Undetectable anti-Mullerian hormone and inhibin B do not preclude the presence of germ cell tumours in 45,X/46,XY or 46,XY gonadal dysgenesis.

OBJECTIVE: Individuals with 45,X/46,XY or 46,XY gonadal dysgenesis are at increased risk of germ cell malignancies. Therefore, prophylactic bilateral gonadectomy is advised in girls and considered in boys with atypical genitalia for undescended, macroscopically abnormal gonads. However, severely dysgenetic gonads may not contain germ cells rendering gonadectomy unnecessary. Therefore, we investigate if undetectable preoperative serum anti-Müllerian hormone (AMH) and inhibin B can predict the absence of germ cells, (pre)malignant or otherwise. DESIGN, PATIENTS AND MEASUREMENTS: Individuals who had undergone bilateral gonadal biopsy and/or gonadectomy because of suspected gonadal dysgenesis in 1999-2019 were included in this retrospective study if preoperative AMH and/or inhibin B were available. Histological material was reviewed by an experienced pathologist. Haematoxylin and eosin and immunohistochemical stainings for SOX9, OCT4, TSPY and SCF (KITL) were used. RESULTS: Thirteen males and 16 females were included, 20 with 46,XY and 9 with 45,X/46,XY DSD. Three females had dysgerminoma alongside gonadoblastoma; two gonadoblastoma, one germ cell neoplasia in situ (GCNIS) and three males had pre-GCNIS and/or pre-gonadoblastoma. Gonadoblastoma and/or dysgerminoma were present in 3/11 individuals with undetectable AMH and inhibin B, one of whom also had non-(pre)malignant germ cells. Of the other 18, in whom AMH and/or inhibin B were detectable, only one had no germ cells. CONCLUSIONS: Undetectable serum AMH and inhibin B cannot reliably predict the absence of germ cells and germ cell tumours in individuals with 45,X/46,XY or 46,XY gonadal dysgenesis. This information should help in counselling about prophylactic gonadectomy, taking into account both the germ cell cancer risk and potential for gonadal function.

Under-reported aspects of diagnosis and treatment addressed in the Dutch-Flemish guideline for comprehensive diagnostics in disorders/differences of sex development.

We present key points from the updated Dutch-Flemish guideline on comprehensive diagnostics in disorders/differences of sex development (DSD) that have not been widely addressed in the current (inter)national literature. These points are of interest to physicians working in DSD (expert) centres and to professionals who come across persons with a DSD but have no (or limited) experience in this area. The Dutch-Flemish guideline is based on internationally accepted principles. Recent initiatives striving for uniform high-quality care across Europe, and beyond, such as the completed COST action 1303 and the European Reference Network for rare endocrine conditions (EndoERN), have generated several excellent papers covering nearly all aspects of DSD. The Dutch-Flemish guideline follows these international consensus papers and covers a number of other topics relevant to daily practice. For instance, although next-generation sequencing (NGS)-based molecular diagnostics are becoming the gold standard for genetic evaluation, it can be difficult to prove variant causality or relate the genotype to the clinical presentation. Network formation and centralisation are essential to promote functional studies that assess the effects of genetic variants and to the correct histological assessment of gonadal material from DSD patients, as well as allowing for maximisation of expertise and possible cost reductions. The Dutch-Flemish guidelines uniquely address three aspects of DSD. First, we propose an algorithm for counselling and diagnostic evaluation when a DSD is suspected prenatally, a clinical situation that is becoming more common. Referral to ultrasound sonographers and obstetricians who are part of a DSD team is increasingly important here. Second, we pay special attention to healthcare professionals not working within a DSD centre as they are often the first to diagnose or suspect a DSD, but are not regularly exposed to DSDs and may have limited experience. Their thoughtful communication to patients, carers and colleagues, and the accessibility of protocols for first-line management and efficient referral are essential. Careful communication in the prenatal to neonatal period and the adolescent to adult transition are equally important and relatively under-reported in the literature. Third, we discuss the timing of (NGS-based) molecular diagnostics in the initial workup of new patients and in people with a diagnosis made solely on clinical grounds or those who had earlier genetic testing that is not compatible with current state-of-the-art diagnostics.

NR5A1 gene variants repress the ovarian-specific WNT signaling pathway in 46,XX disorders of sex development patients.

