A William, Associate Professor
Mayo Cliniccity: Rochester country: United States (us)
Abstract: translational research that will meaningfully reduce the burden of ovarian cancer. A critical part of this process is to increase the quality and depth of the translational investigator base in ovarian cancer. The Career Development Program is based on the conviction that translational research can effectively proceed from the bench/population to the clinic or from the clinic to the bench/population. The ultimate objectives of the Career Development Program of the Ovarian SPORE are to identify and mentor new and developing investigators in ovarian cancer who demonstrate the clear potential to become independent translational researchers as well as attracting established scientists who wish to refocus on ovarian cancer. This will be accomplished through a rigorous review process aimed at identifying the most talented and promising candidate followed by intense effective mentoring, integration into ongoing SPORE activities and close oversight of the individual´s progress. In addition to a primary mentor, awardees will have complementary mentors in clinical, basic or population sciences necessary to ensure development of a translational research career. This capitalizes on numerous strengths present within the Mayo environment. The Career Development Program will maintain close oversight over the mentoring activities and progress of the awardee. In turn, the Program will report to the Executive Committee of the Mayo Clinic Ovarian Cancer SPORE
Keywords: Achievement; Animal Model; anticancer research; Award; base; Basic Science; Biometry; career; career development; Clinic; Clinical Sciences; Development; Disabled Persons; Ensure; Environment; experience; Faculty; Fostering; Funding; Goals; Individual; Malignant neoplasm of ovary; Malignant Neoplasms; meetings; Mentors; Minority; Ovarian; patient registry; Population; Population Sciences; Process; programs; Qualifying; Reporting; Research; Research Infrastructure; Research Personnel; Resources; Scientist; Structure; Translational Research; Travel; Wages; Woman; Women`s Group
Budget start date: 1-SEP-2011
Budget end date: 31-AUG-2012
5P50CA136393-03_7932 (2011): $65176
Sponsored Links Excellgen http://Excellgen.com
Grants awarded to A William
AEROSOL MANAGEMENT PLATFORM WITH EXPOSURE CHAMBER
A William, Associate Professor
University Of Washingtoncity: Seattle country: United States (us)
Grant 1S10RR028041-01 from National Center For Research Resources
Abstract: We propose to purchase the AeroMP aerosol management platform and a whole animal exposure chamber from Biaera Technologies. The AeroMP controls aerosol exposure for up to 24 mice simultaneously via either the whole body exposure chamber or a nose-only InTox exposure system, which is already available in the Center for Lung Biology. The sophisticated computerized control system of the AeroMP monitors and records airflows and pressures during aerosol generation, allowing reproducible aerosol exposures across experiments. A parallel sampling port allows characterization of the aerosol being delivered to the exposure chamber. The AeroMP combined with both the whole body exposure chamber and the InTox nose-only exposure system will be used by a broad group of investigators within the University of Washington Center for Lung Biology as well as associated collaborators. The system will also be available to all researchers within the new University of Washington South Lake Union Research Campus. The whole body exposure system will be used for exposure of mice to a variety of BSL 1 and 2 microbial pathogens such as influenza, pneumonia virus of the mouse, Pseudomonas aeruginosa, and Staphylococcus aureus to facilitate ongoing investigations into host defense, lung injury, and immunity. The whole body exposure system will also be used to deliver particulate aerosols such as diesel exhaust to study mechanisms of cardiopulmonary disease in response to particulate pollution exposure. The InTox nose-only exposure system coupled with the AeroMP will be used to eaerosolize small volumes of valuable reagents such as pharmacological agents or adenoviral vectors containing shRNA for efficient delivery, allowing dissection of mechanistic pathways involved in lung disease and healing
Keywords: Adenovirus Vector; Aerosols; Animals; Biology; computerized; Cor pulmonale; Coupled; Diesel Exhaust; Dissection; Exposure to; Generations; Healed; healing; Host Defense; Immunity; Influenza; Investigation; Lung; Lung diseases; lung injury; microbial; Monitor; Murine pneumonia virus; Mus; Nose; Particulate; pathogen; Pathway interactions; Pollution; pressure; Pseudomonas aeruginosa; Reagent; Records; Research; Research Personnel; research study; response; Sampling; small hairpin RNA; Staphylococcus aureus; System; Technology; Universities; Washington
Project start date: 2010-07-03
Project end date: 2011-07-02
Budget start date: 3-JUL-2010
Budget end date: 2-JUL-2011
PFA/PA: PAR-09-028
1S10RR028041-01 (2010): $118665
FIELD STUDIES OF HUMAN IMMUNITY TO AMEBIASIS IN BANGLADESH
A William, Wade Hampton Frost Professor
University Of Virginia Charlottesvillecity: Charlottesville country: United States (us)
Grant 5R01AI043596-13 from National Institute Of Allergy And Infectious Diseases
Abstract: We propose a genetic study of the human-pathogen-environment relationship that underlies susceptibility to amebiasis. 8 years ago we started a prospective cohort study of Entamoeba histolytica infection in 300 two to five year old children living in the Mirpur urban slum of Dhaka, Bangladesh. We discovered that amebiasis is common and associated with malnutrition and geohelminth infection, that parasite genotypes differ in propensity to cause disease, and that mucosal IgA and systemic IFN-gamma are markers of protective immunity. Despite a substantial burden of disease, we noted that children differ markedly in their susceptibility to amebiasis. Our hypothesis is that susceptibility is determined by (a) host innate and acquired immune responses that vary between individuals in part due to human genetic polymorphisms; (b) environmental influences including malnutrition and concurrent geohelminth infection; and (c) virulence differences of E. histolytica genotypes. Three specific aims are proposed to delineate the interaction of host, parasite and environment leading to disease. Specific Aim (1) will measure the incidence of amebiasis and correlate it with human and parasite genetic polymorphisms, immune responses, and environmental factors such as geohelminth infection and malnutrition. As part of Aim 1 a 2nd cohort of 500 live births will be enrolled to separate innate from acquired immune responses, examine the burden of amebiasis in the first 2 years of life, and independently test genetic polymorphisms found to influence susceptibility in the older child cohort. Specific Aim (2) will test the hypothesis that protective immunity is mediated both by innate immune responses initiated via TLR stimulation as well as by mucosal IgA against the Gal/GalNAc lectin and systemic IFN-gamma. We will measure innate and acquired immune responses to E. histolytica in the 2 cohorts and correlate them with incidence of infection, host genetic polymorphisms and environmental factors. Specific Aim (3) will test for the association of common genetic polymorphisms in host innate and acquired immune genes with incidence of amebiasis. In the older child cohort, 2800 SNPs in 89 candidate genes will be typed and correlated with immune responses and susceptibility. Using a 2-stage study design (original cohort followed by birth cohort) candidate genes associated with susceptibility will be validated in the birth cohort. Successful completion of these studies will identify host, environmental and parasitic determinants of infection, while at the same time establishing the burden of infection by completion of the only prospective community-based study of amebiasis from infancy to age 18. Amebiasis is a common cause of diarrhea and is associated with malnutrition in grade- school aged children in an urban slum of Dhaka, Bangladesh. We wish to determine the contribution of amebiasis to illness in the first 2 years of life when most deaths due to diarrhea occur, and understand the immunologic and genetic factors that protect children from amebiasis. Successful completion of these studies will provide an estimate of the impact of amebiasis on the health of the study children, and could lead to new approaches to prevent and treat this disease
Keywords: 0-11 years old; 5 year old; acquired immunity; Age; Agonist; Amebiasis; anti-IgA; Antibodies; antibody-based immunity; Antigens; Ascaris lumbricoides; ATGN; Bangladesh; base; Birth; Blood, Cord; Breast Feeding; Breast Milk; Breastfeeding; burden of disease; burden of illness; Candidate Disease Gene; Candidate Gene; Cells; Cessation of life; Child; Child Youth; children; Children (0-21); cohort; Cohort Studies; Communities; Concurrent Studies; CSIF; CSIF-10; cytokine; Cytokine formation-inhibiting factor (mouse clone F115 protein moiety reduced); Cytokine Synthesis Inhibitory Factor; Cytotoxic cell; Data; Death; Diarrhea; dietary deficiency; DIF; Disease; disease burden; disease/disorder; Disorder; Dysentery; E. histolytica; Edodekin Alfa; egg; Endamoeba histolytica; enroll; Enrollment; Entamoeba histolytica; Environment; Environmental Factor; environmental risk; Environmental Risk Factor; Experimental Designs; Family member; Feces; fetal cord blood; field study; five year old; Gal-GalNAc; Gamma Globulin, 7S; Gamma interferon; Gastrointestinal Tract, Feces; Genes; Genetic; Genetic Polymorphism; Genotype; Health; heavy metal lead; heavy metal Pb; host response; hToll; Human; Human Genetics; Human Milk; Human Mother`s Milk; Human, Child; Human, General; Humoral Immunities; IFN; IFN-g; IFN-Gamma; IFNG; IgA; IgG; IL-10; IL-12; IL10; IL10A; IL12; Immune; Immune response; Immunities, Humoral; Immunity; Immunity, Innate; Immunity, Native; Immunity, Natural; Immunity, Non-Specific; immunogen; Immunoglobulin A; Immunoglobulin G; Immunologic, Immunochemical; Immunologics; immunoresponse; In Vitro; Incidence; Individual; infancy; infantile; Infection; Infection by T. trichiura; Infection by Trichuris trichiura; Infectious Diarrheal Disease; Interferon Gamma; Interferon gamma (human lymphocyte protein moiety reduced); Interferon Type II; Interferon, Immune; Interferon-gamma; Interferons; Interleukin 10 Precursor; Interleukin-10; Interleukin-12; K lymphocyte; Lead; Lectin; lFN-Gamma; Life; Live Birth; Low Birth Weight Infant; low birth weight infant human; low birthweight; Malnutrition; Mammary Gland Milk; Man (Taxonomy); Man, Modern; maternal serum; Measures; Mediating; Messenger RNA; Milk, Human; Mononuclear; Mothers; mRNA; Natural Immunity; Natural Killer Cell Stimulatory Factor; Natural Killer Cells; new approaches; NK Cells; NKSF; novel approaches; novel strategies; novel strategy; Nutritional Deficiency; Outcome; Parasites; Parturition; pathogen; Pb element; PBMC; Peripheral Blood Mononuclear Cell; polymorphism; Polymorphism (Genetics); Polymorphism, Genetic; Predisposition; prevent; preventing; Principal Investigator; Production; programs; Programs (PT); Programs [Publication Type]; prospective; Research Design; response; RNA, Messenger; Role; Sample Size; Sampling; school age; School-Age Population; Severities; Slum; social role; Source; Staging; Staining method; Stainings; Stains; stool; study design; Study Type; Surrogate Markers; Susceptibility; T-Cells; T-Lymphocyte; T. trichiura; Testing; thymus derived lymphocyte; Thymus-Dependent Lymphocytes; TIL4; Time; TLR2; TLR2 gene; TLR4; TLR4 gene; TNF; TNF A; TNF gene; TNFSF2; TOLL; Toll/Interleukin 1 Receptor-Like 4 Gene; Trichocephaliasis; Trichocephalus trichiura; Trichuriases; Trichuriasis; Trichuris trichiura; Trichuris trichiura infection; Tumor Necrosis Factor Gene; Umbilical Cord Blood; Undernutrition; Virulence; VLBW (human); years of life lost to disability; years of life lost to disease; youngster
Project start date: 1998-09-15
Project end date: 2012-12-31
Budget start date: 1-JAN-2011
Budget end date: 31-DEC-2011
PFA/PA: PA-07-070
5R01AI043596-13 (2011): $225930
CELL BIOLOGY OF THE NEURONAL SODIUM CHANNEL
A William, Professor And Chair
University Of Washingtoncity: Seattle country: United States (us)
Grant 5R01NS025704-23 from National Institute Of Neurological Disorders And Stroke
Abstract: Voltage-gated sodium (Na) channels initiate and conduct action potentials in neurons and are a molecular target for anti-epileptic drugs. Our work on this project has shown that deletions of both a and ¿ subunits of Na channels cause seizure susceptibility and epilepsy in mice. These results show that alterations in the expression and cell biology of Na channels can lead to epilepsy without gain-of-function mutations. Human Severe Myoclonic Epilepsy in Infancy (hSMEI), an intractable childhood epilepsy accompanied by ataxia and other neurological deficits, is caused by heterozygous loss-of-function mutations in Nav1.1 channels. Because loss of Na channels would cause hypoexcitability, it is paradoxical that hSMEI is caused by haploinsufficiency of Nav1.1 channels. We have created a mouse model of SMEI by targeted deletion of Nav1.1 channels in mice, mSMEI. Like hSMEI, heterozygotes with mSMEI have intractable seizures after weaning, ataxia, and often premature death, which are strikingly dependent on genetic background as in humans. Our results reveal a potential basis for hyperexcitability--selective loss of Na currents in GABAergic inhibitory neurons compared to excitatory pyramidal neurons in the hippocampus. Moreover, a dramatic loss of Na current in GABAergic Purkinje neurons in the cerebellum may cause ataxia. Up-regulation of Nav1.3 channels is unable to compensate for loss of Nav1.1 channels. We will probe the cellular and molecular changes in function, localization, regulation, and cell biology of Na channels in mSMEI. We hypothesize that the epilepsy, ataxia, and other deficits in mSMEI result from loss of Na current in GABAergic inhibitory neurons, that genetic background effects on epileptogenesis in MSMEI are caused by differences in compensatory regulation of expression, localization, and function of other Na channels, and that novel combinations of anti-epileptic drugs that enhance GABAergic neurotransmission will be effective in treating mSMEI. Our experiments will be guided by four Specific Aims (1) to determine whether loss of Na current in specific classes of GABAergic neurons is responsible for the pleiotropic effects of mSMEI; (2) to define the molecular basis for genetic background effects on severity of mSMEI due to expression, localization, and function of Na channels; (3) to describe and analyze the molecular and cellular changes in Na channels that lead from loss of excitability of GABAergic interneurons to hyperexcitability and epilepsy in mSMEI; and (4) to explore novel combination therapies for mSMEI that may be relevant for treatment of the human disease. Our results will provide crucial new information on the cell biology and regulation of Na channels in a relevant disease model, define the molecular and cellular mechanisms underlying mSMEI, and yield insights into novel pharmacotherapies that may be effective in this intractable childhood disease. Severe Myoclonic Epilepsy in Infancy, an inherited seizure disorder cause by a gene defect in a sodium channel, is one of the most severe childhood epilepsy disorders. This project will use a mouse genetic model of this disease to determine how this gene defect causes epilepsy and to test new drug combinations for control of this intractable epilepsy syndrome
Keywords: Action Potentials; Affect; Antiepileptic Agents; Ataxia; base; Cell Adhesion Molecules; cell body (neuron); Cell Death; Cell Nucleus; Cell surface; cell type; Cellular biology; Cerebellum; Cerebral cortex; Cessation of life; Child; Childhood; Clinical; Clinical Trials; Combined Modality Therapy; Complex; Data; Defect; Depressed mood; Development; Disease; Disease model; Drug Combinations; Electroencephalography; Epilepsy; Epileptogenesis; excitatory neuron; Extracellular Matrix; gain of function mutation; gamma-Aminobutyric Acid; Gene Deletion; Generalized Epilepsy; Genes; Genetic; Genetic Models; Genetic screening method; Gliosis; Health; Heterozygote; hippocampal pyramidal neuron; Hippocampus (Brain); Human; human disease; Immunohistochemistry; Individual; infancy; Inherited; inhibitory neuron; insight; Interneurons; Intractable Epilepsy; Ion Channel; Lead; loss of function mutation; Measures; Methods; Minor; Molecular; Molecular Genetics; Molecular Probes; Molecular Target; mouse model; Mus; Mutant Strains Mice; Mutation; Myoclonic Epilepsies; Neurologic; Neurons; neurotransmission; novel; Patients; Penetrance; Pharmaceutical Preparations; Pharmacotherapy; postnatal; Predisposition; premature; Process; Property; Pyramidal Cells; Regulation; research study; Resistance; Reverse Transcriptase Polymerase Chain Reaction; Role; SCN1A protein; Seizures; Severities; Signal Transduction; Signaling Protein; Sleep; Sodium; Sodium Channel; Syndrome; Testing; Text; Thalamic structure; Therapeutic; transmission process; Up-Regulation (Physiology); Video Recording; voltage; Weaning; Work
Relevance: Severe Myoclonic Epilepsy in Infancy, an inherited seizure disorder cause by a gene defect in a sodium channel, is one of the most severe childhood epilepsy disorders. This project will use a mouse genetic model of this disease to determine how this gene defect causes epilepsy and to test new drug combinations for control of this intractable epilepsy syndrome
Project start date: 1988-02-01
Project end date: 2013-05-31
Budget start date: 1-JUN-2011
Budget end date: 31-MAY-2012
PFA/PA: PA-07-070
5R01NS025704-23 (2011): $334425
ANCHORED ANTIOXIDANTS TO PROVIDE INCREASED PROTECTIVE EFFECT
A William
Tosk, Inc.city: Santa Cruz country: United States (us)
Grant 1R43HL103278-01 from National Heart, Lung, And Blood Institute
Abstract: Very compelling data are available to suggest oxidative stress is an underlying component of many significant human diseases such as diabetes, heart disease, cerebrovascular disease (eg, stroke), peripheral vascular disease, neurodegeneration (Alzheimer´s, Parkinson´s, ALS and Huntington´s disease) as well as aging. However, results to date with antioxidant treatments to mitigate human diseases with oxygen stress as an underlying component of the pathology have been disappointing. Tosk, Inc. has successfully designed an agent to irreversibly bind a traditional antioxidant to an enzyme located in the region where reactive oxygen species are formed. This was done to make the traditional antioxidant more effective. Now Tosk requests Phase I SBIR funding to further explore this approach to produce even more active antioxidants. A total of 15 novel antioxidants will be synthesized and tested in the same animal model used originally to discover the activity of the new antioxidant designed at Tosk. Three of the synthesized molecules will also be tested in one additional animal model of human disease associated with excess oxidative stress as a significant part of the condition´s underlying pathology. Results to date with antioxidant treatments to mitigate human diseases having oxygen stress as an underlying component of the pathology have been disappointing. Tosk, Inc. has successfully designed an agent to irreversibly bind a traditional antioxidant to an enzyme located in the region where reactive oxygen species are formed. This was done to make the traditional antioxidant more effective. Now Tosk requests Phase I SBIR funding to further explore this approach to produce more active antioxidants as well as to evaluate the approach in one additional animal model of human disease associated with excess oxidative stress as a significant part of the conditions underlying the pathology
Keywords: Affect; Affinity; Aging; Alzheimer`s Disease; Animal Model; Anthracycline Antibiotics; Antioxidants; Applications Grants; Area; base; Binding (Molecular Function); Biological; Biological Assay; Cancer cell line; Cancer Patient; Cardiac; Cardiolipins; Cardiomyopathies; Cerebrovascular Disorders; Clinical; cytotoxicity; Data; design; Development; Diabetes Mellitus; Disease; Dose; Doxorubicin; drug candidate; Elements; Enzymes; Etiology; Evaluation; Funding; Goals; Half-Life; Heart; Heart Diseases; Hematologic Neoplasms; Hour; Human; human disease; Huntington Disease; improved; In Vitro; in vivo; Lead; Life; Malignant Neoplasms; Measurable; Measures; Mediator of activation protein; Membrane; Middle Cerebral Artery Occlusion; Mitochondria; mitochondrial membrane; Modeling; Mus; Nerve Degeneration; No-Observed-Adverse-Effect Level; novel; Organelles; Oxidative Stress; Oxygen; Parkinson Disease; Pathologic; Pathology; Patients; Peripheral Vascular Diseases; Phase; Plasma; pre-clinical; Production; protective effect; prototype; public health relevance; Rattus; Reactive Oxygen Species; Recording of previous events; Regimen; Research; safety study; Small Business Innovation Research Grant; Solid; Stress; stroke; Test Result; Testing; Therapeutic; Thioredoxin-2; Troponin I
Relevance: Narrative. Results to date with antioxidant treatments to mitigate human diseases having oxygen stress as an underlying component of the pathology have been disappointing. Tosk, Inc. has successfully designed an agent to irreversibly bind a traditional antioxidant to an enzyme located in the region where reactive oxygen species are formed. This was done to make the traditional antioxidant more effective. Now Tosk requests Phase I SBIR funding to further explore this approach to produce more active antioxidants as well as to evaluate the approach in one additional animal model of human disease associated with excess oxidative stress as a significant part of the conditions underlying the pathology
Project start date: 2010-07-15
Project end date: 2011-12-31
Budget start date: 15-JUL-2010
Budget end date: 31-DEC-2011
PFA/PA: RFA-OD-09-009
1R43HL103278-01 (2010): $199500
INTEGRIN ASSOCIATED PROTEIN/CD47 IS A THROMBOSPONDIN RECEPTOR
A William, Professor Of Biochemistry And Molec
Washington Universitycity: Saint Louis country: United States (us)
