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Abstract

Glucocorticoids regulate a broad spectrum of physiologic functions essential for life and play an important role in the maintenance of basal and stress-related homeostasis. At the cellular level, glucocorticoids exert their actions through the ubiquitously expressed human glucocorticoid receptor (hGR). Primary Generalized Glucocorticoid Resistance or Chrousos syndrome is a rare, familial or sporadic genetic condition characterized by generalized, partial, target-tissue resistance to glucocorticoids. The molecular basis of the condition has been ascribed to mutations in the human glucocorticoid receptor (hGR) gene, which impair glucocorticoid signal transduction and reduce tissue sensitivity to glucocorticoids. A consequent increase in the activity of the hypothalamic-pituitary-adrenal axis compensates for the reduced sensitivity of peripheral tissues to glucocorticoids at the expense of ACTH hypersecretion-related pathology. In this article, we review the clinical aspects, molecular mechanisms and implications of this disorder.

Introduction

In humans, glucocorticoids regulate a broad spectrum of physiologic functions essential for life and play an important role in the maintenance of basal and stress-related homeostasis ( 1-3 ). Glucocorticoids are involved in almost every cellular, molecular and physiologic network of the organism and play a pivotal role in critical biologic processes, such as growth, reproduction, intermediary metabolism, immune and inflammatory reactions, as well as central nervous system and cardiovascular functions ( 1-4 ). In addition, glucocorticoids represent one of the most widely used therapeutic compounds often employed in the treatment of inflammatory, autoimmune and lymphoproliferative disorders ( 1 ).

The Glucocorticoid Receptor Gene and Protein

At the cellular level, the action of glucocorticoids is mediated by an intracellular protein, the glucocorticoid receptor (GR) ( 3, 5, 6 ). The human (h) GR belongs to the steroid/thyroid/retinoic acid nuclear receptor superfamily of transcription factor proteins and functions as a ligand-dependent transcription factor that regulates the expression of glucocorticoid-responsive genes positively or negatively. The hGR gene consists of 9 exons and is located on chromosome 5q31.3. Alternative splicing of the hGR gene in exon 9 generates two highly homologous receptor isoforms, the hGR  and hGRβ. These are identical through amino acid 727, but then diverge, with hGR  having an additional 50 amino acids and hGRβ having an additional, nonhomologous 15 amino acids ( Figure 1 ). The hGR  resides primarily in the cytoplasm of cells and represents the classic glucocorticoid receptor that functions as a ligand-dependent transcription factor. The hGRβ, on the other hand, does not bind glucocorticoid agonists, may or may not bind the synthetic glucocorticoid antagonist RU486, has intrinsic, hGR  -independent, gene-specific transcriptional activity, and exerts a dominant negative effect upon the transcriptional activity of hGR  ( 3, 5-8 ).

The hGR  mRNA further expresses multiple isoforms by using at least 8 alternative amino-terminal translation initiation sites ( 9 ). All these hGR  isoforms are differentially distributed in the cytoplasm and/or the nucleus in the absence of ligand, have different transcriptional activity following ligand-induced activation and display distinct transactivation or transrepression patterns on global gene expression examined by cDNA microarray analyses ( 9 ). Therefore, these hGR  isoforms may differentially transduce the glucocorticoid signal to target tissues depending on their selective relative expression and inherent activities. Since hGRβ shares a common amino-terminal domain that contains the same translation initiation sites with the hGR  , the hGRβ variant mRNA might also be translated through the same initiation sites to a similar host of hGRβ isoforms. It is likely that differential cell-specific production and functional differences might also be present between the putative hGRβ translational isoforms.

