Purpose
Klinefelter syndrome (KS) is characterized by late adolescence/young adulthood onset of primary hypogonadism. Hypogonadotropic hypogonadism (HH), when congenital, is usually associated with ...absent/incomplete puberty and low/normal gonadotropins. We report the clinical and genetic features of two subjects with KS and an unexpected HH hormone profile.
Methods
Magnetic resonance imaging (MRI) of hypothalamus-pituitary region and next generation sequencing (NGS) of congenital HH-associated genes were obtained. A narrative review of the literature was conducted.
Results
Patients were diagnosed with Klinefelter syndrome following karyotype analysis. Nevertheless, they showed unusual features: both had incomplete puberty, low gonadotropins and testosterone levels, and the first one was anosmic. Sellar lesions were excluded by MRI, and NGS was negative in both subjects. Our data add to those of the only 14 similar cases reported so far. Unexplained HH rarely occurs in KS and is variably associated with anosmia, other pituitary hormones deficiencies and heterogeneous karyotypes. However, most cases show an early, pre-pubertal onset of hypogonadism. If the causes behind this gonadotropins defect are largely unknown, hereby we provide the first review of the literature on this topic and propose some pathogenetic hypotheses, including the coexistence of KS and congenital HH as suggested by overlapping clinical features in some of these patients.
Conclusion
HH is an exceptional occurrence in Klinefelter syndrome and is associated with heterogeneous phenotypes and, probably, aetiologies. Moreover, KS could underlie HH nonresponsive to gonadotropins. An exhaustive diagnostic workup and a tailored clinical management are advisable in these rare forms.
Aneurysmal bone cysts (ABCs) are rare benign pseudotumoral bone lesions with potential aggressive behavior due to the extensive destruction of surrounding bone. Traditionally, these tumors were ...treated with open surgery, but there is more and more a shift to less invasive procedures. In particular, treatment for spinal ABCs is generally unsatisfactory due to the risk of morbidity, neurological impairment and recurrence, and there is a need for innovative therapies. Denosumab has been reported as a useful treatment in giant cell tumors of bone (GCTB), so its efficacy has been tested also in other fibro-osseus lesions affecting children and adolescents, such as spinal aneurysmal bone cysts. The pediatric literature is limited to case reports and small series, all of which highlight the efficacy of this treatment on lesions growth and associated bone pain. Some of these reports have already reported well known side effects associated with denosumab, such as hypocalcemia at the beginning of the treatment, and rebound hypercalcemia at the discontinuation. The latter seems to be more frequent in children and adolescents than in adults, probably due to the higher baseline bone turnover in children. In addition, the use of denosumab in young patients could affect both bone modeling and remodeling, even if the consequences on the growing skeleton have not been reported in detail. Here we describe the case of a spinal ABC diagnosed in an 8-year old young boy which was not accessible to surgery but responded favorably to denosumab. Our aim is to describe the rapid changes in mineral and bone homeostasis in this patient, that required advice from the experts of the European Reference Network (ERN) for rare bone and endocrine diseases.
Objective
The serum calcium (Ca)–to–phosphorus (P) ratio has been proposed to identify patients with primary hyperparathyroidism and chronic hypoparathyroidism (HPT), but it has never been tested in ...pseudohypoparathyroidism (PHP). The aim of this study was to test the performance of Ca/P ratio in PHP diagnosis compared with that in healthy subjects and patients with HPT for differential diagnosis.
Design
A retrospective, cross-sectional, and observational study was carried out.
Methods
Serum Ca, P, creatinine, parathyroid hormone (PTH), and albumin were collected. Ca and P were expressed in mmol/L. Ca/P diagnostic performance was evaluated by receiver operating characteristic curve, sensitivity, specificity, and accuracy.
