Preclinical and clinical evidence that 18FDG-PET/CT is a reliable tool for the detection of early molecular responses to erlotinib in head and neck cancer.

Clin Cancer Res. 2010 Jul 26;

Authors: Vergez S, Delord JP, Thomas F, Rochaix P, Caselles O, Filleron T, Brillouet S, Canal P, Courbon F, Allal BC

PURPOSE: There is a clinical need to identify predictive markers of the responses to EGFR tyrosine-kinase inhibitors (EGFR-TKI). Positron Emission Tomography of deoxy-2-[18F]Fluoro-D-glucose (18FDG-PET/CT) could be a tool of choice for monitoring the early effects of this class of agent on tumor activity.EXPERIMENTAL DESIGN: Using models of human head and neck carcinoma (CAL33 and CAL166 cell lines), we tested first in vitro and in vivo, whether the in vivo changes in 18FDG-PET/CT uptake were associated with the molecular and cellular effects of the EGFR-TKI erlotinib. Then, the pathological and morphological changes and the 18FDG-PET/CT uptake before and after erlotinib exposure in patients were analyzed.RESULTS: Erlotinib strongly inhibited ERK-1/2 phosphorylation in both preclinical models and in patients. Western blotting, immunofluorescence and immunohistochemistry showed that erlotinib did not modify Glut-1 expression at the protein level either in cell line models or in tumor tissue from mouse xenografts or in patients. Phospho-ERK-1/2 inhibition was associated with a reduction in 18FDG uptake in animal and human tumors. The Biological Volume was more accurate than the Standardized Uptake Value for the evaluation of the molecular responses.CONCLUSIONS: These results show that the 18FDG-PET/CT response is a reliable surrogate marker of the effects of erlotinib in head and neck carcinoma.

PMID: 20660574 [PubMed - as supplied by publisher]

Small-Animal PET/CT for Monitoring the Development and Response to Chemotherapy of Thymic Lymphoma in Trp53-/- Mice.

J Nucl Med. 2010 Jul 21;

Authors: Walter MA, Hildebrandt IJ, Hacke K, Kesner AL, Kelly O, Lawson GW, Phelps ME, Czernin J, Weber WA, Schiestl RH

Transgenic mouse models of human cancers represent one of the most promising approaches to elucidate clinically relevant mechanisms of action and provide insights into the treatment efficacy of new antitumor drugs. The use of Trp53 transgenic mice (Trp53 knockout [Trp53(-/-)] mice) for these kinds of studies is, so far, restricted by limitations in detecting developing tumors and the lack of noninvasive tools for monitoring tumor growth, progression, and treatment response. METHODS: We hypothesized that quantitative small-animal PET with (18)F-FDG was able to detect the onset and location of tumor development, follow tumor progression, and monitor response to chemotherapy. To test these hypotheses, C57BL/6J Trp53(-/-) mice underwent longitudinal small-animal PET during lymphoma development and gemcitabine treatment. Trp53 wild-type (Trp53(+/+)) mice were used as controls, and histology after full necropsy served as the gold standard. RESULTS: In Trp53(+/+) mice, the thymic standardized uptake value (SUV) did not exceed 1.0 g/mL, with decreasing (18)F-FDG uptake over time. Conversely, all Trp53(-/-) mice that developed thymic lymphoma showed increasing thymic glucose metabolism, with a mean SUV doubling time of 9.0 wk (range, 6.0-17.5 wk). Using an SUV of 3.0 g/mL as a criterion provided a sensitivity of 78% and a specificity of 100% for the detection of thymic lymphoma. Treatment monitoring with (18)F-FDG PET correctly identified all histologic responses and relapses to gemcitabine. CONCLUSION: (18)F-FDG small-animal PET can be used to visualize onset and progression of thymic lymphomas in Trp53(-/-) mice and monitor response to chemotherapy. Thus, (18)F-FDG small-animal PET provides an in vivo means to assess intervention studies in the Trp53 transgenic mouse model.

PMID: 20660381 [PubMed - as supplied by publisher]

TNF-alpha mRNA Expression Correlates with TGF-beta mRNA Expression In Vivo.

