Brown adipose tissue (BAT) and white adipose tissue (WAT) and adipocytes are targets of Trypanosoma cruzi infection. Adipose tissue obtained from CD-1 mice 15 days after infection, an early stage of infection revealed a high parasite load. There was a significant increase in macrophages in infected adipose tissue and a reduction in lipid accumulation, adipocyte size, and fat mass and increased expression of lipolytic enzymes. Infection increased levels of Toll-like receptor (TLR) 4 and TLR9 and in the expression of components of the mitogen-activated protein kinase pathway. Protein and messenger RNA (mRNA) levels of peroxisome proliferator-activated receptor γ were increased in WAT, whereas protein and mRNA levels of adiponectin were significantly reduced in BAT and WAT. The mRNA levels of cytokines, chemokines, and their receptors were increased. Nuclear Factor Kappa B levels were increased in BAT, whereas Iκκ-γ levels increased in WAT. Adipose tissue is an early target of T. cruzi infection.
High dietary intake of calcium has been classified as a probable cause of prostate cancer although the mechanism underlying the association between dietary calcium and prostate cancer risk is unclear. The vitamin D receptor (VDR) is a key regulator of calcium absorption. In the small intestine, VDR expression is regulated by the CDX-2 transcription factor, which binds a polymorphic site in the VDR gene promoter. We examined VDR Cdx2 genotype and calcium intake, assessed by a food frequency questionnaire, in 533 African American prostate cancer cases (256 with advanced stage at diagnosis, 277 with localized stage) and 250 African American controls who participated in the California Collaborative Prostate Cancer Study. We examined the effects of genotype, calcium intake, and diet-gene interactions by conditional logistic regression. Compared to men in the lowest quartile of calcium intake, men in the highest quartile had an approximately two-fold increased risk of localized and advanced prostate cancer (odds ratio [OR]= 2.20, 95% confidence interval [CI]= 1.40, 3.46), with a significant dose-response. Poor absorbers of calcium (VDR Cdx2 GG genotype) had a significantly lower risk of advanced prostate cancer (OR= 0.41, 95% CI = 0.19, 0.90). The gene-calcium interaction was statistically significant (p=0.03). Among men with calcium intake below the median (680 mg/day), carriers of the G allele had an approximately 50% decreased risk compared to men with the AA genotype. These findings suggest a link between prostate cancer risk and high intestinal absorption of calcium.
Vitamin D receptor; calcium absorption; genetic polymorphism; prostate cancer; African American
Thioredoxin interacting protein (TXNIP) has recently been described as a key regulator of energy metabolism through pleiotropic actions that include nutrient sensing in the mediobasal hypothalamus (MBH). However, the role of TXNIP in neurochemically specific hypothalamic subpopulations, and the circuits downstream from MBH TXNIP engaged to regulate energy homeostasis remain unexplored. To evaluate the metabolic role of TXNIP activity specifically within arcuate Agrp neurons, we generated Agrp-specific TXNIP gain- and loss-of-function mouse models using Agrp-Ires-cre mice, TXNIPflox/flox mice and a lentivector expressing the human TXNIP isoform conditionally in the presence of Cre recombinase. Overexpression of TXNIP in Agrp neurons predisposed to diet-induced obesity and adipose tissue storage by decreasing energy expenditure and spontaneous locomotion, without affecting food intake. Conversely, Agrp neuronal TXNIP deletion protected against diet-induced obesity and adipose tissue storage by increasing energy expenditure and spontaneous locomotion, without affecting food intake. TXNIP overexpression in Agrp neurons did not primarily affect glycemic control, whereas deletion of TXNIP in AgRP neurons improved fasting glucose levels and glucose tolerance independently of its effects on body weight and adiposity. Bidirectional manipulation of TXNIP expression induced reciprocal changes in central leptin sensitivity and the neural regulation of lipolysis. Together these results identify a critical role for TXNIP in Agrp neurons in mediating diet-induced obesity through the regulation of energy expenditure and adipose tissue metabolism, independently of food intake. They also reveal a previously unidentified role for Agrp neurons in the brain-adipose axis.
Vorinostat (V) at levels >2.5 μM enhances chemotherapy in vitro. Yet the approved oral dose of 400 mg inconsistently achieves this level in patients. We developed an intermittent oral pulse-dose schedule of V to increase serum levels. We combined V with the cyclin dependent kinase inhibitor flavopiridol (F) which increases V-induced apoptosis.
