Somatic USP8 mutations are frequent events in corticotroph tumor progression causing Nelson's tumor.

OBJECTIVE: Somatic mutations in the ubiquitin-specific protease 8 (USP8) gene are frequent in corticotroph tumors causing Cushing's disease (CD). Corticotroph tumor progression, the so-called Nelson's syndrome (NS), is a potentially life-threatening complication of bilateral adrenalectomy in patients with refractory CD that is caused by the development of an ACTH-secreting tumor of the pituitary gland. Whether USP8 alterations are also present in progressive Nelson's tumors has not been studied in detail so far. DESIGN AND METHODS: Retrospective, multicenter study involving tumors from 33 patients with progressive corticotroph tumors (29 females) and screening for somatic mutations on the mutational hotspot of the USP8 gene in the exon 14 with Sanger sequencing. RESULTS: Fifteen out of 33 tumors (45%) presented with a mutation in the exon 14 of USP8, with c.2159C>A (p.Pro720Gln) being the most frequent (9/33), followed by c.2155_2157delTCC (p.Ser718del, 4/33) and c.2152T>C (p.Ser718Pro, 2/33). This prevalence is similar to that previously reported for CD. Mutations were found exclusively in females. Other variables, such as age at diagnosis with NS, body mass index, hyperpigmentation, visual field defects, adenoma size or mortality, did not significantly differ between patients with wild-type and mutant tumors. Patients with USP8 mutant tumors exhibited higher levels of plasma ACTH after surgery (median: 640 vs 112 pg/mL, P = 0.03). No differences were observed in ACTH normalization (<50 pg/mL) and tumor control after surgery for Nelson's tumor. CONCLUSION: Somatic mutations in USP8 are common in Nelson's tumors, indicating that they do not drive the corticotroph tumor progression that leads to NS, and may be associated with a less favorable biochemical outcome after surgery for Nelson's tumor.

Possible role of a radiation-induced p53 mutation in a Nelson's syndrome patient with a fatal outcome.

Nelson's syndrome (NS) is characterized by the appearance and/or progression of ACTH-secreting pituitary macroadenomas in patients who had previously undergone bilateral adrenalectomy for the treatment of Cushing's disease. Such corticotroph macroadenomas respond poorly to currently available therapeutic options which include surgery, radiotherapy and chemotherapy. P53 protein accumulation may be detected by immunohistochemistry in pituitary corticotroph adenomas and it has been suggested that it might be causally related to tumor development. Wild type P53 protein plays an important role in the cellular response to ionizing radiation and other DNA damaging agents and is mutated in many human tumors. In this study we report an adult male patient with NS who underwent both transsphenoidal and transcranial pituitary surgeries, conventional and stereotaxic radiotherapy and brachytherapy. Despite of the efforts to control tumor mass and growth, this macroadenoma displayed relentless growth and aggressive behavior. DNA extracted from the first two surgical samples, as well as DNA from peripheral blood leukocytes disclosed normal p53 sequence. DNA extracted from tumor samples obtained at surgeries performed after pituitary irradiation carried a somatic heterozygous mutation, consisting of a deletion of four cytosines between nucleotides 12,144-12,149 in exon 4 of the p53 gene. This frameshift mutation creates a stop codon in exon 4 excluding the expression of a functional protein from the defective allele. These data demonstrate a possible association between the P53 protein loss of function induced by radiotherapy and the aggressive course of the disease in this patient.

Corticosteroid resistance and disease.

Both the unpredictability of side-effects and efficacy of glucocorticoid treatment can be problematic during clinical treatment. Here we discuss the evidence that exists for the hypothesis that individual glucocorticoid sensitivity underlies this problem. We suggest that glucocorticoid sensitivity is actually much more dynamic and common than previously thought. It is postulated that acquired tissue-specific glucocorticoid resistance could play a role in the origin and pathogenesis of depression, autoimmune disorders and AIDS. Moreover, recent genetic research has shown mutations in the glucocorticoid receptor (GR) gene and GR protein which are suggested to play a role in the pathogenesis of leukaemia, hereditary glucocorticoid resistance and Nelson's syndrome. These findings indicate that variations in the GR and in glucocorticoid resistance play a central role in a wide variety of diseases.