The hGR gene has at least three different promoters, A, B and C. Promoter A can be used with three untranslated exons, 1A1, 1A2 and 1A3, that contain unique promoter fragments ( 10 ). Therefore, the hGR gene can produce five different transcripts from different promoters that encode the same hGR proteins. Through differential use of these promoters, the expression levels of hGR proteins may vary considerably among tissues. The splice and translational hGR isoforms expressed from different promoters appear to form up to 256 different combinations of homo- and hetero-dimers with varying transcriptional activities. The marked complexicity in the transcription/translation of the hGR gene enables target tissues to differentially respond to circulating glucocorticoid concentrations and accounts for the highly stochastic nature of the glucocorticoid signaling pathway ( 11 ).

Molecular Mechanisms of hGR Action

In the absence of ligand, hGR  resides primarily in the cytoplasm of cells as part of a hetero-oligomeric complex, which contains chaperone heat shock proteins (HSPs) 90, 70 and 50, immunophilins, as well as other proteins ( 12 ). HSP90 regulates ligand binding, as well as cytoplasmic retention of hGR  by exposing the ligand-binding site and masking the two nuclear localization sequences (NLS), NL1 and NL2, which are located adjacent to the DNA-binding domain (DBD) and in the ligand-binding domain (LBD) of the receptor, respectively ( Figure 2A ). Upon ligand-induced activation, the receptor undergoes a conformational change that results in dissociation from this multiprotein complex and translocation into the nucleus ( 12, 13 ). Within the nucleus, the receptor binds as a dimer to glucocorticoid-response elements (GREs) in the promoter regions of target genes, and regulates their expression positively or negatively depending on GRE sequence and promoter context ( 14, 15 ). The GRE-bound hGR  stimulates the transcription of target genes by facilitating the formation of the transcription initiation complex, including the RNA polymerase II and its ancillary components via its AF-1 and AF-2 domains ( 16 ) ( Figure 2B ). To initiate transcription, hGR  uses its transcriptional activation domains, AF-1 and AF-2, as surfaces to interact with nuclear receptor coactivators and chromatin-remodeling complexes. Several coactivators form a bridge between the DNA-bound hGR  and the transcription initiation complex, and facilitate the transmission of the glucocorticoid signal to the RNA polymerase II ( 17-19 ). These include: (1) The p300 and the homologous cAMP-responsive element-binding protein (CREB)-binding protein (CBP), which also serve as macromolecular docking “platforms” for transcription factors from several signal transduction cascades, including nuclear receptors, CREB, AP-1, NF-  B, p53, Ras-dependent growth factor, and STATs. In view of their central position in many signal transduction cascades, the p300/CBP coactivators are also called co-integrators; (2) The p300/CBP-associated factor (p/CAF), which interacts with p300/CBP, but is also a broad transcription coactivator; and (3) The p160 family of coactivators, which preferentially interact with the steroid hormone receptors, and include the steroid receptor coactivator-1 (SRC-1), SRC-2 and SRC-3 ( 17-19 ) ( Figure 2B ). The hGR  also interacts with several other distinct chromatin modulators through its transactivation domains, such as the mating-type switching/sucrose non-fermenting (SWI/SNF) complex and components of the vitamin D receptor-interacting protein/thyroid hormone receptor-associated protein (DRIP/TRAP) complex ( 17-19 ).

The ligand-activated hGR  can also modulate gene expression independently of binding to GREs, by interacting possibly as a monomer with other transcription factors, such as activator protein-1 (AP-1), nuclear factor-  B (NF-  B), p53 and signal transducers and activators of transcription (STATs) ( 20, 21 ). Therefore, hGR  may affect signal transduction cascades through protein-protein interactions with specific transcription factors by influencing their ability to stimulate or inhibit the transcription rates of respective target genes. This activity may be more important than the GRE-mediated one, given that mice harboring a mutant GR, which is active in terms of protein-protein interactions but inactive in terms of transactivation via DNA, survive and procreate, in contrast to mice with a deletion of the entire GR gene that die immediately after birth from severe respiratory distress syndrome ( 22 ). The protein-protein interactions of GR with other transcription factors may take place on the promoters that do not contain GREs, as well as on the promoters that have both GRE(s) and responsive element(s) of transcription factors that interact with GR (“composite promoters”). Suppression of transactivation of other transcription factors through protein-protein interactions may be particularly important in suppression of immune function and inflammation by glucocorticoids ( 22, 23 ). Most of the effects of glucocorticoids on the immune system may be mediated by the interaction between GR and NF-  B, AP-1 and STATs ( 24, 25 ).

