ranged from 1 month to 15 years, 55 (29.4%) presented in infancy, 104 (55.6%) between 1 and 10 years and 28 (15%) older than 10 years. Twenty five (13.4%) were raised as female and 162 as (86.6%) male. The main complaints were ambiguous genitalia, unilateral cryptorchidism, bilateral cryptorchidism, micropenis, delayed puberty, hypospadias, female like genitalia with gonads, inguinal mass. The karyotype was 46 XY in 183 (97.9%), 46 XX in 2 (1.1%), 47 XXY in 1 (0.5%), 45 X/46 XY in 1 (0.5%) patient. HCG stimulation test showed low testosterone response in 43 (23 %), high testosterone response in 62 (33.2%), partial testosterone response in 32 (17.1%) and normal testosterone response in 50 (26.7%). Genitogram was carried out in 86 (45.98%) patients. Presumptive etiological diagnosis of androgen sensitivity syndrome/ 5-alpha reductase deficiency, testicular biosynthetic defect/ leydig cell hypoplasia, partial gonadal dysgenesis, ovotesticular DSD, XX testicular DSD, mixed gonadal dysgenesis, testicular vanishing syndrome, klinefelter syndrome, hypogonadotropic hypogonadism, isolated hypospadias and isolated micropenis was made. Conclusion: Clinical, chromosomal and hormonal assessment may help in making the presumptive etiological diagnosis. Further molecular genetics analysis are needed in differentiating these abnormalities and to make a final diagnosis. Key Words: Undervirilized male. Disorder of sex development. HCG stimulation test. Testosterone. INTRODUCTION Department of Paediatric Medicine, Unit II, National Institute of Child Health. Karachi. Correspondence: Dr. Irum Atta, A-14, Younus Terrace, Block 13-B, Gulshan-e-Iqbal, Karachi. irumatta@yahoo.com Received: February 20, 2013; Accepted: June 06, Disorders of Sex Development (DSD) defined as a condition in which chromosomal sex is inconsistent with phenotypic sex, or in which the phenotype is not classifiable as either male or female. 1 Human sex development is a highly complex process controlled by multiple genes and hormones. Sexual differentiation follows a sequence of events with predictable development of the gonads, internal genital ducts and the external genitalia. Abnormalities of sexual differentiation may occur at any step along the way. 2 Abnormal differentiation of gonads, internal genital ducts, or the external genitalia produce predictable clinical syndromes. Many defects of sexual differen-tiation are evident at birth, others will not be identified until puberty, at which time the patient may manifest aberrant external maturation or may remain sexually infantile. 3 The new Disorders of Sex Development (DSD) classification includes three main diagnostic categories: sex chromosome DSD, 46, XY DSD (formerly male pseudohermaphroditism / undervirilized male) and 46, XX DSD (formerly female pseudohermaphroditism). The category of sex chromosome DSD embraces not only ovotesticular DSD (formerly true hermaphroditism) and 45, X/46, XY mixed gonadal dysgenesis, but also Turner Syndrome (TS) and Klinefelter syndrome (KS), which are not included in the previous classifications of intersex disorders. 4,5 Because the consensus has taken the karyotype as the primary root, ovotesticular DSD has been classified in three DSD categories - XX, XY and XX/XY. The new classification also includes disorders such as vanishing testes syndrome and TS, which are not associated with genital ambiguity at birth. 714 Journal of the College of Physicians and Surgeons Pakistan 2014, Vol. 24 (10):

2 Etiological diagnosis of undervirilized male Despite the multiple genetic causes of 46, XY DSD, around 30-40% of cases remain without diagnosis. 6 Currently, there is a frequent, non-genetic variant of 46, XY DSD characterized by reduced prenatal growth and lack of clear evidence for any associated malformation or steroidogenic defect. Additionally, other studies in undetermined 46, XY DSD report that around 30% of cases are associated with low birth weight, indicating that adverse events in early pregnancy are frequent causes of congenital non-genetic 46, XY DSD. 7,8 The XY DSD is a treatable pediatric endocrine disorder; there is sparse local published data on this subject in Pakistan. This study in hand may provide baseline data which may help to guide paediatricians and family physicians for initial evaluation of XY DSD. This study may also provide data for future studies. The objective was to do clinical, hormonal and chromosomal analysis in undervirilized male/xy DSD and to make presumptive etiological diagnosis according to the new Disorder of Sex Development (DSD) classification system in children presenting at National Institute of Child Health, Karachi. METHODOLOGY This research was approved by local ethical