Altered brown adipose tissue mitochondrial function in newborn fragile X syndrome mice.

Brown adipose tissue (BAT) mitochondria generate heat via uncoupled respiration due to excessive proton leak through uncoupling proteins (UCPs). We previously found hyperthermia in a newborn mouse model of fragile X syndrome and excessive leak in Fmr1 KO forebrain mitochondria caused by CoQ deficiency. The inefficient thermogenic nature of Fmr1 mutant forebrain mitochondria was reminiscent of BAT metabolic features. Thus, we aimed to characterize BAT mitochondrial function in these hyperthermic mice using a top-down approach. Although there was no change in steady-state levels of UCP1 expression between strains, BAT weighed significantly less in Fmr1 mutants compared with controls. Fmr1 KO BAT mitochondria demonstrated impaired substrate oxidation, lower mitochondrial membrane potentials and rates of respiration, and CoQ deficiency. The CoQ analog decylubiquinone normalized CoQ-dependent electron flux and unmasked excessive proton leak. Unlike mutant forebrain, where such deficiency resulted in pathological proton leak, CoQ deficiency within BAT mitochondria resulted largely in abnormal substrate oxidation. This suggests that CoQ is important in BAT for uncoupled respiration to produce heat during development. Although our data provide further evidence of a link between fragile X mental retardation protein (FMRP) and CoQ biosynthesis, the results highlight the importance of CoQ in developing tissues and suggest tissue-specific differences from CoQ deficiency. Because BAT mitochondria are primarily responsible for regulating core body temperature, the defects we describe in Fmr1 KOs could manifest as an adaptive downregulated response to hyperthermia or could result from FMRP deficiency directly.

Improving diagnostic recognition of primary hyperparathyroidism with machine learning.

BACKGROUND: Parathyroidectomy offers the only cure for primary hyperparathyroidism, but today only 50% of primary hyperparathyroidism patients are referred for operation, in large part, because the condition is widely under-recognized. The diagnosis of primary hyperparathyroidism can be especially challenging with mild biochemical indices. Machine learning is a collection of methods in which computers build predictive algorithms based on labeled examples. With the aim of facilitating diagnosis, we tested the ability of machine learning to distinguish primary hyperparathyroidism from normal physiology using clinical and laboratory data. METHODS: This retrospective cohort study used a labeled training set and 10-fold cross-validation to evaluate accuracy of the algorithm. Measures of accuracy included area under the receiver operating characteristic curve, precision (sensitivity), and positive and negative predictive value. Several different algorithms and ensembles of algorithms were tested using the Weka platform. Among 11,830 patients managed operatively at 3 high-volume endocrine surgery programs from March 2001 to August 2013, 6,777 underwent parathyroidectomy for confirmed primary hyperparathyroidism, and 5,053 control patients without primary hyperparathyroidism underwent thyroidectomy. Test-set accuracies for machine learning models were determined using 10-fold cross-validation. Age, sex, and serum levels of preoperative calcium, phosphate, parathyroid hormone, vitamin D, and creatinine were defined as potential predictors of primary hyperparathyroidism. Mild primary hyperparathyroidism was defined as primary hyperparathyroidism with normal preoperative calcium or parathyroid hormone levels. RESULTS: After testing a variety of machine learning algorithms, Bayesian network models proved most accurate, classifying correctly 95.2% of all primary hyperparathyroidism patients (area under receiver operating characteristic = 0.989). Omitting parathyroid hormone from the model did not decrease the accuracy significantly (area under receiver operating characteristic = 0.985). In mild disease cases, however, the Bayesian network model classified correctly 71.1% of patients with normal calcium and 92.1% with normal parathyroid hormone levels preoperatively. Bayesian networking and AdaBoost improved the accuracy of all parathyroid hormone patients to 97.2% cases (area under receiver operating characteristic = 0.994), and 91.9% of primary hyperparathyroidism patients with mild disease. This was significantly improved relative to Bayesian networking alone (P < .0001). CONCLUSION: Machine learning can diagnose accurately primary hyperparathyroidism without human input even in mild disease. Incorporation of this tool into electronic medical record systems may aid in recognition of this under-diagnosed disorder.