Nelson's syndrome associated with a somatic frame shift mutation in the glucocorticoid receptor gene.

Nelson's syndrome is the appearance and/or progression of ACTH-secreting pituitary macroadenomas in patients who had previously undergone bilateral adrenalectomy for Cushing's disease. Extremely high plasma ACTH levels and aggressive neoplastic growth might be explained by the lack of appropriate glucocorticoid negative feedback due to defective glucocorticoid signal transduction. To study the glucocorticoid receptor (GR) gene in Nelson's syndrome, DNA was extracted from pituitary adenomas and leukocytes of four patients with this condition and amplified by PCR for direct sequence analysis. In one of the tumors, a heterozygous mutation, consisting of an insertion of a thymine between complementary DNA nucleotides 1188 and 1189, was found in exon 2. This frame-shift mutation led to premature termination at amino acid residue 366 of the wild-type coding sequence, excluding the expression of a functioning receptor protein from the defective allele. The mutation was not detected in the sequence of the GR gene in the patient's leukocyte DNA, indicating a somatic origin. By lowering the receptor number in tumorous cells, this defect might have caused local resistance to negative glucocorticoid feedback similar to that caused by the presence of a null allele in a kindred with the generalized glucocorticoid resistance syndrome. P53 protein accumulation, previously reported in 60% of corticotropinomas, could not be detected in any of the four pituitary tumors examined by immunohistochemistry. We suggest that a somatic GR defect might have played a pathophysiological role in the tumorigenesis of the corticotropinoma bearing this mutation.

Monoclonality of corticotroph macroadenomas in Cushing's disease.

The pathophysiological mechanism of pituitary ACTH oversecretion in Cushing's disease remains unclear. The question of whether a collection of corticotroph cells is a primary pituitary event or is driven by increased production of hypothalamic corticotropin releasing factor is still debated. Establishing whether or not there is a clonal nature of such pituitary lesions has important conceptual and practical implications. Clonal composition of corticotroph cell adenomas was determined by X chromosome inactivation analysis using a DNA probe, M27 beta, which detects a multiallelic polymorphism in 90% of females. A first digestion by PstI reveals the polymorphism. A second digestion by MspI or its methylation sensitive isoschizomer HpaII, distinguishes the active from the inactive copy. DNA was extracted from 11 corticotroph macroadenomas responsible for Cushing's disease or Nelson's syndrome. Eight of the 11 female patients were heterozygous for the locus and included in the study. Blood leukocytes were available for 5 females and were used as controls. All 8 tumors demonstrated a monoclonal pattern while the 5 leukocyte DNA were polyclonal. Ours results show that a somatic modification plays an important role in the pathogenesis of corticotroph macroadenomas allowing monoclonal expansion of a genetically aberrant cell.

Quantitative in-situ hybridization histochemistry of anterior pituitary hormone mRNA species in human pituitary adenomas.

We have examined the anterior pituitary hormone messenger (m) RNA species contained in biopsies of 41 pituitary tumours obtained at hypophysectomy using in-situ hybridization histochemistry. The adenoma were grouped clinically into 12 prolactinomas, 8 somatotrope adenomas, 16 non-functioning, 4 Nelson's syndrome, and 1 thyrotrope adenoma. Of these, 10 contained no detectable anterior pituitary hormone mRNA species and 11 appeared to be expressing the gene responsible for the patients' clinical state in isolation. In a number of cases the accumulation of specific mRNA species was not accompanied by an increase in the circulating levels of the corresponding hormone or subunit. Evidence of activation of more than 1 anterior pituitary hormone gene was present in 16 adenomas of which only 7 showed a pattern of activation or amplification of gene expression which would suggest deregulation of either the inositol phospholipid or cAMP second messenger pathway. It was therefore not possible from these data to postulate that isolated deregulation of a single second messenger transduction pathway is a common etiological factor in pituitary tumour formation.