Results
A total of 60 patients with PHP, 60 patients with HPT, and 120 controls were enrolled. The Ca/P ratio was lower in patients with PHP and HPT than that in controls (p < 0.0001). The cutoff of 1.78 (2.32 if Ca and P measured in mg/dL) for Ca/P ratio could identify patients with PHP and HPT among the entire cohort (sensitivity and specificity of 76%). No valid cutoff of Ca/P was found to distinguish patients with PHP from patients with HPT; in this case, PTH above 53.0 ng/dL identified patients with PHP (sensitivity and specificity of 100%). The index (Ca/P × PTH) above 116 ng/L recognized patients with PHP from controls (sensitivity of 84.7% and specificity of 87.4%), whereas (Ca/P × PTH) below 34 ng/L recognized patients with HPT from controls (sensitivity of 88.9% and specificity of 90.8%).
Conclusions
The Ca/P ratio below 1.78 (2.32 CU) is highly accurate to identify patients with PHP and HPT, although it is not reliable to differentiate these two conditions. The index (Ca/P × PTH) is excellent to specifically recognize PHP or HPT from healthy subjects.
The actin binding protein filamin A (FLNA) is required for somatostatin receptor 2 (SSTR2) and dopamine receptor 2 (DRD2) expression and signaling in GH- and PRL-secreting PitNETs, respectively, ...playing a role in tumor responsiveness to somatostatin receptors ligands and dopaminergic drugs. FLNA functions are regulated by several mechanisms, including phosphorylation. It has been shown that in GH-secreting PitNETs FLNA phosphorylation on Ser2152 (P-FLNA) switches FLNA function from a scaffold that allows SSTR2 signal transduction, to a signal termination protein that hampers SSTR2 antitumoral effects. Aims of the present study were to evaluate in PRL- and ACTH-secreting PitNETs cell lines MMQ and AtT-20 the effects of cAMP pathway activation and DRD2 agonist on P-FLNA and the impact of P-FLNA on DRD2 signal transduction. We found that forskolin increased (+2.2 ± 0.8-fold, p < 0.01 in MMQ; +1.9 ± 0.58-fold, p < 0.05 in AtT-20), and DRD2 agonist BIM53097 reduced (-49.4 ± 25%, p < 0.001 in MMQ; -45.8 ± 28%, p < 0.05 in AtT-20), P-FLNA on Ser2152. The overexpression of a phosphomimetic (S2152D) FLNA mutant in both cell lines prevented DRD2 antiproliferative effects, that were comparable in cells transfected with empty vector, wild-type FLNA as well as phosphodeficient FLNA mutant (S2152A) (-20.6 ± 5% cell proliferation, p < 0.001 in MMQ; -36.6 ± 12%, p < 0.01 in AtT-20). Accordingly, S2152D FLNA expression abolished the expected ability of BIM53097 to increase or decrease, in MMQ and in AtT20 respectively, ERK phosphorylation, an effect that was maintained in S2152A FLNA expressing cells (+1.8 ± 0.65-fold, p < 0.05 in MMQ; -55 ± 13%, p < 0.01 in AtT-20). In addition, the inhibitory effects of DRD2 on hormone secretion (-34.3 ± 6% PRL, p < 0.05 in MMQ; -42.8 ± 22% ACTH, p < 0.05 in AtT-20, in cells expressing S2152A FLNA) were completely lost in S2152D FLNA transfected cells. In conclusion, our data demonstrated that cAMP pathway and DRD2 agonist regulated FLNA activity by increasing or decreasing, respectively, its phosphorylation. Moreover, we found that P-FLNA prevented DRD2 signaling in PRL- and ACTH-secreting tumoral pituitary cell lines, suggesting that this FLNA modification might represent a new regulatory mechanism shared by different GPCRs. In PitNETs expressing DRD2, modulation of P-FLNA might suggest new pharmacological strategies to overcome drug resistance, and P-FLNA might represent a new biomarker for tumor responsiveness to dopaminergic agents.
Syndrome of inappropriate antidiuresis (SIAD) can be a complication of hypothalamus-pituitary surgery. The use of tolvaptan in this setting is not well established, hence the primary aim of this ...study was to assess the sodium correction rates attained with tolvaptan compared with standard treatments (fluid restriction and/or hypertonic saline). Furthermore, we compared the length of hospital stay in the two treatment groups and investigated the occurrence of overcorrection and side effects including osmotic demyelination syndrome.