One week before combination treatment, V alone was given daily for 3d (cycle −1). Then V was given on d1-3 and d8-10, and F on d2 and d9, every 21-d. Due to neutropenia, this was modified to V on d1-3 and d15–17, and F on d2 and d16, every 28-d. Bolus and split-dose F schedules were studied.
34 patients were treated. On the 21-d schedule, the maximum tolerated dose (MTD) was V 600 mg/d and F 60 mg/m2 bolus. On the 28-d schedule, the MTD was V 800 mg/d and F 30 mg/m2 over 30 min and 30 mg/m2 over 4 h. V Cmax at the 800 mg dose was 4.8 μM (± 2.8). V Cmax ≥2.5 μM was achieved in 86% of patients at the MTD. F increased the Cmax of V by 27% (95% CI 11%–43%). F Cmax of ≥2 μM was achieved in 90% of patients. 8 patients had stable disease for on average 5.5 m (range 1.6–13.2 m).
Intermittent high dose oral V in combination with F is feasible and achieves target serum levels >2.5 μM. V concentrations higher than previously reported with oral dosing were achieved.
CDKs and CDK inhibitors; Histone deacetylase inhibitors; Phase I trials; Combination chemotherapy; Pharmacokinetics
Growth hormone (GH) exerts diverse tissue-specific metabolic effects that are not revealed by global alteration of GH action. To study the direct metabolic effects of GH in the muscle, we specifically inactivated the growth hormone receptor (ghr) gene in postnatal mouse skeletal muscle using the Cre/loxP system (mGHRKO model). The metabolic state of the mGHRKO mice was characterized under lean and obese states. High-fat diet feeding in the mGHRKO mice was associated with reduced adiposity, improved insulin sensitivity, lower systemic inflammation, decreased muscle and hepatic triglyceride content, and greater energy expenditure compared with control mice. The obese mGHRKO mice also had an increased respiratory exchange ratio, suggesting increased carbohydrate utilization. GH-regulated suppressor of cytokine signaling-2 (socs2) expression was decreased in obese mGHRKO mice. Interestingly, muscles of both lean and obese mGHRKO mice demonstrated a higher interleukin-15 and lower myostatin expression relative to controls, indicating a possible mechanism whereby GHR signaling in muscle could affect liver and adipose tissue function. Thus, our study implicates skeletal muscle GHR signaling in mediating insulin resistance in obesity and, more importantly, reveals a novel role of muscle GHR signaling in facilitating cross-talk between muscle and other metabolic tissues.
Previous evidence indicates that duodenal lipid sensing engages gut-brain neurocircuits to determine food intake and hepatic glucose production, but a potential role for gut-brain communication in the control of energy expenditure remains to be determined. Here, we tested the hypothesis that duodenal lipid sensing activates a gut–brain–brown adipose tissue neuraxis to regulate thermogenesis. We demonstrate that direct administration of lipids into the duodenum increases brown fat temperature. Co-infusion of the local anesthetic tetracaine with duodenal lipids abolished the lipid-induced increase in brown fat temperature. Systemic administration of the CCKA receptor antagonist devazepide blocked the ability of duodenal lipids to increase brown fat thermogenesis. Parenchymal administration of the N-methyl-d-aspartate receptor blocker MK-801 directly into the caudomedial nucleus of the solitary tract also abolished duodenal lipid-induced activation of brown fat thermogenesis. These findings establish that duodenal lipid sensing activates a gut–brain–brown fat axis to determine brown fat temperature, and thereby reveal a previously unappreciated pathway that regulates thermogenesis.
Sirtuin 1 (SIRT1) and its activator resveratrol are emerging as major regulators of metabolic processes. We investigate the site of resveratrol action on glucose metabolism and the contribution of SIRT1 to these effects. Because the arcuate nucleus in the mediobasal hypothalamus (MBH) plays a pivotal role in integrating peripheral metabolic responses to nutrients and hormones, we examined whether the actions of resveratrol are mediated at the MBH.