Grant 5R01HL054390-12 from National Heart, Lung, And Blood Institute
Abstract: Thrombospondin-1 (TSP1) and its receptors have long been thought to have important roles in regulating vascular cells, both circulating and mural. During the preceding grant period we have discovered that TSP1 and CD47 (integrin-associated protein) regulate the dynamic range of nitric oxide (NO) signaling in vascular cells. Thus TSP1-CD47 interactions are important not only in angiogenic regulation but in rapid regulation of tissue perfusion and many other roles where NO maintains the health of the cardiovascular system. We have identified the binding surface on the C-terminal domain of TSP1 that interacts with CD47. We have also discovered a new extracellular mechanism for integrin activation via CD47. This proposal focuses on the molecular interactions among TSP1, CD47 and 23 integrins. The molecular details of these interactions will be deduced by mutagenesis of all three interacting partners. Mutant proteins will be tested in vitro in binding assays and assays of CD47 and integrin function. The extracellular clasp mechanism for integrin activation that we identified within the 1v23 structure will be tested in cultured cells and in 23-null mice repopulated with bone marrow expressing activated 23 integrin constructs. A similar approach will test the ability of CD47 to activate 23 integrins via interaction with the clasp in cultured cells and in mice expressing mutant CD47 transgenes. Conditions for formation of TSP1-CD47 and CD47-integrin complexes will be evaluated using physical methods and several EM approaches. These methods will then be used to evaluate the interactions of mutant proteins in order to identify structural elements of each that are important for their molecular interactions. Based on these results, crystallization and X-ray diffraction studies will be initiated of CD47 in complexes with the TSP1 C-terminal domain and with 1v23 integrin. The sum of these studies will provide for the first time a physical model of TSP1-CD47-integrin interactions. Given our new paradigm for TSP1-CD47 regulation of NO signaling in the vascular system, this information will be vital in designing new strategies to modulate endogenous NO signaling, a new approach to ameliorating cardiovascular disease
Keywords: Address; Affect; Affinity; angiogenesis; Avidity; base; Binding (Molecular Function); Binding Sites; Biological Assay; Blood Platelets; Blood Vessels; Bone Marrow; Bone Resorption; C-terminal; Cardiovascular Diseases; Cardiovascular system; CD36 Antigens; CD36 gene; CD47 Antigen; CD47 gene; cell growth regulation; Cells; Cholesterol; Complex; Crystallization; Cultured Cells; Cyclic GMP; design; Electron Microscopy; Elements; extracellular; Fostering; Grant; Health; Homologous Gene; Hyperplasia; in vitro testing; in vivo; inhibitor/antagonist; Injury; Integrin Binding; Integrins; Ischemia; Knockout Mice; Lateral; Leukocytes; Ligands; Membrane; Methods; Modeling; Molecular; Monitor; Mus; Mutagenesis; mutant; Mutation; Necrosis; Nitric Oxide; novel; novel strategies; Osteoclasts; Oxygen; Peptides; Perfusion; Phenotype; physical model; Physiological; Platelet Activation; Play; Process; Property; Proteins; receptor; Regulation; Resistance; Role; Signal Pathway; Signal Transduction; Site; Smooth Muscle Myocytes; Structure; Sum; Surface; Testing; Thrombospondin 1; Time; Tissues; Transgenes; Vascular constriction (function); Vascular System; X ray diffraction analysis; X-Ray Crystallography; X-Ray Diffraction
Project start date: 2002-07-01
Project end date: 2013-06-30
Budget start date: 1-JUL-2011
Budget end date: 30-JUN-2012
PFA/PA: PA-07-070
5R01HL054390-12 (2011): $380000
POLYPEPTIDE MODIFICATION FOR ENHANCED BRAIN DELIVERY
A William, Parke-davis Prof And Director
University Of Nebraska Medical Centercity: Omaha country: United States (us)
Grant 5R01NS051334-05 from National Institute Of Neurological Disorders And Stroke
Abstract: This proposal is a competing continuation of the NIH grant R01 NS051335. The long term goal is to produce polypeptides that could cross the blood-brain barrier (BBB) and so be developed as central nervous system (CNS) therapeutics. The goal is achieved by modification of polypeptides with Pluronic copolymers (poly(ethylene oxide)-b-poly(propylene oxide)-b-poly(ethylene oxide)). The studies have focused on leptin, a candidate for the treatment of epidemic obesity. Leptin failure is mostly due to peripheral resistance - a loss of its ability to cross the BBB necessary to reach its receptors in the arcuate nucleus of the hypothalamus. The previous work produced a conjugate of leptin with Pluronic P85 that is centrally active, has a long half-life in blood, is enzymatically resistant, crosses the BBB by a non-saturable mechanism independently of the leptin transporter, and can exert effects on feeding and body weight after peripheral administration. This proposal will 1). optimize modifications to produce superior leptin analogs with maximal BBB permeability, improved half-life in blood and high enzymatic resistance in brain and blood; 2). identify mechanisms underlying enhanced permeability of leptin analogs in BBB; 3). determine the permeability of modified leptin analogs across the BBB to select the best analogs for activity testing; and 4) determine the biological activity after intravenous injection of the best leptin analogs 3 in leptin sensitive (ob/ob) and leptin resistant (dietary obese) mouse models of obesity. Analogs completing this development will be a) active in brain, b) have favorable pharmacokinetics, c) be able to cross the BBB in obese mice, and d) reverse obesity after iv injection. Such analogs are candidates for clinical trials in human obesity. The long term goal is to produce polypeptides that could cross the blood-brain barrier (BBB) and so be developed as central nervous system (CNS) therapeutics. The goal is achieved by modification of polypeptides with Pluronic copolymers (poly(ethylene oxide)- b-poly(propylene oxide)-b-poly(ethylene oxide))
Keywords: analog; Biological; Biological Availability; Blood; Blood - brain barrier anatomy; Body Weight; Body Weight decreased; Brain; Caveolae; Cell model; Cells; Clinical Trials; copolymer; Development; Diet; Drug Kinetics; Eating; Endocytosis; Endothelial Cells; Epidemic; Ethylene Oxide; Failure (biologic function); feeding; Goals; Grant; Half-Life; Human; Hypothalamic structure; improved; in vivo; inhibitor/antagonist; Injection of therapeutic agent; intravenous injection; Knockout Mice; Length; Leptin; Link; Measures; Mediating; Modeling; Modification; mouse model; Mus; Neuraxis; Obese Mice; Obesity; Pathway interactions; Perfusion; Peripheral; Peripheral Resistance; Permeability; Pluronics; poly(propylene oxide); polypeptide; public health relevance; receptor; receptor binding; Resistance; Structure of nucleus infundibularis hypothalami; Testing; Therapeutic; United States National Institutes of Health; Work
Relevance: The long term goal is to produce polypeptides that could cross the blood-brain barrier (BBB) and so be developed as central nervous system (CNS) therapeutics. The goal is achieved by modification of polypeptides with Pluronic copolymers (poly(ethylene oxide)- b-poly(propylene oxide)-b-poly(ethylene oxide))
Project start date: 2005-04-01
Project end date: 2015-03-31
Budget start date: 1-APR-2011
Budget end date: 31-MAR-2012
PFA/PA: PA-07-070
5R01NS051334-05 (2011): $332745
THIOREDOXIN INTERACTING PROTEIN´S (TXNIP) ROLE IN GLUCOSE METABOISM IN FAT
A William
Brigham And Women´s Hospitalcity: Boston country: United States (us)
Grant 5K08HL088977-04 from National Heart, Lung, And Blood Institute
Abstract: This proposal details a comprehensive 5-year training program for my career development in academic cardiovascular medicine. I have completed clinical training in cardiovascular medicine at Brigham and Women´s Hospital and plan to embark on a mentored research program to provide additional scientific training necessary for an independent career in biomedical resesarch. I will gain in-depth experience in cellular biology, genetics, molecular biochemistry, metabolism, and in vivo physiology as applied to the redox regulation of glucose metabolism. Dr. Richard T. Lee will mentor my scientific and career development. Dr. Lee is an internationally recognized leader in the fields of redox regulation, mechano-transduction, and cardiovascular diseases, and has a proven track record for successful career mentorship in academic medicine and the basic sciences. Dr. Evan Rosen of Beth Isreal Deaconess Hospital, an expert in metabolic metabolism and isulin-action, will serve as a co-mentor. In addition, an advisory committee of established medical scientists (Drs. Peter Libby, Thomas Michel) will provide scientific and career advice. The central hypothesis of this proposal is that the ubiquitously expressed redox regulatory protein Txnip (thioredoxin interacting protein) is a key mediator of glucose homeostasis. Txnip expression is highly glucose and insulin responsive. I show that forced Txnip expression significantly attenuates glucose uptake, and targeted Txnip gene deletion in mice promotes glucose uptake and blunts liver glucose production. My aims are 1) To explore the mechanisms by which Txnip regulates glucose uptake in a peripheral tissue (fat), using gene deletion and forced overexpression techniques in adipocytes; 2) To test the hypothesis that peripheral glucose uptake is regulated by Txnip expression in fat using mice with fat-selective targeted Txnip gene deletion; and 3) To test the hypothesis that impairment of glucose uptake by oxidative stress is mediated by Txnip using well-defined models of oxidative stress as applied to Txnip gene-deleted mice. Relevance Diabetes mellitus is an expanding worldwide epidemic that already affects greater than 1 in 15 adults. 4 out of 5 diabetic patients will die prematurely from a cardiovascular cause. This research aims to understand the fundamental mechanisms of dysregulated sugar metabolism with the ultimate goal of discovering new therapies to prevent or treat this devastating disease and its cardiovascular consequences. (End of )
Keywords: ing; Adipocytes; Adult; Advisory Committees; Affect; Antioxidants; Attenuated; basal insulin; base; Basic Science; Binding (Molecular Function); Biochemistry; Biology; blood glucose regulation; Cardiovascular Diseases; Cardiovascular system; career; career development; Cells; Cellular biology; Clinical; Cysteine; Data; Defect; Development; Diabetes Mellitus; diabetic patient; Disease; Disulfides; Enzymes; Epidemic; Event; experience; Fatty acid glycerol esters; Ferredoxin; Gene Deletion; Gene Proteins; genetic regulatory protein; Glucose; glucose metabolism; glucose production; glucose transport; glucose uptake; Glutathione; Goals; Hepatic; Hospital Planning; Hospitals; Hypertriglyceridemia; Hypoglycemia; Impairment; in vivo; inhibitor/antagonist; Insulin; Insulin Resistance; insulin signaling; Knockout Mice; Light; Liver; Mammalian Cell; Mediating; Mediator of activation protein; Medical; Medicine; Mentors; Mentorship; Metabolic; Metabolism; Modeling; Molecular Genetics; Mouse Strains; Mus; Mutation; Operating System; overexpression; Oxidation-Reduction; Oxidative Stress; Peripheral; Physiology; Plants; Play; Postabsorptive Hypoglycemia; prevent; Process; programs; protein expression; Protein Overexpression; protein protein interaction; Proteins; Regulation; Reporting; Research; Research Personnel; response; Role; Scientist; Signal Transduction; Signaling Molecule; sugar; Sulfhydryl Compounds; Techniques; Testing; Thioredoxin; Tissues; tool; Training; Training Programs; transcription factor; Woman
Project start date: 2008-03-01
Project end date: 2013-01-31
Budget start date: 1-FEB-2011
Budget end date: 31-JAN-2012
PFA/PA: PA-06-512
5K08HL088977-04 (2011): $136350
ROLES OF NUCLEUS ACCUMBENS CREB AND KAPPA FUNCTION IN DEPRESSION
A William, Professor Of Psychiatry
Mc Lean Hospital (belmont, Ma)city: Belmont country: United States (us)
Grant 2R01MH063266-11 from National Institute Of Mental Health
Abstract: This application is for renewal of a grant that we have had since 2001. Its purpose is to understand how interactions of CREB and kappa-opioid receptor (KOR) systems in the nucleus accumbens shell (NAs) affect behavior in the context of mood and anxiety disorders. Our 9 years of support has led to discoveries that have had a significant impact on the field (i) stress activates CREB in the NAs; (ii) elevated CREB function in the NAs triggers depressive-like behaviors; (iii) depressive-like behaviors are induced by CREB-mediated increases in dynorphin actions at KORs; (iv) KOR antagonists have antidepressant-like effects; and (v) KOR antagonists have anxiolytic-like effects. Our work has provided new insights on the role of the NAs in encoding rewarding and aversive states, and led directly to drug development efforts in academia and industry. Our over-arching hypothesis is that stress-induced CREB activation in the NAs increases dynorphin expression, which enhances feedback inhibition of the mesocorticolimbic system. This process triggers behaviors characteristic of depressive and, as new evidence suggests, anxiety-related disorders. Our continuing studies are designed to determine how modifications to this CREB-KOR cascade can cause and protect against the persistent effects of stress, using models that quantify key signs of depression and co-morbid disorders. In Aim 1, we will test the hypothesis that elevated CREB function in the NAs produces behavioral signs common to depressive and frequently co-morbid anxiety disorders such as post-traumatic stress disorder (PTSD). We will use an attention test to examine if elevated CREB in the NAs produces deficits in concentration and decision-making that are hallmark signs of major depression and PTSD. We will also use an inhibition-of-fear test to examine if elevated CREB in the NAs produces deficits in sensitivity to safety signals that characterize PTSD but not depression. In Aim 2, we will test the hypothesis that disruption of KOR function in the mesocorticolimbic system prevents the development of depressive and anxiety-related behaviors. We have made a line of mice in which the KOR gene is floxed, and found that ablation of KORs in all brain areas expressing dopamine uptake transporter (DAT) attenuates stress-related behavioral adaptations. We will use viral vectors to determine if further restriction of KOR ablation to the mesocorticolimbic system during adulthood produces similar resistance to stress. We will use tests (fear conditioning; intracranial self- stimulation) that quantify hallmark signs of depression and anxiety while providing insight on any non-specific effects of our manipulations. In Aim 3, we will test the hypothesis that disruption of KOR function prevents stress effects via actions that involve reduced sensitivity to corticotropin releasing factor (CRF), and that the long-lasting anti-stress effects of KOR antagonists require activation of c-Jun N-terminal kinase (JNK). We will also test the hypothesis that KOR disruption blocks stress-induced alterations in microRNA (miR) expression in the mesocorticolimbic system. This work may provide a basis for improved diagnostics and therapeutics. This research examines the mechanisms by which stress-induced alterations in the function of the transcription factor CREB and kappa-opioid receptor (KOR) systems trigger depressive and anxiety-related behavior. Our proposed studies are designed to determine how modifications to this CREB-KOR cascade can cause and protect against the persistent effects of stress, using animal models that quantify hallmark signs of depression and anxiety disorders in humans. This work may provide a basis for improved diagnosis, treatment, and prevention of mental illness
Keywords: Ablation; Academia; Adult; Affect; Animal Model; Anti-Anxiety Agents; Antidepressive Agents; Anxiety; Anxiety Disorders; Area; Attention; Attenuated; base; Behavior; Behavioral; Brain; cell body (neuron); Characteristics; conditioned fear; Corticotropin-Releasing Hormone; CREB1 gene; Cues; Decision Making; Depressive disorder; depressive symptoms; Development; Diagnosis; Diagnostic; Disease; Dominant-Negative Mutation; Dopamine; drug development; Dynorphins; Extinction (Psychology); Feedback; Fright; Future; Grant; Human; improved; Industry; insight; Major Depressive Disorder; Mediating; Mental Depression; Mental disorders; MicroRNAs; Modeling; Modification; Molecular; Mood Disorders; Mus; N-terminal; Neurons; Nucleus Accumbens; Opioid; Output; Phosphotransferases; Play; Post-Traumatic Stress Disorders; prevent; Prevention; Procedures; Process; Reaction Time; receptor function; Receptor Gene; Receptors, Opioid, kappa; recombinase; Research; Research Design; research study; Resistance; Rewards; Rodent; Role; Safety; Self Stimulation; Signal Transduction; social; Stress; stress-activated protein kinase 1; System; Testing; Therapeutic; transcription factor; uptake; vector control; Ventral Tegmental Area; Viral Vector; Work
Relevance: This research examines the mechanisms by which stress-induced alterations in the function of the transcription factor CREB and kappa-opioid receptor (KOR) systems trigger depressive and anxiety-related behavior. Our proposed studies are designed to determine how modifications to this CREB-KOR cascade can cause and protect against the persistent effects of stress, using animal models that quantify hallmark signs of depression and anxiety disorders in humans. This work may provide a basis for improved diagnosis, treatment, and prevention of mental illness
Project start date: 2001-06-17
Project end date: 2016-03-31
Budget start date: 17-JUN-2011
Budget end date: 31-MAR-2012
PFA/PA: PA-10-067
2R01MH063266-11 (2011): $359638
OXIDATIVE DYSFUNCTION OF LRP AT THE BLOOD-BRAIN BARRIER IN ALZHEIMER´S DISEASE
A William
Seattle Inst For Biomedical/clinical Rescity: Seattle country: United States (us)
Grant 5R01AG029839-04 from National Institute On Aging
Abstract: The neurovascular hypothesis states that an impaired brain-to-blood efflux of amyloid ¿ protein (ABP) at the vascular blood-brain barrier (BBB) is an important mechanism underlying ABP accumulation and cognitive impairments in patients with Alzheimer´s disease (AD). Low-density lipoprotein receptor-related protein-1 (LRP) has been identified as the major efflux pump at the BBB for ABP. Zlokovic and co-workers have shown that LRP is deficient in the BBB of patients with AD and of Hsiao mice that overexpress amyloid precursor peptide (APP). We have shown that ABP efflux is impaired at the BBB in animals which overexpress ABP and that knockdown of APP expression with antisense restores BBB efflux of ABP. This suggests that ABP poisons its own transporter, LRP. Our goal is to determine whether the mechanism of impaired efflux of ABP is caused by ABP-induced oxidative damage to LRP. ABP, especially in its oligomeric form, induces oxidative stress and by this mechanism impairs the function of transporters other than LRP in non-BBB tissues. LRP in non-BBB tissues is already known to be readily oxidized and its ability to transport its other ligands in those tissues is impaired in its oxidative state. Our hypothesis is that ABP impairs its own efflux at the BBB by oxidizing LRP. We will test this hypothesis in 3 Specific Aims Specific Aim 1 To determine in vivo whether mice that do overexpress APP have an oxidized LRP and whether ABP efflux can be restored to normal rates by treatments which reduce ABP or by antioxidants. Specific Aim 2 To determine the role of ABP and oxidation in impairing BBB efflux of ABP in vitro in a BBB monolayer model that uses brain endothelial cells derived from mice that do not overexpress APP. Specific Aim 3 To determine the status of oxidative modification of LRP in human brain tissue obtained at short post mortem intervals (PMI; specifically, < 4h after death) from patients with AD and mild cognitive impairment (MCI) relative to that from control brain tissue and correlate this information to the level and oligomeric status of ABP1-42 in those same brains and to compare this to a similar analysis for the mice that overepress APP
