HO-1 Limits the Efficacy of Vemurafenib/PLX4032 in BRAFV600E Mutated Melanoma Cells Adapted to Physiological Normoxia or Hypoxia.

Induction of heme oxygenase 1 (HO-1) favors immune-escape in BRAFV600 melanoma cells treated with Vemurafenib/PLX4032 under standard cell culture conditions. However, the oxygen tension under standard culture conditions (~18 kPa O2) is significantly higher than the physiological oxygen levels encountered in vivo. In addition, cancer cells in vivo are often modified by hypoxia. In this study, MeOV-1 primary melanoma cells bearing the BRAFV600E mutation, were adapted to either 5 kPa O2 (physiological normoxia) or 1 kPa O2 (hypoxia) and then exposed to 10 µM PLX4032. PLX4032 abolished ERK phosphorylation, reduced Bach1 expression and increased HO-1 levels independent of pericellular O2 tension. Moreover, cell viability was significantly reduced further in cells exposed to PLX4032 plus Tin mesoporphyrin IX, a HO-1 inhibitor. Notably, our findings provide the first evidence that HO-1 inhibition in combination with PLX4032 under physiological oxygen tension and hypoxia restores and increases the expression of the NK ligands ULBP3 and B7H6 compared to cells exposed to PLX4032 alone. Interestingly, although silencing NRF2 prevented PLX4032 induction of HO-1, other NRF2 targeted genes were unaffected, highlighting a pivotal role of HO-1 in melanoma resistance and immune escape. The present findings may enhance translation and highlight the potential of the HO-1 inhibitors in the therapy of BRAFV600 melanomas.

HO-1 downregulation favors BRAFV600 melanoma cell death induced by Vemurafenib/PLX4032 and increases NK recognition.

Heme oxygenase 1 (HO-1) plays a pivotal role in preventing cell damage. Indeed, through the antioxidant, antiapoptotic and anti-inflammatory properties of its metabolic products, it favors cell adaptation against different stressors. However, HO-1 induction has also been related to the gain of resistance to therapy in different types of cancers and its involvement in cancer immune-escape has been hypothesized. We have investigated the role of HO-1 expression in Vemurafenib-treated BRAFV600 melanoma cells in modulating their susceptibility to NK cell-mediated recognition. Different cell lines, isolated in house from melanoma patients, have been exposed to 1-10 µM PLX4032, which efficiently reduced ERK phosphorylation. In three lines, Vemurafenib was able to induce only a limited decrease in cell viability, while HO-1 expression was upregulated. HO-1 silencing/inhibition was able to induce a further significant reduction of Vemurafenib-treated melanoma viability. Moreover, while NK cell degranulation and killing activity were decreased upon interaction with melanoma exposed to Vemurafenib, HO-1 silencing was able to completely restore NK cell ability to degranulate and kill. Furthermore, melanoma cell treatment with Vemurafenib downregulated the expression of ligands of NKp30 and NKG2D activating receptors, and HO-1 silencing/inhibition was able to restore their expression. Our results indicate that HO-1 downregulation can both improve the efficacy of Vemurafenib on melanoma cells and favor melanoma susceptibility to NK cell-mediated recognition and killing.

Intraoperative parathyroid hormone (PTH) testing in patients with primary hyperparathyroidism and PTH levels in the normal range.