Notch3 expression correlates with thyroid cancer differentiation, induces apoptosis, and predicts disease prognosis.

BACKGROUND: Thyroid tumorigenesis is characterized by a progressive loss of differentiation exhibited by a range of disease variants. The Notch receptor family (1-4) regulates developmental progression in both normal and cancerous tissues. This study sought to characterize the third Notch isoform (Notch3) across the various differentiated states of thyroid cancer, and determine its clinical impact. METHODS: Notch3 expression was analyzed in a tissue microarray of normal and pathologic thyroid biopsies from 155 patients. The functional role of Notch3 was then investigated by upregulating its expression in a follicular thyroid cancer (FTC) cell line. RESULTS: Notch3 expression regressed across decreasingly differentiated, increasingly malignant thyroid specimens, correlated with clinicopathological attributes reflecting poor prognosis, and independently predicted survival following univariate and multivariate analyses. Overexpression of the active Notch3 intracellular domain (NICD3) in a gain-of-function FTC line led to functional activation of centromere-binding protein 1, while increasing thyroid-specific gene transcription. NICD3 induction also reduced tumor burden in vivo and initiated the intrinsic apoptotic cascade, alongside suppressing cyclin and B-cell lymphoma 2 family expression. CONCLUSIONS: Loss of Notch3 expression may be fundamental to the process of dedifferentiation that accompanies thyroid oncogenesis. Conversely, activation of Notch3 in thyroid cancer exerts an antiproliferative effect and restores elements of a differentiated phenotype. These findings provide preclinical rationale for evaluating Notch3 as a disease prognosticator and therapeutic target in advanced thyroid cancer. Cancer 2017;123:769-82. © 2016 American Cancer Society.

Notch1 Signaling Regulates the Aggressiveness of Differentiated Thyroid Cancer and Inhibits SERPINE1 Expression.

PURPOSE: Notch1, a transmembrane receptor, has been recently shown to aid in the determination of thyroid cell fate associated with tumorigenesis. This study aimed to investigate the clinical relevance of Notch1 and its role in the regulation of differentiated thyroid cancer (DTC) behavior. EXPERIMENTAL DESIGN: We examined Notch1 expression level and its relationship with clinicopathologic features and outcomes of DTC. Notch1 intracellular domain (NICD) was further characterized both in vitro and in vivo by gain-of-function assays using an inducible system. RESULTS: Notch1 expression levels were downregulated in primary DTC tissue samples compared with contralateral nontumor and benign thyroid tissues. Decreased Notch1 expression in DTC was associated with advanced patient age (P = 0.032) and the presence of extrathyroidal invasion (P = 0.005). Patients with lower Notch1 expression had a significantly higher recurrence rate (P = 0.038). Restoration of NICD in a stably doxycycline-inducible metastatic DTC cell line reduced cell growth and migration profoundly. Using an orthotopic thyroid cancer model, NICD induction significantly reduced the growth of the primary thyroid tumor and inhibited the development of lung metastasis. Serpin peptidase inhibitor, clade E, member 1 (SERPINE1) was discovered by microarray as the most significant gene downregulated by NICD. Further validation showed that the induction of NICD reduced SERPINE1 expression in a dose-dependent manner, whereas restoration of a relative higher level of SERPINE1 was observed with NICD back to minimal level. In addition, SERPINE1 knock-down inhibited DTC cell migration. CONCLUSIONS: Notch1 regulates the aggressive phenotypes of DTC, which could be mediated by SERPINE1 inhibition. Notch1/SERPINE1 axis warrants further investigation as a novel therapeutic target for advanced DTC. Clin Cancer Res; 22(14); 3582-92. ©2016 AACR.

Radioguided parathyroidectomy for tertiary hyperparathyroidism.