RESEARCH DESIGN AND METHODS
Sprague Dawley (SD) male rats received acute central (MBH) or systemic injections of vehicle, resveratrol, or SIRT1 inhibitor during basal pancreatic insulin clamp studies. To delineate the pathway(s) by which MBH resveratrol modulates hepatic glucose production, we silenced hypothalamic SIRT1 expression using a short hairpin RNA (shRNA) inhibited the hypothalamic ATP-sensitive potassium (KATP) channel with glibenclamide, or selectively transected the hepatic branch of the vagus nerve while infusing resveratrol centrally.
Our studies show that marked improvement in insulin sensitivity can be elicited by acute administration of resveratrol to the MBH or during acute systemic administration. Selective inhibition of hypothalamic SIRT1 using a cell-permeable SIRT1 inhibitor or SIRT1-shRNA negated the effect of central and peripheral resveratrol on glucose production. Blockade of the KATP channel and hepatic vagotomy significantly attenuated the effect of central resveratrol on hepatic glucose production. In addition, we found no evidence for hypothalamic AMPK activation after MBH resveratrol administration.
Taken together, these studies demonstrate that resveratrol improves glucose homeostasis mainly through a central SIRT1-dependent pathway and that the MBH is a major site of resveratrol action.
The topoisomerase I inhibitor, irinotecan, and its active metabolite SN-38 have been shown to induce G2/M cell cycle arrest without significant cell death in human colon carcinoma cells (HCT-116). Subsequent treatment of these G2/M-arrested cells with the cyclin-dependent kinase inhibitor, flavopiridol, induced these cells to undergo apoptosis. The goal of this study was to develop a noninvasive metabolic biomarker for early tumor response and target inhibition of irinotecan followed by flavopiridol treatment in a longitudinal study. A total of eleven mice bearing HCT-116 xenografts were separated into two cohorts where one cohort was administered saline and the other treated with a sequential course of irinotecan followed by flavopiridol. Each mouse xenograft was longitudinally monitored with proton (1H)-decoupled phosphorus (31P) magnetic resonance spectroscopy (MRS) before and after treatment. A statistically significant decrease in phosphocholine (p = 0.0004) and inorganic phosphate (p = 0.0103) levels were observed in HCT-116 xenografts following treatment, which were evidenced within twenty-four hours of treatment completion. Also, a significant growth delay was found in treated xenografts. To discern the underlying mechanism for the treatment response of the xenografts, in vitro HCT-116 cell cultures were investigated with enzymatic assays, cell cycle analysis, and apoptotic assays. Flavopiridol had a direct effect on choline kinase as measured by a 67% reduction in the phosphorylation of choline to phosphocholine. Cells treated with SN-38 alone underwent 83±5% G2/M cell cycle arrest compared to untreated cells. In cells, flavopiridol alone induced 5±1% apoptosis while the sequential treatment (SN-38 then flavopiridol) resulted in 39±10% apoptosis. In vivo 1H-decoupled 31P MRS indirectly measures choline kinase activity. The decrease in phosphocholine may be a potential indicator of early tumor response to the sequential treatment of irinotecan followed by flavopiridol in noninvasive and/or longitudinal studies.
irinotecan; flavopiridol; choline kinase; colon cancer; 1H-decoupled 31P MRS; apoptosis
The prevalence of low serum vitamin D levels in cancer patients with fatigue or poor appetite and their association with symptom burden and other correctable endocrine abnormalities were investigated.
Vitamin D deficiency in noncancer patients is associated with symptoms of fatigue, muscle weakness, and depression. These symptoms are common among advanced cancer patients. We investigated the prevalence of low serum vitamin D levels in cancer patients with fatigue or poor appetite and their association with symptom burden and other correctable endocrine abnormalities.
This was a retrospective review of 100 consecutive cancer patients with appetite or fatigue scores of ≥4 of 10 referred to a supportive care clinic. We investigated serum levels of 25(OH) vitamin D, cortisol, thyroid-stimulating hormone, and bioavailable testosterone. Symptoms were measured by the Edmonton Symptom Assessment Scale. Serum 25(OH) vitamin D <20 ng/mL was considered deficient; ≥20 ng/mL and <30 ng/mL were considered insufficient.
Patients were predominantly male (68%) and white (66%), with a median age of 60 years (range, 27–91 years). Gastrointestinal (30%) and lung (22%) cancers were predominant. Forty-seven patients (47%) were vitamin D deficient and 70 (70%) were insufficient. Thirteen of 70 patients (19%) with vitamin D insufficiency were on supplementation. Vitamin D deficiency was more common among nonwhites (82% versus 36%) and females. No significant association was found between vitamin D and symptoms. Hypogonadic males had a significantly lower mean 25(OH) vitamin D level than eugonadic males.