Keywords: 1, 2-Dithiolane-3-pentanoic acid; 3-mononitrotyrosine; 3-nitrotyrosine; 4 hydroxynal; 4-HNE cpd; 4-hydroxy-2, 3-nal; 4-hydroxy-2-nal; 4-hydroxyn-2-al; ABCB1; Acetylcysteine; Acetylin; Address; Adenosine Triphosphatase; Adenosinetriphosphatase; Affect; Airbron; Allen & Hanburys Brand of Acetylcysteine; alpha-2-Macroglobulin Receptor; alpha-Lipoic Acid; alpha2-Macroglobulin Signaling Receptor; Alzheimer; Alzheimer disease; Alzheimer Disease 1 Protein; Alzheimer sclerosis; Alzheimer syndrome; Alzheimer`s; Alzheimer`s Disease; Alzheimer`s Disease Amyloid Protein; Alzheimers Dementia; Alzheimers disease; Amyloid; amyloid beta (A4) precursor protein (protease nexin-II, Alzheimer disease) protein, human; Amyloid Beta A4 Protein Precursor; Amyloid of Aging and Alzheimer Disease Protein; Amyloid Substance; analog; Animals; anti-oxidant; Antibodies; Antioxidants; ApoE Receptor; APP protein, human; arm; ATP phosphohydrolase; ATP phosphohydrolase (Na+ K+ transporting); ATP-Binding Cassette, Sub-Family B Proteins; ATP-Binding Cassette, Sub-Family B, Member 1; ATPase; base; Binding; Binding (Molecular Function); Blood; Blood - brain barrier anatomy; Blood capillaries; Blood Vessels; Blood-Brain Barrier; Body Tissues; Brain; brain control; brain tissue; Bristol-Myers Squibb Brand of Acetylcysteine; Bristol-Myers Squibb Brand of Acetylcysteine Sodium Salt; Broncholysin; Brunac; Capillaries; capillary; Capillary; Capillary, Unspecified; Cerebralvascular Amyloid Peptide; Cessation of life; Cognitive decline; Cognitive Disturbance; cognitive dysfunction; Cognitive function abnormal; Cognitive Impairment; cognitive loss; cognitively impaired; Death; dementia of the Alzheimer type; Dementia, Alzheimer Type; Dementia, Primary Senile Degenerative; Dementia, Senile; Disturbance in cognition; Dysfunction; efflux pump; electron acceptor; Encephalon; Encephalons; Endothelial Cells; Epidemiologic Research; Epidemiologic Studies; Epidemiological Studies; Epidemiology Research; Event; experiment; experimental research; experimental study; Fabrol; Fluatox; Fluimucetin; Fluimucil; Fluprowit; Functional disorder; gene product; Genes; Genetic Alteration; Genetic Change; Genetic defect; genome mutation; Glutamate Translocase; Glutamate Transport Glycoprotein; Glutamate Transporter; Glutamates; Goals; Hemato-Encephalic Barrier; Hepatic Cells; Hepatic Parenchymal Cell; Hepatocyte; Human; human APP protein; Human, General; Hydrophobicity; Impaired cognition; Impairment; In Vitro; in vitro Model; in vivo; indexing; Inpharzam Brand of Acetylcysteine; intervention therapy; L-Alpha-acetamido-beta-mercaptopropionic Acid; L-Glutamate; L-Methionine; Laboratories; Laboratory mice; LDL-Receptor Related Protein 1; Length; Ligands; Lipid Peroxidation; Lipoic Acid; Lipoprotein Receptor; Liver Cells; Low Density Lipoprotein Receptor-Related Protein; Low-Density-Lipoprotein Receptor-Related Protein-1; Macroglobulins; Mammals, Mice; Mammals, Rodents; Man (Taxonomy); Man, Modern; MDR1 Protein; Measures; meetings; Mercapturic Acid; Methionine; Methionine, L-Isomer; Mice; mild cognitive disorder; mild cognitive impairment; mild neurocognitive disorder; mind control; Modeling; Modification; Molecular Interaction; monolayer; mouse model; Muco Sanigen; Mucocedyl; Mucolator; Mucolyticum; Mucomyst; Mucosolvin; Mucret; Multidrug Resistance 1; Multidrug Resistance Protein 1; Multidrug Resistance Proteins; Multidrug Resistant Proteins; Murine; Mus; mutant; Mutation; N-Acetyl Cysteine; N-acetyl-3-mercaptoalanine; N-Acetyl-L-cysteine; N-Acetylcysteine; Na(+)-K(+)-Exchanging ATPase; Na(+)-K(+)-Transporting ATPase; Na+ K+ ATPase; NAC; NAC Zambon; Neo-Fluimucil; Nervous System, Brain; nitrotyrosine; Norleucine; Optipect Hustengetr??nk; overexpression; Oxidants; oxidation; oxidative damage; Oxidative Stress; Oxidizing Agents; P-Glycoprotein; P-Glycoprotein 1; P-Glycoprotein Transporter; P-Glycoproteins; P-GP; Parvolex; pathophysiology; Patients; Peptides; PGY-1 Protein; Phosphorothioate Oligonucleotide; Physiopathology; Play; poison; Poisons; Potassium Pump; PreA4; primary degenerative dementia; Primary Senile Degenerative Dementia; Produpharm Lappe Brand of Acetylcysteine; protease nexin 2; protease nexin 2, human; Protease Nexin II; Protease Nexin-II, Alzheimer Disease; Proteins; Pump; Receptor, Apo E; Receptor, Apolipoprotein E; Relative; Relative (related person); research study; Resistance; resistant; Respaire; Reticuloendothelial System, Blood; Risk; Roberts Brand of Acetylcysteine; Rodent; Rodentia; Rodentias; Role; senescence; senile dementia of the Alzheimer type; Series; social role; sodium potassium exchanging ATPase; Sodium Pump; Sodium, Potassium Adenosine Triphosphatase; Sodium, Potassium Adenosinetriphosphatase; Sodium, Potassium ATPase; Sodium-Potassium Pump; System; System, LOINC Axis 4; Testing; Therapeutic Intervention; Thiemann Brand of Acetylcysteine; Thioctic Acid; Tissues; Tixair; Toxic Chemical; toxic compound; Toxic Substance; transgenic; Transgenic Organisms; unpublished works; Unpublished Works (PT); Unpublished Works [Publication Type]; UPSA Brand of Acetylcysteine; vascular; Vitamin E; Work; Zambon Brand of Acetylcysteine; Zyma Brand of Acetylcysteine
Project start date: 2008-03-01
Project end date: 2013-02-28
Budget start date: 15-MAR-2011
Budget end date: 29-FEB-2012
PFA/PA: PA-07-070
5R01AG029839-04 (2011): $255767
SOLID STATE RADIATION CHEMISTRY OF DNA
A William, Professor
University Of Rochestercity: Rochester country: United States (us)
Grant 5R01CA032546-36 from National Cancer Institute
Abstract: The long-term objective of this proposed research program is to (i) measure the types, yields and spatial distribution of the biologically relevant DNA lesions produced by the direct effect of ionizing radiation, (ii) determine the chemical mechanisms by which these DNA lesions are formed, and (iii) evaluate how molecules such as histones and other DNA binding proteins modulate the yield and distribution of these DNA lesions. Achievement of these objectives should make it possible to predict the composition and spatial distribution of the DNA lesions within biologically important clusters. This predictive capability is central to making risk/benefit decisions at both low dose rates and low doses of radiation and to constructing biologically relevant models of clustered DNA damage to be used by DNA enzymologist, biochemists and biophysicists in their DNA repair/misrepair studies. The specific aims are 1) to determine the yields of specific sugar and base damage end-products produced by the direct effect as a function of radiation dose, and elucidate the reaction mechanisms giving rise to them, 2) to determine the yield and composition of DNA damage within clusters produced by the direct effect, and 3) to determine how histones and other protein DNA binding complexes modify the yield and composition of clustered DNA damage produced by the direct effect. Damage to DNA by the direct effect occurs via two routes. One is by direct ionization of the DNA. The other is by ionization of that portion of the solvent shell that is tightly bound to the DNA and rapid transfer of that damage to the DNA. Because of these properties, information on the direct effect is optimized by studying DNA in the solid state. The DNA samples, for this proposed research program, will be prepared in the form of crystals and films. Using crystals of known structure, we maximize our knowledge of relevant parameters base sequence, conformation, hydration state, counter ions, packing, and purity. Using films we are able to vary parameters such as base sequence and degree of hydration. Unstable free radical intermediates will be studied by electron paramagnetic resonance (EPR) spectroscopy. Stable end products will be quantified by high performance liquid chromatography (HPLC), gas chromatography/mass spectrometry (GC/MS), and liquid chromatography/mass spectrometry (LC/MS). Stable end products will be analyzed using the same samples as those studied by EPR. Key elements in our experimental design are the use of structurally well defined DNA samples, making it possible to extend our knowledge of free radical reactions to understand the mechanisms of end product formation at low dose. By achieving the goals set down in this proposal, we will improve our ability to make risk/benefit decisions at low dose rates and low doses of radiation. Further, it provides the information needed to construct biologically relevant models of clustered DNA damage that can be used in studies of DNA repair and misrepair. The long-term objective of this proposed research program is to, (i) measure the types, yields and spatial distribution of the biologically relevant DNA lesions produced by the direct effect of ionizing radiation, (ii) determine the chemical mechanism(s) by which these DNA lesions are formed, and (iii) evaluate how molecules such as histones and other DNA binding proteins modulate the yield and distribution of these DNA lesions. Achievement of these objectives should make it possible to predict the composition and spatial distribution of the DNA lesions within biologically important clusters. This predictive capability is central to making risk/benefit decisions at both low dose rates and low doses of radiation and to constructing biologically relevant models of clustered DNA damage to be used by DNA enzymologist, biochemists and biophysicists in their DNA repair/misrepair studies
Keywords: 2-Deoxyribose; Accounting; Achievement; Achievement Attainment; B-DNA; base; Base Sequence; Benefits and Risks; Binding; Binding (Molecular Function); Binding Proteins; Biological Models; Chemicals; Chromatography, Gas-Liquid-Mass Spectrometry; Chromatography, High Performance Liquid; Chromatography, High Pressure Liquid; Chromatography, High Speed Liquid; Complex; conformation; conformational state; D-erythro-Pentose, 2-deoxy-; Deoxyribonucleic Acid; Deoxyribose; design and construction; DNA; DNA Damage; DNA Damage Repair; DNA Injury; DNA lesion; DNA Repair; DNA Sequence; DNA, B Form; DNA-Binding Proteins; Dose; Dose-Rate; Electromagnetic Radiation, Ionizing; Electron Paramagnetic Resonance; electron paramagnetic resonance spectroscopy; Electron Spin Resonance; Electron Spin Resonance Spectroscopy; EPR spectroscopy; experiment; Experimental Designs; experimental research; experimental study; Film; Foundations; Free Radicals; GC MS; GCMS; gene product; Goals; High Pressure Liquid Chromatography; Histones; HPLC; Hydration; Hydration status; improved; In element; in vivo; Indium; ion trap mass spectrometry; ionization; Ionizing radiation; Ions; Knowledge; LC/MS; Lesion; Ligand Binding Protein; liquid chromatography mass spectrometry; Low Dose Radiation; Mass Fragmentographies; Mass Fragmentography; mass fragmentometry; Measures; Model System; Modeling; Models, Biologic; Molecular Configuration; Molecular Conformation; Molecular Interaction; Molecular Stereochemistry; nucleic acid sequence; Nucleotide Sequence; oxidation; Paramagnetic Resonance; Peptides; plasmid DNA; Plasmids; Play; polypeptide; programs; Programs (PT); Programs [Publication Type]; Property; Property, LOINC Axis 2; Proteins; Radiation; Radiation Chemistry; Radiation-Ionizing Total; Radiochemistry; ray (radiation); Reaction; Research; research study; Right-Handed DNA; Role; Route; Sampling; Site; social role; solid state; Solvents; Spatial Distribution; Spectrometry, Mass-Gas Chromatography; Spectroscopy, ESR; Spectrum Analysis, Mass-Gas Chromatography; Structure; sugar; Tail; Testing; tetralysine; Unscheduled DNA Synthesis
Project start date: 1982-02-01
Project end date: 2013-01-31
Budget start date: 1-FEB-2011
Budget end date: 31-JAN-2012
PFA/PA: PA-07-070
5R01CA032546-36 (2011): $430241
Sponsored Links Excellgen http://Excellgen.com
IDENTIFYING THE MEMBRANE PROTEINS OF THE LBRC, A KEY REGULATOR OF INFLAMMATION
A William, Magerstadt Professor And Chair
Northwestern Universitycity: Chicago country: United States (us)
Grant 5R21HL102519-02 from National Heart, Lung, And Blood Institute
Abstract: The lateral border recycling compartment (LBRC) is a recently-discovered membrane compartment located along the borders of vascular endothelial cells. Membrane from the LBRC recycles constitutively and rapidly between the compartment and the plasma membrane at borders between endothelial cells. The physiologic function of this constitutive recycling is not known. However, during leukocyte transmigration, recycling membrane from the LBRC is redirected-targeted to the site at which the leukocyte engages the endothelial cell junctions. It surrounds the leukocyte during the transmigration process providing increased membrane surface area in the junction and several key adhesion molecules that regulate transmigration. Blocking targeted recycling blocks leukocyte transmigration. Hence, the LBRC and its targeted recycling are critical factors regulating leukocyte transmigration in inflammation. Understanding how targeted recycling is regulated would provide insights into the control of the inflammatory response and potentially identify new therapeutic targets. Knowing the composition of the LBRC would allow us to formulate testable hypotheses about the function(s) of constitutive recycling and the regulation of targeted recycling. However, only three components of the LBRC are known Platelet/endothelial cell adhesion molecule-1 (PECAM, CD31), CD99, and Junctional Adhesion Molecule A (JAM-A). In order to obtain an unbiased insight into the composition of the LBRC, we will isolate LBRC membrane from endothelial cell homogenates by density gradient centrifugation. We will analyze the protein composition of this membrane by two dimensional gel electrophoresis and identify membrane proteins unique to or enriched in the LBRC by comparison to the protein profiles of plasma membrane fractionated in parallel. Spots on the gels representing proteins that appear to be unique or markedly enriched in LBRC membrane will be excised, protease digested, and identified by mass spectrometry. As a complementary approach, a total proteomic comparison of LBRC and plasma membrane fractions will be performed. Candidate LBRC proteins will be validated in several ways. In the first step, we will localize these proteins in intact endothelial cells to be certain they have the correct distribution. They should be concentrated at the endothelial cell borders. Immunolocalization by confocal fluorescence microscopy will be performed. We expect most of the proteins will be well known and previously described. We will use existing antibodies against the candidate proteins where they are available and generate FLAG-tagged constructs of the candidate proteins, based on known sequences, where they are not. Those proteins that co-localize with authentic LBRC in intact cells will be further tested to determine whether they are truly components of the LBRC in functional assays. We will determine, using well-established assays in our lab, whether they recycle constitutively in the same manner as LBRC and participate in targeted recycling during leukocyte transmigration. Most diseases (including atherosclerosis, asthma, and autoimmune diseases) involve an inflammatory response that is uncontrolled or misdirected. We have discovered a novel membrane compartment in endothelial cells (the cells that line blood vessels) that is critical for the inflammatory response. We will isolate this membrane compartment and identify its component proteins in order to understand how it functions and design better anti-inflammatory therapies
Keywords: Anti-inflammatory; Anti-Inflammatory Agents; Antibodies; Applications Grants; Area; Asthma; Atherosclerosis; Autoimmune Diseases; base; Biochemical; Biological Assay; Blood Vessels; cadherin 5; CD31 Antigens; Cell Adhesion Molecules; Cell Fractionation; Cell Line; Cell membrane; Cells; Databases; Density Gradient Centrifugation; design; Development; Disease; Electrophoresis, Gel, Two-Dimensional; Endothelial Cells; Ensure; Fluorescence Microscopy; Funding Mechanisms; Future; Gel; Goals; Immunoelectron Microscopy; Immunofluorescence Immunologic; Inflammation; Inflammatory Response; insight; Intercellular Junctions; junctional adhesion molecule; Kinesin; Knowledge; Lateral; Leukocytes; Life; Lymphocyte; Mass Spectrum Analysis; Membrane; Membrane Proteins; Methods; Microtubules; migration; Molecular Motors; monocyte; Names; neutrophil; new therapeutic target; novel; Organelles; PECAM1 gene; Peptide Hydrolases; Peptide Mapping; Physiological; Preparation; Procedures; Process; protein profiling; Proteins; Proteomics; public health relevance; Recycling; Regulation; Research Personnel; Rest; Reticulum; Role; scale up; Side; Site; Spottings; Staging; Structure; Surface; Techniques; Testing; Time; two-dimensional; Vascular Endothelial Cell
Project start date: 2010-05-13
Project end date: 2012-03-31
Budget start date: 1-APR-2011
Budget end date: 31-MAR-2012
PFA/PA: PA-09-164
5R21HL102519-02 (2011): $190625
GLI1 AND CANCER STEM CELLS IN EWING SARCOMA
A William, Associate Professor Of Pediatrics
Children´s Hospital Los Angelescity: Los Angeles country: United States (us)
Grant 5R21CA137485-02 from National Cancer Institute
Abstract: The horizons of cancer biology have been expanded by the concept of cancer stem cells (CSC). In concept, tumors may arise from and continue to harbor a stem cell population which is responsible for ongoing tumor self renewal. This distinct population may hold the key to understanding the development of treatment resistance. Because of these factors, the study of cancer stem cells promises novel insights into the origins and treatment of cancer. The Ewing Sarcoma Family of Tumors (ESFT) represents a particular challenge. Its cell of origin is unknown. While initially very responsive to chemotherapy, the emergence of treatment resistance occurs in over 40%, usually with fatal results. ESFT are united by a chromosomal translocation fusing the EWS gene an ETS transcription factor. The prototype fusion, EWS/FLI11 occurs in over 85% of cases. Abundant evidence supports a crucial role for EWS/FLI1 in ESFT through its function as an aberrant transcription factor. My lab has recently uncovered a novel effect of EWS/FLI1 in enhancing expression of GLI1 in ESFT. GLI1 is the effector of the Hedgehog/GLI (HH/GLI) signaling pathway. This pathway has prominent roles in development and in many malignancies. It has been shown to contribute to the maintenance of stem cell phenotype in other tumor systems. This proposal will investigate the hypothesis that GLI1 upregulation by EWS/FLI1 is critical to maintaining a stem cell compartment in ESFT and whether this CSC compartment is resistant to treatment Since ESFT have been shown to harbor CD133 positive CSC, we will select CD133 positive fractions from ESFT cell lines and perform in vivo tumorigenesis assays. Validated CD133 sorted ESFT cell populations will be assessed by gene expression array to assess the expression of GLI1 and its targets. We will also assess for other potential stem cell markers. The role of GLI signaling in ESFT CSC will be tested by measuring the quantitative effects of GLI1 inhibition on the CSC fraction. Methods of inhibition will include both GLI1 specific shRNA and small molecule inhibitors of GLI signaling. We will also test the results of GLI inhibition on CSC function by measuring the effect of GLI inhibition on CSC tumorigenesis assays. We will also test the hypothesis that stem cell populations are associated with treatment resistance and whether GLI1 contributes to this. This will be tested using validated CD133 positive CSC populations generated in Aim 1. Using systems in place for the Ewings Preclinical Testing Lab, we will measure any differences in sensitivity to a wide range of relevant drugs in already characterized ESFT cell lines. These changes will be documented by high throughput in vitro assessment of chemosensitivity (DIMSCAN). Ewing´s Sarcoma is a deadly cancer of childhood, adolescence and young adulthood which takes the life of over 40% of those diagnosed. This work will detail the role of Gli1 signaling in maintaining cancer stem cells in Ewing´s Sarcoma. If Gli1 is important in maintaining cancer stem cells, these treatment- resistant cells can be specifically targeted with drugs, enhancing our ability to cure patients
Keywords: 12-20 years old; Adolescence; adolescence (12-20); adult youth; Anaplastic; anticancer therapy; Assay; Bioassay; Biologic Assays; Biological Assay; biological signal transduction; bone; Bone; Bone and Bones; Bones and Bone Tissue; Cancer Biology; Cancer stem cell; cancer therapy; Cancer Treatment; Cancers; Cell Communication and Signaling; Cell Fraction; Cell Function; Cell physiology; Cell Process; Cell Signaling; Cells; Cellular Function; Cellular Physiology; Cellular Process; chemotherapy; Childhood Cancers; Chromosomal dislocation; Chromosomal translocation; chromosome dislocation; chromosome translocation; Cytotoxic Chemotherapy; Cytotoxic Therapy; Development; Diagnosis; DNA Binding Domain; drug/agent; Drugs; Erinaceidae; Ewing Sarcoma Breakpoint Region 2; Ewing`s Family of Tumours; Ewing`s Sarcoma/Peripheral Primitive Neuroectodermal Tumor; Ewing`s Tumor; Ewings sarcoma; EWSR2; Exhibits; Family; FLI1; FLI1 gene; Gene Expression; gene product; Genes; GLI; GLI gene; GLI1; GLI1 Gene; Glioma-Associated Oncogene Homolog; Hedgehogs; In Vitro; in vivo; inhibitor; inhibitor/antagonist; insight; intervention development; Intracellular Communication and Signaling; Life; Maintenance; Maintenances; malignancy; Malignant Childhood Neoplasm; Malignant Childhood Tumor; Malignant Neoplasm Therapy; Malignant Neoplasm Treatment; Malignant Neoplasms; Malignant Pediatric Neoplasm; Malignant Pediatric Tumor; Malignant Tumor; Measures; Medication; Methods; Mother Cells; neoplasm/cancer; neoplastic cell; novel; Oncogenesis; pathway; Pathway interactions; Patients; pediatric cancer; pediatric neoplasm/cancer; Pharmaceutic Preparations; Pharmaceutical Preparations; Phenotype; Play; Population; Preclinical Testing; Progenitor Cells; Property; Property, LOINC Axis 2; Protein Motifs, DNA-Binding; Proteins; prototype; public health relevance; Research; Resistance; resistant; Role; self-renewal; short hairpin RNA; shRNA; SIC-1; Signal Pathway; Signal Transduction; Signal Transduction Systems; Signaling; Site; small hairpin RNA; small molecule; social role; soft tissue; sorting; Sorting - Cell Movement; stem cell population; Stem cells; Subcellular Process; System; System, LOINC Axis 4; teenage; Test Result; Testing; therapy development; transcription factor; Translocation, Genetic; treatment development; tumor; Tumor Cell; Tumor Cell Line; tumorigenesis; Undifferentiated; Up-Regulation; Up-Regulation (Physiology); Upregulation; Work; young adult
Relevance: Ewing´s Sarcoma is a deadly cancer of childhood, adolescence and young adulthood which takes the life of over 40% of those diagnosed. This work will detail the role of Gli1 signaling in maintaining cancer stem cells in Ewing´s Sarcoma. If Gli1 is important in maintaining cancer stem cells, these treatment- resistant cells can be specifically targeted with drugs, enhancing our ability to cure patients
Project start date: 2010-01-01
Project end date: 2011-12-31
Budget start date: 1-JAN-2011
Budget end date: 31-DEC-2011
PFA/PA: PA-08-165
5R21CA137485-02 (2011): $156600
PROBLEMS IN MEMBRANE PROTEIN CRYSTALLOGRAPHY: HETERO-OLIGOMERIC CYTOCHROME B6F
A William
Purdue University West Lafayettecity: West Lafayette country: United States (us)
Grant 5R01GM038323-23 from National Institute Of General Medical Sciences
