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Glucocorticoid-remediable aldosteronism
Robert G. Dluhy, Gordon H. Williams *
Department of Medicine, Harvard Medical School and Brigham and Women’s Hospital, Boston, MA, USA
Received 9 August 1995;accepted 12March 1996
Keywords: Glucocorticoid remediable aldosteronism
1. Introduction
2. Discussion
GRA has been known since 1966 to be a form of
mineralocorticoid hypertension characterized by reversal
of the mineralocorticoid-excess state by the exogenous
administration of a glucocorticoid, such as dexamethasone
[ 1,2]. GRA was subsequently shown to be inherited in an
autosomal fashion, establishing the genetic basis for this
disorder. Clinically, the age of onset of hypertension in
GRA patients is in the first two decades of life. This
* Correspondingauthor.
Elsevier Science B.V.
PII SOOOS-6363(96)00069-7
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Blood pressure is known to be influenced by heritable
and environmental factors. The genetic contribution is
complex since multiple genes are known to participate in
the regulation of blood pressure. Once blood pressure is
elevated, structural change also ensues, although the degree of vascular remodeling probably relates to hormonal
and genetic differences. Thus, an attempt to discover ‘hypertensive genes’ or mutations in genes that maintain
normotensionmight be ‘misguided’ owing to the multiplicity of factors that regulate blood pressure. However, the
genetic basis of a hypertensive disorder has recently been
reported, indicating that the discovery of additional mutations in genesthat regulate blood pressurewill be likely in
the future. This disorder - glucocorticoid-remediable aldosteronism(GRA) - illustrates that one successfulstrategy to identify ‘ hypertensive genes’ is to subdivide the
hypertensive syndromeinto intermediatephenotypes.These
phenotypescan then be scrutinized by searching for mutations in candidategenesthat relate to the specific hypertensive phenotypes.
contrastssharply with that of subjectswith other causesof
primary aldosteronism who are usually diagnosed during
the fourth and fifth decadesof life [3]. In contrast to other
etiologies of primary aldosteronism, many patients with
GRA are not hypokalemic and, thus, a potassium level
lacks sensitivity as a screening test for this disorder. GRA
had been reported worldwide, but is more common in
individuals of Scottish and Irish ancestry; no caseshave
yet been reported in Blacks.
In GRA aldosterone secretion is positively and solely
regulated by adrenocorticotropic hormone CACTI-I) [4,5].
As a consequence,exogenous glucocorticoid administration profoundly suppressesaldosterone secretion in affected subjects. In addition, the exogenous administration
of ACTH is associatedwith an exuberant and prolonged
increase in aldosterone secretion and blood pressure in
such subjects. In contrast, in normal individuals ACTH
administration is associated with a rise and subsequent
decline in aldosteronelevels to basal over several days. As
in other etiologies of primary aldosteronism,plasma renin
levels are suppressedin GRA subjects.However, parameters of aldosterone production often overlap the normal
range, consistent with one hypothesis that other unmeasured mineralocorticoids contribute to this mineralocorticoid-excess state. However, it should be recalled that
aldosteronesecretion in GRA is solely regulatedby ACTH.
Thus, there is dysregulation of mineralocorticoid secretion,
since aldosterone does not have a feedback-suppressive
effect on the pituitary production of ACTH.
Two abnormal steroids - 18-hpdroxycortisol (l&OHF)
and 18-oxocortisol (18-OXOF) - were found by Ulick
and others to be greatly overproduced in the urine in
affected GRA subjects [6,7]. Like aldosterone, these 18oxygenated cortisol compounds were found to be positively regulated by ACTH. Moreover, these compounds
R.G. Dluhy, G.H. Williams/Cardiovascular
Al&s&one
synthase
11-0Hase
-3
Unequal crossing over
5
Aldosterone synthase
Chimaeric gene
11-0Hase
Fig. 1. The chimeric gene duplication in GRA, a result of unequal
crossing-overbetween the homologous 11/Shydroxylase and aldosterone
synthase genes. The chimera fuses the S-regulatory sequencesof the
11r3-hydroxylase gene and the 3’-coding sequencesof the aldosterone
synthasegene. From Lifton et al. [14] by permission.
QRA Patlent
Pituitary
v
ACTH
j
cortLsol
Mineralocorticoids
j
JAngiotensin
z
II
f Na
I
Adrenal
Fig. 2. The chimeric gene results in ectopic expression of aldosterone
synthaseactivity in the cortisol-producing zona fasciculata (F) under the
regulation of adrenocorticotropinCACTI-f).This results in overproduction
of aldosterone and other mineralocorticoids (including 18-oxygenated
cortisol compounds),which leads to suppressionof the renht-angiotensin
system and atrophy of the zona glomerulosa (G).
fusing sequencesof the 11P-hydroxylase and aldosterone
synthase genes [14] (Fig. 1). This gene duplication was
shown to contain the 5’ regulatory sequences,confirming
ACTH responsivenessof llP-hydroxylase, fused to more
distal coding sequencesof aldosteronesynthase.Given the
homologies of the 11Phydroxylase and aldosterone synthasegenes,the mechanismthat would likely createsuch a
gene duplication would be a recombination event, an unequal crossing-over between these two genes.