We retrospectively reviewed 308 transsphenoidal surgical procedures performed between 2011 and 2019 at our hospital. We selected adult patients who developed post-operative SIAD and recorded sodium monitoring, treatment modalities and outcomes. Correction rates were adjusted based on pre-treatment sodium levels.
Twenty-nine patients (9.4%) developed post-operative SIAD. Tolvaptan was administered to 14 patients (median dose 15 mg). Standard treatments were employed in 14 subjects (fluid restriction n=11, hypertonic saline n=1, fluid restriction and hypertonic saline n=2). Tolvaptan yielded higher adjusted sodium correction rates (12.0 mmolL
/24h and 13.4 mmolL
/48h) than standard treatments (1.8 mmolL
/24h, p<0.001, and 4.5 mmolL
/48h, p=0.004,
tolvaptan). The correction rate exceeded 10 mmolL
/24h or 18 mmolL
/48h in 9/14 and 2/14 patients treated with tolvaptan, respectively, and in no patient who received standard treatments. No side effects including osmotic demyelination occurred. Tolvaptan was associated with a shorter hospital stay (11
15 days, p=0.01).
Tolvaptan is more effective than fluid restriction (with or without hypertonic saline) and allows for a shortened hospital stay in patients with SIAD after transsphenoidal surgery. However, its dose and duration should be carefully tailored, and close monitoring is recommended to allow prompt detection of overcorrection.
The insulin-like growth factor 2 (IGF2) promotes cell growth by overactivating the IGF system in an autocrine loop in adrenocortical carcinomas (ACCs). The cytoskeleton protein filamin A (FLNA) acts ...as a repressor of IGF2 mitogenic signalling in ACC cells. The aims of this study were to test FLNA expression by immunohistochemistry in 119 ACCs and 26 adrenocortical adenomas (ACAs) and to evaluate its relationship with clinicopathological features and outcome in ACCs. We found that 71.4% of ACCs did not express FLNA, whereas FLNA absence was a rare event in ACAs (15.4%,
< 0.001 vs. ACCs). In addition, the expression of FLNA was associated with a less aggressive tumour behaviour in ACCs. Indeed, the subgroup of ACCs with high FLNA showed a lower ENSAT stage, Weiss score, and S-GRAS score compared to ACCs with low FLNA expression (
< 0.05). Moreover, patients with high FLNA had a longer overall survival than those with low FLNA (
< 0.05). In conclusion, our data suggest that FLNA may represent a "protective" factor in ACCs, and the integration of FLNA immunohistochemical expression in ACC tissues along with other clinical and molecular markers could be helpful to improve diagnostic accuracy and prognosis prediction in ACCs.
The mammalian target of rapamycin (mTOR) inhibitor everolimus has been shown to display antiproliferative effects on a wide spectrum of tumors.
In vitro
studies demonstrated that everolimus inhibited ...pituitary neuroendocrine tumor (PitNET) cell growth in a subset of patients. Sensitivity to everolimus is reduced by an escape mechanism that increases AKT phosphorylation (p-AKT), leading to pro-survival pathway activation. Dopamine receptor type 2 (DRD2) mediates a reduction of p-AKT in a subgroup of non-functioning PitNETs (NF-PitNETs) and in prolactin-secreting tumor cells (MMQ cells) through a β-arrestin 2-dependent mechanism. The aim of this study was to investigate the efficacy of everolimus combined with DRD2 agonist cabergoline in reducing NF-PitNET primary cells and MMQ cell proliferation and to evaluate AKT phosphorylation and a possible role of β-arrestin 2. We found that 9 out of 14 NF-PitNETs were resistant to everolimus, but the combined treatment with cabergoline inhibited cell proliferation in 7 out of 9 tumors (-31.4 ± 9.9%,