Abstract: The hetero-oligomeric cytochrome b6f and bc1 complexes are in the center of the electron transfer chains in photosynthetic and respiratory energy transducing membranes. Such membranes contain the majority of the relatively few hetero-oligomeric integral membrane proteins that have been solved by X-ray crystallography to a resolution d 3.0 E. Studies on crystallization of the dimeric 220 kDa eight subunit integral b6f complex would analyze problems of proteolysis and lipid-protein interactions that are of general relevance to the crystallization of integral membrane proteins. Structure-function analysis would focus on the properties of a unique redox group, heme cn, and on the mechanism of transfer across the membrane of qui (ol) that carries the electrons and protons and is reduced by heme cn. Proposed studies (1) Crystal preparation; proteolysis. b6f complex cannot be isolated from transformable unicellular cyanobacteria because the b6f dimer is monomerized and rendered inactive and non-crystallizable upon extraction from the membrane. Successful crystallization of cyanobacterial b6f has utilized the filamentous M. laminosus, in which the extent of proteolysis is smaller. However, M. laminosus is not transformable. Therefore, the cyanobacterial source of b6f complex will be changed to the filamentous Nostoc (Anabaena) sp. PCC 7120, from which active and crystallizable complex has been obtained. Because this kind of proteolysis problem frequently hinders efforts to crystallize membrane proteins, the critical protease(s) in the unicellular cyanobacteria would be identified by mass spectroscopic and proteomic analysis. (2) Function of phospholipids. The function of intra-protein lipids has been analyzed in only a few multi-subunit membrane proteins. Our novel lipid augmentation procedure resulted in a major increase in the rate of crystallization and improvement in crystal quality. The properties of crystals of the plant thylakoid membrane b6f complex, which contains a ninth (FNR) subunit and whose crystallization depends uniquely on a different (anionic DOPG) lipid, is under study, as is the dependence of electron transfer activity and rate of crystallization on the nature of added lipids. (3) Functions of heme cn; evolution of b6f complex. The function of the unique heme cn, not found in ubiqui-containing cyt bc1 complexes, will be studied by site-directed mutagenesis in Nostoc and, through structure-function analysis, in firmicutes such as Bacillus subtilis that are phylogenetically close to cyanobacteria. His-tagged, promoter-augmented firmicute "qcr" complex will be purified and screened for crystallization and electron transfer reactions with menaqui. (4) Qui transfer though the narrow p-side portal. Electrons and protons are carried across an inter-monomer qui exchange cavity in bc1 and b6f complexes by lipophilic ubi- and plastoqui (PQ). The mechanisms by which PQ/PQH2 finds, enters, and exits a narrow 11 x 12 E p-side portal will be studied through mutagenesis of portal residues and computational analysis of the portal force field. Some of the biomedically relevant aspects of these studies are that they are directed toward an understanding of the detailed internal structure of the proteins that mediate all traffic, including nutrients and drugs, across biological membranes. Via the membrane, the set of energy-transducing proteins determines the level of energy and its regulation in the human cell
Keywords: 1, 2, 3-Propanetriol; 1, 2, 3-Trihydroxypropane; 1, 2-diacylglycerol; 2, 5-Cyclohexadiene-1, 4-dione, 2, 3-dimethyl-5-(3, 7, 11, 15, 19, 23, 27, 31, 35-nonamethyl-2, 6, 10, 14, 18, 22, 26, 30, 34-hexatriacontanonaenyl)-, (all-E)-; 2-Hydroxy-N, N, N-trimethylethanaminium; 4H-1-Benzopyran-4-one, 2-(4, 6-dimethoxy-3, 5, 11-trimethyl-7, 9, 11-tridecatrienyl)-8-hydroxy-5, 7-dimethoxy-3-methyl-; Abbreviations; ing; Acceleration; Active Oxygen; Adrenodoxin Reductase; Algae; Algae, Blue-Green; Aminoethanols; Anabaena; analog; Antibody Fragments; Antiviral Agents; Antiviral Drugs; Antivirals; Appearance; Bacillus subtilis; Benzodiones; Benzoquis; Binding; Binding (Molecular Function); Binding Sites; Biological; Blue-Green Bacteria; body movement; Cardiolipins; Cells; Chlorophyll; Chloroplasts; Choline; Choline Glycerophospholipids; Choline Phosphoglycerides; Coenzyme Q; Combining Site; comparative genomics; Complex; computational analysis; computational methodology; computational methods; Computer Analysis; computer methods; Computing Methodologies; Coupling; Crude Extracts; Crystallization; Crystallographies; Crystallography; Crystallography, X-Ray; Crystallography, X-Ray Diffraction; Crystallography, X-Ray/Neutron; Crystallography, Xray; Cyanobacterium; Cyanophyceae; Cyanophyta; CYP; Cytochrome b(6)-f Complex; Cytochrome b6-f; cytochrome b6f; Cytochrome b6f Complex; Cytochrome bc1; Cytochrome bc1 Complex; Cytochromes; DAG; Data; Dependence; DGDG; diacyl glyceride di Gal; diacylglycerol; Diacylglycerols; digalactosyldiacylglycerol; diglyceride; Diglycerides; dimer; drug/agent; Drugs; Electron Paramagnetic Resonance; electron paramagnetic resonance spectroscopy; Electron Spin Resonance; Electron Spin Resonance Spectroscopy; electron transfer; Electron Transport; Electrons; Electrophoresis; EPR spectroscopy; Esteroproteases; Ethanaminium, 2-hydroxy-N, N, N-trimethyl-; Ethanolamines; Evolution; experience; Extracts, Complex; Fe element; Ferrate(2-), (7, 12-diethenyl-3, 8, 13, 17-tetramethyl-21H, 23H-porphine-2, 18-dipropanoato(4-)-N21, N22, N23, N24)-, dihydrogen, (SP-4-2)-; Ferredoxin-NADP Reductase; Ferredoxin[{..}]NADP+ oxidoreductase; ferroheme; Ferroprotoporphyrin; Fractionation, Electrophoretic; G Protein-Complex Receptor; G Protein-Coupled Receptor Genes; G-Protein-Coupled Receptors; gene product; Genetics-Mutagenesis; Genome; genome sequencing; Glycerin; Glycerol; Goals; GPCR; GPR; H(+) Pump; H+ element; Harvest; Heme; Heme b; Heme Group; his-PG; Human; Human, General; Hydrogen Ions; Hydroquis; Immunoblotting; Immunoglobulin Fragments; improved; in vivo; inhibitor; inhibitor/antagonist; insight; Integral Membrane Protein; Intrinsic Membrane Protein; Iron; Iron-Sulfur Protein Reductase; Iron-Sulfur Proteins; Lecithin; Ligands; Light; Link; Lipids; Man (Taxonomy); Man, Modern; Measures; Mediating; Medical Imaging, Positron Emission Tomography; Medication; Membrane; Membrane Proteins; membrane structure; Membrane-Associated Proteins; Menaqui; Molecular Biology, Mutagenesis; Molecular Interaction; monomer; Movement; Mutagenesis; Mutagenesis, Site-Directed; Na element; NADPH-Ferredoxin Reductase; Nature; Negative Beta Particle; Negatrons; nonyl-4-hydroxyquinoline-N-oxide; Nostoc; novel; NQ-N-oxide; NQNO; Nutrient; oxidation reduction reaction; Oxidation-Reduction; Oxygen Radicals; p-Dihydroxybenzenes; para-Dihydroxybenzenes; Paramagnetic Resonance; pathway; Pathway interactions; PCR; Peptidases; Peptide Hydrolases; PET; PET imaging; PET Scan; PETSCAN; PETT; Pharmaceutic Preparations; Pharmaceutical Preparations; Phosphatides; Phosphatidylcholines; Phospholipids; Photoradiation; photosystem; Physiologic; Physiological; Plant Components; Plant Structures; Plants; Plants, General; Plastocyanin; Plastocyanine; Plastoqui; Plastoqui-9; polyacrylamide; Polymerase Chain Reaction; polypeptide; Positron Emission Tomography Scan; Positron-Emission Tomography; Preparation; Pro-Oxidants; Procedures; progesterone 11-hemisuccinate-(2-iodohistamine); Promoter; Promoters (Genetics); Promotor; Promotor (Genetics); Property; Property, LOINC Axis 2; Prosthesis; Prosthetic device; Prosthetics; Proteases; Protein Cleavage; protein structure; Proteinases; Proteins; Proteolysis; Proteolytic Enzymes; Proteomics; Protoheme; Protoheme IX; Proton Magnetic Resonance Spectroscopic Imaging; Proton Pump; Protons; Quinols; Qui Compound; Qui Reductases; Quis; Rad.-PET; Reaction; Reactive Oxygen Species; Reactive Site; Redox; Regulation; Resolution; respiratory; Rhinovirus; rhomboid; Role; S element; screening; Screening procedure; screenings; Side; Single Crystal Diffraction; Site-Directed Mutagenesis; Site-Specific Mutagenesis; social role; Sodium; Source; Spectroscopy, ESR; stigmatellin; Structure; Sulfur; Surface Proteins; Survey Instrument; Surveys; System; System, LOINC Axis 4; Targeted DNA Modification; Targeted Modification; Thylakoid Membranes; trafficking; Transmembrane Protein; Trees; ubiquinol; Ubiqui; Vitamin K 2; Vitamin K Qui; Vitamin K2; X Ray Crystallographies; X-Ray Crystallography; Yeasts
Relevance: Narrative Some of the biomedically relevant aspects of these studies are that they are directed toward an understanding of the detailed internal structure of the proteins that mediate all traffic, including nutrients and drugs, across biological membranes. Via the membrane, the set of energy-transducing proteins determines the level of energy and its regulation in the human cell
Project start date: 1987-04-01
Project end date: 2013-01-31
Budget start date: 1-FEB-2011
Budget end date: 31-JAN-2012
PFA/PA: PA-07-070
5R01GM038323-23 (2011): $299913
THE ENVIRONMENTAL DETERMINANTS OF DIABETES IN YOUTH: WASHINGTON CLINICAL CENTER
A William, Private Investigator
Pacific Northwest Research Institutecity: Seattle country: United States (us)
Grant 5U01DK063829-09 from National Institute Of Diabetes And Digestive And Kidney Diseases
Abstract: The Environmental Determinants of Diabetes in the Young (TEDDY), a multicenter prospective cohort study, was initiated in 2003 to identify environmental factors that trigger or protect against the development of islet autoimmunity and type 1 diabetes (T1D). A successful study outcome should allow better understanding of the etiology and pathogenesis of T1D and the development of new strategies to prevent, delay, or reverse the disease. Newborns are screened by HLA-DR.DQ genotyping to identify those at increased risk for T1D. Eligible children are followed four times per year until 4 years of age and twice a year thereafter until 15 years of age. The specific aims of this Clinical Center renewal application are to 1. Complete this Center´s contribution to the HLA-DR.DQ screening of 361,000 newborns and enrollment of 7,800 high-risk infants into intensive prospective follow-up to achieve the overall TEDDY goals. 2. Refine strategies to retain subjects enrolled in the follow-up and collect all planned biological specimens and epidemiological data according to the standard protocol including close monitoring of performance and sample/data quality. 3. Ascertain prospectively development of islet autoantibodies and clinical T1D in the study subjects. 4. Perform planned laboratory tests at appropriate times using a nested case-control study design to answer specific scientific questions and hypotheses pertinent to the TEDDY study goals. 5. Analyze and publish laboratory and epidemiological data in collaboration with the TEDDY Data Coordinating Center (funded by a separate contract). 6. Guide the ongoing TEDDY project by participation of the Clinical Center investigators and staff in work of the study Steering Committee and sub-committees
Keywords: 15 year old; 4 year old; Accounting; Area; Autoantibodies; Autoimmune Diabetes; Autoimmune Diseases; Autoimmunity; Beta Cell; Biological; Blood specimen; C-Peptide; Candidate Disease Gene; Cell Communication; Child; Childhood; Clinical; cohort; Cohort Studies; Collaborations; Contracts; Data; Data Coordinating Center; Data Quality; Development; Diabetes Mellitus; diabetes risk; Disease; disorder risk; Eating; endocrine pancreas development; Enrollment; Environment; Environmental Exposure; Environmental Risk Factor; Epidemiology; Etiology; Evaluation; Family; Fasting; Feces; follow-up; Food; Freezing; Funding; Future; gene environment interaction; Genes; Genetic; Genetic Polymorphism; Genetic Predisposition to Disease; Genetic Risk; Genotype; Goals; Hair; high risk; high risk infant; HLA-DR Antigens; Human; Hypersensitivity; Institutional Review Boards; Insulin; Insulin-Dependent Diabetes Mellitus; Laboratories; Measures; Methods; Microsatellite Repeats; Minisatellite Repeats; Monitor; Monozygotic twins; Mycotoxins; Neonatal Screening; Nested Case-Control Study; Newborn Infant; non-diabetic; Outcome Study; Oxidants; Parents; Pathogenesis; Penetrance; Performance; Peripheral Blood Mononuclear Cell; Pharyngeal structure; Play; Population; population based; Prediabetes syndrome; prevent; programs; Promotor (Genetics); prospective; Protocols documentation; Publishing; Recruitment Activity; Research Design; Research Personnel; Risk; Role; Sampling; Screening procedure; Specimen; Staging; Statistical Data Interpretation; Sterility; stressor; Study Subject; Surveys; Swab; Telephone; Testing; Time; time use; Toxin; Twin Studies; Urine; Vaccination; Variation (Genetics); virus culture; Washington; Work; Youth
Project start date: 2003-03-01
Project end date: 2013-04-30
Budget start date: 1-MAY-2011
Budget end date: 30-APR-2012
PFA/PA: RFA-DK-07-500
5U01DK063829-09 (2011): $586783
BIODEFENSE RESEARCH TRAINING AND CAREER DEVELOPMENT
A William, Wade Hampton Frost Professor
University Of Virginia Charlottesvillecity: Charlottesville country: United States (us)
Grant 5T32AI055432-09 from National Institute Of Allergy And Infectious Diseases
Abstract: The Biodefense Research Training Program is in its 5th year. Predoctoral trainees enter the Program at the start of the 2nd graduate year, after completion of research rotations & qualifying exam. Postdoctoral fellows enter the Program after selecting a Program mentor´s lab. Admission is competitive, with 16% and 3% of qualified 2nd year predoc and postdoc applicants accepted in 2006. Research encompasses the immunology, cell biology, pharmacology and microbiology of Category A-C agents of bioterrorism. The training of predoctoral students, M.D., and Ph.D. postdoctoral fellows is enriched by special activities of the Program, including a monthly breakfast research-in-progress meeting solely for the trainees chaired by the Program Director, two graduate courses in Biodefense, an expanded 2 semester course in molecular mechanisms of microbial pathogenesis and virology, an MPH degree program, Career Day, journal club, and coordination of training with the Middle Atlantic Center of Excellence for Biodefense. The Program Director is William Petri, M.D., Ph.D. who conducts Biodefense research on amoebiasis, and is Chief of the Infectious Diseases Division. An Executive Committee provides Program oversight and trainee selection. Six departments contribute a total of 18 faculty members (1 DVM, 5 MD and 16 PhD). All labs are within a 5 minute walk, and all faculty members collaborate, with 80% co-authoring papers. 100% of the preceptors are extramurally supported by the NIH through at least 2008, at an average of $900,000 in annual direct costs/faculty (a 90% increase since 2001). The average Preceptor has trained > 10 pre- and/or postdoctoral students, but select "up and coming" junior faculty with less extensive training records are also included with more experienced faculty provided as co- mentors. Funds are requested to train 4 predoctoral fellows and 2 postdoctoral fellows (an increase in 1 predoctoral position). Progress includes predoctoral trainee GRE scores of 660V/690Q and GPA of 3.5, 100% of trainees in Biodefense labs, and 94% retention of trainees in the Program. 67% (12/18) of the mentors have trained at least one trainee. 17% of trainees are underrepresented minorities. Women comprise 61% of trainees and 33% of faculty. 6 trainees have graduated, with an average of 3 papers published or submitted, with 83% active in research (5/6 - the 6th is currently a stay-at-home mother). Evaluations of the Program include an annual faculty retreat and a trainee questionnaire
Keywords: biodefense; career development; Research Training
Project start date: 2003-08-01
Project end date: 2013-06-30
Budget start date: 1-JUL-2011
Budget end date: 30-JUN-2012
PFA/PA: PA-06-468
5T32AI055432-09 (2011): $222911
BIOCHEMICAL AND BIOPHYSICAL ANALYSIS OF ANGIOPOIETIN SIGNALING
A William, Asst Professor
Virginia Commonwealth Universitycity: Richmond country: United States (us)
Grant 5R01CA127501-04 from National Cancer Institute
Abstract: The Angiopoietin-Tie receptor-ligand system plays a central role in both developmental as well as tumor-induced angiogenesis. Its activation during tumor progression highlights the importance of understanding the complexities of this unique signaling pathway from both a basic as well as therapeutic perspective. In agreement, it has recently been proposed that therapies targeting the tumor vasculature may be less toxic and more efficacious than conventional cancer treatments. Therefore, to gain a more accurate molecular understanding of the events surrounding angiopoietin signaling and to develop the potential for identifying novel therapeutic strategies, we have set forth the following goals for this proposal i) determine the structural principles behind receptor clustering and activation by examining a full length angiopoietin-Tie2 signaling complex, ii) Examine the functional role for Tie1 in angiopoietin signaling, specifically testing the hypothesis that Tie1 functions as a co-receptor for Tie2 signaling and that complementary electrostatic surfaces identified in our Tie2 structure, and Tie1 homology model, allow them to associate, iii) determine the role of alpha5beta1 integrin in angiopoietin signaling by biochemically characterizing the interactions of the angiopoietins and Tie2 with the alpha5beta1 integrin receptor, and iv) identify chemical tools, including small peptides, that will inhibit the formation and signaling of an angiopoietin-Tie2 complex, a prerequisite to the development of Tie2-based anti-angiogenesis therapeutics. The angiopoietins and Tie2 play central roles both in developmental and tumor-induced angiogenesis. Indeed, Tie2 activation during tumor growth, development, and metastasis suggests that the ability to modulate the receptor- ligand interactions would have important medical applications. We propose to extend our present understanding of angiopoietin-Tie signaling by using the tools of X-ray crystallography combined with other biochemical and biophysical techniques to examine how these molecules function at the molecular level
Keywords: Adult; Affinity; Agreement; angiogenesis; Angiopoietin-1; Angiopoietin-2; Angiopoietins; antiangiogenesis therapy; Antibodies; base; Binding (Molecular Function); Biochemical; Biological Assay; cancer therapy; Cell surface; Cells; Chemicals; Complex; design; Development; Dissociation; Electrostatics; Equilibrium; Event; Extracellular Matrix; Family member; Fibrinogen Receptors; Fibronectin Receptors; Fluorescence Resonance Energy Transfer; Goals; Growth and Development function; Growth Factor; Homology Modeling; in vivo; inhibitor/antagonist; Integrin alpha5beta1; Integrins; Investigation; Kinetics; Length; Ligand Binding; Ligand Binding Domain; Ligands; Literature; Measures; Mediating; Mediator of activation protein; Medical; Messenger RNA; Methods; Modeling; Molecular; Mutagenesis; Neoplasm Metastasis; novel therapeutics; Organism; Orphan; Peptides; Physiological; Play; protein function; Protein Tyrosine Kinase; Proteins; receptor; Receptor Activation; receptor binding; Receptor Protein-Tyrosine Kinases; Regulation; Reporting; research study; Resolution; response; Role; Side; Signal Pathway; Signal Transduction; Structure; Surface; System; Techniques; Testing; Therapeutic; three dimensional structure; TIE-2 Receptor; tool; tumor; tumor growth; tumor progression; X-Ray Crystallography
Project start date: 2008-05-01
Project end date: 2013-03-31
Budget start date: 1-APR-2011
Budget end date: 31-MAR-2012
PFA/PA: PA-07-070
5R01CA127501-04 (2011): $251272
SUN-NESPRIN COMPLEXES IN HUMAN LAMINOPATHIES
A William
University Of Floridacity: Gainesville country: United States (us)
Grant 5R01GM084085-02 from National Institute Of General Medical Sciences
Abstract: The A- and B-type nuclear lamins are members of the intermediate filament family of proteins and represent important structural elements of the nuclear envelope (NE). The NE functions as a selective barrier between the nucleus and cytoplasm. Mutations in the A-type lamin gene, LMNA, have been linked to a variety of human disorders, often referred to as laminopathies, that include autosomal dominant Emery-Dreifuss muscular dystrophy (AD-EDMD), dilated cardiomyopathy, Dunnigan type familial lipodystrophy and Hutchinson Gilford progeria syndrome. An X-linked form of EDMD is caused by defects in another NE protein, emerin. Emerin is an integral protein of the inner nuclear membrane and is associated with the A-type lamins. Both the A-type lamins and emerin are widely expressed. It is therefore puzzling why defects in these proteins should give rise to such a bewildering array of diseases. Recent results have shown that disease- linked defects in both A-type lamins and emerin are associated with cytoskeletal changes resulting in reduced mechanical resilience of the cytoplasm in fibroblasts. We have recently defined a complex consisting of members of both the SUN and nesprin protein families that functions as a link between the NE and the cytokeleton. We have termed this the LINC complex (LInker of Nucleoskeleton and Cytoskeleton). The LINC complex is the only known connection between the A-type lamins and the cytoskeleton. We hypothesize that it is the LINC complex that mediates the cytoskeletal changes associated with mutation of the A-type lamin and emerin genes. In this way the LINC complex might have a key role in the etiology of EDMD and other laminopathies. By gaining an improved understanding of the molecular interactions involved in the development of these disorders we may be in a better position to devise novel drug- or gene-based therapeutic strategies. The A-type lamins are important components of he nuclear envelope (NE). Defects in the A-type lamin gene (LMNA) are linked to a variety of human diseases or laminopathies, which include muscular dystrophy, lipodystrophy and progeria. The goal of this proposal is to determine the mechanism by which lamin defects can cause such disorders and to test the notion that SUN and nesprin proteins of the NE are mediators of these lamin-linked pathologies