Low vitamin D levels were highly prevalent among advanced cancer patients with cachexia or fatigue. Vitamin D deficiency was more frequent among nonwhite and female patients. Vitamin D levels were also significantly lower in male patients with hypogonadism.
Vitamin D; Testosterone; Cancer; Symptoms
Flavopiridol, a Cdk inhibitor, potentiates irinotecan-induced apoptosis. In a phase I trial of sequential irinotecan and flavopiridol, 2 patients with advanced hepatocellular carcinoma (HCC) had stable disease (SD) for ≥14 months. We thus studied the sequential combination of irinotecan and flavopiridol in patients with HCC.
Patients with advanced HCC naïve to systemic therapy, Child-Pugh ≤B8, and Karnofsky performance score (KPS) ≥70% received 100 mg/m2 irinotecan followed 7 hours later by flavopiridol 60 mg/m2 weekly for 4 of 6 weeks. The primary end point was an improvement in progression-free survival at 4 months (PFS-4) from 33% to 54%, using a Simon's two-stage design. Tumors were stained for p53.
Only 16 patients in the first stage were enrolled: median age, 64 years; median KPS, 80%; Child-Pugh A, 87.5%; and stage III/IV, 25%/75%. The primary end point was not met; PFS-4 was 20%, leading to early termination of the study. Ten patients were evaluable for response: 1 had SD >1 year and 9 had disease progression. Grade 3 fatigue, dehydration, diarrhea, neutropenia with or without fever, lymphopenia, anemia, hyperbilirubinemia, and transaminitis occurred in ≥10% of the patients. Of the 9 patients who progressed, 5 had mutant p53 and 4 had wild-type p53. The patient with stable disease had wild-type p53.
Sequential irinotecan and flavopiridol are ineffective and poorly tolerated in patients with advanced HCC. Despite our limited assessments, it is possible that the presence of wild-type p53 is necessary but not sufficient to predict response in HCC.
Homeopathic pathogenetic trials usually rely on symptom self report measures. Adding objective biomarkers could enhance detection of subtle initial remedy effects. The present feasibility study examined electroencephalographic (EEG) effects of repeated olfactory administration of two polycrest remedies.
College student volunteers (ages 18–30, both sexes) from an introductory psychology course were screened for good health and relatively elevated Sulphur OR Pulsatilla symptom scores on the Homeopathic Constitutional Type Questionnaire. Subjects underwent a series of 3 once-weekly double-blind sessions during which they repeatedly sniffed the remedy matched to their CTQ type and solvent controls. Each remedy was given in a 6c, 12c, and 30c potency, one potency per week, in randomly assigned order. Solvent controls included both plain distilled water and a water-ethanol (95%) solution. All sniff test solutions were further diluted just prior to laboratory sessions (0.5 ml test solution in 150 ml distilled water). Within a session, remedies and control solvents were administered via 2-second sniffs (8 sniffs of each of 4 different succussion levels for the potency in randomized order). Primary outcome variable was relative EEG power (alpha 1 8–10 hertz; alpha 2 10–12 hertz) averaged over 19 electrode sites, including all succussions for a given potency.
Mixed-effect models revealed significant main effects for remedy type (Sulphur>Pulsatilla) in both alpha bands, controlling for gender, baseline resting EEG alpha, and solvent control responses. Additional analyses showed significant non-linear interactions between dilution and time (weekly session) in alpha 2 for both remedies and alpha 1 for Sulphur.
EEG alpha offers an objective biomarker of remedy effects for future studies and potential method for distinguishing time-dependent effects of specific remedies and remedy potencies from one another.