This gene duplication appears to explain all of the
known physiology and biochemistry previously reported in
GRA. First, the promoter region of this chime& gene
contains regulatory sequences of 11/3-hydroxylase and
would be expected to be regulated by ACTH. In addition,
this chimeric gene would allow ectopic expression of
aldosteronesynthaseand enzymatic activity in the ACTHregulated zona fasciculata which normally only secretes
cortisol (Fig. 2). Thus, the presenceof this gene duplication in the zona fasciculata would also explain the production of the C-18 oxygenated cortisol compounds. Finally,
the sole regulation of aldosterone secretion by ACTH and
the suppressionof aldosteronesecretionby glucocorticoids
is explained by this gene mutation, since the aldosterone
synthase,geneis abnormally regulated by ACTH promoter
sequences.
In an additional study in 11 unrelated GRA pedigrees,
all affected subjectshad chimeric gene duplications arising
from unequal crossing-over [ 151.Chimeric geneswere also
found in a second study of 4 additional patients from
unrelated pedigrees [16]. In both studies, the sites of
crossing-over were variable, indicating that in different
pedigreesthese gene duplications arose independently and
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were shown to provide a specific means of diagnosis.
Although they are also modestly overproduced in patients
with aldosterone-producing adenomas (but not bilateral
idiopathic hyperplasia), the ratio of these compounds to
aldosterone rarely exceeds one. In GRA, these 18oxygenated cortisol compounds are greatly overproduced
both in an absolute sense and in relationship to aldosterone, with a ratio that invariably exceeds two. These
abnormal steroids are hybrid compounds since they share
structural featuresof both the aldosteroneand cortisol-producing zones of the adrenal cortex. Specifically, they are
hydroxylated at carbons 17 and 18, featuresof zona fasciculata and zona glomerulosa steroids, respectively. Although it was felt originally that these compounds may
contribute to the mineralocorticoid-excessstate in evidence
in GRA, they have been found in rodents to have only
weak mineralocorticoid activity [8]. However, it is possible
that these hybrid compounds possesssignificant mineralocorticoid activity in humans.
The molecular basis of GRA was discovered by applying a candidategene approachin a pedigree where affected
and unaffected subjects were identified using as the specific biochemical phenotypethesehybrid compounds.Thus,
18-OHF and TH 18-OXOF were measuredin the urine of
individuals in three living generations in a large GRA
pedigree where a proband had been diagnosed [9]. Of 18
at-risk individuals, GRA was diagnosed in 11 additional
patients. A candidate gene approach was then taken with
the hypothesis that a mutation in the aldosterone gene
could explain GRA. A gene expressing aldosterone synthase activity was known to be closely related to a second
gene involved in adrenal steroidogenesis,steroid 11P-hydroxylase [lo-131. Both genes are 95% identical in DNA
sequenceand have identical intron-exon structures. Both
genes are located in close proximity on chromosome 8.
Genetic markers were developed to the aldosterone synthase gene and segregationof these markers was compared
with segregationof GRA in the pedigree described. What
was found in affected subjects, but not their affected
relatives, was an extra gene - a hybrid or chime& gene
871
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872
R.G. Dluhy, G.H. Williams/Cardiovascular
3. Conclusion
The genetic basis for one hypertensive syndrome has
been discovered using a strategy that could be extended to
essential hypertension. That is, hype’rtensive intermediate
phenotypes can be developed using physiologic markers,
such as non-modulation or insulin resistance. Heritability
of such intermediate phenotypes could then be demon-
strated by pedigree screening (as in GRA) or in sib pair
studies. Then a directed search of the genome could be
performed for mutations, using candidate genes appropriate for each phenotype. Such a search for these gene
mutations would only appear to be ‘misguided’ if the
efforts were random, and not directed, as illustrated by the
strategy used in the case of GRA.
References
[l] SutherlandDJ, Ruse JL, Laidlaw JC. Hypertension, increasedaldosterone secretion and low plasma renin activity relieved by dexamethasone.Can Med Assoc J 1%6;95:1109-l 119.
[2] New MI, PetersonRE. A new form of congenital adrenal hyperplasia. J Clin Endocrinol Metab 1%7;27:300-305.
[3] Bravo E, Tarazi R, Dustan H et al. The changing clinical spectrum
of primary aldosteronism.Am J Med 1983;74:641-651.