BACKGROUND: Primary hyperparathyroidism is a common endocrine disorder. Hypercalcemia with normal PTH levels is very unusual and can lead to diagnostic difficulties. There are very few very few studies in the literature and all with limited numerical samples. The goal of the present study was to determine the real incidence and characteristics of primary hyperparathyroidism with normal PTH and to evaluate if intraoperative PTH testing is useful in these patients. METHODS: We performed a retrospective review of 314 patients who had undergone parathyroidectomy to treat primary hyperparathyroidism between January 2002 and December 2016. Patients were divided in two groups according to biochemical preoperative findings: in Group A were included patients with normal serum PTH, in Group B those with increased serum PTH. RESULTS: Nine patients (3.7%) were included in group A and 235 in group B. Patients in group A were younger (51.5 ± 12.9 years vs 59.6 ± 12.5); preoperative serum calcium and the incidence of coexisting thyroid disease were similar between the two groups. Symptomatic patients were more frequent in Group A (77.8% vs 39.1%; p = 0.048). There were no significant differences regarding preoperative localization studies and surgical procedure. Intraoperative PTH determination demonstrated sensitivity of 86% in group A and 97% in group B, specificity and positive predictive value of 100% in both the groups, negative predictive value of 67% in group A and 79% in group B. Histopathological examination demonstrated a single gland disease in 8 (88.9%) patients in group A and a multi gland disease in 1 (11.1%), in group B single gland disease was found in 218 (92.8%) patients and multi gland disease in 17 (7.2%). Unsuccessful surgery with persistent or recurrent hyperparathyroidism occurred in 1 (11.1%) patient in group A and 4 (1.7%) in group B. CONCLUSIONS: Primary hyperparathyroidism with normal PTH is rare but physicians should be aware of this possibility in patients with hypercalcaemia. Patients with normal PTH levels are younger and more frequently symptomatic. Intraoperative PTH testing plays an important role in the operative management even in such patients.

Parathyroidectomy for primary hyperparathyroidism in the elderly: experience of a single endocrine surgery center.

BACKGROUND: Primary hyperparathyroidism is a common endocrine disease, and its incidence increases with age. AIMS: Our aim was to retrospectively evaluate the impact of age on patient outcomes following parathyroidectomy for primary hyperparathyroidism. METHODS: Two-hundred fifty-six consecutive patients undergoing parathyroidectomy with preoperative diagnosis of primary hyperparathyroidism were divided into three groups according to patient age: group A, ≤64 years; group B, 65-74 years; and group C, ≥75 years. RESULTS: Thyroid disease was associated with the hyperparathyroidism in 44 patients (28.2%) in group A, 34 (44.7%) in B, and 10 (41.7%) in C (p < 0.01). Minimally invasive parathyroidectomy was performed in 104 patients (66.7%) in group A, 35 (46.1%) in B, and 8 (33.3%) in C (p < 0.01). Conversion to bilateral exploration was carried out in five cases in group A (4.6%), three in B (8.3%), and two in C (20%). Multiglandular disease was observed in six patients (3.8%) in group A, seven (9.2%) in B, and five (20.8%) in C (p = 0.012). Mean postoperative stay was similar between groups; no major complications and no cases of mortality occurred. DISCUSSION: Multiglandular disease is more common in older patients than younger individuals, and minimally invasive approaches are less used in this patient group. Increased surgical risk and paucity of symptoms in these patients sometimes result in a delay in surgical treatment. CONCLUSIONS: Parathyroidectomy is a safe and effective procedure to perform in elderly patients. Multiglandular disease was found to be more prevalent in older patients, but minimally invasive parathyroidectomy can be performed safely. Surgeons should consider parathyroidectomy in patients with primary hyperparathyroidism regardless of age.

Feasibility of unilateral parathyroidectomy in patients with primary hyperparathyroidism and negative or discordant localization studies.

