Cytochrome P450 oxidoreductase deficiency (PORD) is an exceptionally rare form of congenital adrenal hyperplasia (CAH) characterized by impaired activity of multiple microsomal cytochrome P450 enzymes. In adult women, PORD frequently presents with nonspecific reproductive manifestations such as menstrual irregularities, infertility, and ovarian cysts, often mimicking polycystic ovary syndrome (PCOS) or premature ovarian insufficiency (POI). To date, successful pregnancies in affected women remain extremely rare. We describe two biological sisters with compound heterozygous POR variants c.1811A>G (p.Tyr604Cys) and c.1952_1966del (p.Gly651_His655del), both presenting with infertility and recurrent ovarian cysts but initially misdiagnosed as PCOS or POI. Both exhibited elevated serum progesterone and 17-hydroxyprogesterone (17-OHP) without overt androgen excess, consistent with the paradoxical hormonal signature of PORD. The elder sister underwent a progestin-primed ovarian stimulation (PPOS) protocol, while the younger received a short GnRH agonist protocol; in both cases, a freeze-all strategy was adopted due to supraphysiologic progesterone levels. Subsequent hormone replacement therapy frozen embryo transfer (HRT-FET) combined with glucocorticoid supplementation resulted in singleton live births in both patients. The younger sister developed preeclampsia requiring preterm cesarean delivery, highlighting potential obstetric risks. These cases represent the first report of two siblings with nonclassic PORD achieving live births through IVF-FET. Moreover, we identified a previously unreported POR variant, c.1952_1966del, p.Gly651_His655del, which expands the known mutational spectrum of PORD in the Chinese population. Our findings highlight the importance of early genetic testing in women with atypical infertility, recognition of the distinctive hormonal profile of PORD, and the value of glucocorticoid-supported artificial-cycle frozen embryo transfer as an effective reproductive strategy.

Cytochrome P450 oxidoreductase deficiency (PORD) is a rare form of congenital adrenal hyperplasia caused by impaired electron transfer to multiple microsomal enzymes. Hypertension in PORD has been attributed to mineralocorticoid excess; however, experimental evidence suggests that POR may also regulate endothelial vasodilatory pathways. The vascular effects of PORD in humans remain poorly defined. We evaluated blood pressure regulation, microvascular structure, and serum vasoactive mediators in patients with PORD. Observational, cross-sectional study. Ten patients with genetically confirmed PORD (4 females, 6 males; age range 8-27 years) and 41 age- and sex-matched healthy controls were evaluated. Office and 24-hour ambulatory blood pressure monitoring (ABPM) were performed. Microvascular structure was assessed using nailfold capillaroscopy (NFC). Serum levels of thromboxane B₂ (TXB₂), prostaglandin E₂ (PGE₂), and nitric oxide (NO) were measured. Hypertension was present in 80% of patients with PORD compared with 7.3% of controls (P < .001). Office and ambulatory systolic and diastolic pressures were significantly higher in PORD. NFC revealed shorter capillary length (P = .001), more frequent crossed capillaries (P < .001), and avascular areas (P = .039). Serum TXB₂ levels were higher in PORD (P = .007), whereas PGE₂ and NO did not differ between groups. PORD is associated with an increased prevalence of hypertension, microvascular capillary abnormalities, and elevated TXB₂, consistent with enhanced cyclooxygenase-mediated vasoconstrictor tone and endothelial dysfunction. These findings suggest that cyclooxygenase-dependent mechanisms may contribute to the vascular alterations observed in PORD.

P450 oxidoreductase deficiency (PORD) affects cytochrome enzyme activities, causing various symptoms, such as adrenal insufficiency, disorders of sex development and skeletal malformations. This study aims to elucidate the clinical manifestations, genotype characteristics, diagnosis and management of 46,XX karyotype patients with PORD in China. A retrospective study included twelve 46,XX PORD patients in a Chinese tertiary medical center from 2004 to 2024. The patients' clinical characteristics were summarized based on manifestations, hormone profiles, and responses to treatments. The age of first visit was 7-31 years. Except for one young girl presenting with ambiguous genitalia since born, 11 patients presented with either abnormal menses or multiple ovarian cysts. Six patients showed masculinization of their external genitalia, and ten patients showed varying degrees of skeletal deformity. Progesterone was elevated and ovarian reserve was poor in all patients. The most frequent POR variant, c.1370G > A, located in exon 11 occurred in 11/12 patients with an allele frequency of 87.5% (21/24). Two novel nonsense mutations, c.1684dupG and c.2040dupC, were identified and assessed as pathogenic and likely pathogenic by ACMG, respectively. The c.1370G > A might be a dominant mutation type of POR in China. Female patients with PORD have a vulnerable ovarian reserve, and their ovarian macrocysts can be managed conservatively for fertility preservation. This study specifically focuses on PORD in 46,XX Chinese individuals, which implies its genetic causes with novel genetic findings and summarizes the puzzling spectrum of clinical manifestations.