Electroencephalography; Homeopathy; Sulphur; Pulsatilla; EEG alpha; repeated measures; sniffing; olfactory administration
Infusions of lipids into the small intestine potently suppress ongoing feeding. Prior work has identified potential roles for gut extrinsic vagal and non -vagal sensory innervation in mediating the ability of gut lipid infusions to reduce food intake, but the local biochemical processes underlying gut lipid sensing at the level of the small intestine remain unclear. This manuscript will summarize recent progress in the identification and characterization of several candidate gut lipid sensing molecules important in the negative feedback control of ingestion, including the fatty acid translocase CD36, peroxisome proliferator-activated receptor alpha (PPAR-alpha), and the fatty acid ethanolamide oleoylethanolamide (OEA). In addition, this manuscript addresses a larger role for gut lipid sensing in the overall control of energy availability by modulating not only food intake but also hepatic glucose production.
gut-brain axis; vagal afferent; food intake; glucose homeostasis; nutrient sensing
Macroautophagy is a lysosomal degradative pathway that maintains cellular homeostasis by turning over cellular components. Here, we demonstrate a role for autophagy in hypothalamic agouti-related peptide (AgRP) neurons in the regulation of food intake and energy balance. We show that starvation-induced hypothalamic autophagy mobilizes neuron-intrinsic lipids to generate endogenous free fatty acids, which in turn regulate AgRP levels. The functional consequences of inhibiting autophagy are the failure to upregulate AgRP in response to starvation, and constitutive increases in hypothalamic levels of pro-opiomelanocortin and its cleavage product α-melanocyte stimulating hormone that typically contribute to a lean phenotype. We propose a new conceptual framework for considering how autophagy-regulated lipid metabolism within hypothalamic neurons may modulate neuropeptide levels to have immediate effects on food intake, as well as long-term effects on energy homeostasis. Regulation of hypothalamic autophagy could become an effective intervention in conditions such as obesity and the metabolic syndrome.
Uveal melanomas possess activation of the mitogen-activated protein kinase (MAPK) and phosphoinositide 3-kinase (PI3K)/AKT/mammalian Target of Rapamycin (mTOR) pathways. MAPK activation occurs via somatic mutations in the heterotrimeric G protein subunits GNAQ and GNA11 for over 70% of tumors and less frequently via V600E BRAF mutations. In this report, we describe the impact of dual pathway inhibition upon uveal melanoma cell lines with the MEK inhibitor selumetinib (AZD6244/ARRY-142886) and the ATP-competitive mTOR kinase inhibitor AZD8055. While synergistic reductions in cell viability were observed with AZD8055/selumetinib in both BRAF and GNAQ mutant cell lines, apoptosis was preferentially induced in BRAF mutant cells only. In vitro apoptosis assay results were predictive of in vivo drug efficacy as tumor regressions were observed only in a BRAF mutant xenograft model, but not GNAQ mutant model. We went on to discover that GNAQ promotes relative resistance to AZD8055/selumetinib-induced apoptosis in GNAQ mutant cells. For BRAF mutant cells, both AKT and 4E-BP1 phosphorylation were modulated by the combination; however, decreasing AKT phosphorylation alone was not sufficient and decreasing 4E-BP1 phosphorylation was not required for apoptosis. Instead, cooperative mTOR complex 2 (mTORC2) and MEK inhibition resulting in downregulation of the pro-survival protein MCL-1 was found to be critical for combination-induced apoptosis. These results suggest that the clinical efficacy of combined MEK and mTOR kinase inhibition will be determined by tumor genotype, and that BRAF mutant malignancies will be particularly susceptible to this strategy.
screening; secondary hypertension; adrenal; diagnostic accuracy
Sphingosine 1-phosphate (S1P) is an important mediator of cancer cell growth and proliferation. Production of S1P is catalyzed by sphingosine kinase 1 (SphK). Safingol, (L-threo-dihydrosphingosine) is a putative inhibitor of SphK. We conducted a phase I trial of safingol (S) alone and in combination with cisplatin (C).
A 3+3 dose escalation was used. For safety, S was given alone 1 week before the combination. S + C were then administered every 3 weeks. S was given over 60–120 minutes (min), depending on dose. 60 min later, C was given over 60 min. The C dose of 75 mg/m2 was reduced in cohort 4 to 60 mg/m2 due to excessive fatigue.
43 patients were treated. 41 were evaluable for toxicity and 37 for response. The maximum tolerated dose (MTD) was S 840 mg/m2 over 120 min C 60 mg/m2, every 3 weeks. DLTs attributed to cisplatin included fatigue and hyponatremia. DLT from S was hepatic enzyme elevation. S pharmacokinetic parameters were linear throughout the dose range with no significant interaction with C. Patients treated at or near the MTD achieved S levels of > 20 µM and maintained levels ≥ 5 µM for 4 hours. The best response was stable disease in 6 patients for on average 3.3 months (range 1.8 – 7.2 m). One patient with adrenal cortical cancer had significant regression of liver and lung metastases and another had prolonged stable disease. S was associated with a dose-dependent reduction in S1P in plasma.
