Test Id : CAH21

This profile provides the simultaneous determination of 17-hydroxyprogesterone, androstenedione, and cortisol. These steroids can also be ordered individually: OHPG / 17-Hydroxyprogesterone, Serum; ANST / Androstenedione, Serum; and CINP / Cortisol, Mass Spectrometry, Serum.

Collection Container/Tube: Red top (serum gel/SST are not acceptable)

Specimen Volume: 0.6 mL

Submission Container/Tube: Plastic vial

Collection Instructions:

1. Morning (8 a.m.) and afternoon (4 p.m.) specimens are preferred.

2. Include time of collection.

3. Centrifuge and aliquot serum into a plastic vial.

Additional Information: If multiple specimens are collected, send separate order for each specimen.

If not ordering electronically, complete, print, and send a Biochemical Genetics Test Request (T798) with the specimen.

0.25 mL

Preferred screening test for congenital adrenal hyperplasia (CAH) caused by 21-hydroxylase deficiency

Part of a battery of tests to evaluate women with hirsutism or infertility, which can result from adult-onset CAH

This is the preferred screening test for congenital adrenal hyperplasia (CAH) caused by 21-hydroxylase deficiency. It is also useful as part of a battery of tests to evaluate females with hirsutism or infertility, which can result from adult-onset CAH.

The cause of congenital adrenal hyperplasia (CAH) is an inherited genetic defect that results in decreased formation of one of the many enzymes that are involved in the production of cortisol. The enzyme defect results in reduced glucocorticoids and mineralocorticoids and elevated 17-hydroxyprogesterone (OHPG) and androgens. The resulting hormone imbalances can lead to life-threatening, salt-wasting crises in the newborn period and incorrect gender assignment of virilized female patients. Adult-onset CAH may result in hirsutism or infertility in women.

The adrenal glands, ovaries, testes, and placenta produce OHPG. It is hydroxylated at the 11 and 21 positions to produce cortisol. Deficiency of either 11- or 21-hydroxylase results in decreased cortisol synthesis, and the feedback inhibition of adrenocorticotropic hormone (ACTH) secretion is lost. Consequently, increased pituitary release of ACTH increases production of OHPG. In contrast, if 17-alpha-hydroxylase (which allows formation of OHPG from progesterone) or 3-beta-ol-dehydrogenase (which allows formation of 17-hydroxyprogesterone formation from 17-hydroxypregnenolone) are deficient, OHPG levels are low with possible increase in progesterone or pregnenolone, respectively.

Most (90%) cases of CAH are due to mutations in the 21-hydroxylase gene (CYP21A2). CAH due to 21-hydroxylase deficiency is diagnosed by confirming elevations of OHPG and androstenedione with decreased cortisol. By contrast, in 2 less common forms of CAH, due to 17-hydroxylase or 11-hydroxylase deficiency, OHPG and androstenedione levels are not significantly elevated and measurement of progesterone (PGSN / Progesterone, Serum) and deoxycorticosterone (DOCS / 11-Deoxycorticosterone, Serum), respectively, are necessary for diagnosis.

OHPG is bound to both transcortin and albumin, and total OHPG is measured in this assay. OHPG is converted to pregnanetriol, which is conjugated and excreted in the urine. In all instances, more specific tests than pregnanetriol measurement are available to diagnose disorders of steroid metabolism.

CORTISOL

5-25 mcg/dL (a.m.)

2-14 mcg/dL (p.m.)

Pediatric reference ranges are the same as adults, as confirmed by peer-reviewed literature.

Petersen KE. ACTH in normal children and children with pituitary and adrenal diseases. I. Measurement in plasma by radioimmunoassay-basal values. Acta Paediatr Scand. 1981;70(3):341-345

ANDROSTENEDIONE

PEDIATRICS*

Premature infants

26-28 weeks, day 4: 92-282 ng/dL

31-35 weeks, day 4: 80-446 ng/dL

Full-term infants

1-7 days: 20-290 ng/dL

1 month-1 year: <69 ng/dL

Males*

Females*

*Soldin SJ, Brugnara C, Wong EC. Androstenedione. In: Pediatric Reference Ranges. 4th ed. AACC Press; 2003:32-34

ADULTS

Males: 40-150 ng/dL

Females: 30-200 ng/dL

17-HYDROXYPROGESTERONE

Children

Preterm infants: Preterm infants may exceed 630 ng/dL, however, it is uncommon to see levels reach 1,000 ng/dL.