Several recent reports have described a missense variant in the gene NR5A1 (c.274C>T; p.Arg92Trp) in a significant number of 46,XX ovotesticular or testicular disorders of sex development (DSDs) cases. The affected residue falls within the DNA-binding domain of the NR5A1 protein, however the exact mechanism by which it causes testicular development in 46,XX individuals remains unclear. We have screened a cohort of 26 patients with 46,XX (ovo)testicular DSD and identified three unrelated individuals with this NR5A1 variant (p.Arg92Trp), as well as one patient with a novel NR5A1 variant (c.779C>T; p.Ala260Val). We examined the functional effect of these changes, finding that while protein levels and localization were unaffected, variant NR5A1 proteins repress the WNT signaling pathway and have less ability to upregulate the anti-testis gene NR0B1. These findings highlight how NR5A1 variants impact ovarian differentiation across multiple pathways, resulting in a switch from ovarian to testis development in genetic females.

Multiparameter Investigation of a 46,XX/46,XY Tetragametic Chimeric Phenotypical Male Patient with Bilateral Scrotal Ovotestes and Ovulatory Activity.

We report on an adult male initially presenting with gynecomastia and a painless scrotal mass without additional genital anomalies. Hyperpigmentation of the skin following the Blaschko's lines was identified. He underwent gonadectomy because of suspected cancer. Histological analyses revealed an ovotestis with ovulatory activity confirmed by immunohistochemistry with multiple markers. Karyotyping of cultured peripheral blood lymphocytes and a buccal smear revealed a 46,XX/46,XY chimeric constitution with different percentages. Multiple molecular analyses as well as blood typing implied a tetragametic origin. After the unilateral gonadectomy, the patient developed recurrent painful cystic swellings of the remaining gonad. Because of the wish to preserve hormonal activity as well as future fertility, the patient underwent surgical resection of a cystic gonadal area. The removed tissue showed ovulation-related features in addition to both testicular and ovarian tissue, diagnosed as an ovotestis. Testosterone therapy was initiated to suppress the persistently elevated gonadotropins and thereby suppress ovarian activity. During treatment, the recurrent pain complaints and cystic swellings ceased, although gonadotropin levels were not fully suppressed. Based on these observations, the importance of a detailed genetic and pathological diagnosis and the clinical dilemmas including the pros and cons of personalized treatment with gonadal preservative surgery are discussed.

Histological Assessment of Gonads in DSD: Relevance for Clinical Management.

Malignant gonadal germ cell tumors, referred to as germ cell cancers (GCC), occur with increased frequency in individuals who have specific types of differences (disorders) of sex development (DSD). Recent population-based studies have identified new environmental and genetic risk factors that have led to a 'genvironment' hypothesis, which may potentially be helpful in risk assessment in DSD-related GCC. In DSD, the malignancy risk is highly heterogeneous, but recent studies allow now to discriminate between high- and low-risk conditions. Gonadal biopsy is in some cases the best procedure of choice to assess the risk, and with the availability of immunohistochemical biomarkers [OCT3/4 (POU5F1), TSPY, SOX9, FOXL2 and KITLG (SCF)], a reliable classification of GCC and its precursors can be made. The opportunities in the field of virtual diagnostic pathology will be presented, having possibilities for rare diseases in general and DSD specifically. It is expected that the International DSD Registry will stimulate international collaborations, facilitating better diagnostic procedures as well as research.

Functional characterization of novel NR5A1 variants reveals multiple complex roles in disorders of sex development.

Variants in the NR5A1 gene encoding SF1 have been described in a diverse spectrum of disorders of sex development (DSD). Recently, we reported the use of a targeted gene panel for DSD where we identified 15 individuals with a variant in NR5A1, nine of which are novel. Here, we examine the functional effect of these changes in relation to the patient phenotype. All novel variants tested had reduced trans-activational activity, while several had altered protein level, localization, or conformation. In addition, we found evidence of new roles for SF1 protein domains including a region within the ligand binding domain that appears to contribute to SF1 regulation of Müllerian development. There was little correlation between the severity of the phenotype and the nature of the NR5A1 variant. We report two familial cases of NR5A1 deficiency with evidence of variable expressivity; we also report on individuals with oligogenic inheritance. Finally, we found that the nature of the NR5A1 variant does not inform patient outcomes (including pubertal androgenization and malignancy risk). This study adds nine novel pathogenic NR5A1 variants to the pool of diagnostic variants. It highlights a greater need for understanding the complexity of SF1 function and the additional factors that contribute.

