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`Endocrine Reviews, 2022, Vol. 43, No. 1, 91–159
`
`doi:10.1210/endrev/bnab016
`
`Review
`
`Review
`
`Congenital Adrenal Hyperplasia—Current
`Insights in Pathophysiology, Diagnostics, and
`Management
`Hedi L. Claahsen - van der Grinten,1 Phyllis W. Speiser,2 S. Faisal Ahmed,3
`Wiebke Arlt,4,5 Richard J. Auchus,6 Henrik Falhammar,7,8 Christa E. Flück,9
`Leonardo  Guasti,10 Angela  Huebner,11 Barbara  B.  M.  Kortmann,12
`Krone,13,14
`P.  Merke,15 Walter 
`L. Miller,16
`Nils 
`Deborah 
`Anna  Nordenström,17,18 Nicole  Reisch,19 David  E.  Sandberg,20
`Nike  M.  M.  L.  Stikkelbroeck,21 Philippe  Touraine,22 Agustini  Utari,23
`Stefan A. Wudy,24 and Perrin C. White25
`
`1Department of Pediatric Endocrinology, Radboud University Medical Centre, Amalia Childrens Hospital,
`Nijmegen, The Netherlands; 2Cohen Children’s Medical Center of NY, Feinstein Institute, Northwell
`Health, Zucker School of Medicine, New Hyde Park, NY 11040, USA; 3Developmental Endocrinology
`Research Group, School of Medicine Dentistry & Nursing, University of Glasgow, Glasgow, UK; 4Institute
`of Metabolism and Systems Research (IMSR), College of Medical and Dental Sciences, University of
`Birmingham, Birmingham, UK; 5Department of Endocrinology, Queen Elizabeth Hospital, University
`Hospitals Birmingham NHS Foundation Trust, Birmingham, UK; 6Division of Metabolism, Endocrinology,
`and Diabetes, Departments of Internal Medicine and Pharmacology, University of Michigan, Ann Arbor,
`MI 48109, USA; 7Department of Molecular Medicine and Surgery, Karolinska Intitutet, Stockholm,
`Sweden; 8Department of Endocrinology, Karolinska University Hospital, Stockholm, Sweden; 9Pediatric
`Endocrinology, Diabetology and Metabolism, Inselspital, Bern University Hospital, University of Bern, 3010
`Bern, Switzerland; 10Centre for Endocrinology, William Harvey Research Institute, Bart’s and the London
`School of Medicine and Dentistry, Queen Mary University of London, London, UK; 11Division of Paediatric
`Endocrinology and Diabetology, Department of Paediatrics, Universitätsklinikum Dresden, Technische
`Universität Dresden, Dresden, Germany; 12Radboud University Medical Centre, Amalia Childrens Hospital,
`Department of Pediatric Urology, Nijmegen, The Netherlands; 13Department of Oncology and Metabolism,
`University of Sheffield, Sheffield, UK; 14Department of Medicine III, University Hospital Carl Gustav Carus,
`Technische Universität Dresden, Dresden, Germany; 15National Institutes of Health Clinical Center and the
`Eunice Kennedy Shriver National Institute of Child Health and Human Development, Bethesda, MD 20892,
`USA; 16Department of Pediatrics, Center for Reproductive Sciences, and Institute for Human Genetics,
`University of California, San Francisco, CA 94143, USA; 17Department of Women’s and Children’s Health,
`Karolinska Institutet, Stockholm, Sweden; 18Pediatric Endocrinology, Karolinska University Hospital,
`Stockholm, Sweden; 19Medizinische Klinik IV, Klinikum der Universität München, Munich, Germany;
`20Department of Pediatrics, Susan B. Meister Child Health Evaluation and Research Center, University of
`Michigan, Ann Arbor, MI 48109, USA; 21Radboud University Medical Centre, Department of Endocrinology,
`Nijmegen, Netherlands; 22Department of Endocrinology and Reproductive Medicine, Center for Rare
`
`ISSN Print: 0163-769X
`
`ISSN Online: 1945-7189
`
`Printed: in USA
`
`© The Author(s) 2021. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved.