Term infants

0-28 days: <630 ng/dL

Levels fall from newborn (<630 ng/dL) to prepubertal gradually within 6 months.

Prepubertal males: <110 ng/dL

Prepubertal females: <100 ng/dL

Adults

Males: <220 ng/dL

Females

Follicular: <80 ng/dL

Luteal: <285 ng/dL

Postmenopausal: <51 ng/dL

Note: For pregnancy reference ranges, see: Soldin OP, Guo T, Weiderpass E, Tractenberg RE, Hilakivi-Clarke L, Soldin SJ. Steroid hormone levels in pregnancy and 1 year postpartum using isotope dilution tandem mass spectrometry. Fertil Steril. 2005;84(3):701-710

Diagnosis and differential diagnosis of congenital adrenal hyperplasia (CAH) always requires the measurement of several steroids. Patients with CAH due to 21-hydroxylase gene (CYP21A2) variants usually have very high levels of androstenedione, often 5- to 10-fold elevations. 17-Hydroxyprogesterone (OHPG) levels are usually even higher, while cortisol levels are low or undetectable. All 3 analytes should be tested.

In the much less common CYP11A variant, androstenedione levels are elevated to a similar extent as in CYP21A2 variant, and cortisol is also low, but OHPG is only mildly, if at all, elevated.

Also less common is 3-beta hydroxysteroid dehydrogenase (HSD) type 2 deficiency, characterized by low cortisol and substantial elevations in dehydroepiandrosterone sulfate (DHEA-S) and 17-alpha-hydroxypregnenolone, while androstenedione is either low, normal, or rarely, very mildly elevated (as a consequence of peripheral tissue androstenedione production by 3-beta HSD-1).

In the very rare StAR (steroidogenic acute regulatory) protein deficiency, all steroid hormone levels are low and cholesterol is elevated.

In the very rare 17-alpha-hydroxylase deficiency, androstenedione, all other androgen-precursors (17-alpha-hydroxypregnenolone, OHPG, DHEA-S), androgens (testosterone, estrone, estradiol), and cortisol are low, while production of mineral corticoid and its precursors, in particular progesterone, 11-deoxycorticosterone, corticosterone, and 18-hydroxycorticosterone, are increased.

The goal of CAH treatment is normalization of cortisol levels and, ideally, of sex-steroid levels also. OHPG is measured to guide treatment, but this test correlates only modestly with androgen levels. Therefore, androstenedione and testosterone should also be measured and used to guide treatment modifications. Normal prepubertal levels may be difficult to achieve, but if testosterone levels are within the reference range, androstenedione levels up to 100 ng/dL are usually regarded as acceptable.

Androstenedione and, to a lesser degree, dehydroepiandrosterone sulfate supplements can result in elevations of serum androstenedione level. With large androstenedione doses of 300 to 400 mg/day, serum androstenedione levels can almost double in some patients. Testosterone levels and, particularly in men, estrone and estradiol levels may also increase but to a much lesser degree.

This test provides merely supplementary information and should, therefore, never be employed as the sole diagnostic tool.