Following transcriptional activation or inhibition of glucocorticoid-responsive genes, the hGR  dissociates from the ligand and has a lower affinity for binding to GREs. The unliganded hGR  remains within the nucleus for a considerable length of time and is then exported to the cytoplasm; both within the nucleus and within the cytoplasm the hGR may be recycled and/or degraded in the proteasome ( 3, 5 ) ( Figure 2A ).

Although the transcriptional activity of GR is primarily governed by ligand binding, accumulating evidence suggests that post-translational modifications (PTMs) play an important additional role. These include phosphorylation, ubiquitination, acetylation and sumoylation of the receptor. These covalent changes may affect receptor stability, subcellular localization, as well as the interaction between GR and other proteins ( 5 ).

Chrousos Syndrome

Alterations in any of the molecular mechanisms of hGR  action may lead to alterations in tissue sensitivity to glucocorticoids, which may take the form of glucocorticoid resistance or glucocorticoid hypersensitivity and may be associated with significant morbidity ( 26, 27 ). One such condition that we have extensively investigated over the years is primary Generalized Glucocorticoid Resistance or Chrousos syndrome ( 28-32 ).

Clinical Manifestations

Primary Generalized Glucocorticoid Resistance is a condition first described and elucidated by Chrousos et al . as a rare, familial or sporadic, genetic disorder characterized by generalized, partial, end-organ insensitivity to glucocorticoids ( 31-34 ). This leads to compensatory activation of the hypothalamic-pituitary-adrenal (HPA) axis, inferred hypersecretion of corticotropin-releasing hormone (CRH) and arginine-vasopressin (AVP) in the hypophysial portal system and increased secretion of adrenocorticotropic hormone (ACTH) in the systemic circulation ( 31-34 ). The excess ACTH secretion results in adrenocortical hyperplasia and increased secretion of cortisol and adrenal steroids with salt-retaining (mineralocorticoid) [deoxycorticosterone (DOC) and corticosterone] and/or androgenic [androstenedione, dehydroepiandrosterone (DHEA) and DHEA-sulfate (DHEAS)] activity ( 31-34 ). In recognition of Professor George P. Chrousos' extensive and ground-breaking research work in this field, it has been proposed that the term “Chrousos Syndrome” is used in place of “Primary Generalized Familial or Sporadic Glucocorticoid Resistance” ( 32 ).

The clinical presentation of Chrousos syndrome reflects the pathophysiologic alterations described above and is mainly associated with, respectively, hypertension and/or hypokalemic alkalosis and hyperandrogenism ( 31-34 ) ( Table 1 ). Clinical manifestations of glucocorticoid deficiency might occur, but are rare and were only reported in a young child with hypoglycemic generalized tonic-clonic seizures during the course of a febrile illness ( 35 ), in a newborn baby with severe hypoglycemia, excessive fatigability with feeding, increased susceptibility to infections and concurrent growth hormone deficiency ( 36 ), and in several adult patients with chronic fatigue. The latter might indicate inadequate glucocorticoid target tissue compensation at the central nervous system (CNS) and/or the skeletal muscles by the increased circulating cortisol concentrations ( 31-34 ). Clinical manifestations of androgen excess include ambiguous genitalia in a karyotypic female at birth and gonadotropin-independent precocious puberty in children of either gender; acne, hirsutism and hypofertility in both sexes; male-pattern hair loss, menstrual irregularities and oligo-anovulation in females; and oligospermia in males ( Table 1 ). The impaired fertility in both sexes has been attributed in part to the feedback inhibition of gonadotropin secretion by the elevated androgen concentrations, while the profound anxiety observed in some subjects is probably due to compensatory increases in hypothalamic CRH and AVP secretion. The latter might also predispose the patients to the development of an ACTH-secreting pituitary adenoma. Finally, the elevated circulating ACTH concentrations may be responsible for the observed growth of intratesticular adrenal rests and oligospermia ( 31-34 ).