`
`For permissions, please e-mail: journals.permissions@oup.com
`
`https://academic.oup.com/edrv 91
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`1
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`NEUROCRINE-1049
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`Endocrine Reviews, 2022, Vol. 43, No. 1
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`Endocrine Diseases of Growth and Development, Center for Rare Gynecological Diseases, Hôpital
`Pitié Salpêtrière, Sorbonne University Medicine, Paris, France; 23Division of Pediatric Endocrinology,
`Department of Pediatrics, Faculty of Medicine, Diponegoro University, Semarang, Indonesia; 24Steroid
`Research & Mass Spectrometry Unit, Laboratory of Translational Hormone Analytics, Division of
`Paediatric Endocrinology & Diabetology, Justus Liebig University, Giessen, Germany; and 25Division of
`Pediatric Endocrinology, UT Southwestern Medical Center, Dallas TX 75390, USA
`
`ORCiD numbers: 0000-0003-0181-0403 (H. L. CLaahsen); 0000-0002-0565-8325 (P. W. Speiser); 0000-0001-5106-9719 (W. Arlt);
`0000-0002-5622-6987 (H. Falhammar); 0000-0003-1338-5192 (A. Huebner); 0000-0002-8462-7753 (P. Touraine); 0000-0001-
`6262-0289 (P. C. White).
`
`Abbreviations: 11KT, 11-ketotestosterone; 11OHD, 11-hydroxylase deficiency; 17OH-Preg, 17-hydroxypregnenolone; 17OHD,
`17-hydroxylase deficiency; 17OHP, 17-hydroxyprogesterone; 21OHD, 21-hydroxylase deficiency;AAV, adeno-associated
`virus; ACTH, adrenocorticotropic hormone, corticotropin; BMD, bone mineral density; BMI, body mass index; CAH,
`congenital adrenal hyperplasia; cAMP, cyclic adenosine monophosphate; cIMT, carotid intima media thickness; COUP-
`TFII, Chicken Ovalbumin Upstream Promotor-Transcription Factor-2; CRF, corticotropin-releasing factor; CRH, corticotropin-
`releasing hormone; CYP21A2, 21-hydroxylase; CYP19A1, Aromatase; Dex, dexamethasone; DHEA, dihydroeoiandrostene
`dione; DHEAS, dehydroepiandrosterone sulfate; DELFIA, dissociation-enhanced lanthanide fluoroimmunoassay; DHT,
`5-dihydrotestosterone; DOC, 11 deoxycorticosterone; DSD, differences in sex development; ESC, embryonic stem cell;
`FSH, follicle-stimulating hormone; GC, gas chromatography; GnRH, gonadotropin-releasing hormone;HC, hydrocortisone;
`HOMA-β, homeostatic model assessment; HSD3B, 3β-hydroxysteroid dehydrogenase; HSD17B, 17β-hydroxysteroid
`dehydrogenase; IL, interleukin; iPSC, inducible pluripotent stem cell; LC, liquid chromatography; LC-MS/MS, liquid
`chromatography-tandem mass spectrometry; LH, luteinizing hormone; MC2R, adrenocorticotropic hormone receptor; MLPA,
`multiplex ligation-dependent probe amplification; MR, mineralocorticoid receptor; MRI, magnetic resonance imaging; MS,
`mass spectrometry; NC, nonclassic; OMM, outer mitochondrial membrane; PGD, preimplantation genetic diagnosis; POR,
`P450 oxidoreductase; TARTs, testicular adrenal rest tumors; SF-1, steroidogenic factor-1; SRD5A1, 5α-reductase type 1;
`StAR, steroidogenic acute regulatory protein; SV, simple virilizing; SW, salt wasting; TNXB, tenascin-X; WMD, weighted
`mean difference.
`
`Received: 30 January 2021; Editorial Decision: 27 April 2021; First Published Online: 7 May 2021; Corrected and Typeset:
`9 September 2021.
`
`Abstract 
`
`Congenital adrenal hyperplasia (CAH) is a group of autosomal recessive disorders af-
`fecting cortisol biosynthesis. Reduced activity of an enzyme required for cortisol pro-
`duction leads to chronic overstimulation of the adrenal cortex and accumulation of
`precursors proximal to the blocked enzymatic step. The most common form of CAH is
`caused by steroid 21-hydroxylase deficiency due to mutations in CYP21A2. Since the last
`publication summarizing CAH in Endocrine Reviews in 2000, there have been numerous
`new developments. These include more detailed understanding of steroidogenic path-
`ways, refinements in neonatal screening, improved diagnostic measurements utilizing
`chromatography and mass spectrometry coupled with steroid profiling, and improved
`genotyping methods. Clinical trials of alternative medications and modes of delivery
`have been recently completed or are under way. Genetic and cell-based treatments are
`being explored. A large body of data concerning long-term outcomes in patients affected
`by CAH, including psychosexual well-being, has been enhanced by the establishment of
`disease registries. This review provides the reader with current insights in CAH with spe-
`cial attention to these new developments.
