Nonclassic congenital lipoid adrenal hyperplasia: a new disorder of the steroidogenic acute regulatory protein with very late presentation and normal male genitalia

Abstract

Context: Lipoid congenital adrenal hyperplasia is a severe disorder of adrenal and gonadal steroidogenesis caused by mutations in the steroidogenic acute regulatory protein (StAR). Affected children typically present with life-threatening adrenal insufficiency in early infancy due to a failure of glucocorticoid (cortisol) and mineralocorticoid (aldosterone) biosynthesis, and 46,XY genetic males have complete lack of androgenization and appear phenotypically female due to impaired testicular androgen secretion in utero.

Objective: The objective of this study was to investigate whether nonclassic forms of this condition exist.

Patients and methods: Sequence analysis of the gene encoding StAR was undertaken in three children from two families who presented with primary adrenal insufficiency at 2-4 yr of age; the males had normal genital development. Identified mutants were tested in a series of biochemical assays.

Results: DNA sequencing identified homozygous StAR mutations Val187Met and Arg188Cys in these two families. Functional studies of StAR activity in cells and in vitro and cholesterol-binding assays showed these mutants retained approximately 20% of wild-type activity.

Conclusions: These patients define a new disorder, nonclassic lipoid congenital adrenal hyperplasia, and represent a new cause of nonautoimmune Addison disease (primary adrenal failure).

Fig. 1

Mutation analysis. A) DNA sequencing: top, wild-type control; middle, patient 1 with the Val187Met mutation; bottom, patient 2 with the Arg188Cys mutation. B) Alignment of the sequences of residues 180-197 of the human, rat, mouse, sheep, chicken and zebrafish StAR proteins; residues 187 and 188 are highlighted. C) Ribbon diagram of N-62 StAR, positioned to permit both Val187 and Arg188 to be seen; the amino- (N) and carboxy- (C) termini are indicated. D) Arg188 normally forms a hydrogen bond with Glu169; changing Arg188 to Cys eliminates hydrogen bonding with Glu169 and creates a weak hydrogen bond with Thr167.

Fig. 2

Activity of StAR mutants. A) Activity in intact cells. COS-1 cells were co-transfected with expression vectors for the cholesterol side-chain cleavage system and the indicated StAR vector, and pregnenolone was measured 48 h later. Addition of the StAR-independent substrate 22R-hydroxycholesterol (22R-OH) in the absence of StAR indicates the maximum steroidogenic capacity of the cells. Data are the mean ± SEM from six transfections, each performed in triplicate. B) Activity on mitochondria in vitro. Mitochondria were isolated from steroidogenic mouse Leydig (testicular) MA-10 cells, the indicated human N-62 StAR proteins were added, and pregnenolone synthesis from endogenous mitochondrial cholesterol stores was measured. Data are mean ±SEM from three experiments, each performed in triplicate. C) Cholesterol-binding capacities. Binding of various concentrations of fluorescent NBD-cholesterol by wild-type (black, solid line), Val187Met (grey, dotted line), and Arg188Cys (black, dotted line) was measured with a protein concentration of 1 μM; control is buffer without protein (grey solid line). Data are mean ± SEM for three experiments, each performed in triplicate.