TABLE 1: Clinical Manifestations and Diagnostic Evaluation of Chrousos Syndrome *

Clinical Presentation

Apparently normal glucocorticoid function in most cases Asymptomatic Hypoglycemia, chronic fatigue (glucocorticoid deficiency?)

Mineralocorticoid excess Hypertension Hypokalemic alkalosis

Androgen excess Children: Ambiguous genitalia at birth**, clitoromegaly, premature adrenarche, gonadotropin-independent precocious puberty Females: Acne, hirsutism, male-pattern hair loss, menstrual irregularities, oligo-anovulation, hypofertility Males: Acne, hirsutism, oligospermia, adrenal rests in the testes, hypofertility

Increased HPA axis activity (CRH/AVP and ACTH hypersecretion) Anxiety Adrenal rests (oligospermia) Pituitary corticotropinoma

Diagnostic Evaluation

Absence of clinical features of Cushing syndrome Normal or elevated plasma ACTH concentrations Elevated serum or plasma cortisol concentrations Increased 24-hour urinary free cortisol excretion Normal circadian and stress-induced pattern of cortisol and ACTH secretion Resistance of the HPA axis to dexamethasone suppression Thymidine incorporation assays: Increased resistance to dexamethasone-induced suppression of phytohemaglutinin-stimulated thymidine incorporation compared to control subjects Dexamethasone-binding assays: Decreased concentration or affinity of the glucocorticoid receptor for the ligand compared to control subjects Molecular studies: Mutations/deletions of the glucocorticoid receptor; functional studies of mutant receptors

* Modified from Reference 32. ** This is the only case of ambiguous genitalia documented in a child with 46,XX karyotype who also harbored a heterozygous mutation of the 21-hydroxylase gene.

The clinical spectrum of Chrousos syndrome is broad, ranging from most severe to mild forms, and a number of patients may be asymptomatic, displaying biochemical alterations only ( 31-34 ) ( Table 1 ). This variable clinical phenotype is due to variations in the tissue sensitivity of the glucocorticoid, mineralocorticoid and/or androgen receptor signaling pathways; variations in the activity of key hormone-inactivating or -activating enzymes, such as the 11β-hydroxysteroid dehydrogenase ( 37 ) and 5  -reductase ( 38 ); and other genetic or epigenetic factors, such as the presence of insulin resistance and visceral obesity ( 34 ).

Molecular Mechanisms

The molecular basis of Chrousos syndrome has been ascribed primarily to mutations in the hGR gene, which impair the molecular mechanisms of hGR action and decrease tissue sensitivity to glucocorticoids. The pathologic hGR gene mutations causing Chrousos syndrome that have been reported to date are shown in Table 2 ( 35, 36, 39-50 ) and Figure 3A . Eight out of 12 of these mutations are heterozygous (4 are homozygous), while 11 out of 12 partially inactivate GR function. Although studies of GR knock-out mice suggested that complete loss-of-function of the GR is incompatible with extrauterine life ( 51 ), one out of 12 of the mutations completely inactivated GR function ( 36 ).

