Premature adrenarche

Premature adrenarche describes the precocious onset of adrenal androgen secretion, traditionally defined as occurring as early as 8 years in girls and 9 years in boys. It is important to note that premature adrenarche is not equivalent to premature pubarche, i.e. the early appearance of pubic hair only. Indeed, there is some inconsistency in the literature where premature pubarche is often used synonymously for premature adrenarche.

A recently proposed definition of premature adrenarche is the concurrent presence of adrenal androgen levels increased above the age- and sex-specific

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reference range and clinical signs of an increase in androgen action, such as adult- type body odour, oily hair and skin and/or premature pubarche, occurring before the age of 8 years in girls and 9 years in boys (Utriainen et al., 2007) (Table 5). The definition of the exact age limits should take influences of ethnicity into account. Two studies in Caucasian populations found pubic hair Tanner stage 2 or above before the age of 8 years in girls in 0.6% and 0.8% of American and Lithuanian girls, respectively (Zukauskaite et al., 2005; Rosenfield et al., 2009). By contrast, a five- fold higher incidence of premature pubarche was reported in girls of Afro-American descent (Herman-Giddens et al., 1997). All these studies focused on the premature appearance of pubic hair only and did not report on the incidence of other signs of increased androgen action such as adult-type body odour or oily skin. Thus the overall incidence of premature adrenarche may well be higher. Generally, the reported prevalence of premature adrenarche is about 10-fold higher in girls than in boys (Rosenfield, 1994).

Constitutional or idiopathic premature adrenarche (IPA) is most often observed, which is a diagnosis of exclusion. Distinct conditions manifesting with premature adrenarche include non-classic variants of CAH that have been diagnosed in 0-40% of children with premature pubarche (Utriainen et al., 2009; Armengaud et al., 2009; Dacou-Voutetakis and Dracopoulou, 1999; Temeck et al., 1987; del Balzo et al., 1992; Balducci et al., 1994), depending on the pre-selection bias applied to the studied cohorts. Other causes potentially underlying a presentation with premature adrenarche are summarized in Table 5.

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Table 5: Definition and differential diagnosis of disorders presenting with childhood androgen excess.

Definition Causes

Premature adrenarche =

Increased adrenal androgen secretion

above the age- and sex-specific normal reference range before the age of 8 years in girls and 9 years in boys (= premature biochemical adrenarche)

AND

Clinical signs of androgen action before

the age of 8 years in girls and 9 years in boys including

- adult-type body odour - oily hair and skin, acne

- pubic hair (= premature pubarche), axillary hair

AND

No signs of pubertal development

(i.e. no development of secondary sexual characteristics including breast

development in girls or testicular/penile growth in boys)

Most frequent:

Idiopathic (constitutional) premature adrenarche

Rare:

Defects in steroid synthesis, metabolism or action:

Congenital adrenal hyperplasia - 21-hydroxylase deficiency - 11β-hydroxylase deficiency

- 3β-hydroxysteroid dehydrogenase deficiency - Cushing’s disease

- Glucocorticoid resistance

- (Apparent) Cortisone reductase deficiency

- Apparent DHEA sulfotransferase (PAPSS2) deficiency

Other:

- Virilizing tumours originating from adrenals or gonad

- Exogenous testosterone treatment

Precocious pseudo-puberty = Development of secondary sexual characteristics (penile growth, breast development, menarche) before the age of 8 years in girls or 9 years in boys

AND

Low gonadotrophins before and after GnRH stimulation

GnRH-independent; usually non-harmonic chronology of sexual characteristics, e.g. lack of bilateral testicular enlargement in boys (exceptions: gonadal tumours, familial testotoxicosis)

also termed peripheral precocious puberty

Rare:

Defects in steroid synthesis, metabolism or action:

Congenital adrenal hyperplasia - 21-hydroxylase deficiency - 11β-hydroxylase deficiency

Other:

- Virilizing tumours originating from adrenals or gonads - Exogenous sex steroid treatment

- LH- or HCG-driven stimulation of adrenal androgen

production due to

- β-HCG-secreting tumours

- Familial testotoxicosis (due to activating LH receptor (LH/CGR) mutations)

- McCune-Albright-syndrome (due to activating Gsα protein (GNAS1) mutations)

Precocious puberty = Development of secondary sexual characteristics (testicular/penile growth, breast development, menarche) before the age of 8 years in girls or 9 years in boys AND

Increase in gonadotrophins after GnRH stimulation

GnRH-dependent; usually harmonic chronology of sexual characteristics, e.g. bilaterally enlarged testicular volume in boys

Most frequent:

Idiopathic (constitutional) precocious puberty

Rare:

- Central nervous system lesions, e.g. glioma, astrocytoma, hypothalamic hamartoma, arachnoid cysts

- post-infectious (e.g. meningitis, encephalitis) - post-traumatic

- Hypothyroidism

- activating mutations in the genes encoding kisspeptin1 (KISS1) and its receptor KISS1R (GPR54)

