Constitutive Activation of PKA Catalytic Subunit in Adrenal Cushing's Syndrome

Abstract

BACKGROUND Corticotropin-independent Cushing’s syndrome is caused by tumors or hyperplasia of the adrenal cortex. The molecular pathogenesis of cortisol-producing adrenal adenomas is not well understood. METHODS We performed exome sequencing of tumor-tissue specimens from 10 patients with cortisol-producing adrenal adenomas and evaluated recurrent mutations in candidate genes in an additional 171 patients with adrenocortical tumors. We also performed genomewide copy-number analysis in 35 patients with cortisol-secreting bilateral adrenal hyperplasias. We studied the effects of these genetic defects both clinically and in vitro. RESULTS Exome sequencing revealed somatic mutations in PRKACA, which encodes the catalytic subunit of cyclic AMP–dependent protein kinase (protein kinase A [PKA]), in 8 of 10 adenomas (c.617A→C in 7 and c.595_596insCAC in 1). Overall, PRKACA somatic mutations were identified in 22 of 59 unilateral adenomas (37%) from patients with overt Cushing’s syndrome; these mutations were not detectable in 40 patients with subclinical hypercortisolism or in 82 patients with other adrenal tumors. Among 35 patients with cortisol-producing hyperplasias, 5 (including 2 first-degree relatives) carried a germline copy-number gain (duplication) of the genomic region on chromosome 19 that includes PRKACA. In vitro studies showed impaired inhibition of both PKA catalytic subunit mutants by the PKA regulatory subunit, whereas cells from patients with germline chromosomal gains showed increased protein levels of the PKA catalytic subunit; in both instances, basal PKA activity was increased. CONCLUSIONS Genetic alterations of the catalytic subunit of PKA were found to be associated with human disease. Germline duplications of this gene resulted in bilateral adrenal hyperplasias, whereas somatic PRKACA mutations resulted in unilateral cortisol-producing adrenal adenomas. (Funded by the European Commission Seventh Framework Program and others.)

Structural insights into mis-regulation of protein kinase A in human tumors

“…1, Table 1, and Table S1): the full-length wild-type C subunit (PKAc), the predominant point mutant (PKAc L205R), the predominant chimeric fusion protein (DnaJ-PKAc), and PKAc with exon 1 residues deleted (PKAc Δexon1). All structures were determined as complexes with ATP and the protein kinase inhibitor (PKI) peptide (5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24). In each, ATP is bound with two metal ions within the cleft formed between the smaller N-terminal and larger C-terminal lobes of the C subunit, associated as previously described (14).…”

“…Previous studies (8)(9)(10)(11) have suggested that the mutation may abolish the binding of PKAc L205R to the R subunit. The structure of PKAc L205R provides an atomic basis for this effect, where steric clash between R205 and the P+1 residue of the regulatory subunit inhibitory sequence is expected (Fig.…”

“…Gene fusions and chromosome translocations that alter promoter elements controlling oncogene expression are documented contributing factors to carcinogenesis and disease (24,25). Overexpression of the PKA C subunit indeed contributes to disease, as discovered with copynumber gains for PRKACA in some cases of adrenal hyperplasia (8). In FL-HCC, a significant increase in C-subunit levels may favor formation of additional type-I holoenzyme complexes as increases in levels of RI, but not RII, are known to compensate for increased C-subunit levels (26).…”

“…In Cushing's syndrome, a disorder arising from overproduction of cortisol from tumors and hyperplasia of the adrenal cortex, mutations in PRKACA have been discovered in tumor specimens from patients with this disease (8)(9)(10)(11), including the predominant L205R mutation in the PKA C subunit. In fibrolamellar hepatocellular carcinoma (FL-HCC), a rare liver tumor affecting young adults, ∼400-kb deletions have been discovered in chromosome 19.…”

DNAJB1-PRKACA is specific for fibrolamellar carcinoma

“…Honeyman et al 3 proposed the chimera formed by DNAJB1 and PRKACA leads to upregulation of PRKACA activity by a promoter switch mechanism. Interestingly, point mutations [24][25][26] in PRKACA and copy number gain 24 of PRKACA have been shown as recurrent genetic abnormalities underlying functional adrenal adenomas and adrenal hyperplasia. The discovery of a translocation involving PRKACA is a third mechanism of activation of the PRKACA gene in tumors.…”