The purpose of this study was to examine the feasibility of unilateral parathyroidectomy in patients with primary hyperparathyroidism and negative or discordant localization studies. We included in our study 72 patients with preoperative diagnosis of primary hyperparathyroidism who had negative or discordant preoperative studies. In 66 patients, studies were discordant while in six were both negative. In 40 (55.6 %) patients initial approach was a bilateral exploration. In 32 cases (44.4 %) initial surgery was a unilateral exploration: in 26 conservative approach was successful, in six mini-invasive surgery failed and a bilateral exploration was necessary due to IOPTH negative test (five cases) or to the impossibility to find a pathological gland during exploration (one case). Intra-operative PTH test showed a sensitivity of 93.2 %, a specificity of 92.3 %, and an accuracy of 93.1 %. Multiple gland disease was found in 8 (11.1 %) patients (two double adenoma and six multiple gland hyperplasia). Mean operative time was lower in unilateral exploration group (87.9 ± 43.8 min). Comparing unilateral surgery in negative or discordant studies with 77 consecutive patients who underwent focused surgery with positive and concordant studies, conversion to bilateral exploration rate was statistically significantly higher in the first group (15.6 %). We believe that unilateral parathyroidectomy can be safely performed also in patients with discordant localization studies with a high cure rate; in these cases, however, the use of intra-operative PTH is absolutely necessary. We suggest the need for referral of these patients to high-volume medical centers for thyroid and parathyroid surgery.

Controversies in the management of parathyroid carcinoma: A case series and review of the literature.

Parathyroid carcinoma is a rare malignancy representing less than 1% of primary hyperparathyroidism cases. Its management is controversial due to lack of large-scale, multicentric studies. We report 8 new cases of parathyroid carcinoma and review the literature. Preoperative diagnosis of carcinoma was possible in 2 (25%) cases. Unclear surgical margins were present in 5 (62.5%) patients; 4 of them underwent subsequent re-exploration and ipsilateral hemithyroidectomy, in one case associated to central lymph node dissection. Recurrent disease is reported in 2 (25%) patients. Considering the high incidence of local recurrence in case of unclear surgical margins, a re-exploration with ipsilateral hemithyroidectomy is indicated in these patients. A neck dissection should be performed only in case of clinically involved lymph nodes, avoiding prophylactic lymphectomy. An aggressive approach is indicated in case of local or distant recurrence to reduce hypercalcemia.

Differentiated thyroid cancer in the elderly: our experience.

INTRODUCTION: The objective of this retrospective study was to investigate clinical and pathologic characteristics of differentiated thyroid cancer in elderly patients and to evaluate the results of surgical treatment in this age group. METHODS: The clinical records of patients who underwent total thyroidectomy between 2002 and 2012 with histopathological diagnosis of differentiated thyroid cancer were analyzed. Patients were divided into two groups: those 65 years old or older were included in group A (101), those younger in group B (354). RESULTS: The mean surgical time was 100.9 ± 30.5 min in group A and 100.7 ± 27.6 in B. Postoperative stay was significantly longer in group A (2.8 ± 1.5 days vs 2.4 ± 0.7; p < 0.01). Classic papillary carcinoma was more frequent in group B, whereas follicular variant of papillary carcinoma and tall cell carcinoma in A. In group B node metastases were nearly twice. In Group A transient hypoparathyroidism occurred in 25 patients (24.8%), permanent hypoparathyroidism in 4 (4%), hematoma in 6 (5.9%), recurrent nerve palsy in 2 (2%), and wound infection in 2 (2%). In group B transient and permanent hypoparathyroidism occurred in 48 and 7 patients respectively (13.6% and 2%), hematoma in 4 (1.1%), recurrent nerve palsy in 5 (1.4%), and wound infection in 1 (0.3%). CONCLUSIONS: Differentiated thyroid carcinoma is more aggressive in elderly patients for biological causes connected to age and to histotype but also for the diagnostic delay. Thyroid surgery in elderly patients is safe when the procedure is carried out by experienced staff. Total thyroidectomy is the surgical operation of choice.

Cervical lymph node sarcoidosis mimicking a parathyroid adenoma: a clinical case.