`
`Key Words: Steroid biosynthesis, 21-hydroxylase deficiency, CYP21A2, glucocorticoid, mineralocorticoid, cortisol,
`aldosterone
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`Endocrine Reviews, 2022, Vol. 43, No. 1
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`Graphical Abstract 
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`ESSENTIAL POINTS
`
` • Congenital adrenal hyperplasia (CAH) is most often caused by de!ciency of steroid 21 hydroxylase encoded by
`CYP21A2
` • Allelic variants are associated with a spectrum of phenotypes
` • CAH in its severe, classic form includes cortisol and aldosterone de!ciencies, as well as androgen excess
` • Newer concepts in steroid biosynthesis, hormonal and genetic diagnostic tools, and novel therapeutics have
`expanded our understanding of CAH
` • Long-term sequelae of this disease have been reported in detail and strategies are being developed to improve
`quality of life for these patients
`
`Congenital adrenal hyperplasia (CAH) is an inherited in-
`ability to synthesize cortisol. Approximately 90% to 99%
`of cases of CAH are caused by 21-hydroxylase de!ciency
`(21OHD) caused by mutations in the CYP21A2 gene (1,
`
`2); the terms CAH and 21OHD will be used interchange-
`ably in this article. The literature has historically de-
`scribed classic and nonclassic (NC) forms of this disorder,
`although current thinking views CYP21A2 allelic variants
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`Endocrine Reviews, 2022, Vol. 43, No. 1
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`and their phenotypic manifestations as a continuum. The
`classic form, occurring in 1 in 14  000 to 18  000 based
`on newborn screening (Table 1), is de!ned by severely re-
`duced or absent enzyme activity with impaired cortisol
`production manifesting clinically in the neonatal period.
`In the most severe, salt-wasting (SW) form of classic CAH,
`there is little or no residual enzymatic activity, resulting in
`cortisol and aldosterone de!ciency. Lack of negative feed-
`back on the hypothalamic–pituitary–adrenal axis leads to
`excess adrenal androgen production as elevated precursor
`steroids are shifted to the nonaffected androgen pathways.
`If not promptly treated, infants with this form of CAH
`quickly develop potentially fatal “salt-wasting crises”
`with hyponatremia, hyperkalemia, acidosis, and shock.
`Those infants who produce slightly more aldosterone are
`less likely to suffer acute SW crisis, but such patients still
`have severe cortisol de!ciency and markedly elevated ad-
`renal androgen production. They are said to have “simple
`virilizing” (SV) CAH, associated with residual enzymatic
`activity of 1% to 5% of normal. All infants affected with
`classic CAH bene!t from glucocorticoid plus adjunctive
`mineralocorticoid treatment at least within the !rst year
`
`Table 1. Incidence of CAH in different countries
`
`of life, when there is relative renal tubular resistance to
`the salt-retaining effects of aldosterone in early infancy
`(28) and low sodium content of infant diets (29).
`Whereas gonadal development is normal, severely in-
`creased prenatal adrenal androgen production leads to
`virilization of the female external genitalia (30), including
`variable degrees of clitoral enlargement and labial fusion.
`The genital appearance of affected 46,XX infants is occa-
`sionally indistinguishable from that of male genitals with
`penis and scrotum but empty of gonads. Müllerian duct
`development is normal, except for the formation of a uro-
`genital sinus with conjoined urethra and vagina. Thus,
`reproductive potential exists in females despite atypical
`external genitalia. Males have normal external genitalia.
`Wolf!an duct development is normal in males but absent
`in females, who continue to produce COUP-TFII (Chicken
`Ovalbumin Upstream Promoter-Transcription Factor-2),
`which induces Wolf!an duct involution (31).
`Adverse sequelae in CAH patients occur as a result of ad-
`renal hormone imbalance and from chronic glucocorticoid
`therapy (32). Androgen excess can cause inappropriately
`rapid somatic growth, accelerated skeletal maturation,
`
`Country
`
`Complete national data?