Keywords: Actins; Affect; base; Binding; Binding (Molecular Function); Biology; Body Tissues; Cardiomyopathies; Cardiomyopathy, Dilated; Causality; Cell Nucleus; Cells; Cellular Matrix; Complex; Congestive Cardiomyopathy; Coupling; Cytoplasm; Cytoskeletal System; Cytoskeleton; Defect; Development; Dilated Cardiomyopathy; Disease; disease causation; disease etiology; disease/disorder; disease/disorder etiology; Disorder; disorder etiology; Disorder of muscle, unspecified; drug/agent; Drugs; Elements; emerin; Emery-Dreifuss Muscular Dystrophy; Emery-Dreifuss Muscular Dystrophy 2; Emery-Dreifuss Syndrome; env Antigens; env Gene Products; env Polyproteins; env Protein; Envelope Protein; Epithelial; Epithelium; Etiology; Face; facial; Fibroblasts; gene product; Gene Products, env; Genes; Genetic Alteration; Genetic Change; Genetic defect; genome mutation; Goals; Hauptmann-Thannhauser Muscular Dystrophy; Human; human disease; Human, General; Hutchinson-Gilford Disease; Hutchinson-Gilford Syndrome; improved; insight; Intermediate Filaments; intracellular skeleton; Lamin A; Lamin B; Lamin Type A; Lamin Type B; Lamins; Light; Link; Lipodystrophy; Man (Taxonomy); Man, Modern; Mechanics; Mediating; Mediator; Mediator of Activation; Mediator of activation protein; Medication; member; Membrane Proteins; Membrane-Associated Proteins; Molecular Interaction; Morphogenesis; Muscle Disease; Muscle disease or syndrome; Muscle Disorders; Muscular Diseases; muscular disorder; Muscular Dystrophies; Muscular Dystrophy, Emery-Dreifuss; Muscular Dystrophy, Emery-Dreifuss Type; Muscular Dystrophy, Emery-Dreifuss, Autosomal Dominant; Mutation; Mycocardium Disease; Myocardial Diseases; Myocardial Disorder; Myocardiopathies; myocardium disorder; Myodystrophica; Myodystrophy; Myopathic Conditions; Myopathic disease or syndrome; Myopathic Diseases and Syndromes; Myopathy; Myopathy, unspecified; Normal Cell; novel; NPC; Nuclear; Nuclear Envelope; Nuclear Inner Membrane; Nuclear Lamin; Nuclear Membrane; Nuclear Outer Membrane; Nuclear Pore Complex; Nucleus; Pathology; Pharmaceutic Preparations; Pharmaceutical Preparations; Photoradiation; Play; Position; Positioning Attribute; Premature Senility Syndrome; Progeria; Protein Family; protein function; Proteins; public health relevance; resilience; Role; Scapuloilioperoneal Atrophy with Cardiopathy; social role; STA protein; Striated Muscle Tissue; Striated Muscles; Structural Protein; Surface Proteins; Syndrome; Testing; The Sun; Therapeutic; Tissues; Type V IF Protein
Project start date: 2008-07-01
Project end date: 2011-06-30
Budget start date: 1-JUL-2009
Budget end date: 30-JUN-2011
PFA/PA: PA-07-070
5R01GM084085-02 (2009): $303711
INFECTIOUS DISEASES TRAINING PROGRAM
A William, Wade Hampton Frost Professor
University Of Virginia Charlottesvillecity: Charlottesville country: United States (us)
Grant 5T32AI007046-35 from National Institute Of Allergy And Infectious Diseases
Abstract: Project Summary The Infectious Diseases Training Program at the University of Virginia is in its 29th year. The objective is to provide a rich interdisciplinary experience in infectious diseases research in order to prepare our trainees for careers as independent investigators. The cornerstone is the side-by-side education of predoctoral, M.D. and Ph.D. postdoctoral fellows. The rationale is that integration of clinical and basic science provides the varied perspectives and techniques required for creative research to combat infections. The design includes a Program Director reporting to an Executive Committee that selects predoctoral trainees from a pool of rising 2nd year graduate students, and selects postdoctoral applicants based on nominations from both mentors and the Infectious Diseases Division. Training is enriched by the monthly "ID Breakfast" for trainees chaired by the Director, graduate courses taught by Program faculty, an M.S. degree in clinical investigation (taken by all 9 fellows doing clinical investigation), seminars, research in progress and journal clubs. The 26 faculty are 100% NIH supported (median annual direct costs $648,000), interactive (65% co-published), senior (19/26 full Professors, although select "up and coming" junior faculty are included), from 7 Departments, with 20 PhDs, 10 MDs, and 1 DVM. The average mentor graduated 8 trainees in the last 10 years. Evaluation of the Program includes an annual faculty retreat and trainee questionnaire. Institutional support includes funding of 1st year predoctoral fellows, the seminar series and the Director´s effort, and construction of a new research building. NIH support is requested for 6 predoctoral, 5 M.D. and 2 Ph.D. postdoctoral fellows (an increase of 1 predoctoral position). Progress since the last funding period includes increases in applicants (95% predoctoral and 43% postdoctoral), average GRE V/Q of 600/703 and GPA of 3.50, 100% retention of trainees in the Program, and a 44% increase in mentor NIH support. Peer-reviewed original publications averaged 4 for postdoctoral and 3 for predoctoral fellows. Academic and research positions are held by 91% (20/22) of postdoctoral and 100% (16/16) of predoctoral graduates, and 8 K08 and 4 foundation fellowships have been awarded. Women constitute 50% of trainees and 23% of mentors; 13% of trainees are underrepresented minorities. Relevance The Program prepares fellows to conduct research on infectious diseases, a paramount threat to international health in the 21st century
Keywords: Communicable Diseases; Training Programs
Project start date: 1977-07-01
Project end date: 2012-06-30
Budget start date: 1-JUL-2011
Budget end date: 30-JUN-2012
5T32AI007046-35 (2011): $647616
MOLECULAR PROPERTIES OF VOLTAGE-SENSITIVE CALCIUM CHANNELS
A William, Professor And Chair
University Of Washingtoncity: Seattle country: United States (us)
Grant 5R01NS022625-26 from National Institute Of Neurological Disorders And Stroke
Abstract: P/Q-type Ca currents conducted by Cav2.1 channels are responsible for the Ca entry that initiates neurotransmitter release at most fast glutamatergic synapses. Ca entering through presynaptic Ca channels forms a local domain of high Ca concentration that activates exocytosis in the near vicinity. Therefore, synaptic vesicles must dock near presynaptic Ca channels to be efficiently released. Neurotransmitter release is dependent on the third or fourth power of the Ca current through the presynaptic Ca channels, so small changes in Ca entry have large effects on synaptic transmission. Ca-dependent facilitation and depression of synaptic transmission is an important determinant of information coding and transmission in the nervous system. Our results in the present project period have given important new insights into the function and regulation of presynaptic Ca channels in synaptic transmission and short-term synaptic plasticity. First, we have further defined the molecular mechanism for interaction of Ca channels with SNARE proteins and the regulation of that interaction by protein phosphorylation. Second, we have shown that the calmodulin-like neuronal Ca sensor (nCaS) protein VILIP-2 regulates Cav2.1 channels by interaction at the same binding site as calmodulin and CaBP1, but has a distinct set of regulatory effects. Third, we have found that N-terminal myristoylation is required for the distinct regulatory effects of CaBP1 and VILIP-2 on Cav2.1 channels and that the N-terminal lobe of these nCaS proteins confers their specificity of regulation. Fourth, we have discovered that nCaS-dependent facilitation and inactivation of Cav2.1 channels is primarily responsible for short-term facilitation and depression of synaptic transmission in transfected superior cervical ganglion (SCG) neuron synapses, providing the first insight into the molecular mechanisms responsible for short-term synaptic plasticity. Finally, we have found unexpectedly that Ca/calmodulin-dependent protein kinase II (CaMKII) regulates Cav2.1 channels by specific binding to a site on the C-terminal domain, potentially positioning the kinase for rapid response to Ca entry and phosphorylation of nearby proteins. In the next project period, we plan to build on these important advances to 1. further define the molecular mechanisms of binding and regulation of Cav2.1 channels by nCaS proteins; 2. determine the functions of nCaS proteins in short-term synaptic plasticity; 3. explore the signaling functions of CaMKII specifically bound to Cav2.1 channels; and 4. determine the functional role of presynaptic CaMKII in synaptic transmission and synaptic plasticity. These experiments will provide novel insights into the regulation of presynaptic Ca channels and the role of this regulation in short-term synaptic plasticity, an essential form of information encoding and transmission in the nervous system. Calcium channels in nerve terminals begin the process of synaptic transmission, which communicates information from one nerve to cell to another as well as to muscle and hormone-secreting cells. Failure of correct function and regulation of these calcium channels contributes to epilepsy, migraine, ataxia, and other neurological diseases. Our proposed research will provide novel insights into the regulation of these presynaptic calcium channels and their function in short-term synaptic plasticity, an essential process for normal coding and transmission of information in the nervous system and a target for neurological disease
Keywords: Affinity; Ataxia; base; Binding (Molecular Function); Binding Sites; Biochemical; Brain; C-terminal; Calcium Channel; Calcium/calmodulin-dependent protein kinase; Calmodulin; calmodulin-dependent protein kinase II; Cells; Chemosensitization; Code; Complex; Docking; Epilepsy; Exocytosis; Failure (biologic function); Family member; Fluorescence Resonance Energy Transfer; Glutamates; Health; Hormones; insight; Lobe; Measures; member; Mental Depression; Mental disorders; Methods; Migraine; Molecular; Muscle; mutant; myristoylation; N-terminal; Nerve; nervous system disorder; Nervous system structure; neurocalcin; Neuroglia; Neurons; neurotransmitter release; novel; Phosphorylation; Phosphotransferases; Positioning Attribute; presynaptic; Process; Property; Proteins; Reagent; Regulation; Relative (related person); Research; research study; response; Role; sensor; Signal Transduction; Site; SNAP receptor; Specificity; Structure of superior cervical ganglion; Surveys; Synapses; synaptic function; Synaptic plasticity; Synaptic Transmission; Synaptic Vesicles; Testing; transmission process; Tube; voltage; voltage clamp; voltage-dependent calcium channel (P-Q type)
Project start date: 1985-09-09
Project end date: 2012-07-31
Budget start date: 1-AUG-2011
Budget end date: 31-JUL-2012
PFA/PA: PA-07-070
5R01NS022625-26 (2011): $334425
DEVELOPMENT OF A HUMANIZED ANTI-CD47 ANTIBODY FOR TREATMENT OF TISSUE ISCHEMIA.
A William, Professor Of Biochemistry And Molec
Vasculox, Inc.city: Saint Louis country: United States (us)
Grant 1R41HL095172-01A1 from National Heart, Lung, And Blood Institute
Abstract: Nitric oxide (NO) is a critical regulator of cardiovascular physiology, coordinating the activities of the endothelium, the vessel wall and circulating cells to optimize the flow of blood and oxygen to tissues. Increasing the bioavailability of NO should provide therapeutic benefit in the treatment of many diseases ascribed to insufficient NO bioavailability. All current approaches are limited by a heretofore unknown regulatory system, discovered by the founders of Vasculox, that limits the effects of NO signaling in all vascular cells. Thrombospondin-1 (TSP1) and its receptor, CD47, are present in all vascular tissues and limit NO signaling, thus worsening tissue ischemia, promoting thrombosis and inflammation and exacerbating the effects of aging on the cardiovascular system. Blocking the interaction of TSP-1 and CD47 with a monoclonal antibody (mAb) prevents these limiting effects on NO signaling in both mouse and porcine models of wound healing and tissue ischemia. Vasculox was founded to bring these discoveries to the healthcare market. Given the many roles of NO in cardiovascular regulation, we anticipate that a blocking anti-CD47 mAb will have therapeutic applications in many diseases such as peripheral artery disease (PAD), ischemia-reperfusion injury, sickle cell disease, myocardial infarction, stroke, thrombosis and others. Here we plan to develop for commercialization an anti-CD47 monoclonal antibody. This STTR proposal will provide a mechanism for Dr. Frazier´s lab at Washington University School of Medicine to transfer to Vasculox, Inc. selected mAbs vs CD47 for subsequent testing and development as therapeutics. Most promising for future development is a panel of mAbs raised in the CD47-null mice to human CD47. These mAbs react with both human and mouse CD47 and will greatly facilitate development of a humanized anti-CD47 function-blocking mAb for use in clinical trials. In this phase I proposal, we plan to identify the most promising candidate mAbs for further development. In subsequent proposals, we will test these candidates in models of human cardiovascular diseases. The phase I specific aims are 1. Determine the epitope specificity, species reactivity and effect on ligand binding of mAbs. 2. Identify those mAbs with efficacy in augmenting NO signaling in mouse and human smooth muscle cells in vitro. 3. Test active candidates in the MacFarlane skin flap mouse model of tissue ischemia, and determine effective dose, pharmacokinetic parameters and side effects of mAb administration. The founder of Vasculox Inc, has discovered a regulatory receptor, CD47, that governs blood flow to all tissues of the body, blood pressure, thrombosis and other cardiovascular functions. Blocking this receptor improves wound healing, blood flow to dying tissues and other vascular parameters. Vasculox was founded to bring this new discovery to clinical practice by identifying antibodies vs CD47 to take forward into clinical development for treatment of peripheral vascular disease and other cardiovascular diseases in the future such as stroke and heart attack
Keywords: 1D8 antigen; Acute; Adverse effects; age effect; aging effect; Animals; Antibodies; Antigenic Determinants; Antigenic Surface Determinant Protein OA3; Antigenic Surface Determinant Protein OA3 Gene; Apoplexy; Area; base; Binding Determinants; Bioavailability; bioavailability of drug; Biologic Availability; Biological Availability; biological signal transduction; Bizzozero`s corpuscle/cell; Blood flow; Blood leukocyte; Blood Platelets; Blood Pressure; Blood Vessels; Body Tissues; brain attack; cardiac infarct; Cardiac infarction; Cardiovascular; Cardiovascular Body System; Cardiovascular Diseases; cardiovascular disorder; cardiovascular function; Cardiovascular Physiology; Cardiovascular system; Cardiovascular system (all sites); CD47; CD47 Antigen; CD47 Antigen (Rh-Related Antigen, Integrin-Associated Signal Transducer); CD47 Antigen (Rh-Related Antigen, Integrin-Associated Signal Transducer) Gene; CD47 gene; CD47 Glycoprotein; CD47 Glycoprotein Gene; Cell Communication and Signaling; Cell Signaling; Cells; Cerebral Stroke; cerebral vascular accident; Cerebrovascular accident; Cerebrovascular Apoplexy; Cerebrovascular Stroke; cGMP; Chimp; Chimpanzee; circulatory system; Clinical; clinical investigation; clinical practice; Clinical Trials; Clinical Trials, Unspecified; commercialization; coronary attack; coronary infarct; coronary infarction; Cyclic GMP; Deetjeen`s body; Development; diabetes; Diabetes Mellitus; dietary fruit; Disease; disease/disorder; Disorder; Dose; Drug Kinetics; EC 1.14.13.39; EDRF Synthase; Endogenous Nitrate Vasodilator; endothelial cell derived relaxing factor; Endothelium; Endothelium-Derived Growth Factor Synthase; Endothelium-Derived Relaxing Factor; Epitopes; Erectile dysfunction; experiment; experimental research; experimental study; Family suidae; Flaps; Fruit; Future; Genetic; guanosine 3`5` monophosphate; Guanosine Cyclic 3`, 5`-Monophosphate; Guanosine Cyclic Monophosphate; Guanosine, cyclic 3`, 5`-(hydrogen phosphate); Guanylyl Cyclase-Activating Factor Synthase; Hayem`s elementary corpuscle; Hb SS disease; HbSS disease; Healthcare Market; heart attack; heart infarct; heart infarction; Hemoglobin S Disease; Hemoglobin sickle cell disease; Hemoglobin sickle cell disorder; Human; Human, General; IAP Gene; IAP-50 antigen; improved; In Vitro; in vivo Model; Inflammation; INFLM; Integrin-Associated Protein; Integrin-Associated Protein Gene; integrin-associated protein IAP, human; integrin-associated protein p50; interest; intervention development; Intracellular Communication and Signaling; Ischemia; Ischemia-Reperfusion Injury; Island Flaps; Knock-out; Knockout; Knockout Mice; L-Arginine, NADPH[{..}]oxygen oxidoreductase (nitric-oxide-forming); Laser-Doppler Flowmetry; Leiomyocyte; Leukocyte Surface Antigen CD47; Leukocyte Surface Antigen CD47 Gene; Leukocytes; Ligand Binding; Ligands; Literature; Mammals, Mice; Man (Taxonomy); Man, Modern; Markets, Health Care; Marrow leukocyte; Marrow platelet; Measurement; medical schools; MER6; MER6 Gene; Mice; Mice, Knock-out; Mice, Knockout; Moab, Clinical Treatment; Modeling; Monoclonal Antibodies; Mononitrogen Monoxide; mouse model; Murine; Mus; Myocardial Infarct; Myocardial Infarction; Myocytes, Smooth Muscle; NADPH-Diaphorase; National Institutes of Health; National Institutes of Health (U.S.); Necrosis; Necrotic; NIH; Nitric Oxide; Nitric Oxide Signaling Pathway; Nitric Oxide Synthase; Nitric Oxide, Endothelium-Derived; Nitric-Oxide Synthetase; Nitrogen Monoxide; Nitrogen oxide; Nitrogen Protoxide; NO Synthase; Null Mouse; O element; O2 element; OA3; OA3 antigen; OA3 Gene; Organ System, Cardiovascular; OVTL3 protein, human; Oxygen; Pan; Pan Genus; Pan Species; Patients; Peripheral Angiopathies; Peripheral arterial disease; peripheral blood vessel disorder; Peripheral Vascular Diseases; Peripheral Vascular Disorder; Pharmacokinetics; Phase; Physiologic; Physiologic Availability; Physiological; Pigs; Platelets; Play; porcine; Pre-Clinical Model; Preclinical Models; preclinical study; prevent; preventing; Procedures; Process; public health relevance; Pulmonary Hypertension; receptor; Receptor Protein; Regulation; Reperfusion Damage; Reperfusion Injury; research study; Reticuloendothelial System, Leukocytes; Reticuloendothelial System, Platelets; Role; Safety; Sickle Cell Anemia; sickle cell disease; sickle disease; sicklemia; side effect; Signal Transduction; Signal Transduction Systems; Signaling; Skin; Small Business Technology Transfer Research; Smooth Muscle Cells; Smooth Muscle Myocytes; Smooth Muscle Tissue Cell; social role; Species Specificity; stroke; Stroke; STTR; suid; Suidae; Surface Antigen Identified by Monoclonal Antibody 1D8; Surface Antigen Identified by Monoclonal Antibody 1D8 Gene; Surgical Flaps; Swine; System; System, LOINC Axis 4; Testing; THBS1; Therapeutic; therapy adverse effect; therapy development; thrombocyte/platelet; Thrombocytes; Thrombosis; Thrombospondin 1; thrombospondin-1 receptor CD47; tissue repair; Tissues; treatment adverse effect; treatment development; Treatment Side Effects; TSP; TSP-1; TSP1; United States National Institutes of Health; Universities; vascular; Vascular Accident, Brain; Vascular, Heart; Vasodilatation; Vasodilation; Vasorelaxation; Washington; white blood cell; White Blood Cells; white blood corpuscle; White Cell; Wound Healing; Wound Repair
Relevance: The founder of Vasculox Inc, has discovered a regulatory receptor, CD47, that governs blood flow to all tissues of the body, blood pressure, thrombosis and other cardiovascular functions. Blocking this receptor improves wound healing, blood flow to dying tissues and other vascular parameters. Vasculox was founded to bring this new discovery to clinical practice by identifying antibodies vs CD47 to take forward into clinical development for treatment of peripheral vascular disease and other cardiovascular diseases in the future such as stroke and heart attack
Project start date: 2009-06-15
Project end date: 2011-05-31
Budget start date: 15-JUN-2009
Budget end date: 31-MAY-2011
PFA/PA: PA-08-051
1R41HL095172-01A1 (2009): $197711
Sponsored Links Excellgen http://Excellgen.com
TARGETING MYCN PROTEIN WITH SMALL MOLECULES
A William, Professor
Children´s Hospital Los Angelescity: Los Angeles country: United States (us)
Abstract: Neuroblastoma, a tumor of peripheral neural crest origin, is a common and lethal tumor of childhood. Amplification of the transcription factor MYCN occurs commonly in children with high-risk disease. We generated a model for high-risk, neuroblastoma by directing expression of a MYCN transgene to the peripheral neural crest of genetically engineered mice, under control of the Tyrosine Hydroxylase (TH) promoter. We hypothesize that 1).Mycn protein plays a central role In high-risk MYCN-ampllfled neuroblastoma. 2). Therapies targeting Mycn will be effective in MYCN-amplified neuroblastoma. 3). Mice transgenic for TH-MYCN and deleted forp53 model high-risk, therapy-resistant neuroblastoma In relapsed patients, and wilt respond to small molecule Inhibitors targeting Mycn. In contrast to applications that propose to screen small molecules to identify one that targets a known molecular lesion, we are starting with two potent and selective phosphatidylinositol-3´kinase (PI3K) inhibitors now in clinical trials that appear ideally suited as therapy against neuroblastoma and Mycn protein. We will characterize the mechanism of action for these agents, analyzing interactions between tumor cells and cells that comprise the microenvironment. We will assess destabilization of Mycn protein, and subsequent impact on tumor burden and survival. We also propose to study and chemically modify a clinical inhibitor of Aurora kinase (AURKA) that already shows promise in neuroblastoma, to build in additional activity against both Mycn protein and drug resistant neuroblastoma. Our aims are Aim 1. To test available clinical PI3K and PI3K/mT0R inhibitors for activity against neuroblastoma and Mycn protein. We hypothesize that these compounds will be effective and safe in patients with MYCN-amplified neuroblastoma. Aim 2. To clarify additional targets and small molecules that cooperate with inhibitors of PI3K to degrade Mycn. We hypothesize 1). That scaffold-dependent and kinase dependent functions of AurkA contribute independently to the activity of this protein In neuroblastoma. 2). That DFG-out and a-C out inhibitors will disrupt both functions and will show efficacy In neuroblastoma
Keywords: 1-Phosphatidylinositol 3-Kinase; aurora kinase; aurora-A kinase; Biology; Cells; Chemistry; chemotherapy; Child; Clinical; Clinical Trials; combinatorial; Combined Modality Therapy; Disease; Drug resistance; Elements; Genetically Engineered Mouse; Goals; high risk; human FRAP1 protein; In Vitro; in vivo; inhibitor/antagonist; insight; kinase inhibitor; Lesion; Measures; Mediating; Modeling; Molecular; mTOR inhibition; mTOR Inhibitor; Mus; mutant; Mutation; MYCN gene; N-Myc Protein; neoplastic cell; Neural Crest; Neuroblastoma; Newly Diagnosed; novel; Patients; Pediatric Neoplasm; Peripheral; Phosphotransferases; Play; pre-clinical; Pre-Clinical Model; Promotor (Genetics); Proteins; Rattus; Relapse; Risk; Role; Safety; scaffold; Scaffolding Protein; Signal Transduction; small molecule; Solid Neoplasm; Testing; therapy resistant; transcription factor; Transgenes; Transgenic Mice; tumor; Tumor Burden; Tyrosine 3-Monooxygenase