Inflammation. 2010 Jul 23;

Authors: Helmig S, Stephan P, Döhrel J, Schneider J

TNF-alpha is postulated to play a significant role in regulating TGF-beta(1) expression. In lung fibroblasts, for example, TNF-alpha is supposed to induce TGF-beta(1) via AP-1 activation. TNF-alpha receptor, knock-out mice are resistant to induced fibrosis and over-expression of TNF-alpha causes increased TGF-beta(1) production in mice. Therefore, we investigated whether TNF-alpha mRNA levels are associated with the TGF-beta(1) mRNA levels of blood leucocytes in humans. Quantitative real-time PCR of TNF-alpha and TGF-beta(1) was performed in 118 Germans. Calculations of expression were made with the 2(-DeltaDeltaCT) method. When the investigated population was divided in two groups (TNF-alpha low and TNF-alpha high) by the median of the determined TNF-alpha expression, highly significant (p < 0.0001) differences of TGF-beta1 mRNA expression were revealed. Additionally, dividing the investigated population into quartiles of the determined TNF-alpha expression showed significantly different TGF-beta1 mRNA expressions. Comparing the determined CT-values of TNF-alpha in context with these of TGF-beta1, a coefficient of determination R (2) = 0.4635 was calculated. In this study we demonstrated in vivo a significant association of the relative TNF-alpha/B2M mRNA expression and the relative TGF-beta(1)/B2M mRNA expression in 118 Germans.

PMID: 20652825 [PubMed - as supplied by publisher]

5-HT1A and 5-HT1B receptors differentially modulate rate and timing of auditory responses in the mouse inferior colliculus.

Eur J Neurosci. 2010 Jul 14;

Authors: Castellan Baldan Ramsey L, Sinha SR, Hurley LM

Abstract Serotonin (5-hydroxytryptamine; 5-HT) is a physiological signal that translates both internal and external information about behavioral context into changes in sensory processing through a diverse array of receptors. The details of this process, particularly how receptors interact to shape sensory encoding, are poorly understood. In the inferior colliculus, a midbrain auditory nucleus, 5-HT1A receptors have suppressive and 5-HT1B receptors have facilitatory effects on evoked responses of neurons. We explored how these two receptor classes interact by testing three hypotheses: that they (i) affect separate neuron populations; (ii) affect different response properties; or (iii) have different endogenous patterns of activation. The first two hypotheses were tested by iontophoretic application of 5-HT1A and 5-HT1B receptor agonists individually and together to neurons in vivo. 5-HT1A and 5-HT1B agonists affected overlapping populations of neurons. During co-application, 5-HT1A and 5-HT1B agonists influenced spike rate and frequency bandwidth additively, with each moderating the effect of the other. In contrast, although both agonists individually influenced latencies and interspike intervals, the 5-HT1A agonist dominated these measurements during co-application. The third hypothesis was tested by applying antagonists of the 5-HT1A and 5-HT1B receptors. Blocking 5-HT1B receptors was complementary to activation of the receptor, but blocking 5-HT1A receptors was not, suggesting the endogenous activation of additional receptor types. These results suggest that cooperative interactions between 5-HT1A and 5-HT1B receptors shape auditory encoding in the inferior colliculus, and that the effects of neuromodulators within sensory systems may depend nonlinearly on the specific profile of receptors that are activated.

PMID: 20646059 [PubMed - as supplied by publisher]

Genetic Sphingosine Kinase 1 Deficiency Significantly Decreases Synovial Inflammation and Joint Erosions in Murine TNF-{alpha}-Induced Arthritis.

J Immunol. 2010 Jul 19;

Authors: Baker DA, Barth J, Chang R, Obeid LM, Gilkeson GS

Sphingosine kinase 1 (SphK1) is an enzyme that converts sphingosine to bioactive sphingosine-1-phosphate. Recent in vitro data suggest a potential role of SphK1 in TNF-alpha-mediated inflammation. Our aims in this study were to determine the in vivo significance of SphK1 in TNF-alpha-mediated chronic inflammation and to define which pathogenic mechanisms induced by TNF-alpha are SphK1 dependent. To pursue these aims, we studied the effect of SphK1 deficiency in an in vivo model of TNF-alpha-induced chronic inflammatory arthritis. Transgenic hTNF-alpha mice, which develop spontaneous inflammatory erosive arthritis beginning at 14-16 wk, were crossed with SphK1 null mice (SphK1(-/-)), on the C57BL6 genetic background. Beginning at 4 mo of age, hTNF/SphK1(-/-) mice had significantly less severe clinically evident paw swelling and deformity, less synovial and periarticular inflammation, and markedly decreased bone erosions as measured quantitatively through micro-CT images. Mechanistically, the mice lacking SphK1 had less articular cyclooxygenase 2 protein and fewer synovial Th17 cells than did hTNF/SphK1(+/+) littermates. Microarray analysis and real-time RT-PCR of the ankle synovial tissue demonstrated that hTNF/SphK1(-/-) mice had increased transcript levels of suppressor of cytokine signaling 3 compared with hTNF/SphK1(+/+) mice, likely also contributing to the decreased inflammation in the SphK1-deficient mice. Finally, significantly fewer mature osteoclasts were detected in the ankle joints of hTNF/SphK1(-/-) mice compared with hTNF/SphK1(+/+) mice. These data indicate that SphK1 plays a key role in hTNF-alpha-induced inflammatory arthritis via impacting synovial inflammation and osteoclast number.