A Rare Case of Embryonal Carcinoma in a Patient with Turner Syndrome without Y Chromosomal Material but Mutations in KIT, AKT1, and ZNF358 Demonstrated Using Exome Sequencing.

Gonadoblastoma and malignant transformations thereof can occur in females with Turner syndrome (TS) and Y chromosomal material. However, in females with TS and no Y chromosomal material, this is rarely seen. We report a female with an apparent 45,X karyotype (in blood and tumor) who was diagnosed with a metastatic embryonal carcinoma. Exome sequencing of blood and the tumor was done, and no Y chromosomal material was detected, while predicted deleterious mutations in KIT (likely driver), AKT1, and ZNF358 were identified in the tumor. The patient was treated with chemotherapy (first-line: cisplatin, etoposide, and bleomycin; second-line: paclitaxel and gemcitabine), and after that surgical debulking was performed. She is currently well and without signs of relapse. We conclude that embryonal carcinoma can apparently occur in 45,X TS without signs of Y chromosomal material.

The biology of germ cell tumors in disorders of sex development.

Development of a malignant germ cell tumor, i.e., germ cell cancer (GCC) in individuals with disorders of sex development (DSD) depends on a number of (epi-)genetic factors related to early gonadal- and germ cell development, possibly related to genetic susceptibility. Fetal development of germ cells is orchestrated by strict processes involving specification, migration and the development of a proper gonadal niche. In this review we will discuss the early (epi-)genetic events in normal and aberrant germ cell and gonadal development. Focus will be on the formation of the precursor lesions of GCC in individuals who have DSD. In our view, expression of the different embryonic markers in, and epigenetic profile of the precursor lesions reflects the developmental stage in which these cells are blocked in their maturation. Therefore, these are not a primary pathogenetic driving force. Progression later in life towards a full blown cancer likely depends on additional factors such as a changed endocrine environment in a susceptible individual. Genetic susceptibility is, as evidenced by the presence of specific risk genetic variants (SNPs) in patients with a testicular GCC, related to genes involved in early germ cell and gonadal development.

Hormonal evaluation in relation to phenotype and genotype in 286 patients with a disorder of sex development from Indonesia.

OBJECTIVE: The objective of this study was to determine the aetiological spectrum of disorders of sex development (DSD) in a large cohort of underprivileged and undiagnosed patients from Indonesia. METHODS: A total of 286 patients with atypical external and/or internal genitalia were evaluated using clinical, hormonal, molecular genetic and histological parameters. RESULTS: The age (years) at presentation was 0-0·5 in 41 (14·3%), >0·5-12 in 181 (63·3%) and >12 in 64 cases (22·4%). 46,XY DSD was most common (68·2%, n = 195), 46,XX DSD was found in 23·4% (n = 67) and sex chromosomal DSD in 8·4% (n = 24). In 61·2% of 46,XX DSD patients, 17·9% of 46,XY DSD patients and all sex chromosome DSD patients (29·4% in total), a final diagnosis was reached based on genetic or histological gonadal tissue evaluation. 17-hydroxyprogesterone and androstenedione levels were the most distinctive parameters in 46,XX DSD patients. In 46,XY DSD, diagnostic groups were identified based on the external masculinization score: androgen action disorder (AAD), unknown male undermasculinization (UMU), and gonadal dysgenesis (GD). LH, FSH and testosterone levels were most informative especially in the older age group. HCG tests were of no additional value as no patients with androgen synthesis disorders were found. Hormonal profiles of patients with sex chromosome DSD and a Y-chromosome sequence containing karyotype showed high levels of LH and FSH, and low levels of AMH, inhibin B and testosterone compared with the normal male range. Gene mutations were found in all patients with CAH, but in only 24·5% and 1·8% of patients with AAD and UMU. In 32% of 46,XY GD patients, copy number variants of different genes were found. CONCLUSION: A stepwise diagnostic approach led to a molecularly or histologically proven final diagnosis in 29·4% of the patients. The most informative parameters were serum levels of 17-hydroxyprogesterone and androstenedione in 46,XX DSD patients, and serum LH, FSH and testosterone levels in 46,XY DSD patients.

Aberrant SOX2 expression in colorectal cancers does not correlate with mucinous differentiation and gastric mucin MUC5AC expression.