TABLE 2: Mutations of the Human Glucocorticoid Receptor Gene Causing Chrousos Syndrome*
	Mutation Position
Author (Reference)	cDNA Amino acid	Molecular Mechanisms	Genotype	Phenotype
Chrousos et al. (33) Hurley et al. (40)	1922 (A ® T) 641 (D ® V)	Transactivation ¯ Affinity for ligand ¯ (x 3) Nuclear translocation: 22 min Abnormal interaction with GRIP1	Homozygous	Hypertension Hypokalemic alkalosis
Karl et al . (41)	4 bp deletion in exon-intron 6	hGR number: 50% of control Inactivation of the affected allele	Heterozygous	Hirsutism Male-pattern hair-loss Menstrual irregularities
Malchoff et al. (42)	2185 (G ® A) 729 (V ® I)	Transactivation ¯ Affinity for ligand ¯ (x 2) Nuclear translocation: 120 min Abnormal interaction with GRIP1	Homozygous	Precocious puberty Hyperandrogenism

Karl et al. (41) Kino et al. (43)	1676 (T ® A) 559 (I ® N)	Transactivation ¯ Decrease in hGR binding sites Transdominance (+) Nuclear translocation: 180 Abnormal interaction with GRIP1	Heterozygous	Hypertension Oligospermia Infertility
Ruiz et al. (44) Charmandari et al. (49)	1430 (G ® A) 477 (R ® H)	Transactivation ¯ No DNA binding Nuclear translocation: 20 min	Heterozygous	Hirsutism Fatigue Hypertension
Ruiz et al. (44) Charmandari et al. (49)	2035 (G ® A) 679 (G ® S)	Transactivation ¯ Affinity for ligand ¯ (x 2) Nuclear translocation: 30 min Abnormal interaction with GRIP1	Heterozygous	Hirsutism Fatigue Hypertension
Mendonca et al. (45)	1712 (T ® C) 571 (V ® A)	Transactivation ¯ Affinity for ligand ¯ (x 6) Nuclear translocation: 25 min Abnormal interaction with GRIP1	Homozygous	Ambiguous genitalia Hypertension Hypokalemia Hyperandrogenism
Vottero et al. (46)	2241 (T ® G) 747 (I ® M)	Transactivation ¯ Transdominance (+) Affinity for ligand ¯ (x 2) Nuclear translocation ¯ Abnormal interaction with GRIP1	Heterozygous	Cystic acne Hirsutism Oligo-amenorrhea
Charmandari et al. (48)	2318 (T ® C) 773 (L ® P)	Transactivation ¯ Transdominance (+) Affinity for ligand ¯ (x 2.6) Nuclear translocation: 30 min Abnormal interaction with GRIP1	Heterozygous	Fatigue Anxiety Acne Hirsutism Hypertension
Charmandari et al. (50	2209 (T ® C) 737 (F ® L)	Transactivation ¯ Transdominance (time-dependent) (+) Affinity for ligand ¯ (x 1.5) Nuclear translocation: 180 min	Heterozygous	Hypertension Hypokalemia
cMahon et al. (36)	2 bp deletion 773 at nt 2318-9	Transactivation ¯ Affinity for ligand: absent No suppression of IL-6	Homozygous	Hypoglycemia Fatigability with feeding Hypertension
Nader et al. (35)	2141 (G ® A) 714 (R ® Q)	Transactivation ¯ Transdominance (+) Affinity for ligand ¯ (x 2) Nuclear translocation ¯ Abnormal interaction with GRIP1	Heterozygous	Hypoglycemia Hypokalemia Hypertension Mild clitoromegaly Advanced bone age Precocious pubarche
Modified from Reference 32.

The molecular mechanisms through which these various natural hGR mutants affected glucocorticoid signal transduction were systematically investigated in all reported cases with the condition. These mechanisms included: i) the transcriptional activity of the mutant receptors; ii) the ability of the heterozygous mutant receptors to exert a dominant negative effect upon the wild-type receptor; iii) the concentrations and affinity of the mutant receptors for the ligand; iv) the subcellular localization of the mutant receptors and their nuclear translocation following exposure to the ligand; v) the ability of the mutant receptors to bind to GREs; vi) the interaction of the mutant receptors with the glucocorticoid receptor-interacting protein 1 (GRIP1) coactivator, which belongs to the p160 family of nuclear receptor coactivators and plays an important role in hGR  -mediated transactivation of glucocorticoid-responsive genes; and vii) the motility of the mutant receptors inside the nucleus ( 39-50 ).

