Comparison of steroid hormone levels measured by immunoassay and liquid chromatography-mass spectrometry (LC-MS) methods in the follow-up and metabolic control of patients with congenital adrenal hyperplasia

Introduction: Congenital adrenal hyperplasia (CAH) is an autosomal recessive inherited disorder resulting from a genetic defect in one of the enzymes involved in the synthesis of cortisol from cholesterol in the adrenal cortex. It is the most common cause of primary adrenal insufficiency in children. The most common form of CAH is 21-hydroxylase deficiency, accounting for approximately 90% of all cases. Other less common forms include 11β-hydroxylase, 3β-hydroxysteroid dehydrogenase, 17α-hydroxylase/17,20-lyase deficiency, congenital lipoid adrenal hyperplasia, and cytochrome P450 oxidoreductase deficiency. Each enzyme deficiency has different effects on steroidogenesis, leading to varied clinical and laboratory findings in affected individuals. Objective: This study aimed to evaluate steroid hormone levels used in the follow-up of CAH patients at the Ankara University Faculty of Medicine, Department of Pediatrics, Division of Pediatric Endocrinology, using both immunoassay and LC-MS methods. The study also investigated the correlation between the two methods and the correlation of steroid hormone levels with clinical control. Patients and Methods: The medical records of 49 patients diagnosed and treated for CAH at the Ankara University Faculty of Medicine, Department of Pediatrics, Pediatric Endocrinology Clinic were reviewed. Patients meeting the appropriate criteria were included in the study. Demographic information, auxology data, clinical symptoms, simultaneous steroid hormone values measured by immunoassay and LC-MS, follow-up, and complications during treatment were evaluated. The correlation between steroid hormone levels measured by both methods and their correlation with clinical control was examined. SPSS 11.5 and MedCalc 22 programs were used for data analysis. Results: A total of 49 patients diagnosed with CAH were included in the study. The age range of the patients was 0.48 to 21.43 years, with a mean age of 10.53±5.90 years. Sixtythree point three percent (63.3%) of the patients were female, and 36.7% were male (31 females and 18 males). Fifty-one percent (51.0%) of the cases were prepubertal, and 49% were pubertal. Among the patients, 91.9% had 21-hydroxylase deficiency as the cause of CAH. Among these patients, 57.8% had salt-wasting, 8.9% had simple virilizing, and 33.3% had non-classical forms of 21-hydroxylase deficiency. 11β-hydroxylase deficiency constituted 6.1% of all cases (3 patients), and one patient (2.0%) had 17-alpha hydroxylase deficiency. The mean age at diagnosis was calculated as 1.87±3.08 years. The mean age at diagnosis was 0.8±2 years for 21-hydroxylase deficiency (SW form), 4.65±5.4 years for simple virilizing CAH, 4.67±9.5 years for non-classical CAH, and 0.67±1.15 years for 11β-hydroxylase deficiency. Complications were observed in 18.4% of the 21 patients with CAH, including obesity, premature puberty in 12.2%, hypertension in 18.4%, polycystic ovary syndrome in 4.1%, short stature in 6.1%, and testicular adrenal rest tumor (TART) in 2%. Seventy-eight samples from patients diagnosed with CAH were evaluated in the study. There was no significant difference between LC-MS and immunoassay values for 17-OHP, DHEA-S, cortisol, testosterone, and estradiol. Due to the limited number of samples sent simultaneously for aldosterone, a complete evaluation could not be performed. Laboratory results of steroid hormones measured by both methods were statistically correlated. The least difference between immunoassay and LC-MS values of steroid metabolites was observed for DHEA-S, and the highest difference was recorded for 17-OHP. CAH patients with 21-hydroxylase deficiency were divided into two groups based on their metabolic control status. Out of 45 patients with 21-hydroxylase deficiency, 25 (55%) were classified as Group 1 (poor clinical control), and 20 patients (45%) were classified as Group 2 (good clinical control). While there was no statistically significant difference between patients classified as clinically good or poor control in terms of age at diagnosis, demographic and auxology data, and hydrocortisone treatment doses, a statistically significant difference was found only for androstenedione among the laboratory parameters. Conclusion: In this study, we compared steroid hormone results obtained by immunoassay and LC-MS methods in the laboratory follow-up of CAH patients. The results of hormone levels measured by both methods were concordant for all metabolites. Our study did not reveal a significant superiority of immunoassay or LC-MS methods over each other in the follow-up of CAH patients. While there was no statistically significant difference between patient groups classified as clinically good or poor control, a significant difference was observed only for androstenedione among laboratory parameters. Keywords: Congenital Adrenal Hyperplasia, 21-hydroxylase deficiency, 11β-hydroxylase deficiency, immunoassay, LC-MS