committee of National Institute of Child Health. Written informed consent was obtained from the parents before participation. A total of 187 patients of suspected undervirilized male/ XY DSD visiting endocrine clinic between January 1, 2007 and December 31, 2012 were enrolled in the study. Age range was one month to 15 years. Criteria suggesting XY DSD included overt genital ambiguity (any case in which the external genitalia do not appear completely male or completely female), apparent female/male genitalia with inguinal/labial mass, apparent male genitalia with unilateral or bilateral non-palpable testes, micropenis (defined as a penis that is normal in terms of shape and function, but more than 2.5 Standard Deviation (SD) smaller than mean size in terms of length) 9 and isolated hypospadias or with undescended testis. 3 The older children who had delayed puberty, defined as failure of development of secondary sexual characteristics beyond 14 years of age in boys, were evaluated with respect to DSD. Patients with 46 XX DSD (except 46 XX testicular DSD) and 46 XX ovotesticular DSD were excluded. A history including age at presentation, main complaints and rearing gender was taken from each patient. A thorough clinical examination consisting of height, weight, blood pressure, presence of hyperpigmentation, assessment of pubertal stage, penile length, associated anomalies or dysmorphic features was made and recorded for each patient. Measurements of penile length were taken with a rigid rule, graded millimeters. As a part of evaluation of XY, DSD, we performed abdominopelvic ultrasound, karyotype and hormone measurement (testosterone, FSH, LH). We did FISH analysis with SRY probing in children with phenotypically male genitalia and had 46 XX karyotype on work-up. We proceeded for genitogram (to display the type of urethra, presence of vagina, cervix and urogenital sinus) before laparoscopy and gonadal biopsy in patients with cryptorchidism, ambiguity and in patients who had mullerian duct structure on ultrasonography. To demonstrate the presence and absence of functioning testicular tissue, HCG stimulation test was performed following the Basildon and Thurrock University Hospitals protocol. A response was defined as a rise in testosterone levels of 20 ng/ml. If the preand post-hcg testosterone level was lower than 20 ng/ml, there was a defect in testosterone biosynthesis or Leydig cell hypoplasia. A significant rise in serum testosterone level before and after HCG was in favor of androgen insensitivity syndrome or 5-alpha reductase deficiency. If the pre-hcg testosterone level was low and post-hcg response between 20 to 100 ng/ml, this was partial response of gonadal dysgenesis. If pretestosterone was low and post-testosterone > 100 ng/ml, it was considered a normal response. 11 deoxycortisol (11-DOC), dehydroepiandrosterone sulphate (DHEA-S), androstenedione, dihydrotestosterone (DHT), antimullerian hormone (AMH) and Inhibin B and molecular genetic testing related to specific mutations were not performed, as these test facilities are not available in Pakistan. Vanishing testes syndrome was defined as normal virilized external genitalia with non-palpable gonads, no testosterone response to HCG and absent testis with blind ending cord structure on laparoscopy. 10,11 Persistent Mullerian Duct Syndrome (PMDS) was labeled in normal male external genitalia with inguinal mass, normal response to HCG and presence of mullerian duct structure detected by USG and laparoscopy. 10 Androgen insensitivity syndrome (AIS) was diagnosed in undervirilized male who had high testosterone response with HCG and absence of mullerian duct structure on USG. Those with normal female external genitalia were considered as complete AIS (CAIS) and rest as partial AIS (PAIS) or 5-alpha reductase 2 deficiency. 10,12 Testicular biosynthesis defect / leydig cell hypoplasia was labeled in children with female phenotype, male phenotype with micropenis and or hypospadias, cryptorchidism, ambiguous genitalia with absent mullerian duct structure and subnormal response on HCG stimulation test. 11 Partial gonadal dysgenesis was diagnosed in patients who presented with ambiguity, micropenis, hypospadias, with or without mullerian duct structure with streak gonads detected by USG/ laparoscopy and had positive HCG response. 6,11 Mixed Journal of the College of Physicians and Surgeons Pakistan 2014, Vol. 24 (10):

