Congenital adrenal hyperplasia can be due to 11β-hydroxylase deficiency (11βOHD). Sporadic reports of 11βOHD are frequent but overviews on molecular landscape in some populations are lacking. The aim of this research was to compile a genetic landscape from an 11βOHD cohort, and to report a novel yet recurrent splice variant. An overview of CYP11B1 variants in a cohort of 11βOHD is presented here. The functional effect of NM_000497.4(CYP11B1):c.954 + 148C > G was studied in silico and in vitro, and a genotype-phenotype correlation study ("SPLICYP" study, No. 22_1787) was conducted. Patients with 11βOHD who underwent genetic testing at the biochemistry and molecular biology department were considered for inclusion. A total of 250 patients, diagnosed from 1990 to 2024, underwent CYP11B1 sequencing. Forty-four patients carried a novel deep intronic variant (NM_000497.4(CYP11B1):c.954 + 148C > G). Four were excluded from genotype-phenotype correlation due to missing criteria. Functional validation was performed using a Minigene Reporter Assay. We retrospectively analyzed genetic findings, clinical features of 11βOHD, and hormonal assays. The Minigene study confirmed that c.954 + 148C > G disrupts splicing by activating a cryptic donor site. Patients carrying this variant had significantly lower steroid precursor levels (P < .034) and delayed pubertal onset (P = .005) compared to severe variant carriers. This retrospective study provides genetic data in a wide cohort of 11βOHD, and identifies c.954 + 148C > G as the most recurrent variant in our Caucasian participant recruitment. Screening of deep intronic regions, coupled with functional in vitro tools, must not be overlooked in the strategy to avoid diagnostic failure.

11β-hydroxylase deficiency (11βOHD) is the second most common cause (5%) of congenital adrenal hyperplasia (CAH). The CYP11B1 gene shares 95% of genomic sequence homology with CYP11B2, and therefore Sanger sequencing remains the gold standard. We present a case of 11βOHD due to an intragenic inversion in CYP11B1 that was missed by both the Sanger sequencing and massive parallel sequencing (MPS) methods. The child was born with virilised genitalia at Prader stage 4 and the biological findings showed a hydromineral retention pattern and a pathognomonic increase in steroid precursors suggestive of 11βOHD. Standard trio analysis revealed only one heterozygous pathogenic variation inherited from the father. The study using MPS showed similar outcomes. Careful observation of the alignment BAM files revealed breaks in sequencing depth, incomplete alignments and systematic paradoxical read-pairs orientation. A specifically designed amplification and Sanger protocol confirmed the novel NM_000497.4(CYP11B1):c.[892_1121+7 inv;1121+8_1121+9del]; p.(Glu298HisfsTer113) variant at heterozygous state in the proband and his mother, fulfilling the diagnosis. The present case reports the first short intragenic inversion in CAH and illustrates the pitfalls that must always be kept in mind when using sequencing methods. When the phenotype is unequivocal, a thorough investigation of the locus should be carried out with cross-use of different techniques.

11β-hydroxylase (11β-OH) deficiency is the second most frequent cause of classic congenital adrenal hyperplasia (CAH) (5%-8% of cases). Clinically, it is characterized by virilization and arterial hypertension. The objective of this study was to describe the clinical, biochemical and genetic characteristics classic 11β-OH deficiency in patients managed in our hospital and its outcomes. Retrospective longitudinal, observational and descriptive study. Patients with clinical features of virilization, high levels of 11-deoxycortisol and study of CYP11B1 gene with detection of pathogenic and likely pathogenic variants. We identified 6 patients (1 male, 5 female) from 4 families. In the 4 index cases, the median age at diagnosis was 2.3 years. The 46,XX patients exhibited a variable degree of virilization at diagnosis, with a predominance of Prader stage V, and one case of male sex assignment at birth. All patients had elevated serum concentrations of 17-hydroxyprogesterone and testosterone. Fifty percent of the patients had developed arterial hypertension during the follow-up, with onset at a median age of 9.3 years. Three 46,XX patients reached a median final height of 154 cm. Six different variants of theCYP11B1 gene were identified, 5 of which were novel variants (c.595 G > A, c.710 T > C, c.1156delG, c.395 + 2dupT, c.1159dupA). There is considerable heterogeneity in the clinical presentation of patients with CAH due to 11β-OH deficiency. Early diagnosis and treatment are important to prevent complications and improve long-term outcomes. We report 6 different variants of the CYP11B1 gene, including 5 novel variants.

分析1例经典型11β-羟化酶缺陷症合并双侧肾上腺髓样脂肪瘤患者的临床资料，发现1种可能致病的新突变c.600C>A（p.Ser200Arg），且侧面证实了切除双侧肾上腺对于治疗11β-羟化酶缺陷症患者难治性高血压的有效性，提高对临床罕见的11β-羟化酶缺陷症的认识。.

The clinical manifestations of nonclassical 11beta-hydroxylase deficiency are very similar to those of non-classical 21-hydroxylase deficiency. For this study, we investigated the relationship between the clinical and molecular features of congenital adrenal hyperplasia caused by 11beta-hydroxylase deficiency and reviewed the related literature, which are expected to provide assistance for the clinical diagnosis and analysis of congenital adrenal hyperplasia. Clinical data for 10 Chinese patients diagnosed with congenital adrenal hyperplasia in our hospital from 2018 to 2022 were retrospectively analyzed. We examined the effects of gene mutations on protease activity and constructed three-dimensional structure prediction models of proteins. We describe 10 patients with 11beta-hydroxylase gene mutations (n = 5, 46,XY; n = 5, 46,XX), with 10 novel mutations were reported. Female patients received treatment at an early stage, with an average age of 2.08 ± 1.66 years, whereas male patients received treatment significantly later, at an average age of 9.77 ± 3.62 years. The most common CYP11B1 pathogenic variant in the Chinese population was found to be c.1360C > T. All mutations lead to spatial conformational changes that affect protein stability. Our study found that there was no significant correlation between each specific mutation and the severity of clinical manifestations. Different patients with the same gene pathogenic variant may have mild or severe clinical manifestations. The correlation between genotype and phenotype needs further study. Three-dimensional protein simulations may provide additional support for the physiopathological mechanism of genetic mutations.