A novel, homozygous mutation in desert hedgehog (DHH) in a 46, XY patient with dysgenetic testes presenting with primary amenorrhoea: a case report

Abstract

Background: Desert hedgehog (DHH) mutations have been described in only a limited number of individuals with 46, XY disorders of sex development (DSD) presenting as either partial or complete gonadal dysgenesis. Gonadal tumours and peripheral neuropathy have been associated with DHH mutations. Herein we report a novel, homozygous mutation of DHH identified through a targeted, massively parallel sequencing (MPS) DSD panel, in a patient presenting with partial gonadal dysgenesis. This novel mutation is two amino acids away from a previously described mutation in a patient who presented with complete gonadal dysgenesis. Adding to the complexity of work-up, our patient also expressed gender identity concern.

Case presentation: A 14-year-old, phenotypic female presented with primary amenorrhoea and absent secondary sex characteristics. Investigations revealed elevated gonadotrophins with low oestradiol, testosterone of 0.6 nmol/L and a 46, XY karyotype. Müllerian structures were not seen on pelvic ultrasound or laparoscopically and gonadal biopsies demonstrated dysgenetic testes without neoplasia (partial gonadal dysgenesis). The patient expressed gender identity confusion upon initial notification of investigation findings. Formal psychiatric evaluation excluded gender dysphoria. Genetic analysis was performed using a targeted, MPS DSD panel of 64 diagnostic and 927 research candidate genes. This identified a novel, homozygous mutation in exon 2 of DHH (DHH:NM_021044:exon2:c.G491C:p.R164P). With this finding our patient was screened for the possibility of peripheral neuropathy which was not evident clinically nor on investigation. She was commenced on oestrogen for pubertal induction.

Conclusion: The evaluation of patients with DSD is associated with considerable psychological distress. Targeted MPS enables an affordable and efficient method for diagnosis of 46, XY DSD cases. Identifying a genetic diagnosis may inform clinical management and in this case directed screening for peripheral neuropathy. In addition to the structural location of the mutation other interacting factors may influence phenotypic expression in homozygous DHH mutations.

Keywords: DHH; Desert hedgehog; Disorder of sex development; Gonadal dysgenesis; Massively parallel sequencing.

Fig. 1

Normal male phenotypic sex development highlighting the role of DHH. Testis determination requires SRY and one DAX1 copy, whilst with two DAX1 copies and a lack of SRY an ovary is determined. In the bipotential gonad, SF1, WT1 and GATA4 upregulate SRY which in turn upregulates SOX9; SOX9 is pivotal to testis determination. Upregulation of SRY and SOX9 overcomes the action of genes promoting female sex development including DAX1 and β-catenin. With upregulation of SOX9, multiple feed-forward loops then accelerate male pathway commitment. Other genes, including DHH, as well as growth factors also have roles in promoting testis determination. Following testis determination, Sertoli cells secrete AMH leading to Müllerian duct regression; Müllerian ducts would otherwise form the fallopian tubes, uterus, cervix and upper vagina. Sertoli cells also secrete DHH which is important for Leydig cell development and Sertoli-peritubular cell interaction. Responding to luteinizing hormone (via LHCGR), a functional Leydig cell produces testosterone via steroidogenesis, requiring SF1 and steroidogenesis enzymes. Converted to DHT via 5α-reductase, testosterone and DHT induce Wolffian duct differentiation (leading to vas deferens, seminal vesicle and prostate formation) and external genitalia masculinization through actions on the androgen receptor. In the absence of functional Leydig cells female external genitalia form and testes fail to descend. Blue block arrows indicate DHH’s involvement in male sex development. Lines ending in an arrow denote a positive/up-regulating effect whilst lines ending in a perpendicular bar indicate a negative/down-regulating effect. Abbreviations: AMH– Anti-Müllerian hormone, ATRX– X-linked alpha thalassemia and mental retardation, DAX1– dosage-sensitive sex reversal, adrenal hypoplasia critical region, on chromosome X, gene 1, DHH– desert hedgehog, DHT– dihydrotestosterone, FGF9– fibroblast growth factor 9, INSL3– insulin-like protein 3, LHCGR– Luteinizing hormone common G-protein receptor, NGFs– nerve growth factors, PDGFs– platelet derived growth factors, PGD2– prostaglandin D2, RSPO1– R-spondin-1, SF1– steroidogenic factor-1, SRY– sex determining region on the Y chromosome, WT1– Wilms’ tumour suppressor gene 1

Fig. 2

Surgical photos. a Laparoscopic view of left and right gonad within the pelvis. b Blind ending vagina at vaginoscopy. c Left gonad and epididymal flanking structure. d Right gonad and epididymal flanking structure

Fig. 3

Histopathology. a Right gonad excision biopsy, H&E stain, demonstrating seminiferous tubules lined by Sertoli cells without spermatogenesis. There is intervening fibrosis with no definite Leydig cells. b Left gonad excision biopsy, H&E stain, demonstrating seminiferous tubules lined by Sertoli cells without spermatogenesis. There is intervening fibrosis with occasional, unusually large clusters of Leydig cells. c Left gonad excision, inhibin stain, demonstrating Sertoli and Leydig cell appearance. d Normal testis of 15-year-old, showing seminiferous tubules with spermatogenesis

Fig. 4

Three dimensional model of DHH. a SWISS-MODEL generated 3D model of DHH showing residue Leu162, the position of a previously published mutation [12] and b Arg164, the position of our novel mutation. c Close-up of the mutation L162P and d R164P with the side chains of both the wild-type and mutant residue shown and coloured green and red, respectively. On panels a and b, black arrows point to position of mutations