Congenital adrenal hyperplasia (CAH) represents a group of inherited disorders affecting steroidogenesis. Early and accurate diagnosis is crucial for effective treatment, particularly for preventing adrenal insufficiency and minimizing androgen excess. This study aims to develop and validate an ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for the simultaneous quantification of 21 steroid hormones, including 11-oxygenated androgens, which are critical for diagnosing and monitoring various forms of CAH. We utilized a microbore column UPLC combined with hydroxylamine derivatization, which enabled excellent chromatographic separation and enhanced sensitivity of all target compounds. The method was evaluated for precision, linearity, recovery, ion suppression, and carryover according to FDA and CLSI guidelines. Steroid profiles from healthy controls and CAH patients were compared using Mann-Whitney tests. The UPLC-MS/MS method demonstrated excellent precision (<20 % except for 11-ketoandrostenedione), linearity (R 2 > 0.99), low limits of detection and quantification, and satisfactory recovery (57-86 % absolute, 99-111 % relative). Our method showed good correlation with proficiency testing group means, although significant negative biases were noted for androstenedione, progesterone, and 11-deoxycortisol. In a clinical setting, significant increases in pregnenolone, progesterone, 17-hydroxyprogesterone, dehydroepiandrosterone, and other key steroids were observed in patients with 21-hydroxylase deficiency, while distinct profiles were identified for patients with 17-hydroxylase deficiency, cytochrome P450 oxidoreductase deficiency, and lipoid CAH. Our UPLC-MS/MS method provides a sensitive and specific tool for the comprehensive profiling of adrenal steroids, offering improved diagnostic accuracy for CAH. Its ability to differentiate between various CAH subtypes highlights its potential clinical utility in both diagnosis and monitoring.

P450 oxidoreductase (POR) facilitates electron flux to type 2 microsomal P450 cytochrome enzymes (CYPs), including the adrenal steroidogenic enzymes CYP17A1 and CYP21A2. Due to the combined impairment of these enzymes, POR deficiency (PORD), an autosomal recessive condition, results in congenital adrenal hyperplasia characterised by combined glucocorticoid and postnatal sex steroid deficiency. This study focuses on urinary steroid excretion in infants affected by PORD in the first week of life. We report on three neonatal PORD cases from two families. One family had two affected babies born three years apart who were stillborn and first-day deceased, respectively. DNA sequencing revealed a homozygous 3 bp deletion in exon six leading to an glutamic acid deletion (p.[Glu217del]). Bladder contents were obtained from the stillborn baby, and excreted urine was obtained from the second baby. In a second family, their second affected newborn, antenatally diagnosed carrying the common homozygous p.(Ala287Pro) mutation, had urine collected daily during the first week of life. Steroid excretions were quantified by gas chromatography-mass spectrometry (GC-MS). The birth-day excretions were very similar in all babies. Most notable and unusual was a large excretion of unmetabolised corticosterone, suggesting inhibited catabolism to allow maximum active gluco- and mineralocorticoid availability at birth. Because CYP3A7 (16α-hydroxylase) requires POR, there was an almost complete absence of usually dominant 3β-hydroxy-Δ5 steroids (16α-OH-DHEA and 16α-OH-pregnenolone) and the usually characteristic precursor pregnenolone metabolite 5-pregnene-3β,20α-diol (pregnenediol, 5PD). In the baby sequentially studied over a week, we observed gradual maturation to the typical and familiar PORD neonatal metabolome. At the end of the period, the minimally catabolised corticosterone had diminished, and steroid excretion was completely dominated by 5PD, excreted as both mono- and disulphate conjugates. Whether this metabolome is distinctive of all PORD infants, not just those with severe manifestation, is not known. On the first day of life, standard diagnostic markers are compromised due to fetal-placental-maternal contribution and unique neonatal steroid metabolism. However, the Day 1 PORD steroid metabolome remains distinctive, and we propose using additional biochemical markers reflective of the near complete reduction of POR-dependent CYP3A7 (16α-hydroxylase) activity to improve diagnostic yield.