Relevance: Successful completion of this proposal establishes a preclinical rationale for trials in children using clinical PI3K inhibitors, and provides insights into mechanisms of action for inhibitors of PI3K and of AURKA in MYCN-amplified neuroblastoma
Budget start date: 1-JUN-2011
Budget end date: 31-MAY-2012
5P01CA081403-12_6822 (2011): $301193
TARGETING THE MULLERIAN INHIBITING SUBSTANCE PATHWAY IN GYNECOLOGIC CANCER
A William, Consultant
Mayo Cliniccity: Rochester country: United States (us)
Grant 5R01CA148747-02 from National Cancer Institute
Abstract: MISRII is a member of the TGF-2 family of receptors. MIS receptor expression is embryologically important in the regression of the m|llerian ducts in the male fetus after activation by MIS (ligand) secreted by the testis. This tissue-specific pattern of MISRII expression in gynecologic tissues persists in the adult. We further show this specific pattern of expression persists in the majority of gynecologic cancers supporting its relevance as a specific target of anti-cancer therapy. To that end we have been investigating the general hypothesis that MISRII is a tissue-specific receptor (target) present in gynecologic cancers which, additionally, is fundamentally linked to serine/threonine receptor-mediated pathway(s) resulting in growth inhibition and/or cell death. These concepts suggest two primary and independent therapeutic strategies 1) utilize ligand to target MISRII directly resulting in decreased proliferation or potentiation of cytotoxic therapy, and/or 2) using the MISRII receptor as a target to specifically deliver therapy or to improve imaging, either of which would be independent of downstream signaling. Despite the appeal of this approach prior investigations have suffered from key limitations (i) lack of delineation of the relevant T1R and downstream signaling components necessary for MIS activity; (ii) lack of attempt to interpret MIS response with expression of relevant T1Rs; (iii) expression studies based on continuous cancer cell lines fail to reflect the expression pattern in primary cancers; (iv) lack of models to adequately assess rhMIS bioactivity in cancer cell model. This information is vital for rational design and application of any MIS-based strategy absent this, meaningful interpretation of prior in vitro studies using MIS is impossible. In this revised application we show preliminary data and experiments designed to address these limitations. Based on our work with other TGF-2 family receptors we have designed experiments to determine the role(s) and status of T1R and T2R in MIS signaling (Aim 1). The required receptor complexes regulating Smad signaling and cell growth inhibition can will then be tested in vitro in primary cancer cultures (Aim 2) and allow us to define the specific parameters in which MIS directed therapy may be effective. Finally, we will pursue the alternative therapeutic strategy which is not predicated upon MIS- dependent signaling using MISRII as a highly specific target to bring other therapeutic agents or imaging molecules to specific gynecologic cancers (Aim 3). Collectively, this application will provide the necessary insight into the biology of MIS signaling in gyncecologic cancers, and provide the essential data for design and interpretation of human trials targeting MISRII. Relevance to the public health (lay language) Management of advanced ovarian and uterine cancers is inadequate and most patients with advanced disease will die of their cancer. To make progress in the management of these cancers, novel and specific targets that distinguish cancer cells from normal cells are needed. MISRII is a receptor present specifically in the majority of these cancers. This provides a unique target that distinguishes the cancer cells from normal cells. Some evidence suggests that activating this receptor by exposing it to its hormone (MIS) results in growth arrest and increased sensitivity to chemotherapy. While some work has been done in this field it has been limited by (i) key gaps in knowledge of the underlying signaling pathways needed for activation; and (ii) lack of appropriate models to test outcomes in human cancers. We will determine the conditions necessary for this response and test the ability to treat human cancers with MIS. A second, non-overlapping approach will target the receptor directly as a ´bait´ to deliver toxic agents to the cancer cells. Such specific delivery would increase treatment effect and decrease associated toxicities in normal tissues. This projects brings together two investigators with distinctly different areas of expertise to address both the fundamental biologic aspects of the pathway and the translational aspects of targeting this receptor in cancer
Keywords: 21+ years old; Activin Receptor-Like Kinases, Type I; Activin Receptors, Type I; Address; Adult; adult human (21+); advanced disease; Affect; AMH receptor; AMH Type II Receptor; AMHR; AMHR2; animal data; Anti-Cancer Agents; anti-mullerian factor; anti-mullerian hormone; Anti-Mullerian Hormone Receptor; Anti-Mullerian Hormone Receptor Type II; Anti-Tumor Agents; Anti-Tumor Drugs; Antibodies; anticancer agent; anticancer drug; anticancer therapy; antimullerian hormone; Antineoplastic Agents; Antineoplastic Drugs; Antineoplastics; Antiproliferative Agents; Antiproliferative Drugs; Area; base; Binding; Binding (Molecular Function); Biologic Therapy; Biological; Biological Response Modifier Therapy; biological signal transduction; Biological Therapy; Biology; biotherapeutics; biotherapy; Body Tissues; cancer cell; Cancer cell line; Cancer Drug; Cancer of the Ovary; cancer therapy; Cancer Treatment; Cancers; Cause of Death; Cell Communication and Signaling; Cell Culture Techniques; Cell Death; cell growth; Cell model; Cell Signaling; Cells; Cellular Expansion; Cellular Growth; Cellular model; Chemosensitization; Chemosensitization/Potentiation; Chemotherapeutic Agents, Neoplastic Disease; chemotherapy; Chemotherapy-Hormones/Steroids; Complex; Connective Tissue Sarcoma; cytotoxic; Cytotoxic agent; Cytotoxic Chemotherapy; Cytotoxic drug; Cytotoxic Therapy; Data; design; designing; Development; Dose-Limiting; Duct; Duct (organ) structure; effective intervention; Elements; Endocrine Gland Secretion; Endometrial; Epithelial ovarian cancer; experiment; experimental research; experimental study; Family; Female; Female Reproductive Cancer; fetal; Fetus; Funding; G protein-coupled receptor TR2; Generalized Growth; Genital System, Male, Testis; Goals; Growth; Gynecologic; Gynecologic Cancer; Hormones; Human; Human, Adult; Human, General; Image; imaging; improved; In Vitro; in vitro testing; in vivo; Individual; Inhibition of Apoptosis; insight; Intervention; Intervention Strategies; intervention therapy; interventional strategy; Intracellular Communication and Signaling; Investigation; Investigators; Knowledge; L-Serine; L-Threonine; Language; Ligands; Link; Locales; male; malignancy; Malignant Cell; Malignant Female Reproductive System Neoplasm; Malignant Gynecologic Neoplasm; Malignant Gynecologic Tumor; Malignant neoplasm of ovary; Malignant Neoplasm of the Uterus; Malignant Neoplasm Therapy; Malignant Neoplasm Treatment; Malignant Neoplasms; Malignant Ovarian Neoplasm; Malignant Ovarian Tumor; Malignant Soft Tissue Neoplasm; Malignant Tumor; Malignant Tumor of the Female Reproductive System; Malignant Tumor of the Ovary; Malignant Tumor of the Soft Tissue; Malignant Tumor of the Uterus; Malignant Uterine Neoplasm; Malignant Uterine Tumor; Mammals, Mice; Man (Taxonomy); Man, Modern; Measures; Mediating; member; Membrane; membrane structure; Methylation; Mice; Minority; MIS; MIS Type II Receptor; MISRII; Modeling; Molecular Interaction; Mother Cells; mouse model; MRII; Muellerian inhibiting hormone; Mullerian duct inhibiting substance; Mullerian Ducts; mullerian inhibiting substance; mullerian regression factor; Mullerian-inhibiting factor; Mullerian-inhibiting factor receptor; mullerian-inhibiting hormone; Mullerian-inhibiting substance receptor; Mullerian-inhibiting substance type II receptor; mullerian-inhibitory substance; Murine; Mus; Nature; necrocytosis; neoplasm/cancer; Normal Cell; Normal Tissue; Normal tissue morphology; novel; ontogeny; Operation; Operative Procedures; Operative Surgical Procedures; Outcome; Ovarian; ovarian cancer; Ovarian Tissue; Paramesonephric Duct; pathway; Pathway interactions; Patients; Pattern; Population; Potentiation; progenitor; Progenitor Cells; programs; Programs (PT); Programs [Publication Type]; Promoter; Promoters (Genetics); Promotor; Promotor (Genetics); Property; Property, LOINC Axis 2; Protein Methylation; Public Health; public health medicine (field); public health relevance; Publications; Publishing; receptor; receptor expression; Receptor Protein; Receptor Signaling; receptor, MIS; receptor, T2R; Recurrence; Recurrent; reproductive; Research Personnel; Research Specimen; research study; Researchers; Resistance; resistant; response; Role; sarcoma; Sarcoma of the Soft Tissue and Bone; Sarcoma, Soft Tissue; Scientific Publication; Serine; Signal Pathway; Signal Transduction; Signal Transduction Systems; Signaling; social role; Specificity; Specimen; Staging; stem; stem cell population; Stem cells; Structure of paramesonephric duct; surgery; Surgical; Surgical Interventions; Surgical Procedure; System; System, LOINC Axis 4; T2R taste receptors; Testicles; Testing; Testis; Therapeutic; Therapeutic Agents; Therapeutic Effect; therapeutic efficacy; Therapeutic Hormone; Therapeutic Intervention; therapeutically effective; Threonine; Tissue Growth; Tissues; Toxic effect; Toxicities; TR2 gene product (taste-specific); trafficking; treatment effect; Treatment Efficacy; Tumor-Specific Treatment Agents; Type I Activin Receptors; United States; Uterine Cancer; Uterus Cancer; Woman; Work; Xenograft Model
Relevance: Relevance to the public health (lay language): Management of advanced ovarian and uterine cancers is inadequate and most patients with advanced disease will die of their cancer. To make progress in the management of these cancers, novel and specific targets that distinguish cancer cells from normal cells are needed. MISRII is a receptor present specifically in the majority of these cancers. This provides a unique target that distinguishes the cancer cells from normal cells. Some evidence suggest that activating this receptor by exposing it to its hormone (MIS) results in growth arrest and increased sensitivity to chemotherapy. While some work has been done in this field it has been limited by: (i) key gaps in knowledge of the underlying signaling pathways needed for activation; and (ii) lack of appropriate models to test outcomes in human cancers. We will determine the conditions necessary for this response and test the ability to treat human cancers with MIS. A second, non-overlapping approach will target the receptor directly as a ´bait´ to deliver toxic agents to the cancer cells. Such specific delivery would increase treatment effect and decrease associated toxicities in normal tissues. This projects brings together two investigators with distinctly different areas of expertise to address both the fundamental biologic aspects of the pathway and the translational aspects of targeting this receptor in cancer
Project start date: 2010-07-01
Project end date: 2014-12-31
Budget start date: 1-JAN-2011
Budget end date: 31-DEC-2011
PFA/PA: PA-07-070
5R01CA148747-02 (2011): $294505
A William, Professor
University Of California San Franciscocity: San Francisco country: United States (us)
Grant 5R01CA133091-03 from National Cancer Institute
Abstract: Medulloblastoma, a tumor of the central nervous system, is a common and frequently lethal tumor of childhood. Abnormalities in Sonic Hedgehog (SHH) and Wingless signaling contribute to genetics, however these pathways are represented in a minority of tumors. The proto-oncogene Mycn is the major factor driving proliferation in the developing cerebellum, and is expressed at high levels in SHH-associated medulloblastoma. A role for Mycn in the pathogenesis of medulloblastoma in the absence of SHH aberrations is supported by three independent studies showing a majority of human medulloblastomas express Mycn, whereas Mycn is not expressed in normal cerebellum after fetal stages. We hypothesize that aberrant expression of Mycn contributes to the pathogenesis of medulloblastoma, and that targeted therapies against Mycn will show efficacy in this disease. Our long term objectives are to clarify the role of Mycn in the initiation and progression of medulloblastoma, and to determine the impact of therapies directed against Mycn in murine and human medulloblastoma. We will characterize a genetically engineered mouse (GEM) model for medulloblastoma developed in our laboratory that co-expresses both Mycn and luciferase in the CNS, and that differs from existing GEM models for medulloblastoma in that p53 mutation (which is uncommon in human tumors) is not required to achieve high penetrance. Mycn is stabilized through activation of the lipid kinase PI3K. We have shown that small molecule inhibitors of PI3K lead to degradation of this oncoprotein in-vitro and in-vivo. We will therefore test the role of Mycn in maintaining tumors and in supporting tumor vasculature by treating MYCN-driven medulloblastomas using doxycyline to turn off MYCN transcriptionally, or using small molecule PI3K) inhibitors to degrade Mycn. We will compare genetic and epigenetic abnormalities between murine and human tumors, including microRNA profiling, and we will treat cultured and xenotransplated tumorspheres derived from primary human medulloblastomas to assess the importance of Mycn as a therapeutic target in human medulloblastoma. Aim 1. Evaluation of mice transgenic for Glt1-tTATRE-MYCN/Luc as a model for human medulloblastoma. Aim 2. Preclinical testing of isoform-selective PI3K inhibitors in mice transgenic for Glt1-tTATREMYCN/Luc, and in human medulloblastoma tumorspheres. Aim 3. Use of PI3K inhibitors to address a role for Mycn and microRNA targets regulated by Mycn, in maintaining medulloblastoma angiogenesis in vivo. Medulloblastoma is a common and frequently lethal tumor of children, for which current therapies are often ineffective. The transcription factor Mycn is expressed in the majority of these tumors, an event we hypothesize is linked to tumor formation. The long term objective of this application is to characterize a mouse model for medulloblastoma developed in our laboratory and driven by MYCN, to clarify the importance of Mycn as a therapeutic target in this disease. We have also identified a small molecule inhibitor of MYCN, that will be tested in this and derivative models. This study will clarify the importance of Mycn as a therapeutic target in medulloblastoma, and will evaluate a Mycn inhibitor that could potentially be translated to children with this important tumor
Keywords: 0-11 years old; 2-Naphthacenecarboxamide, 4-(dimethylamino)-1, 4, 4a, 5, 5a, 6, 11, 12a-octahydro-3, 5, 10, 12, 12a-pentahydroxy-6-methyl-1, 11-dioxo-, (4S-(4alpha, 4aalpha, 5alpha, 5aalpha, 6alpha, 12aalpha))-; Address; Age; Agreement; alpha-6-Deoxyoxytetracycline; Anaplasia; Anaplastic Change; Anaplastic Lesion; angiogenesis; Apoptosis; Apoptosis Pathway; Automobile Driving; Autopsy; biological signal transduction; Biology; Blood Vessels; Brain; Brain Neoplasia; Brain Neoplasms; Brain Tumors; c myc; c-myc Genes; c-ONC; cancer progression; Cancer Staging; Cancers; Catalytic Core; Catalytic Domain; Catalytic Region; Catalytic Site; Catalytic Subunit; Cell Communication and Signaling; Cell Death, Programmed; Cell Signaling; Cellular Oncogene; Central Nervous System Neoplasms; Cerebellum; Characteristics; Child; Child Youth; Childhood Neoplasm; Childhood Tumor; children; Children (0-21); Clinical; CNS neoplasm; CNS Tumor; Collaborations; Development; Diagnostic Neoplasm Staging; Disease; disease/disorder; Disorder; Doxycycline; driving; Drivings, Automobile; EC 2.7; Encephalon; Encephalons; Epigenetic; Epigenetic Change; Epigenetic Mechanism; Epigenetic Process; Erinaceidae; Evaluation; Event; fetal; Gene Expression; gene product; Genes; Genes, p53; Genetic; Genetic Alteration; Genetic Change; Genetic defect; Genetically Engineered Mouse; genome mutation; Genomics; Health; heavy metal lead; heavy metal Pb; Hedgehogs; Heterograft; high risk; Human; human disease; Human, Child; Human, General; Immunologic, Luciferase; In Vitro; in vivo; inhibitor; inhibitor/antagonist; Intracellular Communication and Signaling; Isoforms; Kinases; Laboratories; language translation; Lead; Link; Lipids; Luciferases; Maintenance; Maintenances; malignancy; Malignant Neoplasms; Malignant Tumor; Mammals, Mice; Man (Taxonomy); Man, Modern; Measures; Mediating; medulloblastoma; Mice; Micro RNA; MicroRNAs; Minority; miRNA; Modeling; mouse model; Murine; Mus; Mutation; MYCN; MYCN gene; Natural History; Natural regeneration; necropsy; neoplasm progression; Neoplasm Staging; neoplasm/cancer; neoplastic progression; Nervous System, Brain; NMYC; NMYC Gene; Oncogene Products; Oncogene Proteins; Oncogenesis; Oncoproteins; Outcome; P53; Pathogenesis; pathway; Pathway interactions; Patients; Pb element; Pediatric Neoplasm; Pediatric Tumor; Penetrance; Phosphotransferases; Play; postmortem; Preclinical Testing; Protein Isoforms; Proteins; Proto-Oncogenes; protooncogene; regenerate; Regeneration; Role; Safety; Signal Transduction; Signal Transduction Systems; Signaling; small molecule; social role; Staging; System; System, LOINC Axis 4; T-Stage; Testing; Tet; Tetanus Helper Peptide; therapeutic target; Toxic effect; Toxicities; TP53; TP53 gene; transcription factor; Transgenic Mice; Translating; Translatings; Transphosphorylases; Transplantation, Heterologous; TRP53; tumor; tumor initiation; tumor progression; Tumor Protein p53 Gene; Tumor stage; Tumor Staging; tumorigenesis; tumors in the brain; tumors in the central nervous system; Tumors, Central Nervous System; v-myc Avian Myelocytomatosis Viral Oncogene Cellular Homolog; vascular; Vibramycin; WNT signaling; WNT Signaling Pathway; Xenograft; Xenograft procedure; Xenotransplantation; youngster
Relevance: Narrative: Medulloblastoma is a common and frequently lethal tumor of children, for which current therapies are often ineffective. The transcription factor Mycn is expressed in the majority of these tumors, an event we hypothesize is linked to tumor formation. The long term objective of this application is to characterize a mouse model for medulloblastoma developed in our laboratory and driven by MYCN, to clarify the importance of Mycn as a therapeutic target in this disease. We have also identified a small molecule inhibitor of MYCN, that will be tested in this and derivative models. This study will clarify the importance of Mycn as a therapeutic target in medulloblastoma, and will evaluate a Mycn inhibitor that could potentially be translated to children with this important tumor. (
Project start date: 2009-01-01
Project end date: 2013-12-31
Budget start date: 1-JAN-2011
Budget end date: 31-DEC-2011
PFA/PA: PA-07-070
5R01CA133091-03 (2011): $288529
CELLULAR AND GENETIC BASIS OF ANAPLASTIC MEDULLOBLASTOMA
A William
Sanford-burnham Medical Research Institcity: La Jolla country: United States (us)
Grant 1R01CA159859-01 from National Cancer Institute
Abstract: Medulloblastoma (MB) is the most common malignant brain tumor in children. Patients whose tumors exhibit large cell/anaplastic (LCA) histology usually fail therapy and die from their disease. Improved approaches to treating these patients are likely to come from a deeper understanding of LCA tumors, including their aggressive growth properties and their invasive and metastatic behavior. Unfortunately, human LCA MB tissue is difficult to obtain, and existing genetically engineered mouse (GEM) models of MB rarely display anaplasia or metastasis. To address this problem, we have collected >100 human LCA MBs, including paired samples of primary/metastatic tumors. In addition, we have generated GEM and transplant-based models of LCA MB, and mobilized the transposable element Sleeping Beauty (SB) to promote anaplasia and metastasis in these models. Through analysis of our human and murine datasets, we propose to identify the cells and genes that drive progression in LCA medulloblastoma. This application brings together investigators from three institutions, with collective experience in neural development, stem cell biology and genomics of both human and murine MB. Our complementary backgrounds and expertise uniquely position us to investigate the cellular and molecular basis of LCA MB, and will ultimately allow us to develop more effective approaches to targeting these aggressive tumors. The proposed studies focus on large cell/anaplastic medulloblastoma, a highly malignant pediatric brain tumor with an extremely poor prognosis. By identifying the cells and genes responsible for the aggressive behavior of this tumor, we hope to develop more effective approaches to therapy
Keywords: Address; Age; Aggressive behavior; Anaplasia; Anaplastic Cell; Animal Model; base; Behavior; Candidate Disease Gene; Cells; Cerebellum; Cerebrospinal Fluid; Child; Childhood Brain Neoplasm; Complementary DNA; Data; Data Set; Disease; DNA Transposable Elements; Epigenetic Process; Exhibits; experience; Freezing; Gender; Gene Expression; Genes; Genetic; Genetically Engineered Mouse; genome-wide; Genomics; Growth; Histology; Human; improved; in vivo; Institution; Malignant - descriptor; Malignant neoplasm of brain; Mediating; medulloblastoma; Metastatic Neoplasm to the Leptomeninges; Methylation; Modeling; Molecular; Monitor; mouse model; mRNA Expression; Mus; Neoplasm Metastasis; neoplastic cell; neurodevelopment; Outcome; outcome forecast; Pathway interactions; Patients; Phenotype; Positioning Attribute; prevent; Prevention; Primary Neoplasm; Property; public health relevance; Research Personnel; Retroviridae; Sampling; Signaling Pathway Gene; Site; Sleeping Beauty; small hairpin RNA; stem cell biology; Subgroup; Tissues; tool; Transplantation; tumor; tumor growth