Safingol, the first putative SphK inhibitor to enter clinical trials, can be safely administered in combination with cisplatin. Reversible dose-dependent hepatic toxicity was seen, as expected from preclinical data. Target inhibition was achieved with downregulation of S1P. The recommended phase 2 dose is S 840 mg/m2 and C 60 mg/m2, every 3 weeks.
Drug-mediated stimulation of cell death pathways; Pharmacokinetics and pharmacodynamics; Kinase and phosphatase inhibitors; Novel antitumor agents; Sphingosine Kinase; Sphingolipids
Tyrosine kinase inhibitors (TKI’s) of the epidermal growth factor receptor (EGFR) have activity in solid tumors. We evaluated an oral EGFR TKI, erlotinib, in patients with previously treated esophageal cancer.
Thirty patients with measurable, metastatic esophageal and gastroesophageal junction cancer were treated with 150 mg of erlotinib daily. EGFR negative (6, 20%) and EGFR overexpressing (24, 80%) tumors were treated. The majority were male (70%) with adenocarcinoma (57%) and had received prior chemotherapy (97%).
Two partial responses were seen in the EGFR+ cohort (2/24, 8%), and no responses in the EGFR- cohort (0/6). Reponses were limited to squamous cell cancer (2/13, 15%, duration 5.5–7 months). Time to tumor progression was greater in squamous cell (3.3 months, range 1–24 months) compared to adenocarcinoma (1.6 months, range 1–6 months, p = 0.026). Therapy was tolerable with the expected toxicity of skin rash (grade 1–2, 67%, grade 3, 10%).
Erlotinib has limited activity in esophageal cancer, with responses and some protracted stable disease observed in squamous cancer. Efficacy by EGFR status could not be assessed given the rarity of EGFR- tumors. Further evaluation of this agent in squamous cell carcinoma is warranted.
Esophageal cancer; squamous cancer; erolotinib
To determine whether office, home, ambulatory daytime and nighttime blood pressure (BP) responses to antihypertensive drug therapy measure the same signal and which method provides greatest power to identify genetic predictors of BP response.
We analyzed office, home, ambulatory daytime and nighttime BP responses in hypertensive adults randomized to atenolol (N = 242) or hydrochlorothiazide (N = 257) in the Pharmacogenomic Evaluation of Antihypertensive Responses Study. Since different measured BP responses may have different predictors, we tested the "same signal" model by using linear regression methods to determine whether known predictors of BP response depend on the method of BP measurement. We estimated signal-to-noise ratios and compared power to identify a genetic polymorphism predicting BP response measured by each method separately and by weighted averages of multiple methods.
After adjustment for pretreatment BP level, known predictors of BP response including plasma renin activity, race, and sex were independent of the method of BP measurement. Signal-to-noise ratios were more than 2-fold greater for home and ambulatory daytime BP responses than for office and ambulatory nighttime BP responses and up to 11-fold greater for weighted averages of all four methods. Power to identify a genetic polymorphism predicting BP response was directly related to the signal-to-noise ratio and, therefore, greatest with the weighted averages.
Since different methods of measuring BP response to antihypertensive drug therapy measure the same signal, weighted averages of the BP responses measured by multiple methods minimize measurement error and optimize power to identify genetic predictors of BP response.
hypertension; blood pressure monitoring; antihypertensive drug therapy; beta-blocker; thiazide diuretic; plasma renin activity
Serotonin reuptake transporter (SERT) is a key regulator of serotonin neurotransmission and a major target of antidepressants. Antidepressants, such as selectively serotonin reuptake inhibitors (SSRIs), that block SERT function are known to affect food intake and body weight. Here, we provide genetic evidence that food intake and metabolism are regulated by separable mechanisms of SERT function. SERT-deficient mice ate less during both normal diet and high fat diet feeding. The reduced food intake was accompanied with markedly elevated plasma leptin levels. Despite reduced food intake, SERT-deficient mice exhibited glucose intolerance and insulin resistance, and progressively developed obesity and hepatic steatosis. Several lines of evidence indicate that the metabolic deficits of SERT-deficient mice are attributable to reduced insulin-sensitivity in peripheral tissues. First, SERT-deficient mice exhibited beta-cell hyperplasia and islet-mass expansion. Second, biochemical analyses revealed constitutively elevated JNK activity and diminished insulin-induced AKT activation in the liver of SERT-deficient mice. SERT-deficient mice exhibited hyper-JNK activity and hyperinsulinemia prior to the development of obesity. Third, enhancing AKT signaling by PTEN deficiency corrected glucose tolerance in SERT-deficient mice. These findings have potential implications for designing selective SERT drugs for weight control and the treatment of metabolic syndromes.