p
< 0.001
vs
. basal) and reduced cyclin D3 expression. In the everolimus-unresponsive NF-PitNET group, everolimus determined a significant increase of p-AKT/total-AKT ratio (2.1-fold,
p
< 0.01,
vs
. basal) that was reverted by cabergoline cotreatment. To investigate the molecular mechanism involved, we used MMQ cells as a model of everolimus escape mechanism. Indeed everolimus did not affect MMQ cell proliferation and increased the p-AKT/total-AKT ratio (+1.53 ± 0.24-fold,
p
< 0.001
vs
. basal), whereas cabergoline significantly reduced cell proliferation (-22.8 ± 6.8%,
p
< 0.001
vs
. basal) and p-AKT. The combined treatment of everolimus and cabergoline induced a reduction of both cell proliferation (-34.8 ± 18%,
p
< 0.001
vs
. basal and
p
< 0.05
vs
. cabergoline alone) and p-AKT/total-AKT ratio (-34.5 ± 14%,
p
< 0.001
vs
. basal and
p
< 0.05
vs
. cabergoline alone). To test β-arrestin 2 involvement, silencing experiments were performed in MMQ cells. Our data showed that the lack of β-arrestin 2 prevented the everolimus and cabergoline cotreatment inhibitory effects on both p-AKT and cell proliferation. In conclusion, this study revealed that cabergoline might overcome the everolimus escape mechanism in NF-PitNETs and tumoral lactotrophs by inhibiting upstream AKT activation. The co-administration of cabergoline might improve mTOR inhibitor antitumoral activity, paving the way for a potential combined therapy in β-arrestin 2-expressing NF-PitNETs or other PitNETs resistant to conventional treatments.
Patients with adrenal insufficiency (AI) may be exposed to supraphysiological glucocorticoids levels during standard treatment with cortisone acetate (CA) or immediate-release hydrocortisone (IR-HC). ...Recent studies, predominantly including patients in IR-HC treatment, suggested that modified-release hydrocortisone (MRH) provide a more physiological cortisol rhythm, improving metabolic control and quality of life. Our primary aim was to assess clinical and biochemical modifications in patients shifted from CA to MRH.
We designed a retrospective longitudinal study, enrolling 45 AI patients (22 primary and 23 secondary AI) treated exclusively with CA thrice daily, shifted to MRH once daily; 29/45 patients concluded at least 18-months follow-up (MRH-group). We recruited 35 AI patients continuing CA as a control group (CA-group). Biochemical and clinical data, including metabolic parameters, bone quality, and symptoms of under- or overtreatment were collected. In 24 patients, a daily salivary cortisol curve (SCC) performed before and one month after shifting to MRH was compared to healthy subjects (HS).
No significant changes in glycometabolic and bone parameters were observed both in MRH and CA-groups during a median follow-up of 35 months. A more frequent decrease in blood pressure values (23.1% vs 2.8%, p=0.04) and improvement of under- or overtreatment symptoms were observed in MRH vs CA-group. The SCC showed a significant steroid overexposure in both CA and MRH-groups compared to HS AUC (area under the curve) = 74.4 ± 38.1 nmol×hr/L and 94.6 ± 62.5 nmol×hr/L respectively, vs 44.1 ± 8.4 nmol×hr/L, p<0.01 for both comparisons, although SCC profile was more similar to HS in MRH-group.
In our experience, patients shifted from CA to equivalent doses of MRH do not show significant glycometabolic modifications but blood pressure control and symptoms of over-or undertreatment may improve. The lack of amelioration in glucose metabolism and total cortisol daily exposure could suggest the need for a dose reduction when shifting from CA to MRH, due to their different pharmacokinetics.
