Severe combined adrenal and gonadal deficiency caused by novel mutations in the cholesterol side chain cleavage enzyme, P450scc

Abstract

Context: Mitochondrial cytochrome P450scc converts cholesterol to pregnenolone in all steroidogenic tissues. Although progesterone production from the fetally-derived placenta is necessary to maintain pregnancy to term, four patients with mutations in the gene encoding P450scc (CYP11A1), have been described, one in a 46,XX female and three in underandrogenized 46,XY individuals, all with primary adrenal failure.

Objective: Our aim was to determine whether P450scc mutations might be found in other children and to explore genotype/phenotype correlations.

Methods and patients: We performed mutational analysis of CYP11A1 in individuals with 46,XY disorders of sex development and primary adrenal failure, followed by functional studies of P450scc activity and of P450scc RNA splicing.

Results: Among nine 46,XY infants with adrenal failure and disordered sexual differentiation, two infants had compound heterozygous mutations in CYP11A1. One patient harbored the novel P450scc missense mutations L141W and V415E, which retained 38 and 0% activity, respectively. The other carried a CYP11A1 frameshift mutation c835delA (0% activity) and a splice site mutation [IVS3+(2-3)insT] that prevented correct splicing of P450scc mRNA.

Conclusions: P450scc deficiency is a recently recognized disorder that may be more frequent than originally thought. The phenotypic spectrum ranges from severe loss-of-function mutations associated with prematurity, complete underandrogenization, and severe, early-onset adrenal failure, to partial deficiencies found in children born at term with clitoromegaly and later-onset adrenal failure. In contradistinction to congenital lipoid adrenal hyperplasia caused by steroidogenic acute regulatory protein mutations, adrenal hyperplasia has not been reported in any of the six patients with P450scc deficiency.

Figure 1

Activities of P450scc mutations. COS-1 cells were transfected with either wild-type or mutant F2 plasmid. Pregnenolone secreted into the culture medium was determined by RIA. Empty vector was used as a negative control, and the StAR-independent substrate 22R-hydroxycholesterol was added to the culture medium. Values are the mean ± sem from four independent transfection experiments, each performed in triplicate.

Figure 2

Minigene experiment. HeLa cells were transfected with empty pcDNA3.1 vector, or vector expressing the wild-type or mutant CYP11A1 minigene construct. Total RNA from the transfected HeLa cells were used for RT-PCR, showing the expected 580-bp product from the wild-type construct but no correctly spliced product from the mutant. PCR of GAPDH RNA was used as an internal control, producing a 247-bp PCR fragment.