The molecular defects that have been elucidated in cases with Chrousos syndrome are summarized in Table 2 . Compared with the wild-type receptor, all mutant receptors demonstrated variable reduction in their ability to transactivate glucocorticoid-responsive genes following exposure to dexamethasone, with the most severe impairment observed in the cases of R477H, I559N, V571A and D641V mutations ( 39-50 ). Furthermore, the mutant receptors hGR  I559N, hGR  R714Q, hGR  F737L, hGR  I747M and hGR  L773P exerted a dominant negative effect upon the wild-type receptor, which might have contributed to manifestation of the disease at the heterozygote state ( 35, 39, 43, 46, 48, 50 ). All mutant receptors in which the mutations were located in the ligand-binding domain (LBD) of the receptor showed a variable reduction in their affinity for the ligand, with the most severe reduction observed in the cases of I559N, I747M and V571A mutations ( 39-50 ). The only mutant receptor that demonstrated normal affinity for the ligand was the hGR  R477H, in which the mutation was located at the DNA-binding domain (DBD) ( 49 ).

In subcellular localization and nuclear translocation studies, the pathologic mutant receptors were observed primarily in the cytoplasm of cells in the absence of ligand, except for the hGR  V729I and hGR  F737L receptors, which were localized both in the cytoplasm and the nucleus of cells. Exposure to dexamethasone induced a slow translocation of the mutant receptors into the nucleus, which ranged from 20 min (R477H) to 180 min (I559N and F737L) compared with the wild-type hGR  , which required only 12 min for complete translocation ( 39-50 ). These findings suggest that all hGR mutations affect the nucleocytoplasmic shuttling of the receptor, probably through impairment of the nuclear localization signal (NL)-1 and/or NL2 functions ( 52 ).

All mutant receptors in which the mutations were located in the LBD preserved their ability to bind to DNA ( 39-50 ). The only mutant receptor that failed to bind to DNA was the hGRR477H, in which the mutation was located at the C-terminal zinc finger of the DBD ( 49 ). A major function of the C-terminal zinc finger of the DBD of hGR  is to contribute to receptor homodimerization, a prerequisite for potent receptor binding to GREs and efficient transactivation of glucocorticoid-responsive genes ( 53 ). All mutant receptors except hGR  R477H displayed an abnormal interaction with the GRIP1 (SRC-2) coactivator in vitro ( 39-50 ). Finally, all mutant receptors had dynamic motility defects inside the nucleus of living cells, possibly caused by their inability to properly interact with key partner nuclear molecules of the transcription initiation complex necessary for full activation of glucocorticoid-responsive genes ( 54 ).

Clinical Evaluation of the Patients

The first step in evaluating a patient with suspected Chrousos syndrome is to obtain a complete personal and family history, with particular attention to evidence suggesting hyperactivity of the HPA axis and ACTH hypersecretion-related pathology. In addition, any evidence suggesting possible CNS dysfunction, such as headaches, visual impairment or seizures, should be noted. In female subjects, the regularity of menstrual cycles should be documented. In children and adolescents, growth and sexual maturation should be evaluated carefully, given that progressive hyperandrogenism is almost invariably associated with an increased growth velocity, an advanced bone age and changes in pubertal development.

The physical examination should include an assessment for signs of hyperandrogenism and/or virilization, such as acne, hirsutism, pubic and axillary hair development, male-pattern hair loss and clitoromegaly. Hirsutism should be assessed using the Ferriman-Gallwey score ( 55 ), while pubic hair development should be classified according to Tanner ( 56, 57 ). Arterial blood pressure should be recorded and preferably monitored over a 24-hour period. All subjects should be screened for signs suggestive of Cushing syndrome and undergo a complete neurologic examination.