To determine whether treatment response to the Aurora B kinase inhibitor, AZD1152, could be monitored early in the course of therapy by non-invasive [18F]FDG and/or [18F]FLT PET imaging.
AZD1152-treated and control HCT116 and SW620 xenograft-bearing animals were monitored for tumor size and by [18F]FDG and [18F]FLT PET imaging. Additional studies assessed the endogenous and exogenous contributions thymidine synthesis in the two cell lines.
Both xenografts showed a significant volume-reduction to AZD1152. In contrast, [18F]FDG uptake did not demonstrate a treatment response. [18F]FLT uptake decreased to less than 20% of control values in AZD1152-treated HCT116 xenografts, whereas [18F]FLT uptake was near background levels in both treated and untreated SW620 xenografts. The EC50 for AZD1152-HQPA was ~10 nM in both SW620 and HCT116 cells; in contrast, SW620 cells were much more sensitive to Methotrexate (MTX) and 5-Fluorouracil (5FU) than HCT116 cells. Immunoblot analysis demonstrated marginally lower expression of thymidine kinase in SW620 compared to HCT116 cells. The above results suggest that SW620 xenografts have a higher dependency on the de novo pathway of thymidine utilization than HCT116 xenografts.
AZD1152 treatment showed anti-tumor efficacy in both colon cancer xenografts. Although [18F]FDG PET was inadequate in monitoring treatment-response, [18F]FLT PET was very effective in monitoring response in HCT116 xenografts, but not in SW620 xenografts. These observations suggest that de novo thymidine synthesis could be a limitation and confounding factor for [18F]FLT PET imaging and quantification of tumor proliferation, and this may apply to some clinical studies as well.
Colon cancer; HCT116; SW620; Positron-Emission Tomography; AZD1152; Methotrexate - MTX; 5-Fluorouracil – 5-FU; Fluorodeoxythymidine – FLT; Fluorodeoxyglucose – FDG
Aurora kinases are mitotic serine/threonine protein kinases and are attractive novel targets for anticancer therapy. Many small-molecule inhibitors of Aurora kinases are currently undergoing clinical trials. Aurora A kinase is essential for successful mitotic transition. MK8745 is a novel and selective small-molecule inhibitor of Aurora A kinase. MK8745 induced apoptotic cell death in a p53-dependent manner when tested in vitro in cell lines of multiple lineages. Cells expressing wild-type p53 showed a short delay in mitosis followed by cytokinesis, resulting in 2N cells along with apoptosis. However, cells lacking or with mutant p53 resulted in a prolonged arrest in mitosis followed by endoreduplication and polyploidy. Cytokinesis was completely inhibited in p53-deficient cells, as observed by the absence of 2N cell population. The induction of apoptosis in p53-proficient cells was associated with activation of caspase 3 and release of cytochrome c but was independent of p21. Exposure of p53 wild-type cells to MK8745 resulted in the induction of p53 phosphorylation (ser15) and an increase in p53 protein expression. p53-dependent apoptosis by MK8745 was further confirmed in HCT 116 p53−/− cells transfected with wild-type p53. Transient knockdown of Aurora A by specific siRNA recapitulated these p53-dependent effects, with greater percent induction of apoptosis in p53 wild-type cells. In conclusion, our studies show p53 as a determining factor for induction of apoptosis vs. polyploidy upon inhibition of Aurora A.
Aurora A kinase; polyploidy; apoptosis; p53; cell cycle
Malignant peripheral nerve sheath tumors (MPNSTs) are soft tissue tumors with a very poor prognosis and largely resistant to chemotherapy. MPNSTs are characterized by activation of the ras pathway by loss of tumor suppressor neurofibromatosis type I (NF1). In view of this MPNST may be susceptible to inhibition of the activated Ras/Raf/MAPK pathway by the B-Raf inhibitor sorafenib.