3 Irum Atta, Mohsina Ibrahim, Arit Parkash, Saira Waqar Lone, Yasir Naqi Khan and Jamal Raza gonadal dysgenesis was defined in children presented with ambiguity, hypospadias, cryptorchidism and had XY/XO karyotyping. 8,11 46 XY ovotesticular DSD was defined as patient with ambiguous genitalia to almost normal male genitalia with ovotestis +testis/ovotestis +ovary/bilateral ovotestis/ovary+testis on gonadal biopsy. 8,11 46 XX testicular DSD was labeled in phenotypic male genitalia with hypospadias, had normal HCG response with 46 XX karyotyping and on FISH analysis with SRY probing was 46 XX SRY+. 11 Idiopathic micropenis was diagnosed in children of normal male genitalia with micropenis and had normal FSH, LH and HCG response. 11,14 Isolated hypospadias considered in children with normal male genitalia with hypospadias and had normal HCG response. 6 Hypogonadotropic hypogonadism was labeled in children who present with delayed puberty and have infantile genitalia, micropenis with no HCG response and low FSH and LH. 11 Data was analyzed by using Statistical Package for Social Sciences (SPSS) version 17. Frequencies and percentages were determined for categorical data whereas mean, median and standard deviation was calculated for continuous data. RESULTS A total of 187 patients met the criteria of XY DSD. There was wide variation in age at presentation ranging from 1 month to 15 years with a mean age of 4.71 ± 4.65 years. Age distribution is mentioned in Table I. At the time of presentation, 25 (13.4%) patients were raised as female and 162 (86.6%) as male. The height mean was ± cm and mean weight was ± kg. The median height SDS was and median weight SDS was The main complaints in XY DSD patients were ambiguous genitalia, unilateral cryptorchidism, bilateral cryptorchidism, micropenis, delayed puberty, hypospadias, female like genitalia with gonads, inguinal mass as shown in Table II. The karyotype was 46 XY in 183 (97.9%), 46 XX in 2 (1.1%), 47 XXY in 1 (0.5%) and 45 X/46 XY in 1 (0.5%) patient. HCG stimulation test performed in 187 patients showed low testosterone response in 43 (23%), high testosterone response in 62 (33.2%), partial testosterone response in 32 (17.1%) and normal testosterone response in 50 (26.7%). Genitogram was carried out in 86 (45.98%) patients. Out of 86, 50 (58.1%) patients had male urethra, 9 (10.46%) had female like urethra and 27 (31.3%) had male urethra with mullerian duct remnant. Laparoscopy performed in 35 (18.71%) patients with ambiguity and cryptorchidism. Mullerian duct remnant, gonads, vas vessels were identified and biopsy taken in 20 (10.69%) patients. Histological examination revealed ovotesticular tissue in 5 (25%), dysgenetic gonads in 9 (45%) and immature testicular tissue in 6 (30%) patients. No evidence of malignancy was detected on histopathology. Presumptive diagnosis of androgen sensitivity syndrome/ 5-alpha reductase deficiency, testicular biosynthetic defect/ Leydig cell hypoplasia, partial gonadal dysgenesis, Table I: Age distribution. Age Frequency Percentage < 1 year % 1-10 years % > 10 years % Table II: Presenting complaints of XY DSD children. Presenting complaint Frequency Percentage Ambiguous genitalia % Unilateral cryptorchidism 7 3.7% Bilateral cryptorchidism % Micropenis % Delayed puberty 3 1.6% Hypospadias % Female like genitalia with gonads % Inguinal mass 2 1.1% Table III: Etiological diagnosis of XY DSD. Etiological diagnosis Frequency Percentage Isolated hypospadias % Partial gonadal dysgenesis % Vanishing testes syndrome 3 1.6% 46 XX testicular DSD 2 1.1% Complete androgen insensitivity syndrome 9 4.8% Hypogonadotropic hypogonadism 3 1.6% Isolated micropenis % Klinefelter's syndrome 1 0.5% Mixed gonadal dysgenesis 1 0.5% Ovotesticular DSD 4 2.1% Partial androgen insensitivity syndrome / 5 alpha reductase deficiency % Persistent mullerian duct syndrome 2 1.1% Testicular biosynthetic defect/ leydig cell hypoplasia % Figure 1: Presenting complaints of patients with XY DSD according to etiological diagnosis. 716 Journal of the College of Physicians and Surgeons Pakistan 2014, Vol. 24 (10):

4 Etiological diagnosis of undervirilized male ovotesticular DSD, XX testicular DSD, mixed gonadal dysgenesis, testicular vanishing syndrome, Klinefelter's syndrome, hypogonadotropic hypogonadism, isolated hypospadias and isolated micropenis was made as shown in Table III. Figure 1 shows the distribution of complaints with different etiological diagnosis. DISCUSSION The Disorders of Sex Development (DSD) with sex ambiguity and 46 XY karyotype can be classified in three main groups: (1) disorders of gonadal development (ovotesticular DSD and partial gonadal dysgenesis), (2) disorders of testosterone synthesis (testosterone biosynthesis defects like steroidogenic acute regulatory protein (STAR) deficiency, side-chain cleavage (CYP11A1) deficiency, 3 β-hydroxysteroid dehydrogenase type-ii (HSD3B2) deficiency, 17 β-hydroxylase/17, 20-lyase (CYP17A1) deficiency, 17 β-hydroxysteroid dehydrogenase-iii (HSD17B3) deficiency, P450 oxidoreductase (POR) defect; cytochrome b5 (CYB5) defect, and defects in luteinizing hormone action (LHCGR defect)), and (3) disorders of testosterone action [partial (PAIS) androgen insensitivity syndromes] or metabolization (5 α-reductase type-2 deficiency). 