BACKGROUND: Tertiary hyperparathyroidism (3HPT) is defined as the persistent hyperproduction of parathyroid hormone and resulting hypercalcemia after renal transplantation. Here, we examine the utility of radioguided parathyroidectomy (RGP) in patients with 3HPT. MATERIALS AND METHODS: We reviewed a prospective surgery database containing 80 3HPT patients who underwent RGP from January 2001-July 2014 at our institution. We evaluated patient demographics, operative management, radioguided neoprobe utilization, and operative outcomes. Data are reported as mean ± standard error of the mean. RESULTS: The mean age of the patients was 52 ± 1 y, and 46% were male. A total of 69 patients had hyperplasia and received subtotal parathyroidectomy, whereas 5 patients had double adenomas and 6 patients had single adenomas. The average calcium level among 3HPT patients was 10.8 ± 0.1 mg/dL preoperatively and 8.7 ± 0.1 mg/dL postoperatively. In vivo radioguided counts normalized to background counts averaged 145 ± 4%, whereas ex vivo counts normalized to background counts averaged 69 ± 5%. All but one ex vivo count was >20%. Ectopically located glands were successfully localized in 38 patients using the gamma probe. Ex vivo percentage did not correlate with parathyroid gland weight, preoperative parathyroid hormone, or preoperative calcium. Our radioguided approach achieved normocalcemia in 96% of 3HPT patients undergoing RGP; two patients developed recurrent disease. CONCLUSIONS: In this series, all enlarged parathyroid glands were localized and resected using the gamma probe. Thus, RGP reliably localizes adenomatous, hyperplastic, and ectopically located glands in patients with 3HPT, resulting in high cure rate after resection.

The effect of cinacalcet on intraoperative findings in tertiary hyperparathyroidism patients undergoing parathyroidectomy.

INTRODUCTION: Tertiary hyperparathyroidism (3HPTH) patients who undergo parathyroidectomy (PTX) are often managed with calcium lowering medications such as cinacalcet (Sensipar) before surgery. Here, we assess how cinacalcet treatment influences intraoperative parathyroid hormone (IOPTH) kinetics and surgical findings in 3HPTH patients undergoing PTX. METHODS: We reviewed retrospectively 113 patients 3HPTH who underwent PTX, 14 of whom were taking cinacalcet and 112 who were not taking the drug. IOPTH levels fitted to linear curves versus time were used to evaluate the role of cinacalcet. RESULTS: Cinacalcet did not correlate with rates of cure (P = .41) or recurrence (P = .54). Patients taking cinacalcet experienced a steeper decrease in IOPTH compared with those not taking the medication (P = .005). Cinacalcet treatment was associated with an increase in rate of hungry bones (P = .04). Weights of the heaviest glands resected (P = .02) and preoperative PTH levels (P = .0004) were greater among patients taking cinacalcet. CONCLUSION: Perioperative cinacalcet treatment in patients with 3HPTH alters IOPTH kinetics by causing a steeper decrease in IOPTH, but does not require modification of the standard IOPTH protocol. Although cinacalcet use does not adversely affect cure rates, it is associated with greater preoperative PTH and an increased incidence of hungry bones, hence serving as an indicator of more severe disease. Cinacalcet does not need to be held before operation.

Thyroid: Medullary Carcinoma.

Medullary thyroid cancers (MTC) are rare neuroendocrine tumors arising from the parafollicular C-cells of the thyroid. In this review, we provide a general overview of the classification, pathology, and clinical management of MTC. In the latter half, we survey the underlying genetic framework of MTC and its potential implications within a diagnostic and therapeutic context.

The Phosphatidylinositol 3-kinase/Akt Signaling Pathway in Neuroendocrine Tumors.

The phosphatidylinositol 3-kinase (PI3K)-Akt pathway is often aberrantly activated in neuroendocrine-derived cancers. Therefore, selectively targeting this pathway using small-molecule inhibitors may reduce neuroendocrine tumor burden, potentiate adjunct therapies, and achieve symptomatic control for patients with hormonally active and inoperable disease. Here, we discuss the role of the PI3K-Akt pathway in the malignant transformation of neuroendocrine tumors, specifically carcinoids and small cell lung cancers. The collective findings presented in this review propose that selective targeting of the PI3K-Akt pathway may mitigate neuroendocrine tumor progression, thus offering a viable therapeutic approach for managing systemic disease.