`
`Sample size
`
`1/Incidence
`
`PPV (term infants or overall)
`
`Reference
`
`Argentina (Buenos Aires)
`Australia*
`Australia (New South Wales)
`Australia (Western Australia)*
`Brazil
`Brazil (Goias state)
`Brazil (Minas Gerais state)
`Brazil (Rio Grande do Sul state)
`China
`China (Beijing)
`Croatia
`Cuba
`Czech Republic
`France
`Germany (Bavaria)
`India
`Israel
`Japan (Sapporo)
`Japan (Tokyo)
`Netherlands
`New Zealand
`Sweden
`Turkey
`United Arab Emirates
`United Kingdom*
`Uruguay
`
`No
`Yes
`No
`No
`No
`No
`No
`No
`No
`No
`Yes
`Yes
`Yes
`Yes
`No
`No
`Yes
`No
`No
`Yes
`Yes
`Yes
`No
`Yes
`Yes
`Yes
`
`80 436
`
`185 854
`550 153
`748 350
`82 603
`159 415
`108 409
`30 000
`44 360
`532 942
`621 303
`888 891
`6 012,798
`1 420,102
`55 627
`1 378,132
`498 147
`2 105,108
`2 235,931
`1 175,988
`2 737,932
`241 083
`750 365
`
`190 053
`
`8937
`18 034
`15 488
`14 869
`14 967
`10 325
`19 927
`13 551
`6084
`7393
`14 403
`15 931
`12 520
`15 699
`12 457
`6334
`16 910
`20 756
`21 264
`17 468
`26 727
`14 260
`15 067
`9030
`18 248
`15 800
`
`50
`N/A
`1.8
`N/A
`
`28.6
`2.1
`1.6
`
`3.0
`
`0.3
`1.6
`2.3
`5
`
`16.5
`8
`25.8
`24.7
`
`25.1
`1.9
`
`N/A
`
`(3)
`(4)
`(4)
`(5)
`(6)
`(7)
`(8)
`(9)
`(10)
`(11)
`(12)
`(13)
`(14)
`(15)
`(16)
`(17)
`(18)
`(19)
`(20)
`(21)
`(22)
`(2)
`(23)
`(24)
`(25)
`(26)
`
`Data are from studies published in 2008 and later; Earlier studies are summarized by (27) and (2). Data are from newborn screening except those marked with an
`asterisk (*),which are from national case registries.
`Abbreviation: PPV, positive predictive value.
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`and reduced adult height. A  systematic review and meta-
`analysis for >1000 classic CAH patients found shorter than
`average stature for mid-parental heights (–1.03 standard
`deviations, corresponding to ~7  cm) (33), but many of
`these children were diagnosed before the implementation
`of neonatal screening and did not receive the bene!t of
`early initiation of treatment.
`Elevated levels of adrenal androgens affect the hypothal-
`amic–pituitary–gonadal axis. Central precocious puberty is
`a risk in patients experiencing prolonged periods of poor
`hormonal control. Young women with well-controlled
`CAH usually experience normal menarche (34), but poor
`control is associated with acne, female hirsutism, male pat-
`tern baldness, altered body habitus, irregular menses, and
`subnormal fertility (35). Males with poor hormonal con-
`trol may develop small testes and benign testicular adrenal
`rest tumors (TARTs) (see section “Long-term sequelae,”
`“Gonadal function in males,” “Testicular adrenal rest tu-
`mors”) (36).
`Individuals affected with milder allelic variants (ie, NC
`CAH) tend to present to medical attention after infancy, hence
`the former term “late-onset” CAH. The associated alleles en-
`code enzymes with residual activity of 20% to 50%. Thus,
`these individuals typically have normal basal cortisol and al-
`dosterone production but mildly elevated levels of adrenal
`androgens; however, suboptimal cortisol levels after adreno-
`corticotropic hormone (ACTH) stimulation are reported in
`up to 30% of patients (37). Children may present with symp-
`toms due to elevated adrenal androgens such as premature
`adrenarche, acne, and accelerated skeletal maturation but
`many, especially males, are asymptomatic. Adolescent girls or
`adult women may present with hirsutism, oligomenorrhea,
`acne, and subnormal fertility (37). Because NC CAH is not
`the primary target of neonatal screening and is rarely de-
`tected by that strategy, the true prevalence of this milder dis-
`order is unclear. The estimated prevalence is ~1 in 200 in the
`Caucasian population (38).
