A luteinizing hormone-, alpha-subunit- and prolactin-secreting pituitary adenoma responsive to somatostatin analogs: in vivo and in vitro studies.

OBJECTIVE: Evaluation of the efficiency of somatostatin analogues in the treatment of a mixed luteinizing hormone (LH)-, alpha-subunit-, prolactin (PRL)-secreting pituitary adenoma. DESIGN: A 30-year-old woman, with amenorrhaea-galactorrhaea, presented with a pituitary macroadenoma. The endocrine evaluation showed high plasma levels of PRL, LH, and alpha-subunit inhibited by 65%, 65% and 33% respectively under octreotide test (200 microg, s.c.). Long-term treatment with slow release (SR) lanreotide (30 mg/10 days, i.m.) restored menstrual cycles and normalized PRL values. Due to persisting supranormal levels of LH and alpha-subunit, and to the absence of tumoral shrinkage, the adenoma was resected by the transsphenoidal route. METHODS: In vitro characterization of the somatostatin receptor subtypes (SSTR) expression and functionality. Real-time polymerase chain reaction was performed to quantify the expression of SSTR mRNAs and functionality of the SSTRs was assessed in cell culture studies with various concentrations of native somatostatin (SRIF-14) and of analogues preferential for SSTR2 or SSTR5. RESULTS: This adenoma presented with high levels of SSTR2, SSTR3 and SSTR5 mRNAs, as compared with a series of gonadotroph adenomas. In cell culture studies, PRL, LH and alpha-subunit were inhibited by 60%, 47% and 33% respectively by SRIF-14 at a concentration of 10 nmol/l. The SSTR2 (BIM-23197, lanreotide) and SSTR5 (BIM-23268) preferential analogues both produced a partial 21-38% inhibition of PRL, LH, and alpha-subunit release. DISCUSSION: In this plurihormonal-secreting adenoma, the high efficacy of somatostatin analogues to inhibit PRL, LH and alpha-subunit secretion in vivo may be explained by the unusually high level of expression and by the functionality of both SSTR2 and SSTR5 receptor subtypes.

Squelching of ETS2 transactivation by POU5F1 silences the human chorionic gonadotropin CGA subunit gene in human choriocarcinoma and embryonic stem cells.

The subunit genes encoding human chorionic gonadotropin, CGA, and CGB, are up-regulated in human trophoblast. However, they are effectively silenced in choriocarcinoma cells by ectopically expressed POU domain class 5 transcription factor 1 (POU5F1). Here we show that POU5F1 represses activity of the CGA promoter through its interactions with ETS2, a transcription factor required for both placental development and human chorionic gonadotropin subunit gene expression, by forming a complex that precludes ETS2 from interacting with the CGA promoter. Mutation of a POU5F1 binding site proximal to the ETS2 binding site does not alter the ability of POU5F1 to act as a repressor but causes a drop in basal promoter activity due to overlap with the binding site for DLX3. DLX3 has only a modest ability to raise basal CGA promoter activity, but its coexpression with ETS2 can up-regulate it 100-fold or more. The two factors form a complex, and both must bind to the promoter for the combination to be transcriptionally effective, a synergy compromised by POU5F1. Similarly, in human embryonic stem cells, which express ETS2 but not CGA, ETS2 does not occupy its binding site on the CGA promoter but is found instead as a soluble complex with POU5F1. When human embryonic stem cells differentiate in response to bone morphogenetic protein-4 and concentrations of POU5F1 fall and hCG and DLX3 rise, ETS2 then occupies its binding site on the CGA promoter. Hence, a squelching mechanism underpins the transcriptional silencing of CGA by POU5F1 and could have general relevance to how pluripotency is maintained and how the trophoblast lineage emerges from pluripotent precursor cells.

D-amino acid substitution of residues 32 to 46 of the glycoprotein hormone common alpha-subunit: development of a synthetic glycoprotein hormone antagonist.

We have used single- or double-point D-amino acid substitutions to study the structure-function relationships involving residues 32 to 46 of the glycoprotein hormone common alpha-subunit (GPHa) and the testicular follicle-stimulating hormone (FSH) and luteinizing hormone (LH/hCG) receptors. D-Amino acid substitution analogs of GPHa(32-46) were synthesized and tested in both FSH and hCG radioligand receptor assays using bovine calf testis membranes as receptor source. Correct orientation of the amino acid side chains was generally of paramount importance for peptide interaction with receptor and bioactivity. Most substitutions with corresponding D-amino acids did not enhance the potency of native GPHa(32-46). A significant increment in peptide potency, however, was observed by inversion of configuration at positions Ser-34 and Thr-39 with D-amino acid isoforms. Based on the relative potency of each peptide analog. [D-Ser-34, D-Thr-39]GPHa(32-46) was approximately twofold more potent than native peptide GPHa(32-46) in both FSH and hCG radioligand receptor assays. [D-Ser-34, D-Thr-39]GPHa(32-46) also markedly inhibited FSH-stimulated estradiol biosynthesis in cultured rat Sertoli cells. The present study is unique in that it represents the first report of utilizing D-amino acid substitution to develop more potent peptide analogs related to the glycoprotein hormone common alpha-subunit region 32-46. Our results offer hope for the development of more potent and stabile peptide antagonists of possible usefulness in fertility regulation and control.