PMID: 20644167 [PubMed - as supplied by publisher]

Substitution of 5-HT1A receptor signaling by a light-activated G protein-coupled receptor.

J Biol Chem. 2010 Jul 19;

Authors: Oh E, Maejima T, Liu C, Deneris ES, Herlitze S

Understanding serotonergic (5-HT) signaling is critical for understanding human physiology, behavior and neuropsychiatric disease. 5-HT mediates its actions via ionotropic and metabotropic 5-HT receptors. The 5-HT(1A) receptor is a metabotropic G protein-coupled receptor (GPCR) linked to the G(i/o) signaling pathway and has been specifically implicated in the pathogenesis of depression and anxiety. In order to understand and precisely control 5-HT(1A) signaling, we created a light-activated GPCR, which targets into 5-HT(1A) receptor domains and substitutes for endogenous 5-HT(1A) receptors. To induce 5-HT(1A)-like targeting, vertebrate rhodopsin was tagged with the C-terminal domain (CT) of 5-HT(1A) (Rh-CT(5-HT1A)). Rh-CT(5-HT1A) activates G protein-coupled inward rectifying K(+) channels (GIRK) in response to light and causes membrane hyperpolarization in hippocampal neurons, similar to the agonist-induced responses of the 5-HT(1A) receptor. The intracellular distribution of Rh-CT(5-HT1A) resembles that of the 5-HT(1A) receptor; Rh-CT(5-HT1A) localizes to somatodendritic sites and is efficiently trafficked to distal dendritic processes. Additionally, neuronal expression of Rh-CT(5-HT1A), but not Rh, decreases 5-HT(1A) agonist sensitivity, suggesting that Rh-CT(5-HT1A) and 5-HT(1A) receptors compete to interact with the same trafficking machinery. Finally, Rh-CT(5-HT1A) is able to rescue 5-HT(1A) signaling of 5-HT(1A) KO mice in cultured neurons and in slices of the dorsal raphe showing that Rh-CT(5-HT1A) is able to functionally compensate for native 5-HT(1A). Thus, as an optogenetic tool, Rh-CT(5-HT1A) has the potential to directly correlate in vivo 5-HT(1A) signaling with 5-HT neuron activity and behavior in both normal animals and animal models of neuropsychiatric disease.

PMID: 20643652 [PubMed - as supplied by publisher]

Effects of Central Airway Shunting on the Mechanical Impedance of the Mouse Lung.

Ann Biomed Eng. 2010 Jul 17;

Authors: Schwartz BL, Anafi RC, Aliyeva M, Thompson-Figueroa JA, Allen GB, Lundblad LK, Bates JH

The mechanical properties of the lung are embodied in its mechanical input impedance, which it is interpreted in physiological terms by being fit with a mathematical model. The normal lung is extremely well described by a model consisting of a single uniformly ventilated compartment comprised of tissue having a constant-phase impedance, but to describe the abnormal lung it frequently becomes necessary to invoke additional compartments. To date, all evidence of regional mechanical heterogeneity in the mouse lung has been assumed to be of the parallel variety. We therefore investigated the use of a serial heterogeneity model, relative to parallel heterogeneity and homogeneous models, for describing impedance spectra in mice subjected to a variety of interventions designed to make their lungs heterogeneous. We found that functional evidence of the finite stiffness of the airway wall in mice with airways obstruction can sometimes be apparent in lung impedance below 20 Hz. The model estimates of airway stiffness were smaller than direct estimates obtained from micro-CT images of the lung in vivo, suggesting that the conducting airways alone are likely not the precise anatomical correlate of proximal functional stiffness in the lung. Nevertheless, we conclude that central airway shunting in mice can sometimes be an important physiological phenomenon.

PMID: 20640513 [PubMed - as supplied by publisher]

18[F]FDG small animal PET study of sorafenib efficacy in lymphoma preclinical models.