`in
`Since the
`last publication summarizing CAH
`Endocrine Reviews in 2000 (1), there have been nu-
`merous new developments. These include more detailed
`understanding of steroidogenic pathways, re!nements in
`neonatal screening, improved diagnostic measurements
`utilizing high-throughput liquid chromatography-tandem
`mass spectrometry (LC-MS/MS) coupled with steroid pro-
`!ling, and improved genotyping methods. Clinical trials of
`alternative medications and modes of delivery have been
`recently completed or are under way, with the nearer pro-
`spect of genetic and cell-based treatments and a large body
`of data concerning long-term outcomes in patients affected
`by CAH, including psychosexual well-being, enhanced by
`the establishment of disease registries.
`Much remains to be learned in several other domains
`spanning fetal life through adulthood. Both human and
`
`animal studies have illuminated risks of antenatal dexa-
`methasone (Dex) treatment. Noninvasive prenatal diag-
`nosis of CAH in families with known CYP21A2 pathogenic
`genotypes has been accomplished by analysis of circulating
`free fetal DNA in maternal blood in proof-of-concept
`studies, but is not yet widely available. Genital recon-
`structive surgery in affected females is no longer viewed
`as an emergency procedure, and indeed the practice of
`genital surgery in infancy has been questioned. Shared de-
`cision making among parents, patients, surgeons, endocrin-
`ologists, mental health providers, and support groups has
`been promoted as model for optimal care. Bene!t-to-risk
`ratio for no surgery, or early or late genital surgery for fe-
`males with CAH remains to be determined. Unfortunately,
`even in advanced societies, medical care for CAH is neg-
`lected, increasing the risk for cardiovascular or metabolic
`morbidities due to suboptimal corticosteroid therapy.
`Methods to improve transition of care from pediatric to
`adult healthcare, as well as patient and provider education,
`are important goals.
`This multiauthored review is the result of a planned
`European CAH Symposium, which was postponed due
`to the Covid-19 pandemic. The large international group
`of authors contributed innovative approaches to under-
`standing and managing this condition.
`
`Basic Principles of Steroid Synthesis and
`Adrenal Enzymatic Defects
`
`Physiology and Pathophysiology of
`Steroidogenesis
`
`Steroidogenesis in the adrenal cortex takes place in 3
`concentric zones: the outermost zona glomerulosa (min-
`eralocorticoid biosynthesis), the zona fasciculata (gluco-
`corticoid biosynthesis), and the innermost zona reticularis
`(sex steroid precursor biosynthesis). It entails conversion
`of cholesterol to active steroid hormones, and involves
`many enzymes, cofactors, and accessory proteins (Fig. 1).
`Most of these are expressed in the appropriate zones of the
`adrenal cortex, with others expressed in the gonads, pla-
`centa, and some “peripheral” tissues; these factors and the
`conditions caused by their mutations have been studied in
`detail (39). Mutations have been described in most of the
`genes encoding these proteins; those that disrupt cortisol
`synthesis with compensatory elevations in ACTH cause
`CAH, but in common parlance “CAH” refers to 21OHD.
`This section describes all enzymatic conversions required to
`synthesize cortisol.
`
`Cholesterol side-chain cleavage
`Steroidogenesis is initiated by the conversion of chol-
`esterol to pregnenolone, catalyzed by the cholesterol
`
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`Figure 1. Adrenal steroidogenesis. Enzymes are boxed with dotted lines extending to arrows denoting each enzymatic conversion; 2 enzymes,
`
`CYP11B2 and CYP17, catalyze several successive enzymatic conversions. Accessory proteins required for activity of cytochrome P450 enzymes are
`
`shown next to each such enzyme: POR, P450 oxidoreductase, required by CYP enzymes in the endoplasmic reticulum; FDXR/FDX1, ferredoxin re-
`
`ductase and ferredoxin, required by mitochondrial CYP enzymes. Cytochrome B5 (CYP5A) is required for full 17,20-lyase activity of CYP17A1. There
`
`are 2 11β-hydroxysteroid dehydrogenase isozymes; HSD11B1, expressed mainly in the liver, catalyzes reduction (eg, cortisone to cortisol), whereas
`HSD11B2, expressed mainly in the kidney, catalyzes oxidation (eg, cortisol to cortisone). The steps affected by 21OHD, including steroids secreted in
`
`increased amounts in this disease, are denoted by red lines and red lettering. Steps taking place only in the adrenal glands are in unshaded boxes;
`
`steps taking place partly or predominantly outside the adrenal cortex are denoted by shaded boxes. Planar structures of cholesterol, aldosterone,
`
`cortisol, and testosterone are illustrated; the position of the 11-oxo (11-keto) group in 11-ketotestosterone is illustrated in green. Colored rectangles
`
`indicate the following: gray, early steps of steroidogenesis common to all zones of the cortex; orange, steps in the zona glomerulosa leading to aldos-
`
`terone; blue, steps in the zona fasciculata leading to cortisol; magenta; steps in the zona reticularis and extra-adrenal tissues leading to androgens;
`
`purple, the “backdoor” or alternate pathway from 17-OH progesterone to dihydrotestosterone (for clarity, the alternative pathway from progesterone
`
`is not shown); green, conversions leading to 11-oxo androgens.