Relevance: The proposed studies focus on large cell/anaplastic medulloblastoma, a highly malignant pediatric brain tumor with an extremely poor prognosis. By identifying the cells and genes responsible for the aggressive behavior of this tumor, we hope to develop more effective approaches to therapy
Project start date: 2011-04-04
Project end date: 2016-03-31
Budget start date: 4-APR-2011
Budget end date: 31-MAR-2012
PFA/PA: PA-10-067
1R01CA159859-01 (2011): $640572
MECHANISMS OF INNATE IMMUNE RESPONSE MODULATION BY MECHANICAL VENTILATION
A William, Associate Professor
University Of Washingtoncity: Seattle country: United States (us)
Grant 5R01HL086883-04 from National Heart, Lung, And Blood Institute
Abstract: Mechanical ventilation is frequently used to support critically ill patients even in the absence of overt lung injury. A recent study, however, has linked exposure to mechanical ventilation with subsequent development of the acute respiratory distress syndrome (ARDS). The mechanism linking mechanical ventilation with initiation of lung injury is unknown. We have shown that mechanical ventilation can alter the transcriptional response to low-dose bacterial product exposure leading to excessive inflammation and early development of lung injury. Our long-term goal is to identify how mechanical ventilation modulates the transcriptional response to microbial pathogens as a first step toward developing specific interventions to improve survival in mechanically ventilated patients. The specific hypothesis for this proposal is that mechanical ventilation activates the activator protein-1 (AP-1) transcription factor, which augments gene transcription in response to pathogen-associated molecular pattern (PAMP) recognition by toll-like receptors (TLRs). We base this hypothesis on the observations that A) mechanical ventilation preferentially induces transcription of genes with a promoter region containing the AP-1 binding sequence, B) mechanical ventilation causes nuclear translocation of AP-1, C) mechanical ventilation synergistically augments mRNA levels of genes transcribed in response to lipopolysaccharide (LPS), a cell wall component of gram-negative bacteria, and D) inhibition of c-Jun amino-terminal kinase (JNK - a kinase activator of AP-1) is protective against other types of lung injury. The specific aims of this proposal are to 1) Determine whether ventilator augmentation of PAMP-induced inflammation is (a) specific for individual TLR- signaling pathways and/or (b) dependent on generation of endogenous TLR ligands. This aim is accomplished through the combination of highly specific toll-like receptor agonists and genetically modified mice, which lack either the MyD88 or TRIF adapter protein necessary for specific TLR signaling. 2) Determine the lung cell type primarily responsive to ventilator-induced immunomodulation by (a) measuring response to TLR ligands and ventilation in chimeric mice, which lack MyD88 in either myeloid or non- myeloid cells and (b) measuring response to TLR ligands and ventilation in transgenic mice in which distal lung epithelial cells have an isolated defect in downstream TLR signaling pathways. 3) Determine the role of AP-1 activation in ventilator-augmentation of LPS-induced inflammation by (a) using chromatin immunoprecipitation assay to identify which protein subunits of AP-1 are recruited to target gene promoters by mechanical ventilation, (b) inhibiting AP-1 activation during mechanical ventilation and LPS exposure, and (c) using RNA interference to determine the function of specific AP-1 subunits identified in (a). Relevance for public health - The incidence of acute lung injury (ALI) is 78.9 per 100,000 patient-years with a mortality rate of 38.5%. Based on these values, there are an estimated 190,600 cases of ALI in the U.S. each year with 74,500 associated deaths. Additionally, the economic impact of ALI is high secondary to prolonged ICU care and overall length of hospitalization and to chronic post-hospitalization disability. These data highlight the importance of further research to help understand the factors, which promote the development of ALI. This project will help identify mechanisms by which mechanical ventilation support causes lung injury; thereby, providing guidance for the development of future interventions to decrease the incidence and severity of acute lung injury
Keywords: activating transcription factor; Acute; adapter protein; Adult Respiratory Distress Syndrome; Agonist; Alveolar; Attenuated; Bacteremia; base; Binding (Molecular Function); Biological Assay; Caring; cell type; Cell Wall; Cessation of life; chemokine; chromatin immunoprecipitation; Chronic; Critical Illness; Data; Defect; design; Development; disability; Distal; DNA Binding; Dose; economic impact; Environmental air flow; Epithelial Cells; Exposure to; Future; Gene Targeting; Generations; Genes; Genetic Transcription; Goals; Gram-Negative Bacteria; Hospitalization; human TLR3 protein; Immune response; immunoregulation; improved; in vivo; Incidence; Individual; Inflammation; Inflammatory Response; injured; Interferons; Interleukin-1; Intervention; JNK-activating protein kinase; JUN gene; Knowledge; Lead; Length; Ligands; Link; Lipopolysaccharides; Lung; lung injury; Lung Injury, Acute; MAPK8 gene; Measures; Mechanical ventilation; Messenger RNA; microbial; Mitogen-Activated Protein Kinases; Modeling; Molecular; Mortality Vital Statistics; mouse model; Mus; Myelogenous; Myeloid Cells; novel; Nuclear Translocation; pathogen; Pathogenesis; Pathway interactions; Patients; Pattern; Pattern Recognition; Phosphotransferases; Pneumonia; Principal Investigator; programs; Promoter Regions (Genetics); Promotor (Genetics); Protein Subunits; public health relevance; Receptor Signaling; Recruitment Activity; Reporting; Research; research study; response; RNA Interference; Role; Safety; Secondary to; Severities; Signal Pathway; Signal Transduction; Source; Specificity; standard of care; Testing; Tissue-Specific Gene Expression; Toll-Like Receptor 2; toll-like receptor 4; Toll-like receptors; transcription factor; Transcription Factor AP-1; Transgenic Mice; Ventilator; Ventilator-induced lung injury
Project start date: 2008-02-01
Project end date: 2013-01-31
Budget start date: 1-FEB-2011
Budget end date: 31-JAN-2012
PFA/PA: PA-07-070
5R01HL086883-04 (2011): $390000
3R01HL086883-02S1 (2009): $257006
REGULATION OF CARDIAC CALCIUM CHANNELS BY AN AUTOINHIBITORY SIGNALING COMPLEX
A William, Professor And Chair
University Of Washingtoncity: Seattle country: United States (us)
Grant 5R01HL085372-05 from National Heart, Lung, And Blood Institute
Abstract: L-type Ca currents conducted by Cav1.2 channels are responsible for Ca entry that initiates contraction in cardiac muscle, and these channels are therefore the final common pathway for regulation of Ca signaling and contractile force by many different effectors such as neurotransmitters, hormones and drugs, and their receptors. Alterations in L-type Ca currents are crucially involved in cardiovascular disease and therapy. Misregulation of L-type Ca currents contributes to hypertension, and Ca channel antagonist drugs are an important mode of therapy. Ischemic heart disease is often accompanied by angina pectoris, which is also treated with Ca antagonist drugs. Arrhythmias can be generated by altered regulation of L-type Ca currents and by inappropriately timed Ca transients generating early and delayed afterdepolarizations, and Ca antagonist drugs are important in treatment of atrial arrhythmias, (-adrenergic regulation of L-type Ca currents is altered in heart failure. Surprisingly, despite their importance in cardiovascular physiology and pathophysiology, regulation of Cav1.2 channels in cardiac myocytes is not well understood. Because of their key role in regulation of contraction, many intracellular regulators and second messengers converge on these Ca channels and regulate their function, including Mg, cAMP, Ca, and calmodulin. Our work has shown that the sites of action of these second messengers are in the large intracellular C-terminal domain, which represents approximately 30% of the mass of the al subunit. In addition, the C-terminal domain is subject to proteolytic processing, which modulates its function. Thus, the C-terminal domain integrates many kinds of cellular regulatory signals, which together form an integrated intracellular signaling network controlling Ca channel activity. In this project we propose to determine the molecular mechanism and physiological significance of Cav1.2 channel autoinhibition the C-terminal domain, define the mechanism and physiological significance of proteolytic processing of the C-terminal of Cav1.2 channels, and determine the molecular mechanism of regulation of the Cav1.2 channel by the (-adrenergic receptor pathway acting through PKA bound to the channel´s distal C-terminal domain by AKAP15. The results of our experiments will be crucial for understanding regulation of Ca and cAMP signaling in the cardiac myocyte and its dysfunction in cardiovascular disease. This information will provide the essential basic science background for translational research aimed at preventing and treating cardiovascular disease
Keywords: 1, 2-diacylglycerol; A kinase anchoring protein; adrenergic; Adrenergic Agents; Adrenergic Receptor; Amino Acids; Angina Pectoris; Arrhythmia; base; Basic Science; Binding (Molecular Function); C-terminal; Calcium; Calcium Channel; Calcium Channel Blockers; Calcium Signaling; Calmodulin; calpain 10; Cardiac; Cardiac Myocytes; Cardiovascular Diseases; cardiovascular disorder therapy; Cardiovascular Physiology; Cells; Cleaved cell; Complex; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Diglycerides; Distal; Functional disorder; Heart Atrium; Heart failure; Hormones; Hypertension; Light; Macromolecular Complexes; Magnesium; Molecular; Muscle Cells; Myocardial Ischemia; Myocardium; Neurotransmitters; Pathway interactions; Pharmaceutical Preparations; Physiological; prevent; Process; Proteolytic Processing; Published Comment; receptor; reconstitution; Regulation; research study; second messenger; Second Messenger Systems; Signal Pathway; Signal Transduction; Site; Testing; Time; Translational Research; Ventricular; Work
Project start date: 2007-05-15
Project end date: 2012-04-30
Budget start date: 1-MAY-2011
Budget end date: 30-APR-2012
5R01HL085372-05 (2011): $390000
LEPTIN REGULATION OF INTESTINAL INFLAMMATION AND INFECTION
A William, Wade Hampton Frost Professor
University Of Maryland Baltimorecity: Baltimore country: United States (us)
Abstract: Hypothesis Leptin is an important regulator of the intestinal inflammatory response to infection, and does so through its effects on either hematopoietic or non-hematopoietic cells of the gut. Leptin and leptin receptor are expressed in the intestinal epithelium and in infiltrating mononuclear cells in the lamina propria. Leptin controls expression of epithelial sodium/glucose and peptide transporters, regulates apoptosis, and induces intestinal inflammation via T lymphocytes. We and others have observed that mice deficient in leptin (ob/ob) or functional leptin receptor (db/db) have altered susceptibility to amebiasis and Clostridium difficile, as well as experimentally-induced inflammatory bowel disease. These results suggest that leptin may have broad regulatory roles in enteric infection and inflammation. Intriguingly, a common (present in one half of the CephUtah population analyzed by HapMap) single amino acid polymorphismm in the leptin receptor (that alters its affinity for leptin) is associated with resistance in children to amebiasis, and in adults to amebic liver abscess. We propose to study the mechanisms by which leptin and its receptor regulate intestinal defense against infection in mice and in humans. First we will test how general the observation is of the link of leptin to intestinal infection and inflammation. With collaborating investigators within MARGE we will determine if ob/ob (leptin deficient) and db/db (leptin receptor deficient) mice have increased susceptibility to infection and inflammation due to Giardia lamblia, Cryptospordium pan/urn, enteroaggregative E. coli and C. difficile. We will then determine the contribution to infection and inflammation of leptin and leptin receptor in intestinal epithelium vs. bone marrow-derived cells for a single infectious agent, and its dependency upon STATS signaling. Finally we will extend these observations to humans by testing for associations of protection from Giardia lamblia and enteroaggregative E. coli with the leptin receptor polymorphism found to be protective from amebiasis
Keywords: 0-11 years old; 21+ years old; Abscess, Hepatic, Amebic; Adult; adult human (21+); Affinity; Amebiasis; Amebic Liver Abscess; Amino Acids; aminoacid; Apoptosis; Apoptosis Pathway; biodefense; biological signal transduction; Body Tissues; Bone Marrow; Bone Marrow Transplant; Bone Marrow Transplantation; bowel; C. difficile; C. parvum; C.difficile; Cell Communication and Signaling; Cell Death, Programmed; Cell Signaling; cell type; Cells; Child; Child Youth; children; Children (0-21); Clostridium difficile; Cryptosporidium; Cryptosporidium parvum; D-Glucose; db/db mouse; Dependency; Dependency (Psychology); Dextrose; E. histolytica; EAEC; EAggEC; Effector Cell; emotional dependency; Endamoeba histolytica; Entamoeba histolytica; Enteral; Enteric; enteroaggregative E. coli; enteroaggregative Escherichia coli; Epithelial; Epithelium, Intestinal; experiment; experimental research; experimental study; Foundations; Genetic Polymorphism; Giardia; Giardia intestinalis; Giardia lamblia; Glucose; Grafting, Bone Marrow; Gut Inflammation; Hematopoietic; Hepatic Amebiasis; host response; HuB219; Human; Human, Adult; Human, Child; Human, General; Immune response; immunoresponse; Infection; Infectious Agent; infectious organism; Inflammation; Inflammatory Bowel Diseases; Inflammatory Bowel Disorder; Inflammatory disease of the intestine; Inflammatory Diseases of the Intestinal Tract; Inflammatory disorder of the intestine; Inflammatory Intestinal Disease; Inflammatory Intestinal Disorder; Inflammatory Response; INFLM; Intestinal; intestinal epithelium; Intestinal Inflammation; Intestines; Intracellular Communication and Signaling; Investigators; Lamblia; Lamblia intestinalis; Lamina Propria; LEP-R; LEPR; LEPR Protein; Leptin; leptin receptor; Leptin receptor mutation; leptin-binding protein; Link; Liver Abscess, Amebic; Mammals, Mice; Man (Taxonomy); Man, Modern; Marrow Transplantation; Maryland; Mediating; Mice; Modeling; Mononuclear; Murine; Mus; Na element; new approaches; novel; novel approaches; novel strategies; novel strategy; Ob Gene Product; Ob Protein; OB Receptor; OB-R; ob/ob mouse; Obese Gene Product; Obese Protein; pathogen; Peptides; polymorphism; Polymorphism (Genetics); Polymorphism, Genetic; Population; Predisposition; programs; Programs (PT); Programs [Publication Type]; receptor; Receptor Protein; Regulation; Research Personnel; research study; Researchers; Resistance; resistant; response; Reticuloendothelial System, Bone Marrow; Role; Signal Pathway; Signal Transduction; Signal Transduction Systems; Signaling; social role; Sodium; STAT3; STAT3 gene; Structure of intestinal epithelium; Susceptibility; T-Cells; T-Lymphocyte; Testing; thymus derived lymphocyte; Thymus-Dependent Lymphocytes; Tissues; Universities; Virginia; Work; youngster
Relevance: Successful completion of these studies will provide (1) an understanding of the role of leptin in the intestinal response to infection; (2) a mechanistic understanding of how leptin acts in the gut; and (3) the extent to which common genetic polymorphisms in the leptin signaling pathway sensitize humans to enteric infection. Novel management of enteric infection and inflammation via modulation of leptin is a promise of this work
Budget start date: 1-MAR-2011
Budget end date: 29-FEB-2012
5U54AI057168-08_5604 (2011): $313800
SHORT-TERM RESEARCH EDUCATION PROGRAM TO INCREASE DIVERSITY IN HEALTH-RELATED RES
A William, Associate Dean For Multicultur
University Of Chicagocity: Chicago country: United States (us)
Grant 5R25HL096383-03 from National Heart, Lung, And Blood Institute
Abstract: For the past 15 years, the University of Chicago Pritzker School of Medicine has been strongly committed to increasing the diversity within the biomedical research workforce. Minority and disadvantaged scientists are particularly underrepresented in modern clinical and basic science research and few such individuals are encouraged to embark upon academic careers. Early exposure to research opportunities through the Short Term Research Education Program to Increase Diversity in Health-Related Research provides a mechanism to enrich the experiences of minority and disadvantaged students at the college and medical student level to establish a foothold in science that may provide the underpinnings for further career development in biomedical research. Support is requested to fund an 8-week undergraduate program in research focused on the diseases and treatments thereof of the heart, blood vessels, blood, and lungs. Minority and disadvantaged students will be recruited from a national pool and an online application will be used to collect data for evaluation of candidates by the PSOMER Steering Committee. The selected participants will be of exceptional scholastic ability and motivation for biomedical research. Support is also requested to fund a 12-week medical student program in research in the scientific areas mentioned above. Minority and disadvantaged medical students enrolled at Pritzker who express an interest in considering an academic medicine career will be selected for a research opportunity by the Pritzker Summer Research Steering Committee on the basis of the merit of the research proposal, strength of the mentor, and the soundness of the research plan to be accomplished in the time available to the proposed project. The goals of both programs are to help to initiate careers in biomedicine among minority and disadvantaged students, and thus a rigorous tracking plan is to be implemented to evaluate the success of the program. Continuation of the mentored environment at the University of Chicago is fostered through the association of the programs with the Bowman Society at Pritzker. The aim of this measure is to support the development of inclined minority individuals to pursue academic career pathways. It meets bimonthly and has the potential for nurturing participants who have been exposed to biomedical research early in their careers in medicine. Pipeline programs that give an early introduction to motivated and well-prepared minority and disadvantaged students have the potential to address the lack of diversity in biomedical research. Two programs supported by the NHLBI Short Term Research Education Program to Increase Diversity in Health-Related Research conducted at the University of Chicago will provide undergraduate and medical students with the opportunity to become involved in high-quality, hypothesis-driven research in hopes of fostering their desire for pursuit of academic/research careers. (End of )
Keywords: ing; Address; Area; base; Basic Science; Biomedical Research; Blood; Blood Vessels; career; career development; Chicago; Clinical Sciences; Commit; Data; Development; Disadvantaged; Disease; Educational aspects; Enrollment; Environment; Evaluation; experience; Exposure to; Fostering; Funding; Goals; Health; Heart; Individual; interest; Lung; Measures; medical schools; Medical Students; Medicine; meetings; Mentors; Minority; Motivation; National Heart, Lung, and Blood Institute; Participant; Pathway interactions; programs; Recruitment Activity; Research; Research Proposals; Science; Scientist; Societies; Students; success; Time; Universities; university student
Relevance: Pipeline programs that give an early introduction to motivated and well-prepared minority and disadvantaged students have the potential to address the lack of diversity in biomedical research. Two programs supported by the NHLBI Short Term Research Education Program to Increase Diversity in Health-Related Research conducted at the University of Chicago will provide undergraduate and medical students with the opportunity to become involved in high-quality, hypothesis-driven research in hopes of fostering their desire for pursuit of academic/research careers
Project start date: 2009-05-01
Project end date: 2014-04-30
Budget start date: 1-MAY-2011
Budget end date: 30-APR-2012
PFA/PA: RFA-HL-08-016
5R25HL096383-03 (2011): $112163
VOLTAGE SENSITIVE SODIUM CHANNELS IN BRAIN
A William, Professor And Chair
University Of Washingtoncity: Seattle country: United States (us)
Grant 5R01NS015751-30 from National Institute Of Neurological Disorders And Stroke