Colorectal cancer (CRC) can be divided into non-mucinous and mucinous subtypes, of which the latter portends to have a worse clinical prognosis. A previous study suggested a putative link between SOX2 expression observed selectively in mucinous CRC and the induction of the gastric mucin MUC5AC. In this study, we re-evaluated the expression behavior of SOX2, MUC5AC, and CDX2 in both types of CRC. We performed immunohistochemical analysis on 90 cases of non-mucinous CRCs, 57 cases of mucinous CRCs, and 15 case-matched normal intestinal mucosa. In contrast to the previously suggested link between SOX2 and mucinous CRC, we observe aberrant expression of SOX2 at equal levels in both subtypes. Fluorescence in situ hybridization (FISH) analysis shows that expression is not attributed to genomic amplification. While SOX2 and CDX2 are normally expressed in a reciprocal manner, SOX2-positive tumor cells co-express CDX2. Furthermore, we show that MUC5AC is expressed independently of SOX2. In conclusion, we show that aberrant SOX2 expression is specifically linked neither to mucinous CRCs nor to the induction of MUC5AC, in contrast to previous suggestions.

Gonadal malignancy in 13 consecutive collected patients with disorders of sex development (DSD) from Semarang (Indonesia).

AIMS: Caucasian patients with disorders of sex development (DSD) are at a high risk of developing germ cell cancer (GCC). GCC is prominent in young adults in Western countries, while the incidence is significantly lower in Asia. So far, the risk of GCC in Asian DSD patients is unknown. METHODS AND RESULTS: A detailed study of gonad histology , morphology and immunohistochemistry (OCT3/4, testis-specific protein Y-encoded, VASA, SCF/KITLG, SOX9, FOXL2) of 16 Indonesian DSD patients was undertaken. 13 cases could be analysed, including ovarian tissue (n=3), streak gonad (n=1), undifferentiated gonad (n=1) and testicular tissue (n=8), diagnosed as 46, XX (n=1), 46, XY (n=7) and sex chromosome DSD (n=5). The precursor lesion gonadoblastoma or carcinoma in situ, or GCC was diagnosed in four cases (30.8%; three 46, XY and one sex chromosome DSD ). A hormone producing ovarian Leydig cell tumour was identified in a 46, XX patient, supposed to be a late onset congenital adrenal hyperplasia. CONCLUSIONS: In spite of the significantly lower risk of GCC in the general Asian population, DSD is a dominant risk factor. The study demonstrates the power of immunohistochemical markers for (early) diagnosis. This knowledge will deepen understanding of the pathobiology of GCC and clinical handling of patients with DSD, globally.

SRY mutation analysis by next generation (deep) sequencing in a cohort of chromosomal Disorders of Sex Development (DSD) patients with a mosaic karyotype.

BACKGROUND: The presence of the Y-chromosome or Y chromosome-derived material is seen in 4-60% of Turner syndrome patients (Chromosomal Disorders of Sex Development (DSD)). DSD patients with specific Y-chromosomal material in their karyotype, the GonadoBlastoma on the Y-chromosome (GBY) region, have an increased risk of developing type II germ cell tumors/cancer (GCC), most likely related to TSPY. The Sex determining Region on the Y gene (SRY) is located on the short arm of the Y-chromosome and is the crucial switch that initiates testis determination and subsequent male development. Mutations in this gene are responsible for sex reversal in approximately 10-15% of 46,XY pure gonadal dysgenesis (46,XY DSD) cases. The majority of the mutations described are located in the central HMG domain, which is involved in the binding and bending of the DNA and harbors two nuclear localization signals. SRY mutations have also been found in a small number of patients with a 45,X/46,XY karyotype and might play a role in the maldevelopment of the gonads. METHODS: To thoroughly investigate the presence of possible SRY gene mutations in mosaic DSD patients, we performed next generation (deep) sequencing on the genomic DNA of fourteen independent patients (twelve 45,X/46,XY, one 45,X/46,XX/46,XY, and one 46,XX/46,XY). RESULTS AND CONCLUSIONS: The results demonstrate that aberrations in SRY are rare in mosaic DSD patients and therefore do not play a significant role in the etiology of the disease.

Prevalence of c-KIT mutations in gonadoblastoma and dysgerminomas of patients with disorders of sex development (DSD) and ovarian dysgerminomas.