`
`side-chain cleavage enzyme, CYP11A1 (P450scc). To
`initiate steroidogenesis, cholesterol from cytoplasmic
`storage depots must reach CYP11A1 on the inner mito-
`chondrial membrane; this cholesterol in&ux requires the
`steroidogenic acute regulatory protein (StAR), acting on
`the outer mitochondrial membrane (OMM) (40). The
`action of StAR requires its phosphorylation and inter-
`action with some other proteins, but the exact mech-
`anism of StAR’s action remains under investigation (41,
`42). Mutations in StAR cause another rare form of CAH,
`congenital lipoid adrenal hyperplasia, in which virtu-
`ally no steroid hormones are made and 46,XY fetuses
`are phenotypically female due to impaired testicular
`
`steroidogenesis (43, 44). CYP11A1 defects were once
`considered incompatible with term pregnancy; however,
`more than 30 cases of such defects have been reported
`(40). These 2 conditions are clinically and hormonally
`indistinguishable, but lipoid CAH is typically associ-
`ated with very large adrenals, whereas CYP11A1 de!-
`ciency is not; gene sequencing is needed for de!nitive
`diagnosis. Milder “nonclassical” forms of these condi-
`tions have been reported with intermediate phenotypes
`(45-48). CYP11A1 is one of 7 human mitochondrial
`cytochrome P450 (CYP) enzymes, all of which require
`electron donation via ferredoxin and ferredoxin re-
`ductase (49). Mutations in ferredoxin have not been
`
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`reported, but several patients have been described with
`ferredoxin reductase mutations that disrupt synthesis of
`iron/sulfur centers, causing neuropathic deafness, optic
`atrophy, encephalopathy, and developmental delay (50-
`52); impaired steroidogenesis is to be expected but not
`yet reported.
`
`3β-Hydroxysteroid dehydrogenase
`Once pregnenolone is produced, it may be converted to pro-
`gesterone by 3β-hydroxysteroid dehydrogenase (HSD3B,
`3β-HSD). There are 2 human HSD3B genes: HSD3B1 en-
`codes an isozyme found in the placenta, brain, liver, and
`elsewhere; HSD3B2 encodes an isoenzyme found in the
`adrenals and gonads. Both of these isozymes can convert
`the ∆ 5-steroids (pregnenolone, 17-hydroxypregnenolone
`[DHEA],
`and
`[17OHPreg], dehydroepiandrosterone
`androstenediol) to the corresponding ∆ 4-steroids (proges-
`terone, 17OH-progesterone [17OHP], androstenedione,
`testosterone) (53), but the placental/hepatic HSD3B1 has
`a low Michaelis–Menten constant (Km), permitting it to
`act on low concentrations of steroids in the circulation
`(54), whereas the Km for the adrenal/gonadal HSD3B2 is
`10-fold higher (55), so it acts only on locally produced,
`intraglandular steroids. Mutations in HSD3B2 cause a rare
`form of CAH, characterized by elevated ratios of ∆ 5/∆ 4 ster-
`oids, notably 17OH-Preg/17OH-progesterone (17OHP),
`that are >8 SD above normal (56, 57). The low Km of hep-
`atic HSD3B1 permits it to convert some of the elevated
`17OH-Preg to 17OHP, engendering false positives in new-
`born screening programs for 21OHD (58). HSD3B2 de!-
`ciency causes DSD in both sexes: genetic females are mildly
`virilized because some fetal adrenal DHEA is converted to
`testosterone by HSD3B1; genetic males synthesize some an-
`drogens by peripheral conversion of DHEA, but these are
`insuf!cient for complete male genital development (59).