Endocrinologic Evaluation of the Patients

The concentrations of plasma ACTH, renin activity (recumbent and upright) and aldosterone, as well as those of serum cortisol, testosterone, androstenedione, DHEA and DHEAS should be recorded in the morning. Determination of the 24-hour urinary free cortisol (UFC) excretion on 2 or 3 consecutive days is central to the diagnosis, given that patients with the condition demonstrate increased 24h UFC excretion in the absence of clinical manifestations suggestive of hypercortisolism. Plasma ACTH concentrations may be normal or high. However, the circadian pattern of ACTH and cortisol secretion and their responsiveness to stressors are preserved, albeit at higher concentrations.

The responsiveness of the HPA axis to exogenous glucocorticoids should also be tested with dexamethasone suppression testing. Increasing doses of dexamethasone should be given orally at midnight every other day, and a serum sample should be drawn at 0800h the following morning for determination of serum cortisol and dexamethasone concentrations. Affected subjects demonstrate resistance of the HPA axis to dexamethasone suppression, which may vary depending on the severity of the condition. The concurrent measurement of serum dexamethasone concentrations is suggested in order to exclude the possibility of increased metabolic clearance or decreased absorption of this medication.

Cellular and Molecular Studies in the Patients

Thymidine incorporation assays and dexamethasone-binding assays on peripheral blood mononuclear cells in association with sequencing of the hGR gene are necessary to confirm the diagnosis and to provide genetic counseling ( 39-50 ) ( Table 1 ). In affected subjects, the thymidine incorporation assays reveal resistance to dexamethasone-induced suppression of phytohemaglutinin-stimulated thymidine incorporation, while the dexamethasone-binding assays often show d ecreased affinity of the hGR receptor for the ligand compared to control subjects. Sequencing of the coding region of the hGR gene, including the intron/exon junctions, will reveal mutations or deletions in most ( 39-50 ) but not all ( 58 ) cases with Chrousos syndrome. Finally, once the structural defect is determined, its adverse effects on receptor function should be confirmed using in vitro mutagenesis and standardized assays that examine the ability of the mutant receptor to transactivate glucocorticoid-responsive genes.

Management of the Patients

The aim of treatment in Chrousos syndrome is to suppress the excess secretion of ACTH, thereby suppressing the increased production of adrenal steroids with mineralocorticoid and/or androgenic activity. Treatment involves administration of high doses of mineralocorticoid-sparing synthetic glucocorticoids, which activate the mutant and/or wild-type hGR  , and suppress the endogenous secretion of ACTH in affected subjects ( 31-34 ). Adequate suppression of the HPA axis is of particular importance in cases of severe impairment of hGR  action, given that long-standing corticotroph hyperstimulation in association with decreased glucocorticoid negative feedback inhibition at the hypothalamic and pituitary levels may lead to the development of an ACTH-secreting adenoma ( 39 ). Long-term dexamethasone treatment should be carefully titrated according to the clinical manifestations and biochemical profile of the affected subjects ( 31-34 ).

Conclusions and Recommendations

The glucocorticoid receptor is a ubiquitously expressed intracellular, ligand-dependent transcription factor, which mediates the action of glucocorticoids and influences physiologic functions essential for life. Mutations in the hGR gene impair one or more of the molecular mechanisms of glucocorticoid action, thereby altering tissue sensitivity to glucocorticoids. A consequent increase in the activity of the HPA axis compensates for the reduced sensitivity of peripheral tissues to glucocorticoids at the expense of ACTH hypersecretion-related pathology. The variable clinical phenotype of Chrousos syndrome, including chronic fatigue, mild hypertension and hyperandrogenism, in association with the difficulties encountered in establishing the correct diagnosis may account for the low reported prevalence of the condition, given that many cases may be unrecognized and misclassified. We recommend screening with 24h UFC excretion and sequencing of the hGR gene in patients with manifestations of mineralocorticoid and androgen excess (hypertension, hirsutism, menstrual irregularities, oligo-anovulation, impaired fertility), in whom detailed investigations fail to reveal an underlying etiology.