Q J Nucl Med Mol Imaging. 2010 Jul 16;

Authors: Ambrosini V, Quarta C, Zinzani PL, Nanni C, Fini M, Torricelli P, Giavaresi G, D'errico-Grigioni A, Malvi D, Franchi R, Fanti S

AIM: Kinase inhibitors have been proposed as novel therapeutic agents in different forms of solid tumours. The Food and Drug Administration (FDA) approved the use of Sorafenib, an oral multikinase inhibitor, for advanced renal carcinoma and unresectable hepatocellular carcinoma. On-going studies are investigating the efficacy of Sorafenib in other solid tumours such as melanoma and non-small cells lung carcinoma and pre-clinical models showed the efficacy of treatment with Sorafenib in murine models of renal cells carcinoma, breast cancer, colon carcinoma and melanoma. To our knowledge, Sorafenib has never been employed in human lymphoma. The aim of the present study was to assess the efficacy of Sorafenib in murine models of human anaplastic large cells lymphoma (ALCL) and Hodgkin lymphoma (HD). METHODS: Sorafenib cytotoxicity was assessed in vitro and growth inhibition (IC50) was calculated. Cells were assayed for Caspase-3 to measure apoptosis. Human ALCL and HD xenografts in NOD/SCID mice were monitored by small animal positron emission tomography (PET) and computed tomography (CT) over time. Tumour bearing animals were randomly selected to receive treatment with Sorafenib or no treatment. Pathology was available in all cases. RESULTS: Sorafenib efficacy on cells proliferation and apoptosis (IC50: HD=0.0343mg/L; ALCL=0.319 mg/L) was confirmed in vitro. Caspase-3 production showed a dose-dependent trend reaching significantly higher values for 0.046mg/L and 0.465mg/L drug concentrations in both cell lines. In vivo experiments showed a progressive increase of tumour lesions metabolism and dimensions regardless treatment. CONCLUSION: Sorafenib showed a good citotoxic effect in vitro especially on human HD cell line, but these findings were not confirmed in vivo. The strong discrepancy between in vitro and in vivo results suggests that further studies are needed to better acknowledge the biodistribution and metabolism of Sorafenib in NOD/SCID mice. Factors influencing drug availability at tumour site or differences in the downstream pathways may be responsible for the scarse effect of treatment.

PMID: 20639808 [PubMed - as supplied by publisher]

Antiangiogenic Effect of Licochalcone A.

Biochem Pharmacol. 2010 Jul 14;

Authors: Kim YH, Shin EK, Kim DH, Lee HH, Park JH, Kim JK

To date, no antiangiogenic activity has been demonstrated for licochalcone A (LicA), a major phenolic constituent of Glycyrrhiza inflata, although it shows significant antitumor activity in human malignant cell lines. Our previous work demonstrated that LicA down-regulates inflammatory responses to lipopolysaccharide in murine macrophages. The purpose of the present study was to evaluate whether LicA inhibits angiogenesis, which is crucial for cancer development and progression. LicA significantly inhibited proliferation (20muM), migration (5-20muM), and tube formation (10-20muM) of human umbilical vascular endothelial cells (HUVECs) as well as microvessel growth from rat aortic rings (10-20muM). Furthermore, LicA significantly inhibited the growth of CT-26 colon cancer implants in BALB/c mice, with fewer CD31- and Ki-67-positive cells but more apoptotic cells. The underlying antiangiogenic mechanism of LicA correlated with down-regulation of vascular endothelial growth factor receptor (VEGFR)-2 activation. Our findings provide the first evidence that LicA inhibits angiogenesis in vitro and in vivo, perhaps by blocking VEGF/VEGFR-2 signaling. Inhibition of tumor growth may be attributed, at least in part, to decreased angiogenesis in LicA-treated mice. These findings emphasize the potential use of LicA against tumor development and progression in which angiogenesis is stimulated.

PMID: 20637733 [PubMed - as supplied by publisher]

TARP phosphorylation regulates synaptic AMPA receptors through lipid bilayers.

Neuron. 2010 Jun 10;66(5):755-67

Authors: Sumioka A, Yan D, Tomita S

Neurons use neurotransmitters to communicate across synapses, constructing neural circuits in the brain. AMPA-type glutamate receptors are the predominant excitatory neurotransmitter receptors mediating fast synaptic transmission. AMPA receptors localize at synapses by forming protein complexes with transmembrane AMPA receptor regulatory proteins (TARPs) and PSD-95-like membrane-associated guanylate kinases. Among the three classes of ionotropic glutamate receptors (AMPA, NMDA, and kainate type), AMPA receptor activity is most regulatable by neuronal activity to adjust synaptic strength. Here, we mutated the prototypical TARP, stargazin, and found that TARP phosphorylation regulates synaptic AMPA receptor activity in vivo. We also found that stargazin interacts with negatively charged lipid bilayers in a phosphorylation-dependent manner and that the lipid interaction inhibited stargazin binding to PSD-95. Cationic lipids dissociated stargazin from lipid bilayers and enhanced synaptic AMPA receptor activity in a stargazin phosphorylation-dependent manner. Thus, TARP phosphorylation plays a critical role in regulating AMPA receptor-mediated synaptic transmission via a lipid bilayer interaction.