`
`17α-Hydroxylase/17,20-lyase
`Pregnenolone can also be converted to 17OH-Preg by
`17α-hydroxylase (CYP17A1, P450c17). CYP17A1 cata-
`lyzes both 17  α -hydroxylase and 17,20-lyase activities.
`The 17  α -hydroxylase activity converts pregnenolone to
`17OHPreg and progesterone to 17OHP. The 17,20-lyase
`activity can convert 17OH-Preg to DHEA, but very little
`17OHP is converted to androstenedione because the human
`enzyme catalyzes this reaction poorly (60, 61). The activ-
`ities of CYP17A1 are expressed in a zone-speci!c fashion:
`the enzyme is absent in the adrenal zona glomerulosa, hence
`pregnenolone yields mineralocorticoids; only the 17  α
`-hydroxylase activity is found in the zona fasciculata, thus
`pregnenolone yields cortisol; both activities are present in
`the zona reticularis, hence pregnenolone yields 19-carbon
`(C19) precursors of sex steroids (Fig. 1). The principal
`
`factor regulating 17,20-lyase activity is electron transport
`from NADPH via cytochrome P450 oxidoreductase (POR)
`with the assistance of cytochrome b5 (b5). CYP17A1 muta-
`tions causing 17-hydroxylase de!ciency (17OHD) are rare
`except in Brazil and China. Lack of CYP17A1 prevents sex
`steroid biosynthesis, yielding a female phenotype in 46,XY
`males and sexual infantilism in both sexes; overproduction
`of 11-deoxycorticosterone (DOC) in the zona fasciculata
`typically causes mineralocorticoid hypertension; cortisol
`is not produced, but corticosterone substitutes for gluco-
`corticoid requirements (62). Rare cases of apparently
`isolated 17,20-lyase de!ciency may be attributable to mu-
`tations in CYP17A1, b5 (CYB5 gene) or POR (63-65).
`The enzymology of adrenal 21-hydroxylase (CYP21A2,
`P450c21, encoded by CYP21A2 within the HLA locus),
`is discussed in section “Basic principles of steroid syn-
`thesis and adrenal enzymatic defects,” “Enzymology of
`CYP21A2.”
`
`P450 oxidoreductase
`All microsomal cytochrome P450 (CYP) enzymes, including
`CYP17A1, CYP21A2, CYP19A1 (aromatase, P450aro), as
`well as the drug-metabolizing CYP enzymes of the liver,
`require the activity of POR, a &avoprotein that transfers
`electrons from NADPH to all microsomal CYP enzymes (49).
`Mutations in POR cause POR de!ciency; patients have been
`described with highly variable clinical and hormonal !nd-
`ings depending on the underlying mutations (66-72). Most
`POR mutations impair CYP17A1, especially 17,20-lyase ac-
`tivity (including the G539R POR variant with a phenotype
`simulating isolated 17,20 lyase de!ciency) (63, 68, 73), with
`CYP21A2 and CYP19A1 being affected variably, depending
`on the POR mutation. It is dif!cult to reach de!nitive con-
`clusions about phenotype–genotype correlations with such
`rare disorders, although there is a suggestion that compound
`heterozygotes carrying R457H in trans with null muta-
`tions tend to have a more severe phenotype (72). Findings
`range from severely affected infants with 46,XX and 46,XY
`disorders/differences of sex development (DSD), cortisol
`de!ciency, and the Antley–Bixler skeletal malformation syn-
`drome to mildly affected women who appear to have poly-
`cystic ovary syndrome, or mildly affected men with gonadal
`insuf!ciency. The skeletal phenotype probably results from
`diminished activity of CYP26B1, a POR-dependent enzyme
`that degrades retinoic acid (74). POR mutations also re-
`sult in clinically relevant disruption of hepatic CYP enzyme
`activity (75). Patients with POR de!ciency typically have
`normal electrolytes and mineralocorticoid function, nearly
`normal cortisol levels that respond poorly to ACTH stimu-
`lation, increased levels of 17OHP that respond variably to
`ACTH, and low levels of sex steroids. Impaired CYP21A2
`activity may generate levels of 17OHP detected by newborn
`
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`
`screening for 21OHD (66, 76). Atypical genital develop-
`ment occurs in both sexes, with considerable variability.
