ESPE Abstracts

Background: Biochemical ratios of precursor-to-product urinary steroid metabolites have been proposed as surrogate markers of steroidogenic enzyme activity to aid the differential diagnosis of inborn disorders of steroidogenesis. Using ratios rather than total amounts facilitates analysis of single spot urine samples, more convenient than 24-h urine collections for young children. Previous studies examining the utility of these biochemical ratios have been limited by small numbers and primarily compared single patient groups to healthy controls. Here, we established sex- and age-dependent variability in urinary steroid ratios from birth to adulthood, established their discriminative ability to differentiate healthy from inborn steroidogenic disorders and also differentiate distinct steroidogenic disorders from each other.

Methods: Urine samples were collected from 829 healthy controls and 178 patients with genetically confirmed inborn disorder of steroidogenesis (CYP21A2 n=27, CYP11B1 n=12, POR n=36, CYP17A1 n=30, HSD3B2 n=22, SRD5A2 n=51). 32 urinary steroid metabolites were quantified using GC-MS. 14 previously published diagnostic biochemical ratios of urinary metabolites were examined. To characterise age-related variability, 10 age groups were compared: <1month (n=115), 1-3months (n=72), 3-6months (n=76), 6-12months (n=42), 1-4years (n=96), 4-10years (n=44), 10-18years (n=24), 18-30years (n=67), 30-50years (n=165), >50years (n=94). To characterise age- and sex-related variability, 3 age groups were compared: <6months (n=249; 126 male, 123 female), 6months-16years (n=220; 125 male, 95 female) and >16years (n=360; 138 male, 222 female). The 97.5th percentile of healthy was used as an upper reference limit for the purposes of diagnosing abnormalities.

Results: Data distribution for all ratios was highly skewed. No subgroups showed a normal distribution, less than half showed a log-normal distribution. Distinct ratio patterns were observed for childhood and adulthood, some also showing distinct patterns in early infancy; times of major changes in steroid ratios were 3 months of life and onset of puberty. Observed age-related differences were of greater statistical significance than sex-related differences. Biochemical ratios showed a lack of specificity when values for all patient groups were compared for each biochemical ratio. Discriminative ability was improved through scaling calculated ratio values to a healthy reference value. Using a simple interpretation algorithm, ratios performed with 100% sensitivity (Patient vs. healthy), 89.9% sensitivity (correct diagnosis) and 80% specificity. CYP21A2, POR and CYP17A1 deficiencies were challenging to differentiate.

Conclusion: We present considerations required for establishing appropriate reference ranges, challenges of using established biochemical ratios for the analysis of urinary steroid metabolome data and propose a simple algorithm for the diagnosis of inborn steroidogenic disorders.