Oral Presentation The Joint Annual Scientific Meetings of the Endocrine Society of Australia and the Society for Reproductive Biology 2017

LC-MS/MS diagnostic criteria for 21 hydroxylase deficient non-classic congenital adrenal hyperplasia (#156)

Jennifer Ng 1 2 , Ee Mun Lim 3 4 , Narelle Hadlow 3 , John Beilby 5 , Gerald Watts 6 7 , Bronwyn Stuckey 2 4 8
  1. Hollywood Medical Centre, Nedlands, WA, Australia
  2. The keogh Institute for Medical Research, Nedlands, WA, Australia
  3. Chemical Pathology, PathWest QEII Laboratory, Nedlands, WA, Australia
  4. Endocrinology and Diabetes, Sir Charles Gairdner Hospital, Nedlands, WA, Australia
  5. Diagnostic Genomics, PathWest QEII Laboratory, Nedlands, WA, Australia
  6. Metabolic Research Centre, Royal Perth Hospital, Perth, WA, Australia
  7. Medicine and Pharmacology, University of Western Australia, Perth, WA, Australia
  8. Medicine and Pharmacology, University of Western Australia, Perth, WA, Australia

Non-classic congenital adrenal hyperplasia is clinically indistinguishable from PCOS but accurate diagnosis is therapeutically important. Inheritance is recessive but heterozygotes can be symptomatic1. Immunoassay-defined diagnostic thresholds for the synacthen stimulation test are imperfect. Mismatch with positive biochemistry and negative genetics occurs in up to 75%(1) Liquid chromatography and tandem mass spectometry (LC-MS/MS) offers greater accuracy for steroid hormone quantitation. However, diagnostic thresholds for LC-MS/MS have not been determined.


We aimed to define LC-MS/MS -specific SST diagnostic criteria and determine whether PCOS and NCCAH can be distinguished from a basal LC-MS/MS- androgen profile.


We identified females >15yrs who had undergone CYP21A2 mutation analysis at PathWest QEII from Jan 2010 to June 2017. Biochemistry was compared among normal genotype, CYP21A2 heterozygotes and NCCAH patients. ROC analysis was conducted to determine optimal 17 OHP thresholds to identify CYP21A2 heterozygotes.


Of 84 genetic studies, 72 (85.7%) had been requested to investigate hyperandrogenic symptoms. SSTs were available in 32/54 with normal genotype, 9/17 heterozygotes and 2/8 with NCCAH. Basal 17 OHP was at least 5 times above the follicular phase upper limit in all NCCAH cases. However basal 17 OHP did not distinguish heterozygotes from normal genotype patients. A peak stimulated 17 OHP threshold of 8.4 nmol/L identified heterozygotes with 100% sensitivity and 90.6% specificity. The AUC for the ROC analysis was 0.957, 95% CI (0.902, 1.000), p<0.001. Median serum testosterone, androstenedione and di-hydrotestosterone did not differ significantly among the 3 groups.  Importantly, 4/12 (33%) heterozygotes investigated for hyperandrogenic symptoms carried severe mutations.


We conclude that NCCAH should be considered in hyperandrogenic patients whether or not testosterone is elevated. An SST should be considered in those desiring fertility to exclude CYP21A2 heterozygosity. A stimulated 17 OHP threshold of 8.4 nmol/L reliably identifies CYP21A2 mutation carriers with 100% sensitivity and 90.6% specificity.

  1. Neocleous V, Shammas C, Phedonos AA, Phylactou LA, Skordis N. Phenotypic variability of hyperandrogenemia in females heterozygous for CYP21A2 mutations. Indian journal of endocrinology and metabolism. 2014;18(Suppl 1):S72-9.
  2. Ambroziak U, Kepczynska-Nyk A, Kurylowicz A, Malunowicz EM, Wojcicka A, Miskiewicz P, et al. The diagnosis of nonclassic congenital adrenal hyperplasia due to 21-hydroxylase deficiency, based on serum basal or post-ACTH stimulation 17-hydroxyprogesterone, can lead to false-positive diagnosis. Clinical endocrinology. 2016;84(1):23-9.