Activating c-KIT mutations (exons 11 and 17) are found in 10-40% of testicular seminomas, the majority being missense point mutations (codon 816). Malignant ovarian dysgerminomas represent ~3% of all ovarian cancers in Western countries, resembling testicular seminomas, regarding chromosomal aberrations and c-KIT mutations. DSD patients with specific Y-sequences have an increased risk for Type II Germ Cell Tumor/Cancer, with gonadoblastoma as precursor progressing to dysgerminoma. Here we present analysis of c-KIT exon 8, 9, 11, 13 and 17, and PDGFRA exon 12, 14 and 18 by conventional sequencing together with mutational analysis of c-KIT codon 816 by a sensitive and specific LightCycler melting curve analysis, confirmed by sequencing. The results are combined with data on TSPY and OCT3/4 expression in a series of 16 DSD patients presenting with gonadoblastoma and dysgerminoma and 15 patients presenting pure ovarian dysgerminomas without DSD. c-KIT codon 816 mutations were detected in five out of the total of 31 cases (all found in pure ovarian dysgerminomas). A synonymous SNP (rs 5578615) was detected in two patients, one DSD patient (with bilateral disease) and one patient with dysgerminoma. Next to these, three codon N822K mutations were detected in the group of 15 pure ovarian dysgerminomas. In total activating c-KIT mutations were found in 53% of ovarian dysgerminomas without DSD. In the group of 16 DSD cases a N505I and D820E mutation was found in a single tumor of a patient with gonadoblastoma and dysgerminoma. No PDGFRA mutations were found. Positive OCT3/4 staining was present in all gonadoblastomas and dysgerminomas investigated, TSPY expression was only seen in the gonadoblastoma/dysgerminoma lesions of the 16 DSD patients. This data supports the existence of two distinct but parallel pathways in the development of dysgerminoma, in which mutational status of c-KIT might parallel the presence of TSPY.

A 46,XY female DSD patient with bilateral gonadoblastoma, a novel SRY missense mutation combined with a WT1 KTS splice-site mutation.

Patients with Disorders of Sex Development (DSD), especially those with gonadal dysgenesis and hypovirilization are at risk of developing malignant type II germ cell tumors/cancer (GCC) (seminoma/dysgerminoma and nonseminoma), with either carcinoma in situ (CIS) or gonadoblastoma (GB) as precursor lesion. In 10-15% of 46,XY gonadal dysgenesis cases (i.e., Swyer syndrome), SRY mutations, residing in the HMG (High Mobility Group) domain, are found to affect nuclear transport or binding to and bending of DNA. Frasier syndrome (FS) is characterized by gonadal dysgenesis with a high risk for development of GB as well as chronic renal failure in early adulthood, and is known to arise from a splice site mutation in intron 9 of the Wilms' tumor 1 gene (WT1). Mutations in SRY as well as WT1 can lead to diminished expression and function of SRY, resulting in sub-optimal SOX9 expression, Sertoli cell formation and subsequent lack of proper testicular development. Embryonic germ cells residing in this unfavourable micro-environment have an increased risk for malignant transformation. Here a unique case of a phenotypically normal female (age 22 years) is reported, presenting with primary amenorrhoea, later diagnosed as hypergonadotropic hypogonadism on the basis of 46,XY gonadal dygenesis with a novel missense mutation in SRY. Functional in vitro studies showed no convincing protein malfunctioning. Laparoscopic examination revealed streak ovaries and a normal, but small, uterus. Pathological examination demonstrated bilateral GB and dysgerminoma, confirmed by immunohistochemistry. Occurrence of a delayed progressive kidney failure (focal segmental glomerular sclerosis) triggered analysis of WT1, revealing a pathogenic splice-site mutation in intron 9. Analysis of the SRY gene in an additional five FS cases did not reveal any mutations. The case presented shows the importance of multi-gene based diagnosis of DSD patients, allowing early diagnosis and treatment, thus preventing putative development of an invasive cancer.

Delayed Recognition of Disorders of Sex Development (DSD): A Missed Opportunity for Early Diagnosis of Malignant Germ Cell Tumors.