1. Von Schnakenburg K, Bidlingmaier F, Knorr D. 17-hydroxyprogesterone, androstenedione, and testosterone in normal children and in prepubertal patients with congenital adrenal hyperplasia. Eur J Pediatr. 1980;133(3):259-267

2. Sciarra F, Tosti-Croce C, Toscano V. Androgen-secreting adrenal tumors. Minerva Endocrinol. 1995;20(1):63-68

3. Collett-Solberg P. Congenital adrenal hyperplasia: from genetics and biochemistry to clinical practice, part I. Clin Pediatr. 2001;40(1):1-16

4. Speiser PW, Azziz R, Baskin LS, et al. Congenital adrenal hyperplasia due to steroid 21-hydroxylase deficiency: an Endocrine Society Clinical Practice Guideline. J Clin Endocrinol Metab. 2010;95(9):4133-4160

5. Nordenstrom A, Falhammar H. Management of endocrine disease: Diagnosis and management of the patient with non-classic CAH due to 21-hydroxylase deficiency. Eur J Endocrinol. 2019;180(3):R127–R145

6. Young WF Jr. Primary aldosteronism: A common and curable form of hypertension. Cardiol Rev. 1999;7(4):207-214

7. Young WF Jr. Pheochromocytoma and primary aldosteronism: diagnostic approaches. Endocrinol Metab Clin North Am. 1997;26(4):801-827

8. Wudy SA, Hartmann M, Svoboda M. Determination of 17-hydroxypregnenolone in plasma by stabile isotope dilution/benchtop liquid chromatography-tandem mass spectrometry. Horm Res. 2000;53(2):68-71

9. Therrell BL. Newborn screening for congenital adrenal hyperplasia. Endocrinol Metab Clin North Am. 2001;30(1):15-30

10. Bachega TA, Billerbeck AE, Marcondes JA, Madureira G, Arnhold IJ, Mendonca BB. Influence of different genotypes on 17-hydroxyprogesterone levels in patients with nonclassical congenital adrenal hyperplasia due to 21-hydroxylase deficiency. Clin Endocrinol. 2000;52(5):601-607

11. Kao PC, Machacek DA, Magera MJ, Lacey JM, Rinaldo P. Diagnosis of adrenal cortical dysfunction by liquid chromatography-tandem mass spectrometry. Ann Clin Lab Sci. 2001;31(2):199-204

12. Young WF Jr. Management approaches to adrenal incidentalomas. A view from Rochester, Minnesota. Endocrinol Metab Clin North Am. 2000;29(1):159-185

13. Ibanez L, DiMartino-Nardi J, Potau N, Saenger P. Premature adrenarche-normal variant or forerunner of adult disease? Endocr Rev. 2000;21(6):671-696

14. Allolio B, Arlt W. DHEA treatment: myth or reality? Trends Endocrinol Metab. 2002;13(7):288-294

15. Lin CL, Wu TJ, Machacek DA, Jiang NS, Kao PC. Urinary free cortisol and cortisone determined by high performance liquid chromatography in the diagnosis of Cushing's syndrome. J Clin Endocrinol Metab. 1997;82:151-155

16. Findling JW, Raff H. Diagnosis and differential diagnosis of Cushing's syndrome. Endocrinol Metab Clin North Am. 2001;30(3):729-747

17. Buchman Al. Side effects of corticosteroid therapy. J Clin Gastroenterol. 2001;33(4):289-297

18. Dodds HM, Taylor PJ, Cannell GR, Pond SM. A high-performance liquid chromatography-electrospray-tandem mass spectrometry analysis of cortisol and metabolites in placental perfusate. Anal Biochem. 1997;247(2):342-347

19. Cengiz H, Demirci T, Varim C, Cetin S. Establishing a new screening 17 hydroxyprogesterone cut-off value and evaluation of the reliability of the long intramuscular ACTH stimulation test in the diagnosis of nonclassical congenital adrenal hyperplasia. Eur Rev Med Pharmacol Sci. 2021;25(16):5235-5240. doi:10.26355/eurrev_202108_26537

Deuterated stable isotopes (d4-cortisol, d7-androstenedione, d8 17-hydroxyprogesterone) are added to the serum sample as internal standards. Cortisol, androstenedione, 17-hydroxyprogesterone, and the internal standards are extracted from specimens online using a guard cartridge. The analytes are transferred online to an analytical column and are analyzed by liquid chromatography tandem mass spectrometry.(Unpublished Mayo method)

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This test was developed and its performance characteristics determined by Mayo Clinic in a manner consistent with CLIA requirements. It has not been cleared or approved by the US Food and Drug Administration.

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