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If in addition to the clinical signs and symptoms associated with premature adrenarche, progressive signs of pubertal development are present, such as breast development or menarche in girls and penile or testicular growth in boys, a diagnosis of precocious puberty or precocious pseudo-puberty has to be considered (Table 5). The clinical picture of these two distinct conditions is relatively similar. However, they can be distinguished by analysing the gonadotrophin response to GnRH stimulation. Accordingly, in precocious pseudo-puberty the gonadotrophins are low and remain low after GnRH stimulation. The condition is rare and most of the identified cases in boys are due to simple virilizing and non-classic CAH variants. In CAH, transition from precocious pseudo-puberty to secondary central precocious puberty has been observed and is thought to be due to the induction of the GnRH pulse generator by persistently increased sex steroids (Pescovitz et al., 1984).

1.3.2.1. Metabolic risk in premature adrenarche

Traditionally, idiopathic premature adrenarche (IPA) has been considered to be an extreme variation of normal (Silverman et al., 1952; Voutilainen et al., 1983; Ibanez et al., 1992). However, a series of studies in children with early onset androgen excess provide increasing evidence for the notion that IPA in girls may precede the development of the polycystic ovary syndrome (PCOS). PCOS manifesting during adolescence or early adulthood carries a significantly increased risk of developing the metabolic syndrome (Rosenfield, 2007; Ibáñez et al., 2000; 2009; Ehrmann, 2005) and represents the leading cause of female infertility (Ibanez et al., 1993; Ibáñez et al., 1999; Asuncion et al., 2000). IPA and adolescent PCOS are linked by two similarities, early onset androgen excess and an increased prevalence of insulin resistance.

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The first evidence for a link between premature androgen excess and metabolic syndrome emerged in 1995 from a study in American-Hispanic girls who presented with premature pubarche and evidence of insulin resistance (Oppenheimer et al., 1995). Later on, Ibanez and co-workers published a series of case-control studies in lean Spanish (Catalonian) girls with a history of premature pubarche (PP) that showed evidence of insulin resistance based both on fasting insulin levels and the insulin response to a standard oral glucose tolerance test (oGTT) (Ibanez et al., 1996; Ibáñez et al., 1997; 1998; 2002; Potau et al., 2003). In PP girls from North America, an inverse correlation between insulin sensitivity and ACTH-stimulated Δ5- androgen precursors (17-hydroxypregnenolone, DHEA) has been found, however, the study population was not BMI matched (Vuguin et al., 1999). Overall, impaired insulin sensitivity is a consistent finding in the majority of studies. However, an association of PA/PP with dyslipidaemia, PCOS and neonatal growth restriction is a matter controversy with reported as an inconsistent finding in various study cohorts [see (Idkowiak et al., 2011) for an extensive review].

In conclusion, more research and in particular well-controlled longitudinal studies are required to confidently correlate early-onset androgen excess with metabolic risk in later life.

1.4. A

The aim of this thesis is to investigate the role and the regulation of androgen producing enzymes of the adrenal gland in human disease states characterized by androgen deficiency or androgen excess. Of particular interest in this work are co- factors or co-enzymes supporting steroid modifying enzymes, like P450

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oxidoreductase, cytochrome b5 and PAPS synthases (see chapters 1.1.3 and 1.1.9.1).

This will be examined in the first part of this thesis (chapter 3-6) by clinical, biochemical, genetic and in vitro experimental studies of individual children presenting with a distinct clinical phenotype. In all studies, we will explore ex vivo steroid mass-spectrometry techniques to elucidate underlying conditions and hypothesize that they provide powerful diagnostic tools to diagnose, and differentiate between, distinct steroidogenic defects and co-factor deficiencies.

Secondly, in chapter 7 we will investigate in in vitro studies the regulation of DHEA sulfation by the two known PAPS synthase isoforms, PAPSS1 and PAPSS2. As only deficient PAPSS2 has been previously shown to cause androgen excess (Noordam et al., 2009), we addressed the question why ubiquitously expressed PAPSS1 can not compensate for defect PAPSS2. We hypothesize that a different pattern of subcellular localization of the two isoforms explains the observed superiority of PAPSS2 in the context of DHEA sulfation.

In chapter 8 of this thesis, we will further investigate the concept of DHEA sulfation in pre-receptor androgen metabolism. We have conducted a clinical study in boys and young men with a genetic defect in the steroid sulfatase (STS) enzyme. STS counteracts DHEA sulfation and thus works in the opposite direction of DHEA sulfotransferase and PAPS synthases. The aim of this study is to explore androgen metabolism in these individuals and we hypothesize, in line with previous observations (Hammer, 2005), that STS does not play a major role in the regulation of active androgen levels. Ultimately, we anticipate gaining a deeper understanding of the physiological role of STS during childhood adrenarche.