PMID: 20547132 [PubMed - indexed for MEDLINE]

Viperin is highly induced in neutrophils and macrophages during acute and chronic lymphocytic choriomeningitis virus infection.

J Immunol. 2010 May 15;184(10):5723-31

Authors: Hinson ER, Joshi NS, Chen JH, Rahner C, Jung YW, Wang X, Kaech SM, Cresswell P

Although most cells are thought to respond to IFNs, there is limited information regarding specific cells that respond in vivo. Viperin is an IFN-induced antiviral protein and, therefore, is an excellent marker for IFN-responsive cells. In this study, we analyzed viperin expression in vivo during acute lymphocytic choriomeningitis virus Armstrong infection, which induces high levels of type I IFNs, and in persistently infected lymphocytic choriomeningitis virus carrier mice, which contain low levels of type I IFNs. Viperin was induced in lymphoid cells and dendritic cells (DCs) during acute infection and highly induced in neutrophils and macrophages. The expression kinetics in neutrophils, macrophages, and T and B cells paralleled IFN-alpha levels, but DCs expressed viperin with delayed kinetics. In carrier mice, viperin was expressed in neutrophils and macrophages but not in T and B cells or DCs. For acutely infected and carrier mice, viperin expression was IFN dependent, because treating type I IFNR knockout mice with IFN-gamma-neutralizing Abs inhibited viperin expression. Viperin localized to the endoplasmic reticulum and lipid droplet-like vesicles in neutrophils. These findings delineate the kinetics and cells responding to IFNs in vivo and suggest that the profile of IFN-responsive cells changes in chronic infections. Furthermore, these data suggest that viperin may contribute to the antimicrobial activity of neutrophils.

PMID: 20410488 [PubMed - indexed for MEDLINE]

A Dual Radiologic Contrast Agent Protocol for (18)F-FDG and (18)F-FLT PET/CT Imaging of Mice Bearing Abdominal Tumors.

Mol Imaging Biol. 2010 Jul 15;

Authors: Aide N, Kinross K, Beauregard JM, Neels O, Potdevin T, Roselt P, Dorow D, Cullinane C, Hicks RJ

PURPOSE: The aim of the study was to improve abdominal tumor detection by use of a dual radiologic contrast protocol. PROCEDURES: eXia160(R) (Benitio international) was mixed with 2-deoxy-2-[(18)F]fluoro-D: -glucose or 3'-[(18)F]fluoro-3'-deoxythymidine for intravenous (IV) injections. Omnipaque(R) 300 (GE healthcare) was used for intraperitoneal (IP) injections. Positron emission tomography/computed tomography (PET/CT) scans were acquired on a Siemens Biograph(R) equipped with point spread function reconstruction. The optimal concentration and injection schedule of IP contrast agent was studied in 12 mice. The impact of IP contrast media on PET quantitative accuracy was investigated by phantom studies and by imaging six mice before and after IP injection of Omnipaque(R). The impact of a dual contrast media protocol on tumor delineation and quantitation was evaluated in 15 tumor-bearing mice using ex vivo counting as the reference. RESULTS: The optimal sequence was a mixture of tracer plus IV contrast agent followed by 1 mL of IP contrast agent (20 mg iodine/mL) administered 10 min before PET/CT acquisition. Phantom studies showed that the use of a 20-mg iodine/mL concentration of Omnipaque(R) led to a 4.8% overestimation of radioactivity concentration, as compared to saline. This was confirmed by animal studies that demonstrated a 4.3% overestimation. Tumor detection was excellent and correlation between PET/CT quantitative data and ex vivo counting was good (r (2) = 0.91, slope = 0.7). CONCLUSIONS: A dual radiologic contrast protocol is useful in PET/CT scanning of mice bearing abdominal tumors. Contrast agents used in this manner lead to a small but acceptable overestimation of quantitative PET data.

PMID: 20632117 [PubMed - as supplied by publisher]

[Study on the anti-NTHi infection of Hap recombinant protein in vivo.]

Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2010 Jul;26(7):635-7

Authors: Li WY, Wang BN, Zuo FQ, Zeng W, Feng F, Kuang Y, Jiang ZH, Li MY

AIM: To observe the immune effect of Hap recombinant protein on murine model of bronchopneumonia infected with NTHi, and explore the mechanism about the anti-NTHi infection. METHODS: The C57BL/6 mice intranasally immunized with purified Hap recombinant protein and CT-B were challenged by NTHi encased in agar beads. The immunifaction of anti-infection was observed through encocyte counting of BALF, bacteria detection of lung and the pathologyical change of lung tissue. RESULTS: In the challenge with NTHi experiment, the inflammatory exudation of the infected murine and pathological change of lung tissue was relieved by combined immunization of Hap recombinant protein and CT-B, and quantity of NTHi in lung of the infected murine was reduced obviously. CONCLUSION: The Hap recombinant protein also had good ability of anti-NTHi infection in the murine model of NTHi bronchopneumonia. This study could offer the oretical and experimental basis for development of new vaccine against NTHi.

PMID: 20619085 [PubMed - in process]

Dantrolene mitigates caerulein-induced pancreatitis in vivo in mice.

Am J Physiol Gastrointest Liver Physiol. 2010 Jul;299(1):G196-204

Authors: Orabi AI, Shah AU, Ahmad MU, Choo-Wing R, Parness J, Jain D, Bhandari V, Husain SZ

Acute pancreatitis is a painful, inflammatory disorder for which adequate treatments are lacking. An early, critical step in its development is the aberrant signaling of Ca(2+) within the pancreatic acinar cell. This Ca(2+) release is modulated by the intracellular Ca(2+) channel the ryanodine receptor (RYR). We have previously shown that RYR inhibition reduces pathological intra-acinar protease activation, an early marker of pancreatitis. In this study, we examined whether pretreatment with the RYR inhibitor dantrolene attenuates the severity of caerulein-induced pancreatitis in mice. Immunofluorescent labeling for RYR from mouse pancreatic sections showed localization to the basolateral region of the acinar cell. After 1 h of caerulein hyperstimulation in vivo, dantrolene 1) reduced pancreatic trypsin activity by 59% (P < 0.05) and 2) mitigated early ultrastructural derangements within the acinar cell. Eight hours after pancreatitis induction, dantrolene reduced pancreatic trypsin activity and serum amylase by 61 and 32%, respectively (P < 0.05). At this later time point, overall histological severity of pancreatitis was reduced by 63% with dantrolene pretreatment (P < 0.05). TUNEL-positive cells were reduced by 58% (P < 0.05). These data suggest that the RYR plays an important role in mediating early acinar cell events during in vivo pancreatitis and contributes to disease severity. Blockade of Ca(2+) signals and particularly RYR-Ca(2+) may be useful as prophylactic treatment for this disease in high-risk settings for pancreatitis.

PMID: 20448143 [PubMed - indexed for MEDLINE]

Pharmacodynamics and pharmacokinetics of the gamma-secretase inhibitor PF-3084014.

J Pharmacol Exp Ther. 2010 Jul;334(1):269-77

Authors: Lanz TA, Wood KM, Richter KE, Nolan CE, Becker SL, Pozdnyakov N, Martin BA, Du P, Oborski CE, Wood DE, Brown TM, Finley JE, Sokolowski SA, Hicks CD, Coffman KJ, Geoghegan KF, Brodney MA, Liston D, Tate B

PF-3084014 [(S)-2-((S)-5,7-difluoro-1,2,3,4-tetrahydronaphthalen-3-ylamino)-N-(1-(2-methyl-1-(neopentylamino)propan-2-yl)-1H-imidazol-4-yl)pentanamide] is a novel gamma-secretase inhibitor that reduces amyloid-beta (Abeta) production with an in vitro IC(50) of 1.2 nM (whole-cell assay) to 6.2 nM (cell-free assay). This compound inhibits Notch-related T- and B-cell maturation in an in vitro thymocyte assay with an EC(50) of 2.1 microM. A single acute dose showed dose-dependent reduction in brain, cerebrospinal fluid (CSF), and plasma Abeta in Tg2576 mice as measured by enzyme-linked immunosorbent assay and immunoprecipitation (IP)/mass spectrometry (MS). Guinea pigs were dosed with PF-3084014 for 5 days via osmotic minipump at 0.03 to 3 mg/kg/day and exhibited dose-dependent reduction in brain, CSF, and plasma Abeta. To further characterize Abeta dynamics in brain, CSF, and plasma in relation to drug exposure and Notch-related toxicities, guinea pigs were dosed with 0.03 to 10 mg/kg PF-3084014, and tissues were collected at regular intervals from 0.75 to 30 h after dose. Brain, CSF, and plasma all exhibited dose-dependent reductions in Abeta, and the magnitude and duration of Abeta lowering exceeded those of the reductions in B-cell endpoints. Other gamma-secretase inhibitors have shown high potency at elevating Abeta in the conditioned media of whole cells and the plasma of multiple animal models and humans. Such potentiation was not observed with PF-3084014. IP/MS analysis, however, revealed dose-dependent increases in Abeta11-40 and Abeta1-43 at doses that potently inhibited Abeta1-40 and Abeta1-42. PF-3084014, like previously described gamma-secretase inhibitors, preferentially reduced Abeta1-40 relative to Abeta1-42. Potency at Abeta relative to Notch-related endpoints in vitro and in vivo suggests that a therapeutic index can be achieved with this compound.