[4] Gill JR, Barter FC. Overproduction of sodium-retaining steroids by
the zona glomemlosa is adrenocorticotropindependentand mediate
hypertension in dexamethasone-suppressiblealdosteronism. J Clin
Endocrinol Metab 1981;53:331-337.
[51 Obertield SE, Levine LS, Stoner E et al. Adrenal glomendosa
function in patients with dexamethasone-suppressiblehyperaldosteronism. J Clin Endocrinol Metab 1981;53:158-164.
[6] Ulick S, Chu MD, Land MI. Biosynthesis of I&oxocortisol by
aldosterone-producingadrenal tissue. 1983; J Biol Chem 258:54985502.
[7] Gomez-SanchezCE, Montgomery M, Ganguly A et al. Elevated
urinary excretion of 18-oxocortisol in glucocorticoid-suppressible
aldosteronism.J Clin Endocrinol Metab 1984;59:1022-1024.
[8] Gomez-SanchezCE, Gomez-SanchezEP, Smith JS et al. Receptor
binding and biological activity of ll-oxocortisol. Endocrinology
1985;116:6-10.
[9] Rich GM, Ulick S, Cook S et al. Glucocorticoid-remediable aldosteronism in a large kindred: clinical spectrum and diagnosis using a
characteristic biochemical phenotype. Ann Intern Med
1992;116:813-820.
[ 101 Chua SC, SzaboP, Vitek A et al. Cloning of cDNA encoding steroid
11-hydroxylase (P-450,,,). Pmc Nat1 Acad Sci USA 1987;
84:7193-7197.
[ 11I Kawamoto T, Mitsuuchi Y, Ckhnishi T et al. Cloning and expression
of a cDNA for human cytochrome P-450,, as related to primary
aldosteronism.Bicchem Biophys Res Commun 1990,173:309-316.
1121Momet E, Dupont B, Vitek A et al. Characterization of two genes
encoding human steroid 1I-hydroxylase (P-450,,). J Biol Chem
1989;264:20%1-20967.
[13] Ogishima T, Shibata H, Shimada H et al. Aldosterone synthase
cytochromeP-450 expressedin the adrenalsof patients with primary
aldosteronism.J Biol Cbem 1991;266:10731-10734.
1141Lifton RP, Dluhy RG, Powers M et al. A chimeric llhydroxylase/aldosterone synthase gene causes glucocorticoid-remediable aldosteronism and human hypertension. Nature
1992;355:262-265.
[15] Lifton RP, Dluhy RG, Powers M et al. Hereditary hypertension
caused by chimeric gene duplications and ectopic expression of
aldosteronesynthase.Nature Genet 1992266-74.
[161 PascoeL, Cumow KM, Slutsker L et al. Glucocorticoid-suppressible
hyperaldosteronism results from hybrid genes created by unequal
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did not descend from a single ancestral mutation. However, in all studies, the sites of crossing-over were upstream of exon 5 of aldosterone synthase, suggesting that
encoded amino acids in exon 5 are essential for aldosterone synthase enzymatic functions. This hypothesis is
strengthened by the construction of chimeric genes and
expression of aldosterone synthase enzymatic activity in
vitro [16]. In this study, aldosterone synthase enzymatic
activity was retained, provided that the site of fusion
between 11-hydroxylase and aldosterone synthase genes
was upstream of exon 5 of aldosteronesynthase.
As a result of these studies, direct genetic screeningfor
GRA is now possible. Moreover, the presenceof the gene
duplication appears to be 100% sensitive and specific for
diagnosing GRA and is concordant with the measurement
of the abnormal steroids, TH 18-OXOF, and 18-OHF. It is
recommendedthat patients with aldosteronism without radiographic evidence of tumors, as well as individuals with
suppressedlevels of PRA (especially children and young
adults) should be screenedfor GRA. Moreover, since this
is an autosomal dominant disorder, extended screening of
at-risk individuals in affected families will yield many
additional cases.
Treatment of this disorder is gratifying since therapy is
directed. Traditionally, the suppression with glucocorticoids has been the preferred treatment. However, many
subjects appear to have been overtreated with excessive
glucocorticoid dosing, resulting in Cushing’s syndrome. It
is urged that if this modality of treatment is used, the
lowest dose of a shorter-acting glucosteroid be used and
patients monitored carefully for signs of glucocorticoid
excess. Alternative treatments include the aldosterone antagonist, spironolactone, and amiloride, an agent which
inhibits sodium reabsorption by the renal epithelial sodium
channel. Long-term treatment in males with spironolactone
is usually deemedunsatisfactory becauseof the antiandrogenie effects of this drug which causegynecomastia,impotency, or both. Potassium-wasting diuretics can be used
cautiously, but they may precipitate profound hypokalemia. They are probably best used in combination
with potassium-sparing diuretics, with potassium levels
being carefully monitored.
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