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DAX1 (NR0B1) is an orphan nuclear receptor, which plays an important role in development and function of the adrenal glands and gonads. Mutations in DAX1 cause X-linked adrenal hypoplasia congenita (X-linked AHC), which is characterized by adrenal insufficiency (AI) and hypogonadotropic hypogonadism (HHG). Affected boys present with adrenal failure usually in childhood and, later in life, with delayed puberty. However, patients with a late-onset form of X-linked AHC have also been described in the past years. We report a male patient who presented with symptoms of an adrenal crisis at the age of 38 years and was later diagnosed with HHG. Family history was positive with several male relatives diagnosed with AI and compatible with the assumed X-chromosomal inheritance of the trait. Direct sequencing of DAX1 of the patient revealed a hemizygous cytosine-to-thymine substitution at nucleotide 64 in exon 1, which creates a novel nonsense mutation (p.(Gln22*)). In order to compare the clinical presentation of the patient to that of other patients with X-linked AHC, we searched the electronic database MEDLINE (PubMed) and found reports of nine other cases with delayed onset of X-linked AHC. In certain cases, genotype–phenotype correlation could be assumed.
X-linked adrenal hypoplasia congenita (X-linked AHC) is caused by a mutation of the DAX1 gene (1). Patients with this disease, mainly males whereas also a few females have been described, usually present themselves with signs and symptoms of primary AI in early infancy or anytime throughout childhood. The adequate therapy consists of a lifelong glucocorticoid and mineralocorticoid replacement therapy. At the expected time of puberty, patients present with hypogonadotropic hypogonadism (HHG) and delay or lack of sexual maturation (2). Next to the typical form of presentation in early infancy or childhood, there have been some X-linked AHC patients described that show an onset of symptoms in late adolescence or even adulthood (3, 4, 5, 6, 7, 8, 9). These late-onset cases suggest a milder form of X-linked AHC; however, they are also susceptible to adrenal crisis often triggered by an unrelated severe illness or other environmental stress (8).
The patient was born with a length of 53cm and a weight of 4200g. As a child, the patient grew fast until the age of 14 years – his growth continued until the age of 16 years (final height 186cm). In this period, vertigo attacks started and the patient needed more rest. Pubic hair growth started at the age of 15 years, but otherwise his puberty was delayed. The growth of a beard started with 17 years. The patient reported that muscle composition had ever been slight, and muscle buildup had been hard to achieve. The patient did not report any olfactory problems in terms of a Kallmann’s syndrome. He affirmed a question regarding salt craving.
At the age of 38 years, he complained about fatigue, vertigo, an inclination of collapses. It worsened with nausea and vomiting during a feverish episode (38.3°C) due to acute tonsillitis. Upon clinical examination, low blood pressure (105/65mmHG), increased heart rate (104bpm) and a higher respiratory rate (22breaths/min) were measured. Darker skin pigmentation was noticed. Testicular volume was reported with 12 mL bilaterally (Tanner stage IV). Except for dehydration and painful, swollen tonsils, clinical examination did not show any other pathologic findings. Upon admission, serum sodium was low (122mmol/L); potassium was in normal range (3.8mmol/L).
Due to a positive familial history regarding primary AI and the current clinic, an adrenal crisis due to tonsillitis was assumed and a therapy with hydrocortisone, fludrocortisone and rehydration was initiated. Upon treatment, the patient showed clinical recovery within a day. Hormonal analysis confirmed primary AI with an increased ACTH (>2000ng/L; normal range (NR): 7.2–63ng/L), low basal serum cortisol (52µg/L; (NR): 62–194µg/L), decreased aldosterone (11ng/L; (NR): 20–150ng/L) and an increased renin concentration (1697ng/L, (NR): 1.5–18ng/L). Adrenal gland antibodies (21-hydroxylase antibodies) could not be detected; hepatitis serology and HIV antibodies were negative. Thyroid function and blood glucose values were normal. A normal calcium level excluded a hypoparathyroidism. Sonography of the kidneys and adrenal glands was performed but did not show any abnormalities. Chest X-ray was normal.
Two years later, the patient reported of impaired libido, which had been decreasing since one year. At that time point, luteinizing hormone (LH) and prolactin were tested normal. Follicle-stimulating hormone (FSH), sex hormone-binding globulin (SHBG) and testosterone have been increased. Inhibin B was lowered, but testis-sonography was without pathological findings, despite a testicular volume of 6mL bilaterally (Tanner stage II). Semen analysis revealed azoospermia (semen volume: 3.4mL; sperm count: 0×106per mL; motility: 0%; abnormal forms: not done). On physical examination, bilateral gynecomastia (2cm), sparse beard growth, reduced pubic hair and enhanced skin pigmentation were found. Testosterone replacement was started with dermal testosterone (testogel) and was switched to testosterone-undecanoat injections every 12 weeks during follow-up.
The combined presentation of primary AI and progressive HHG, together with a positive family history (four male cousins on mother’s side of the family with primary AI) raised suspicion of DAX1-related X-linked AHC and mutation analysis was initiated.
Direct sequencing of the DAX1 gene revealed a hemizygous cytosine-to-thymine substitution at nucleotide 64 in codon 22, exon 1 (Fig. 1), which creates a nonsense mutation (TAA) at position 22, instead of glutamine (p.(Gln22*)). The mutation lies within the first repetitive motif of the amino terminus. This finding suggests a mutant NR0B1, resulting in X-linked AHC.