3 In 2006, the LWPES and the ESPE published a consensus statement on the management of 'intersex' disorders and proposed a new classification for DSD. 3,13 To authors' knowledge, there is no study in Pakistan regarding the etiological classification of XY DSD in children according to the new consensus document todate. XY DSD presents at a variable age. It may present from the first day of life to late adolescence. The varied age at presentation, in different geographic areas, could be due to socio-economic, cultural and educational factors. In this study, majority (55.6%) presents between 1 and 10 years. The present results also indicate that the clinical manifestation of XY DSD are not limited to ambiguous genitalia, but show a broad spectrum including isolated hypospadias, micropenis, cryptorchidism, delayed puberty, female like genitalia with gonads and inguinal mass similar to the study by Migeon et al. 14 In this study, a higher number of patients (86.6%) were being reared as males, even though 21.5% had genital ambiguity. A general preference for male gender in the Pakistan society at large which could explain the desire to rear majority of the patients as males. This data demonstrated that the most common cause of XY DSD was AIS (33.1%), either PAIS/5 alpha reductase deficiency (28.3%) or CAIS (4.8%). The percentage of PAIS patients in this study is similar to the study by Erdogan et al. 10 and is higher than the results of previous studies In the 46, XY DSD patient's diagnoses of PAIS and 5 α-reductase deficiency need to be confirmed by molecular genetic analyses. In addition, our 46 XY DSD patients included isolated hypospadias (10.2%) and micropenis (11.8%) without overt genital ambiguity. In these patients, probability of other causes of XY DSD was ruled out. Such subjects may have been either cases of isolated anatomical defect of unknown etiology. In this study, the major limitations encountered are the inability to make definite etiological diagnoses for 46, XY DSD patients. The frequency of vanishing testes syndrome among 46 XY DSD patients was 1.6% while the study presented by Erdogan et al. showed it to 13.3% which was high. 10 These patients presented with undescended testes had no genital ambiguity. Because vanishing testes syndrome was not in the former etiological classifications of intersex disorders, 4,5 its frequency within DSD is unknown. Cryptorchidism, together with hypospadias, is among the most common genitourinary anomalies in male children, affecting 1.6% to 9% live births. 18 In this study, cryptorchidism was seen in 20.8% patients, unilateral in 3.7% and bilateral in 17.1%. In another study, the prevalence of cryptorchidism at birth was found to be 5.9%, and approximately one fourth of the patients had non-palpable testes. 19 Another new population-based study carried out in boys aged less than one year to 19 years has established that the prevalence of cryptorchidism was 1.52%. 20 A very recent study has shown that about half of the patients with impalpable testes had vanishing testes syndrome. 21 Mutations in genes present in X, Y or autosomal chromosomes can cause abnormalities of testis determination or disorders of sex differentiation leading to 46, XY DSD. 6 Clinical and laboratorial investigation must include careful and precise anatomical, chromosomal, hormonal studies (both basal and after stimulation) and molecular genetics analyses prior to gender assignment in XY patients with under masculinization, which can be difficult in most cases. 3,9,22 In Pakistan, there is a general neglect, social taboos and lack of awareness regarding DSD. Reaching an etiological diagnosis is often difficult in limited resource settings because of variability of individual cases. CONCLUSION Phenotypic presentation of XY DSD depends on the underlying defects. Defect in androgen action on the target tissues or production of active metabolite share common morphological features. Clinical, chromosomal and hormonal assessment may help in making the presumptive etiological diagnosis. Further molecular genetics analyses are needed in differentiating these abnormalities and to make a final diagnosis. REFERENCES 1. Warne G, Zajac JD. Disorders of sexual differentiation. Endocrinol Metab Clin North Am 1998; 27: Journal of the College of Physicians and Surgeons Pakistan 2014, Vol. 24 (10):

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