`The 17,20-lyase activity of CYP17A1 is especially sensitive
`to disrupted electron transport (77), thus POR defects typ-
`ically affect fetal testicular steroidogenesis. Virilization of
`46,XX females has 2 causes. First, POR de!ciency diverts
`steroids into the “backdoor pathway” of dihydrotestos-
`terone biosynthesis (Fig. 1), contributing to the prenatal fe-
`male virilization (69, 78-80). Second, as placental CYP19
`(aromatase) requires POR, pregnant women carrying a fetus
`with the POR mutation R457H (but not POR A287P) may
`experience virilization during pregnancy (66-68), similarly
`to women carrying an aromatase-de!cient fetus (81, 82).
`The POR polymorphism A503V, which mildly affects many
`P450 activities, is found commonly—from 19% among
`African Americans to 37% of Chinese Americans (83)—but
`does not affect the presentation of 21OHD (84).
`
`11β-Hydroxylase and aldosterone synthase
`Steroid 11-hydroxylase (CYP11B1, P450c11β) and aldos-
`terone synthase (CYP11B2, P450c11AS, P450aldo) are
`closely related enzymes that catalyze the !nal steps in the
`synthesis of glucocorticoids and mineralocorticoids, re-
`spectively; they are encoded by duplicated genes (39, 85).
`Like CYP11A1, these are mitochondrial enzymes that
`require ferredoxin and ferredoxin reductase to receive
`electrons from NADPH. CYP11B1 is expressed abundantly
`in the zona fasciculata, where it converts 11-deoxycortisol
`to cortisol and DOC to corticosterone, and also in the
`zona reticularis, where it initiates the 11-oxo-pathway (see
`later) (86). CYP11B2 expression is less abundant and con-
`!ned to the zona glomerulosa where it catalyzes the 11 β
`-hydroxylase, 18-hydroxylase, and 18-methyloxidase ac-
`tivities needed to convert DOC to aldosterone (87, 88).
`Mutations in CYP11B1 cause 11 β -hydroxylase de!ciency
`(11OHD), with de!cient cortisol, increased adrenal sex
`steroids, female virilization, and increased DOC, causing
`mineralocorticoid hypertension; 17OHP may be elevated in
`the newborn, leading to misdiagnosis of 21OHD (85, 89).
`Mutations in CYP11B2 selectively impair aldosterone syn-
`thesis, causing hyponatremia and hyperkalemia with normal
`cortisol production (39, 85). However, hyponatremia is
`typically less severe than in 21OHD because of continued
`DOC and cortisol secretion.
`
`17β-Hydroxysteroid dehydrogenases
`The synthesis of sex steroids requires the action of 1 of
`the 17  β -hydroxysteroid dehydrogenases (17  β -HSD,
`HSD17B). These enzymes differ in their structures, co-
`factor requirements, reactions catalyzed, and tissue-speci!c
`expression (39). Several are important in steroidogenesis.
`
`HSD17B1 is required for the synthesis of ovarian estra-
`diol and placental estrogens (90-92). No genetic de!ciency
`syndrome for HSD17B1 has been described. HSD17B2
`inactivates estradiol to estrone and testosterone to andro-
`stenedione in the placenta, liver, small intestine, prostate,
`secretory endometrium, and ovary. Whereas HSD17B1 is
`found in placental syncytiotrophoblast cells, HSD17B2 is
`expressed in endothelial cells of placental intravillous ves-
`sels, consistent with a role in defending the fetal circula-
`tion from transplacental passage of maternal estrogens and
`androgens. No de!ciency state for 17  β HSD2 has been
`reported. HSD17B3 is the testicular form of 17  β HSD
`that completes the synthesis of testosterone from andro-
`stenedione; its mutations cause a form of 46,XY DSD
`(93, 94). HSD17B5 (AKR1C3, an aldo-keto reductase en-
`zyme), which is also a 3α-hydroxysteroid dehydrogenase,
`reduces androstenedione to testosterone (95) in the ovary
`and several nonsteroidogenic tissues. AKR1C3 is expressed
`at low levels in the zona reticularis, accounting for the
`small amount of adrenally produced testosterone (96).
`HSD17B6, also known as RoDH for its homology to ret-
`inol dehydrogenases (97), is expressed at low levels in the
`fetal testes, where it appears to catalyze oxidative 3 α HSD
`activities in the alternative or “backdoor” pathway to 5α
`-dihydrotestosterone (DHT) synthesis (79, 98)(see later).
`
`Aromata