Cushing’s Disease (CD) is a rare condition characterized by an overproduction of ACTH by an ACTH-secreting pituitary tumor, resulting in an excess of cortisol release by the adrenal glands. Somatic ...mutations in the deubiquitinases USP8 and USP48, and in BRAF genes, have been reported in a subset of patients affected by CD. The aim of this study was to characterize the genetic profile of a cohort of 60 patients with ACTH-secreting tumors, searching for somatic mutations in USP8, USP48, and BRAF hotspot regions. Seven patients were found to carry USP8 somatic mutations in the well-characterized 14-3-3 protein binding motif (n = 5 P720R, n = 1 P720Q, n = 1 S718del); 2 patients were mutated in USP48 (M415I); no mutation was identified in BRAF. In addition, a novel USP8 variant, G664R, located in exon 14, upstream of the 14-3-3 protein binding motif, was identified in 1 patient. Functional characterization of USP8 G664R variant was performed in murine corticotroph tumor AtT-20 cells. Transient transfection with the USP8 G664R variant resulted in a significant increase of ACTH release and cell proliferation (+114.5 ± 53.6% and +28.3 ± 2.6% vs. empty vector transfected cells, p < 0.05, respectively). Notably, USP8 proteolytic cleavage was enhanced in AtT-20 cells transfected with G664R USP8 (1.86 ± 0.58–fold increase of N-terminal USP8 fragment, vs. WT USP8, p < 0.05). Surprisingly, in situ Proximity Ligation Assay (PLA) experiments showed a significant reduction of PLA positive spots, indicating USP8/14-3-3 proteins colocalization, in G664R USP8 transfected cells with respect to WT USP8 transfected cells (−47.9 ± 6.6%, vs. WT USP8, p < 0.001). No significant difference in terms of ACTH secretion, cell proliferation and USP8 proteolytic cleavage, and 14-3-3 proteins interaction was observed between G664R USP8 and S718del USP8 transfected cells. Immunofluorescence experiments showed that, contrary to S718del USP8 but similarly to WT USP8 and other USP8 mutants, G664R USP8 displays an exclusive cytoplasmic localization. In conclusion, somatic mutations were found in USP8 (13.3% vs. 36.5% incidence of all published mutations) and USP48 (3.3% vs. 13.3% incidence) hotspot regions. A novel USP8 variant was identified in a CD patient, and in vitro functional studies in AtT-20 cells suggested that this somatic variant might be clinically relevant in ACTH-secreting tumor pathogenesis, expanding the characterization of USP8 functional domains.
Hypoparathyroidism (HypoPT) or pseudo-hypoparathyroidism (pseudo-HypoPT) during pregnancy may cause maternal and fetal/neonatal complications. In this regard, only a few case reports or case series ...of pregnant or lactating women have been published. The purpose of this study was to describe clinical and biochemical course, pharmacological management, and potential adverse events during pregnancy and post-partum in pregnant women with HypoPT or pseudo-HypoPT. This was a retrospective, observational, multicenter, study involving nine Italian referral centers for endocrine diseases affiliated with the Italian Society of Endocrinology and involved in "Hypoparathyroidism Working Group". This study identified a cohort of 28 women (followed between 2005 and 2018) with HypoPT (n = 25, 84% postsurgical, 16% idiopathic/autoimmune) and pseudo-HypoPT (n = 3). In HypoPT women, the mean calcium carbonate dose tended to increase gradually from the first to third trimester (+ 12.6%) in pregnancy. This average increase in the third trimester was significantly greater compared to the pre-pregnancy period (p value = 0.03). However, analyzing the individual cases, in 44% the mean calcium dosage remained unchanged throughout gestation. Mean calcitriol doses tended to increase during pregnancy, with a statistically significant increase between the third trimester and the pre-pregnancy period (p value = 0.02). Nevertheless, analyzing the individual cases, in the third trimester most women with HypoPT (64%) maintained the same dosage of calcitriol compared to the first trimester. Both mean calcium carbonate and calcitriol doses tended to decrease from the third trimester to the post-partum six months. Most identified women (~ 70%) did not display maternal complications and (~ 90%) maintained mean serum albumin-corrected total calcium levels within the low-to-mid normal reference range (8.5 + or - 0.8 mg/dl) during pregnancy. The main complications related to pregnancy period included: preterm birth (n = 3 HypoPT women), and history of miscarriages (n = 6 HypoPT women and n = 2 pseudo-HypoPT women). This study shows that mean serum albumin-corrected total calcium levels were carefully monitored during pregnancy and post-pregnancy, with limited evaluation of other biochemical parameters, such as serum phosphate, 24 h urinary calcium, 25-OH vitamin D, and creatinine clearance. To avoid complications in mothers affected by (HypoPT) or (pseudo-HypoPT) and offspring, intense biochemical, clinical and pharmacological monitoring during pregnancy and breastfeeding is highly recommended.
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