The patient’s treatment is shown in Table 1.
Follow-up (2009–2016) was performed regularly every six months with laboratory tests and physical examinations (Table 1).
X-linked AHC characterized by AI and hypogonadotropic hypogonadism (HHG) and is due to mutations in DAX1, the gene encoding an orphan nuclear receptor. Affected boys present with adrenal failure usually in childhood and, later in life, with delayed puberty (2). However, patients with a late-onset form of X-linked AHC have also been described in the past years. DAX1 protein acts as transcriptional repressor.
The electronic database MEDLINE (PubMed) was searched and we identified nine other cases of late-onset X-linked AHC with six different mutations (Table 2) (3, 4, 5, 6, 7, 8, 9): Two nonsense mutations (p.(Gln37*), p.(Trp39*) (3, 4, 8)), three missense mutations (p.(Tyr380Asp), p.(Iso439Ser) and (p.(Ser259Pro) (5, 6, 9)), and a deletion of three nucleotides resulting in the loss of an amino acid (p.305delGly) (7). Locations of the different patients’ mutations are shown in Fig. 2.
Investigation of clinical presentation revealed that seven patients with late-onset X-linked AHC presented with primary AI (Table 2), but only four of the patients have complete HHG. One subject has partial HHG (case 3) and two did not develop HHG up to now (case 9 and 10). Azoospermia was found in four patients, oligospermia was described in two. In the other patients, semen analysis was not performed; however, normal fertility was stated in two patients (case 9 and 10). An interesting aspect of the clinical history of the patient (reported by us) is the progressive course of HHG with shrinkage of testicular volume, decreasing libido and appearance of gynecomastia after the diagnosis of AI.
Only three patients had a positive family history for primary AI with a segregation pattern compatible with maternal inheritance. Cases 7 and 8 are important to highlight because these two brothers presented first with hypogonadism and later on with an impaired cortisol response to co-syntropin stimulation. Case 8 passed away suddenly during intensive physical activity and before hydrocortisone medication was started (8).
The cases with missense mutations seem similar in phenotype, because all cases (cases 4, 5, 9 and 10) initially reported signs of adrenal insufficiency (Table 2). Case 5 had a missense mutation in the putative LBD (5), thus alternating the structure within the carboxy-terminal region of the protein. Missense mutations tend to cluster in this region and account for approx. 25% of all DAX1 mutations. Also the other known mutations causing late-onset X-linked AHC are also found in the putative LBD region. These mutations seem to alter protein folding, thereby impairing nuclear localization and cofactor recruitment.
The cases with nonsense mutations, which are located in the part of the gene encoding the first amino-terminal nuclear receptor repeat, do not share a similar clinical pattern, because two patients presented first with signs of hypogonadism (cases 7 and 8), whereas the other two (cases 2 and 3) with signs of AI. This suggests some other additional DAX1-unrelated genetic, epigenetic and/or nongenetic factors involved in the phenotype.
The DAX1 gene consists of an internal in-frame translation start site at the methionine codon 83 with a partially conserved Kozak consensus sequence (4). The data of Ozisik et al. strongly suggest that in patients with nonsense mutations in the gene encoding the extreme amino terminus of DAX1, the internal in-frame translation generates a shorter DAX1 protein that partially rescues the clinical phenotype of the patients (4). In addition, low levels of truncated proteins were found in comparison to the wildtype thus indicating lower translation efficiency from the alternate start site (4). However, some of the naturally occurring nonsense or frameshift mutations that cause a premature termination codon 5′ to the putative alternative start site (codon 83) are associated with the classical AHC phenotype (4).
Based on these findings, we hypothesize a similar mechanism with an alternate initiation codon in our patient with the p.(Gln22*) mutation, otherwise the largely truncated protein would be nonfunctional with total loss of repression activity of DAX1 and a classical X-linked AHC with clinical manifestation in early childhood could be expected.
Up to date, there is no clear genotype–phenotype correlation of the DAX1 mutations, and especially, no explanation for the difference between a late-onset and a classical X-linked AHC, as well as X-linked AHC with only primary AI or in combination with HHG are poorly understood. This phenotypic variability might be due to other factors, such as modifier genes, compensation from other genes, epigenetic or nongenetic factors. Therefore, it seems crucial to further investigate families harboring DAX1 mutations, thus identifying those modulators.
The history of the presented patient shows that the detection of a rare disease often takes a long time even though there are affected family members in former generations. As can be seen in this case study, it is important to identify DAX1 mutations for genetic counseling regarding conscious family planning, pregnancies and especially for the early identification of patients at risk for primary AI. Immediate steroid hormone replacement in family members that reveal impaired adrenal function can prevent those patients from adrenal crises as first manifestation of the disease.
The authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported.
This research did not receive any specific grant from any funding agency in the public, commercial or not-for-profit-sector.
Written informed consent has been obtained from the patient for publication of the submitted article and any accompanying images.
Judith Gerards is a medical student. She did the research for this article, assembled patient’s medical data, compared late-onset forms of X-linked AHC and wrote the manuscript. Michael M Ritter identified further family members with this DAX1 mutation, and helped writing the manuscript. Elke Kaminsky did the mutational analysis, and helped writing the manuscript. Andreas Gal and Wolfgang Hoeppner were our counselor in genetic issues and helped writing the manuscript. Marcus Quinkler treated the patient, coordinated the research, analyzed the literature and wrote the manuscript.