PMID: 20363853 [PubMed - indexed for MEDLINE]

Potent intestinal Th17 priming through peripheral lipopolysaccharide-based immunization.

J Leukoc Biol. 2010 Jul;88(1):21-31

Authors: McAleer JP, Liu B, Li Z, Ngoi SM, Dai J, Oft M, Vella AT

Lipopolysaccharide (LPS) is a potent natural adjuvant, commonly used to amplify Th1 responses. Here, we report that systemic immunization using LPS generates large numbers of specific Th17 cells in murine small intestinal lamina propria. The priming of these Th17 cells required IL-23p19 production by bone marrow-derived cells. In contrast, IL-23 had no impact on Th1 differentiation or overall numbers of Ag-specific regulatory T cells. Experiments using T-cell adoptive transfers revealed a previously unappreciated mechanism for how Th17 responses are amplified in vivo: stimulation through LPS expanded precommitted Th17 cells rather than causing Th17 differentiation. Second, LPS drove Th17 cell expansion independently of IL-23, demonstrating that this cytokine is not necessary for expansion and possibly functions at an earlier stage in Th17 priming. Our data provide an impetus for using LPS-based peripheral vaccination to augment specific T-cell-mediated immunity in the gut mucosa.

PMID: 20130220 [PubMed - indexed for MEDLINE]

Important roles of PI3K{gamma} in osteoclastogenesis and bone homeostasis.

Proc Natl Acad Sci U S A. 2010 Jul 2;

Authors: Kang H, Chang W, Hurley M, Vignery A, Wu D

G protein-coupled receptor-regulated PI3Kgamma is abundantly expressed in myeloid cells and has been implicated as a promising drug target to treat various inflammatory diseases. However, its role in bone homeostasis has not been investigated, despite the fact that osteoclasts are derived from myeloid lineage. We therefore carried out thorough bone phenotypic characterization of a PI3Kgamma-deficient mouse line and found that PI3Kgamma-deficient mice had high bone mass. Our analyses further revealed that PI3Kgamma deficiency did not affect bone formation because no significant changes in osteoblast number and bone formation rate were observed. Instead, the lack of PI3Kgamma was associated with decreased bone resorption, as evidenced by decreased osteoclast number in vivo and impaired osteoclast formation in vitro. The decreased osteoclast formation was accompanied by down-regulated expression of osteoclastogenic genes, compromised chemokine receptor signaling, and an increase in apoptosis during osteoclast differentiation. Together, these data suggest that PI3Kgamma regulates bone homeostasis by modulating osteoclastogenesis. Our study also suggests that inhibition of PI3Kgamma, which is being considered as a potential therapeutic strategy for treating chronic inflammatory disorders, may result in an increase in bone mass.

PMID: 20616072 [PubMed - as supplied by publisher]

Targeted overexpression of Dkk1 in osteoblasts reduces bone mass but does not impair the anabolic response to intermittent PTH treatment in mice.

J Bone Miner Metab. 2010 Jul 3;

Authors: Yao GQ, Wu JJ, Troiano N, Insogna K

Parathyroid hormone (PTH) is a potent anabolic agent, but the cellular mechanisms by which it increases bone mass are not fully understood. Dickkopf 1 (Dkk1) is an endogenous inhibitor of Wnt signaling and suppresses bone formation in vivo. We sought to determine if Dkk1 and anabolic PTH treatment interact in regulating bone mass. PTH treatment of primary murine osteoblasts for 24 h reduced Dkk1 expression by 90% as quantified by real-time PCR, whereas PTH treatment in vivo reduced Dkk1 expression by 30% when given as a single daily subcutaneous dose. To directly determine whether Dkk1 modulates the anabolic response of PTH in vivo, we engineered transgenic (TG) mice expressing murine Dkk1 under the control of the 2.3-kb rat collagen alpha-1 promoter. TG mice had significantly reduced bone mass, which was accompanied by reduced histomorphometric parameters of bone formation (reduced OV/TV, ObS/OS, and NOb/TAR). Treatment of TG mice and wild-type (WT) littermates with 95 ng/g body weight of human (1-34) PTH daily for 34 days resulted in comparable increases in bone mass at all skeletal sites. Histomorphometric analyses indicated that PTH treatment increased the numbers of both osteoblasts and osteoclasts in WT mice but only increased the numbers of osteoblasts in TG mice. We conclude that overexpression of Dkk1 does not attenuate the anabolic response to PTH in vivo.