We report a case of cervical lymph node sarcoidosis misdiagnosed as parathyroid adenoma. This is the second case described in the literature in which lymph node sarcoidosis was misdiagnosed as parathyroid adenoma on Tc-99m sestamibi (MIBI) scan, the first case localized in the neck. A 64-year-old woman presented with a hypercalcemia. Neck ultrasonography revealed a paratracheal hypoechoic mass of 15 mm with peripheral vascularization. MIBI scan and SPECT/CT identified a MIBI-positive area corresponding to the nodule detected by ultrasonography suggestive for a lower right parathyroid adenoma. A mass interpreted as the lower parathyroid was excised associated to a total thyroidectomy. Pathologic examination revealed a granulomatous lymph node consistent with active sarcoidosis. Sarcoidosis should be suspected as a cause of unexplained hypercalcemia and the differential diagnosis of hypercalcemia, even in presence of MIBI uptake, must include sarcoidosis localized in an isolated cervical lymph node.

Surgery for primary hyperparathyroidism in patients with preoperatively negative sestamibi scan and discordant imaging studies: the usefulness of intraoperative parathyroid hormone monitoring.

The aim of this study was to evaluate the impact of intraoperative parathyroid hormone (PTH) monitoring on surgical strategy, intraoperative findings, and outcome in patients with negative sestamibi scintigraphy and with discordant imaging studies. We divided our 175 patients into 3 groups: group A was methoxyisobutylisonitrile (MIBI)-positive and ultrasonography positive and was concordant (114 patients), group B was MIBI-positive and ultrasonography-negative (50 patients), and group C was MIBI-and ultrasonography-negative (11 patients). The overall operative success was 99.12% in group A, 98% in group B, and 90.91% in group C, with an incidence of multiglandular disease of 3.5% in group A, 12% in group B, and 9.09% in group C. Intraoperative PTH monitoring changed the operative management in 2.63% of patients in group A and 14% in group B. The use of intraoperative PTH achieves to obtain excellent results in the treatment of primary hyperparathyroidism in high-volume centers, even in the most difficult cases, during MIBI-negative and discordant preoperative imaging studies.

Intraoperative parathyroid hormone assay during focused parathyroidectomy: the importance of 20 minutes measurement.

BACKGROUND: Parathyroid hormone (PTH) monitoring during the surgical procedure can confirm the removal of all hyperfunctioning parathyroid tissue, as the half-life of PTH is approximately 5 min. The commonly applied Irvin criterion is reported to correctly predict post-operative calcium levels in 96-98% of patients. However, the PTH baseline reference concentration is markedly influenced by surgical manipulations during preparation of the affected glands, interindividual variability of the PTH half-life and modifications in the physiological state of the patient during surgery. The aim of this study was to evaluate the possible impact of the measurement of intraoperative PTH 20 minutes after surgery. METHODS: Between 2003 and 2012, 188 patients underwent a focused parathyroidectomy associated to rapid intraoperative PTH assay monitoring. Blood samples were collected: 1) at pre-incision time, 2) at 10 min after gland excision and 3) at 20 min after excision, if a sufficient reduction of PTH value was not observed. On the bases of the Irvin criterion, an intra-operative PTH drop>50% from the highest either pre-incision or pre-excision level after parathyroid excision was considered a surgical success. RESULTS: A >50% decrease of PTH after gland excision compared to the highest pre-excision value occurred in 156/188 patients (83%) within 10 min and in further 12/188 after 20 minutes (6.4%). In the remaining 20 patients (10.6%) values of PTH remained substantially unchanged or decreased less than 50% and for this reason bilateral neck exploration was performed. An additional pathologic parathyroid was removed in 9 cases, a third in one. In the other 10 cases further neck exploration by a standard cervical approach was negative and in four of these persistent postoperative hypercalcemia was demonstrated. The overall operative success was 97.3%. Intraoperative PTH monitoring was accurate in predicting operative success or failure in 96.3% of patients. CONCLUSIONS: The 20 minutes PTH measurement appears very useful, avoiding unnecessary bilateral exploration and the related risk of complications with only a slight increase of the duration of surgery and of the costs. PTH values decreasing appeared to be influenced by surgical manipulations during minimally invasive parathyroidectomy.