Inhibition of mTOR radiosensitizes soft tissue sarcoma and tumor vasculature.

PURPOSE: The PI3K/Akt/mTOR prosurvival pathway is frequently up-regulated in soft tissue sarcoma. Mammalian target of rapamycin (mTOR) inhibitors, such as rapamycin, have recently shown clinical benefit in soft tissue sarcoma, and mTOR inhibition has also been associated with radiosensitization of carcinoma and endothelial cells. This study tested the hypothesis that rapamycin radiosensitizes soft tissue sarcoma and endothelial cells in vitro and in vivo through the inhibition of mTOR. EXPERIMENTAL DESIGN: Colony formation assays were done to determine the radiosensitizing properties of rapamycin on three human soft tissue sarcoma cell lines (SK-LMS-1, SW-872, and HT-1080) and human dermal microvascular endothelial cells (HDMEC). The functional effects of rapamycin and radiation on the endothelial compartment were evaluated with microvascular sprouting assays. The in vivo radiosensitizing activity of rapamycin was assessed with s.c. SK-LMS-1 nude mice xenografts treated with concurrent daily rapamycin, radiation, or both for three weeks. RESULTS: In vitro radiosensitization was shown in all three soft tissue sarcoma cell lines with minimally cytotoxic doses of rapamycin. SK-LMS-1 xenografts displayed significant tumor growth delay with rapamycin and radiation compared with either treatment alone. Radiation resulted in transient increased mTOR function, whereas rapamycin abolished this signaling in irradiated and unirradiated samples. In HDMEC, rapamycin and radiation reduced microvessel sprouting, but did not alter colony formation. CONCLUSIONS: Minimally cytotoxic concentrations of rapamycin inhibited the mTOR cascade in culture and in vivo while radiosensitizing soft tissue sarcoma, and produced synergistic effects with radiation on HDMEC microvessel formation. By targeting both tumor and endothelial compartments, rapamycin produced potent radiosensitization of soft tissue sarcoma xenografts. Clinical trials combining rapamycin and radiotherapy in soft tissue sarcoma are warranted.

Gemcitabine-mediated radiosensitization of human soft tissue sarcoma.

BACKGROUND/PURPOSE: Local and systemic control of soft tissue sarcoma (STS) remains a clinical challenge, particularly for retroperitoneal, deep truncal, or advanced extremity disease. 2',2'-Difluoro-2'-deoxycytidine (gemcitabine) is a potent radiosensitizer in many tumor types, but it has not been studied in human STS. The purpose of this study was to determine the radiosensitizing potential of gemcitabine in preclinical models of human STS. MATERIALS AND METHODS: The in vitro radiosensitizing activity of gemcitabine was assessed with clonogenic survival assay on three human STS cell lines: SK-LMS-1 (leiomyosarcoma), SW-872 (liposarcoma), and HT-1080 (fibrosarcoma). Cell cycle distribution was determined using dual-channel flow cytometry. The in vivo radiosensitizing activity of gemcitabine was assessed with subcutaneous SK-LMS-1 nude mice xenografts. Tumor-bearing mice were treated with concurrent weekly gemcitabine and fractionated daily radiotherapy (RT) (2 Gy daily) for 3 weeks (a total dose of 30 Gy). RESULTS: The 50% inhibitory concentration (IC(50)) of gemcitabine for the human STS cell lines ranged from 10 to 1000 nM. Significant in vitro radiosensitization was demonstrated in all three human STS cell lines using gemcitabine concentrations at and below the IC(50). Maximal radiosensitization was associated with accumulation of cells in early S-phase. SK-LMS-1 xenografts displayed significant tumor growth delay with combined gemcitabine and RT compared to either treatment alone. Treatment related toxicity was greatest in the gemcitabine plus RT arm, but remained at an acceptable level. CONCLUSIONS: Gemcitabine is a potent radiosensitizer in preclinical models of human STS. Clinical trials combining gemcitabine and RT in human STS are warranted.