PMID: 20602130 [PubMed - as supplied by publisher]

Sensory detection and responses to toxic gases: mechanisms, health effects, and countermeasures.

Proc Am Thorac Soc. 2010 Jul;7(4):269-77

Authors: Bessac BF, Jordt SE

The inhalation of reactive gases and vapors can lead to severe damage of the airways and lung, compromising the function of the respiratory system. Exposures to oxidizing, electrophilic, acidic, or basic gases frequently occur in occupational and ambient environments. Corrosive gases and vapors such as chlorine, phosgene, and chloropicrin were used as warfare agents and in terrorist acts. Chemical airway exposures are detected by the olfactory, gustatory, and nociceptive sensory systems that initiate protective physiological and behavioral responses. This review focuses on the role of airway nociceptive sensory neurons in chemical sensing and discusses the recent discovery of neuronal receptors for reactive chemicals. Using physiological, imaging, and genetic approaches, Transient Receptor Potential (TRP) ion channels in sensory neurons were shown to respond to a wide range of noxious chemical stimuli, initiating pain, respiratory depression, cough, glandular secretions, and other protective responses. TRPA1, a TRP ion channel expressed in chemosensory C-fibers, is activated by almost all oxidizing and electrophilic chemicals, including chlorine, acrolein, tear gas agents, and methyl isocyanate, the highly noxious chemical released in the Bhopal disaster. Chemicals likely activate TRPA1 through covalent protein modification. Animal studies using TRPA1 antagonists or TRPA1-deficient mice confirmed the role of TRPA1 in chemically induced respiratory reflexes, pain, and inflammation in vivo. New research shows that sensory neurons are not merely passive sensors of chemical exposures. Sensory channels such as TRPA1 are essential for maintenance of airway inflammation in asthma and may contribute to the progression of airway injury following high-level chemical exposures.

PMID: 20601631 [PubMed - in process]

Loss of lsc/p115-protein leads to neuronal hypoplasia in the esophagus and an achalasia-like phenotype in mice.

Gastroenterology. 2010 Jun 20;

Authors: Zizer E, Beilke S, Bäuerle T, Schilling K, Möhnle U, Adler G, Fischer KD, Wagner M

BACKGROUND & AIMS:: lsc/p115 was originally described as hematopoietic Rho-GEF regulating leukocyte migration, adhesion and marginal zone B-cell homeostasis. Here we investigate the expression pattern of lsc/p115 in the gastrointestinal tract and the consequences of lsc/p115-deficiency in lsc/p115-knockout mice. METHODS:: The phenotype of lsc/p115-deficient mice was analyzed in vivo with small animal CT scans and esophageal manometry. The morphology and myenteric plexus were evaluated with immunohistochemistry, morphometry, western blot analyses and qRT-PCR. RESULTS:: lsc/p115 is expressed in the gastrointestinal tract sparing the segment of the small intestine. Immunohistochemical staining detects lsc/p115 in the muscle layer and the GFAP positive glia in the esophagus. Esophageal manometry uncovers a severe motor dysfunction in lsc/p115-deficient mice. This achalasia like phenotype is characterized by disturbed peristalsis, hypertension of the lower esophageal sphincter (LES) and impaired relaxation of the LES. lsc/p115-deficient mice develop a progressive dilatation of the esophagus and decrease of the muscle layer. The muscle cell differentiation is not altered in lsc/p115-deficient mice. However, the density of inhibitory and excitatory neurons and glia cells in the myenteric plexus and the muscle layer are reduced in morphometric analyses. This reduced number of glia cells is accompanied by reduced expression of the neurotrophic factor NGF. CONCLUSIONS:: lsc/p115-deficiency results in impaired neuronal innervation and in motor dysfunction recapitulating several aspects of esophageal achalasia. Reduced expression of NGF and reduced number of glia cells most likely contribute to this phenotype.

PMID: 20600037 [PubMed - as supplied by publisher]