Disorders of sex development (DSD) are defined as a congenital condition in which development of chromosomal, gonadal or anatomical sex is atypical. DSD patients with gonadal dysgenesis or hypovirilization, containing part of the Y chromosome (GBY), have an increased risk for malignant type II germ cell tumors (GCTs: seminomas and nonseminomas). DSD may be diagnosed in newborns (e.g., ambiguous genitalia), or later in life, even at or after puberty. Here we describe three independent male patients with a GCT; two were retrospectively recognized as DSD, based on the histological identification of both carcinoma in situ and gonadoblastoma in a single gonad as the cancer precursor. Hypospadias and cryptorchidism in their history are consistent with this conclusion. The power of recognition of these parameters is demonstrated by the third patient, in which the precursor lesion was diagnosed before progression to invasiveness. Early recognition based on these clinical parameters could have prevented development of (metastatic) cancer, to be treated by systemic therapy. All three patients showed a normal male 46,XY karyotype, without obvious genetic rearrangements by high-resolution whole-genome copy number analysis. These cases demonstrate overlap between DSD and the so-called testicular dysgenesis syndrome (TDS), of significant relevance for identification of individuals at increased risk for development of a malignant GCT.

Application of the new classification on patients with a disorder of sex development in indonesia.

Disorder of sex development (DSD) patients in Indonesia most often do not receive a proper diagnostic evaluation and treatment. This study intended to categorize 88 Indonesian patients in accordance with the new consensus DSD algorithm. Diagnostic evaluation including clinical, hormonal, genetic, imaging, surgical, and histological parameters was performed. Fifty-three patients were raised as males, and 34 as females. Of 22 patients with 46, XX DSD, 15 had congenital adrenal hyperplasia, while in one patient, an ovarian Leydig cell tumor was found. In all 58 46, XY DSD patients, 29 were suspected of a disorder of androgen action (12 with an androgen receptor mutation), and in 9, gonadal dysgenesis was found and, in 20, severe hypospadias e.c.i. Implementation of the current consensus statement in a resource-poor environment is very difficult. The aim of the diagnostic workup in developing countries should be to end up with an evidence-based diagnosis. This is essential to improve treatment and thereby to improve the patients' quality of life.

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

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

Genome-wide gene expression profiling reveals aberrant MAPK and Wnt signaling pathways associated with early parthenogenesis.

Mammalian parthenogenesis could not survive but aborted during mid-gestation, presumably because of lack of paternal gene expression. To understand the molecular mechanisms underlying the failure of parthenogenesis at early stages of development, we performed global gene expression profiling and functional analysis of parthenogenetic blastocysts in comparison with those of blastocysts from normally fertilized embryos. Parthenogenetic blastocysts exhibited changes in the expression of 749 genes, of which 214 had lower expression and 535 showed higher expressions than fertilized embryos using a minimal 1.8-fold change as a cutoff. Genes important for placenta development were decreased in their expression in parthenote blastocysts. Some maternally expressed genes were up-regulated and paternal-related genes were down-regulated. Moreover, aberrantly increased Wnt signaling and reduced mitogen-activated protein kinase (MAPK) signaling were associated with early parthenogenesis. The protein level of extracellular signal-regulated kinase 2 (ERK2) was low in parthenogenetic blastocysts compared with that of fertilized blastocysts 120 h after fertilization. 6-Bromoindirubin-3'-oxime, a specific glycogen synthase kinase-3 (GSK-3) inhibitor, significantly decreased embryo hatching. The expression of several imprinted genes was altered in parthenote blastocysts. Gene expression also linked reduced expression of Xist to activation of X chromosome. Our findings suggest that failed X inactivation, aberrant imprinting, decreased ERK/MAPK signaling and possibly elevated Wnt signaling, and reduced expression of genes for placental development collectively may contribute to abnormal placenta formation and failed fetal development in parthenogenetic embryos.

Gonadal tumours and DSD.

Disorders of sex development (DSD), previously referred to as intersex, has been recognised as one of the main risk factors for development of type II germ cell tumours (GCTs), that is, seminomas/dysgerminomas and non-seminomas (e.g., embryonal carcinoma, yolk sac tumour, choriocarcinoma and teratoma). Within the testis, this type of cancer is the most frequent malignancy in adolescent and young adult Caucasian males. Although these males are not known to have dysgenetic gonads, the similarities in the resulting tumours suggest a common aetiological mechanism(s),--genetically, environmentally or a combination of both. Within the group of DSD patients, being in fact congenital conditions, the risk of malignant transformation of germ cells is highly heterogeneous, depending on a number of parameters, some of which have only recently been identified. Understanding of these recent insights will stimulate further research, with the final aim to develop an informative clinical decision tree for DSD patients, which includes optimal (early) diagnosis without overtreatment, such as prophylactic gonadectomy in the case of a low tumour risk.