The high relevance of 21DOF, (4AD + 17αOHP)/F, and 11DOF/17αOHP for newborn screening of 21 hydroxylase deficiency

Abstract Background There are limited reports on the detailed examination of steroid profiles for setting algorithms for 21OHD screening by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Aim To define an algorithm for newborn screening of 21OHD by LC-MS/MS measuring a total of 2,077 dried blood spot samples in Tokyo. Methods Five steroids [17α-hydroxyprogesterone (17αOHP), 21-deoxycortisol (21DOF), 11-deoxycortisol (11DOF), Androstenedione (4AD), and Cortisol (F)] were included in the panel of LC-MS/MS. The samples from the two cohorts were assayed; A: 63 “screening positive” neonates who were referred to an endocrinologist (21OHD: 26, false-positive: 37, obtained from 2015 to 2020); B: 2,014 samples whose 17αOHP values were in the 97th percentile or above, in the first-tier test with 17αOHP ELISA from 2020 to 2021. Results Analysis of cohort-A revealed that the three indexes, 21DOF, 11DOF/17αOHP, and (4AD + 17αOHP)/F had higher AUC (area under the curve) values (0.999, 0.997, 0.989, respectively), while the 17αOHP AUC was lower (0.970). Accordingly, in addition to 17αOHP, the three markers were included for defining the screening algorithm. The assay of cohort-B revealed that the new algorithm gave 92% of predicted positive predictive value without false-negative cases. We also determined the reference values for the five steroids at 4 - 7 days after birth according to sex and gestational age (GA), revealing extremely low levels of 21DOF at any GA irrespective of sex differences. Conclusion Our study demonstrated the high relevance of 21DOF, (4AD + 17αOHP)/F, and 11DOF/17αOHP rather than 17αOHP for 21OHD screening.