MPNST (MPNST and ST8814) and dedifferentiated liposarcoma (LS141 and DDLS) human tumor cell lines were characterized for Ras activation and B-Raf expression. Tumor cells were treated with sorafenib and examined for growth inhibition, inhibition of phospho-MEK (p-MEK), phospho-ERK (p-ERK), cell cycle arrest, and changes in cyclin D1 and pRb expression.
MPNSTs were sensitive to sorafenib at nanomolar concentrations. This appeared to be due to inhibition of p-MEK, p-ERK, suppression of cyclin D1, hypophosphorylation of pRb at the CDK4 specific sites, resulting in a G1 cell cycle arrest. These effects were not seen in the liposarcoma cells, which either did not express B-Raf or showed decreased Ras activation. SiRNA-mediated depletion of B-Raf in MPNSTs also induced a G1 cell cycle arrest in these cells, with a marked inhibition of cyclin D1 expression and Rb phosphorylation, whereas depletion of C-Raf did not affect either.
With growth inhibition at the low nano-molar range, sorafenib, by inhibiting the MAPK pathway, may prove to be a novel therapy for patients with MPNST.
sorafenib; MPNST; MAPK; B-Raf
Solitary Fibrous Tumor (SFT) is a mesenchymal neoplasm composed of CD34-positive fibroblastic cells. The pathogenesis driving this neoplasm remains unclear, with no recurrent genetic aberrations described to date. Previous reports suggest a role for IGF2 overexpression in the pathogenesis of these tumors, implicated in triggering hypoglycemia in some patients. The expression profiling of 23 SFTs was investigated using an Affymetrix U133A platform. The transcriptional signature was compared to a set of 34 soft tissue sarcomas spanning 7 subtypes. Potential candidate genes were then further investigated for activating mutations or loss of imprinting (LOI). SFT had a distinct expression signature and clustered in a tight genomic cluster, separate from all other sarcoma subtypes. A number of overexpressed receptor tyrosine kinase (RTK) genes were identified in SFT, including DDR1, ERBB2 and FGFR1, however no mutations were identified by cDNA sequencing. Overexpression of IGF2 was uniformly detected in SFT, regardless of anatomic location and was related to LOI. In contrast, IGF1 and JUN overexpression was seen in pleural, but not meningeal location. SFT shows a distinctive expression signature, with overexpression of DDR1, ERBB2, and FGFR1. Despite of lack of activating mutations in these RTKs, therapy with specific inhibitors targeting these kinases might be considered in advanced/metastatic cases. Our results confirm LOI in several tumors expressing high levels of IGF2, which may explain the observed paraneoplastic hypoglycemia.
solitary fibrous tumor; expression profiling; IGF2; DDR1
The objective of this study was to examine the association between calcium intake and prostate cancer risk. We hypothesized that calcium intake would be positively associated with lower risk for prostate cancer.
We used data from a case-control study conducted among veterans between 2007 and 2010 at the Durham Veterans Affairs Medical Center. The study consisted of 108 biopsy-positive prostate cancer cases, 161 biopsy-negative controls, and 237 healthy controls. We also determined whether these associations differed for blacks and whites or for low-grade (Gleason score <7) and high-grade prostate cancer (Gleason score ≥7). We administered the Harvard food frequency questionnaire to assess diet and estimate calcium intake. We used logistic regression models to obtain odds ratios (ORs) and 95% confidence intervals (CIs).
Intake of calcium from food was inversely related to risk for prostate cancer among all races in a comparison of cases and biopsy-negative controls (P = .05) and cases and healthy controls (P = .02). Total calcium was associated with lower prostate cancer risk among black men but not among white men in analyses of healthy controls. The highest tertile of calcium from food was associated with lower risk for high-grade prostate cancer in a comparison of high-grade cases and biopsy-negative controls (OR, 0.37; 95% CI, 0.15-0.90) and high-grade cases and healthy controls (OR, 0.38; 95% CI, 0.17-0.86).
Calcium from food is associated with lower risk for prostate cancer, particularly among black men, and lower risk for high-grade prostate cancer among all men.