Abstract: Electrical signaling is a nearly universal process in cell physiology, initiating and controlling movement, secretion, enzyme activity, and gene expression. Electrical excitability in most vertebrate cells depends on voltage-gated sodium (Na) channels, which open briefly in response to depolarization to allow rapid influx of Na and depolarize the cell to begin the action potential. Impairment of Na channel function causes periodic paralysis, epilepsy, chronic pain, migraine, and cardiac arrhythmia. Elucidation of the molecular basis for electrical signaling is a major challenge for molecular biology and membrane biophysics. Previous studies supported by this research grant led to purification and reconstitution of the Na channel protein and determination of many of its structural and functional properties. In this project period, we have made several advances. We have probed the function of the pore-lining S6 segments in opening of the pore and in slow inactivation. We have shown that regulation by protein phosphorylation by cAMP-dependent protein kinase (PKA) and protein kinase C (PKC) requires specific Na channel/kinase signaling complexes and is dependent on regulation of the intrinsic slow inactivation process of Na channels. In addition, we have analyzed the alterations of activation gating of Na channels by gating modifier toxins, developed a precise molecular model for the function of the voltage sensors in channel activation, and discovered that mutations in the voltage sensor module cause gating pore current-a voltage-gated leak of ions through the mutant voltage sensor into the cell. Remarkably, we found that gating pore current is the pathophysiological mechanism of mutations that cause Hypokalemic Periodic Paralysis (HypoPP) in skeletal muscle. These results focus attention on voltage sensing and activation of Na channels as an important and universal mechanism of cellular excitation and control of cell function, which is often impaired in disease. In the proposed experiments, we will use structure- guided site-directed mutagenesis, ab initio molecular modeling of protein structure, disulfide-crosslinking, and high-resolution biophysical analysis of gating pore currents in order to define the molecular interactions of the gating charges during activation, develop a refined Rosetta sliding helix model for voltage sensor activation, correlate voltage sensor function in Nav1.2/domain IV with fast inactivation and correlate voltage sensor function with slow inactivation and regulation by protein phosphorylation. These studies will reveal the structural basis for voltage-dependent gating by providing a detailed functional map of the Na channel voltage sensors as they initiate channel activation, fast inactivation, slow inactivation, and regulation of slow inactivation by protein phosphorylation. As defects in voltage sensor function are implicated in inherited forms of periodic paralysis, cardiac arrhythmia, epilepsy, and chronic pain, these advances will have direct implications for understanding and treatment of widespread diseases. Sodium channels initiate electrical signals required for information processing in the nervous system, contraction of skeletal muscle and the heart, and secretion of hormones, and their normal function is impaired in periodic paralysis, epilepsy, chronic pain, and cardiac arrhythmia. This project will determine the molecular mechanism of sodium channel activation and inactivation, which are impaired in disease. This basic science information will be essential to understand and treat periodic paralysis, epilepsy, chronic pain, and cardiac arrhythmia
Keywords: Action Potentials; Activation Analysis; Adenosine Cyclic Monophosphate-Dependent Protein Kinases; Amino Acid Substitution; Analyses, Activation; Arrhythmia; Attention; base; Basic Research; Basic Science; Binding; Binding (Molecular Function); biological signal transduction; Biophysics; body movement; Brain; Calcium Phospholipid-Dependent Protein Kinase; Calcium-Activated Phospholipid-Dependent Kinase; cAMP-Dependent Protein Kinases; Cardiac Arrhythmia; Cell Communication and Signaling; Cell Function; Cell physiology; Cell Process; Cell Signaling; Cells; Cellular Function; Cellular Physiology; Cellular Process; Charge; Chemotherapy-Hormones/Steroids; chronic pain; chronic painful condition; Complex; constriction; Coupled; cross-link; crosslink; Cyclic AMP-Dependent Protein Kinases; Cysteine; Defect; Dependence; Disease; disease/disorder; Disorder; Disulfides; DNA Molecular Biology; EC 2.7; Encephalon; Encephalons; Endocrine Gland Secretion; enzyme activity; epilepsia; Epilepsy; Epileptic Seizures; Epileptics; epileptiform; epileptogenic; experiment; experimental research; experimental study; Extensive Disease; Familial Hypokalemic Periodic Paralysis; Gene Expression; gene product; Generalized Disease; Genetic Alteration; Genetic Change; Genetic defect; genome mutation; Half-Cystine; Headache, Migraine; Health; Heart; Heart Arrhythmias; Hereditary; Hormones; Hypokalemic periodic paralysis; Impairment; information processing; Inherited; Intracellular Communication and Signaling; Ion Channels, Sodium; Ions; Kinases; L-Cysteine; Maps; Measures; Membrane; membrane structure; Migraine; millisecond; Modeling; Molecular; Molecular Biology; Molecular Interaction; molecular modeling; Molecular Models; Movement; Msec; Muscle, Skeletal; Muscle, Voluntary; Mutagenesis, Site-Directed; mutant; Mutation; Na element; na(v)1.2; Nav1.2; Nervous System; Nervous system structure; Nervous System, Brain; Neurologic Body System; Neurologic Organ System; NRVS-SYS; Nucleic Acid Biochemistry, Molecular Modeling; Palsy; Paralysed; paralysis; paralytic; pathway; Pathway interactions; Peripheral Nerves; Phospholipid-Sensitive Calcium-Dependent Protein Kinase; Phosphorylation; Phosphotransferases; PKA; PKC; Plegia; Primary Hypokalemic Periodic Paralysis; Process; Property; Property, LOINC Axis 2; Protein Kinase A; Protein Kinase C; protein kinase C kinase; Protein Phosphorylation; protein structure; Protein/Amino Acid Biochemistry, Molecular Modeling; Proteins; R01 Mechanism; R01 Program; reconstitute; reconstitution; Regulation; Research Grants; Research Project Grants; Research Projects; Research Projects, R-Series; research study; Resolution; response; RPG; SCN2A protein; Seizure Disorder; sensor; Signal Transduction; Signal Transduction Systems; Signaling; Site; Site-Directed Mutagenesis; Site-Specific Mutagenesis; Skeletal muscle structure; Skeletal Muscle Tissue; Slide; Sodium; Sodium Channel; Staging; Structure; Subcellular Process; Targeted DNA Modification; Targeted Modification; Testing; Text; Therapeutic Hormone; Time; Toxin; Transphosphorylases; TXT; V (voltage); voltage; Widespread Disease
Relevance: Sodium channels initiate electrical signals required for information processing in the nervous system, contraction of skeletal muscle and the heart, and secretion of hormones, and their normal function is impaired in periodic paralysis, epilepsy, chronic pain, and cardiac arrhythmia. This project will determine the molecular mechanism of sodium channel activation and inactivation, which are impaired in disease. This basic science information will be essential to understand and treat periodic paralysis, epilepsy, chronic pain, and cardiac arrhythmia
Project start date: 1979-12-01
Project end date: 2013-12-31
Budget start date: 1-JAN-2011
Budget end date: 31-DEC-2011
PFA/PA: PA-07-070
5R01NS015751-30 (2011): $307125
Sponsored Links Excellgen http://Excellgen.com
THE ROLES OF ENDOTHELIAL PECAM AND THE LBRC IN LEUKOCYTE TRANSMIGRATION
A William, Magerstadt Professor And Chair
Northwestern Universitycity: Chicago country: United States (us)
Grant 5R01HL046849-21 from National Heart, Lung, And Blood Institute
Abstract: Diapedesis is a critical step in the inflammatory response in which leukocytes migrate across endothelial cells (EC) out of the circulation and into inflamed tissues. We have found that the molecule PECAM (CD31), expressed at the borders of endothelial cells and on leukocytes, is a critical regulator of diapedesis. We also described a novel PECAM-containing membrane compartment, the lateral border recycling compartment (LBRC), a reticulum of interconnected vesicles in continuity with the plasma membrane at the borders of endothelial cells. During diapedesis, the LBRC moves to surround the transmigrating leukocyte as it moves through the endothelial cell border. Over the past funding period, we demonstrated that this "targeted recycling" was necessary for diapedesis to proceed. Furthermore, the critical role of PECAM in transmigration relates to its presence in the LBRC. Mutations that block its ability to enter the LBRC and function in targeted recycling block diapedesis. The aims of this application are to investigate how PECAM and the LBRC regulate diapedesis, with special attention to the role of PECAM in LBRC function. During diapedesis, the junctional adhesion molecule VE-cadherin is removed from the membrane at the site of diapedesis, while the PECAM and other molecules from the LBRC are recruited. Blocking either VE-cadherin removal or targeted recycling of the LBRC blocks diapedesis. Aim 1 will test the hypothesis that removal of VE-cadherin is functionally linked to targeted recycling of the LBRC. Targeted recycling requires intact microtubules and microtubule molecular motor activity. Aim 2 will test the hypothesis that a specific microtubule molecular motor (kinesin) is required for targeted recycling of the LBRC and will identify that kinesin (or kinesins if there is more than one involved to a significant extent.) Mutations in the cytoplasmic tail of PECAM that interfere with its ability to support transmigration also interfere with its ability to enter and leave the LBRC. Since the signaling properties of the cytoplasmic tail were not affected, we believe that the effects of this mutation on transmigration were due to the effect on PECAM´s ability to function in the LBRC. Aim 3 will test the hypothesis that the cytoplasmic tail of PECAM contains a localization sequence critical for entry into the LBRC. While we are dissecting the role of PECAM in diapedesis in these in vitro studies, we will study the role of PECAM in diapedesis in vivo. Aim 4 will use intravital microscopy to examine the relationship of leukocytes with endothelial cell border in real time in living mice. We will first test the hypothesis that blocking PECAM arrests leukocyte transmigration in vivo the same site as it does in vitro (at the luminal side of the endothelial cell border in all mouse strains exceptC57Bl/6). We will next study the route of transmigration taken by leukocytes under PECAM-independent inflammatory conditions. The results of these studies will contribute significantly to our understanding of the regulation of diapedesis and provide new targets for anti-inflammatory therapy. Most diseases, including atherosclerosis, asthma, and autoimmune disorders, involve an inflammatory response that is uncontrolled or misdirected and actually harms the body instead of helping it. We have discovered a novel membrane compartment in endothelial cells (the cells that line blood vessels) that is critical for a key step in the inflammatory response. We will investigate how this compartment functions so that we can design better anti-inflammatory therapies
Keywords: Affect; Anti-inflammatory; Anti-Inflammatory Agents; Antibodies; Asthma; Atherosclerosis; Attention; Autoimmune Diseases; base; Blood Circulation; Blood Platelets; Blood Vessels; cadherin 5; Calcium; CD31 Antigens; Cell Line; Cell membrane; Cells; Chimera organism; Chronic; Cytoplasmic Tail; design; Disease; Endothelial Cells; Endothelium; Excision; Extracellular Domain; Funding; Genetic; Glycoproteins; Immunoglobulin Genes; In Vitro; in vivo; Inflammation; Inflammatory; Inflammatory Response; Intercellular adhesion molecule 1; intravital microscopy; junctional adhesion molecule; Kinesin; Lateral; Left; Leukocytes; Life; Light; Link; Lymphocyte; Membrane; Membrane Protein Traffic; Microtubules; migration; Modeling; Molecular Motors; Motor; Motor Activity; Mouse Strains; Mus; Mutate; Mutation; novel; PECAM1 gene; Play; Property; public health relevance; Recruitment Activity; Recycling; Regulation; Reticulum; Role; Route; Series; Side; Signal Transduction; Site; Structure; Surface; Testing; Time; Tissues; trafficking; Transmembrane Domain; Tyrosine; Tyrosine Phosphorylation; Vesicle; Work
Project start date: 1991-08-01
Project end date: 2015-05-31
Budget start date: 1-JUN-2011
Budget end date: 31-MAY-2012
PFA/PA: PA-07-070
5R01HL046849-21 (2011): $381250
MITOCHONDRIAL CARBONIC ANHYDRASES AND DIABETIC BLOOD-BRAIN BARRIER DISRUPTION
A William, Research Professor
Saint Louis Universitycity: Saint Louis country: United States (us)
Grant 1R01DK083485-01A2 from National Institute Of Diabetes And Digestive And Kidney Diseases
Abstract: Diabetes mellitus is associated with deterioration of the brain´s microvasculature and the blood-brain barrier (BBB) that it forms. The BBB prevents the unrestricted leakage of plasma proteins into the brain. The vascular BBB is physically formed by specially modified brain endothelial cells (BECs), but other cell types interact with it to form the neurovascular unit (NVU). Pericytes in particular are important in maintaining BBB function in the face of glycemic insult. We have shown in vitro that the insulin transporter is lost in the face of high glucose concentrations unless BECs are co-cultured with pericytes; astrocyte co-culture does not preserve BEC insulin transporter function. However, pericytes are themselves susceptible to glucose toxicity and the accompanying oxidative stress. Work with retinal pericytes and the blood-retinal barrier (BRB) shows that hyperglycemia induces apoptosis in retinal pericytes; with pericyte loss the BRB deteriorates. Mitochondrial carbonic anhydrases (CAs) are important in the generation of reactive oxygen species (ROS) and the subsequent oxidative stress that arises and is accelerated during hyperglycemia. In brief, mitochondrial CAs generate mitochondrial bicarbonate that is necessary for the oxidative catabolism of glucose, the major source of ROS. Hyperglycemia accelerates this process, increasing production of ROS. Blocking mitochondrial CAs shuttles pyruvate through anaerobic metabolism and so prevents excessive ROS production. We hypothesize that inhibition of mitochondrial carbonic anhydrases (CAs) will protect pericytes and BECs from ROS induced apoptosis and slow the development of disruption of the BBB in STZ-induced diabetes. Wewill test this hypothesis in two specific aims. SA1 will measure oxidative stress induced by high glucose media and the resulting mitochondrial leakage and apoptosis in primary cultures of brain pericytes and BEC isolated from mitochondrial CA KO & WT mice and in pericytes and BEC that overexpress CA or are treated with clinically used CA inhibitors. SA 2 will measure in diabetic and control mice BBB disruption and oxidative stress in BEC from CA KO mice, WT mice, and mice treated with the CA inhibitors of SA1. We will investigate a mechanism that could explain why diabetics develop problems with the small vessels of the brain and drugs that could reverse those problems. If successful, a major complication of diabetes might be treatable with a class of drugs already on the market
Keywords: 2-Deoxy-2-((methylnitrosoamino)carbonyl)amino-D-glucose; 2-deoxy-2-(3-methyl-3-nitrosoureido)-D-glucopyranose; 2-deoxy-2-[[(methylnitrosamino)-carbonyl]amino]-D-glucopyranose; Acetyl CoA; Acetyl Coenzyme A; Active Oxygen; Adventitial Cell; aerobic glycolysis; aerobic metabolism; aerobic respiration; Affect; Apoptosis; Apoptosis Pathway; Astrocytes; Astrocytus; Astroglia; Bicarbonates; Binding; Binding (Molecular Function); Blood; Blood - brain barrier anatomy; Blood Vessels; Blood-Brain Barrier; Blood-Retinal Barrier; Brain; carbonate dehydratase; Carbonate Dehydratase Inhibitors; Carbonate hydro-lyase; Carbonic Anhydrase Inhibitors; Carbonic Anhydrases; Carboxyanhydrase Inhibitors; Catabolism; Cell Death, Programmed; Cell Respiration; cell type; Cells; Cellular Respiration; Central Nervous System; Citric Acid; Citric Acid Cycle; Co-culture; Cocultivation; Coculture; Coculture Techniques; Coenzyme A, S-acetate; Communication; Complications of Diabetes Mellitus; Cytoplasm; D-Glucose; Deterioration; Development; Dextrose; diabetes; Diabetes Complications; Diabetes Mellitus; Diabetes-Related Complications; diabetic; Diabetic Complications; Drug usage; drug use; drug/agent; Drugs; Dysfunction; electron transfer; Electron Transport; Encephalon; Encephalons; Endothelial Cells; experiment; experimental research; experimental study; Extravasation; Functional disorder; Generations; Glucose; glucose metabolism; HCO3; heavy metal lead; heavy metal Pb; Hemato-Encephalic Barrier; Hexosamines; Humulin R; Hydrogen Carbonates; Hyperglycemia; hyperglycemic; In Vitro; in vitro Model; in vivo; in vivo Model; inhibitor; inhibitor/antagonist; Insulin; Insulin (ox), 8A-L-threonine-10A-L-isoleucine-30B-L-threonine-; Insulin Resistance; insulin resistant; Insulin, Regular; Intermediary Metabolism; Ketosuccinates; Knock-out; Knockout; Krebs Cycle; Laboratories; Lead; Leakage; Mammals, Mice; Marketing; Measures; Medication; Membrane; membrane structure; Metabolic Processes; Metabolism; METBL; Methods; Mice; Mitochondria; mitochondrial; mitochondrial membrane; Modeling; Molecular Interaction; Murine; Mus; nervous system function; Nervous System Physiology; Nervous System, Brain; Nervous System, CNS; Neuraxis; Neurologic function; Neurological function; neurovascular unit; Novolin R; overexpression; Oxaloacetates; oxidative metabolism; Oxidative Stress; Oxosuccinates; Oxygen Radicals; pathophysiology; pathway; Pathway interactions; Pb element; Pericapillary Cell; Pericytes; Perivascular Cell; Pharmaceutic Preparations; Pharmaceutical Preparations; Physiopathology; Plasma Proteins; polyol; prevent; preventing; Pro-Oxidants; Process; Production; Property; Property, LOINC Axis 2; public health relevance; Pyruvate; Pyruvates; Reactive Oxygen Species; research study; Resistance; resistant; Reticuloendothelial System, Blood; Retinal; Role; Rouget Cells; Serum Proteins; social role; Source; Spillage; Streptozocin; Streptozotocin; STZ; System; System, LOINC Axis 4; TCA cycle; Testing; Toxic effect; Toxicities; translation research enterprise; Translational Research; Translational Research Enterprise; Translational Science; Tricarboxylic Acid Cycle; vascular; Wild Type Mouse; Work; Zanosar
Relevance: We will investigate a mechanism that could explain why diabetics develop problems with the small vessels of the brain and drugs that could reverse those problems. If successful, a major complication of diabetes might be treatable with a class of drugs already on the market
Project start date: 2011-02-01
Project end date: 2015-01-31
Budget start date: 1-FEB-2011
Budget end date: 31-JAN-2012
PFA/PA: PA-10-067
1R01DK083485-01A2 (2011): $324178
A William, Professor Of Psychiatry
Mount Sinai School Of Medicinecity: New York country: United States (us)
Abstract: Center investigators are studying numerous genes and their protein products in the brain´s reward circuits to define their role in the regulation of mood and motivation under normal circumstances and in animal models of depression and antidepressant action. To accomplish this goal, the Behavioral Core has established an extremely broad battery of behavioral tests in rats and mice. This battery includes routine measures of locomotor activity and anxiety-like behavior,, as well as several standard depression-related tests such as the forced swim and learned helplessness tests. The battery also incorporates several additional assays that provide complementary information about an animal´s affective state; these include measures of sexual behavior, incentive motivation for food, intracranial self-stimulation, social interaction, and sophisticated cognitive tasks, to name some examples. In addition, Core personnel continually work to extend this battery to additional tests. For example, we recently adapted the social defeat paradigm, where certain behavioral abnormalities respond to chronic (but not acute) antidepressant treatment, to study stress-induced neuroadaptations in mouse brain. The imperative to employ such a large battery of behavioral tests is that it is difficult to infer something about complex behavior from a single test or even a limited number of tests. Rather, by utilizing numerous complementary measures we are able to infer, with much greater accuracy, the role of a given gene in complex behavior related to depression. By consolidating these behavioral tests within a centralized Core, we can ensure rigorous control over the data as well as facilitate comparisons and contrasts of experimental results from the individual Projects. This consolidation also makes financial sense, since we can concentrate and maximize efficient use of our behavioral expertise. The role of specific molecular targets in behavioral responses related to mood and motivation are tested with a variety of approaches, including advanced mouse mutagenesis and viral gene transfer in conjunction with the Transgenic Core. The Behavioral Core then provides routine, relatively high throughput behavioral tests for investigators in the Center´s preclinical Projects (1-4). Encouraging findings are pursued with more sophisticated behavioral tests also via this Core. In addition, the Core obtains routine neuroendocrine measurements (e.g., plasma corticosterone levels) and antidepressant blood levels in behaving animals as needed for particular experiments
Keywords: Acute; Affective; Animal Model; Animals; Antidepressive Agents; Anxiety; Appetitive Behavior; Behavior; behavior test; Behavioral; Biological Assay; Blood; Brain; Chronic; Cognitive; Complex; Corticosterone; Data; Ensure; Food; Gene Transfer; Genes; Goals; Human Resources; Incentives; Individual; Learned Helplessness; Measurement; Measures; Mental Depression; Molecular Target; mood regulation; Moods; Motivation; Motor Activity; Mus; Mutagenesis; Names; neuroadaptation; Neurosecretory Systems; Plasma; pre-clinical; Proteins; Rattus; relating to nervous system; Research Personnel; research study; response; Rewards; Role; Self Stimulation; Sex Behavior; social; Social Interaction; Stress; Swimming; Testing; Transgenic Organisms; Viral Genes; Work
Budget start date: 1-AUG-2011
Budget end date: 31-JUL-2012
5P50MH066172-10_9002 (2011): $352257
MEDULLOBLASTOMA AND METASTASES
A William, Professor
University Of California San Franciscocity: San Francisco country: United States (us)
Grant 5R01CA148699-02 from National Cancer Institute
Abstract: Brain tumors are the most common solid malignancies and the leading cause of cancer- related death in children. Medulloblastoma (MB) is the most common pediatric brain tumor. Dissemination (metastasis) of MB through the cerebrospinal fluid seeds the leptomeningeal membranes that cover the brain and spinal cord. Metastases in MB are refractory to treatment, essentially defining children with incurable tumor. We mobilized the transposable element Sleeping Beauty (SB) in a genetically engineered mouse model of MB driven by Sonic Hedgehog (Shh) signaling and observed robust metastases. Genetic analyses of matched primary and metastatic lesions indicate that tumors undergo parallel evolution and harbor distinct, clonally selected mutations generated by transposition. This is among the first mouse models enabling identification of genes driving metastases. We hypothesize that SB can uncover clonal organization and genes underlying progression/metastasis, and that these data will inform human MB. We propose experiments characterizing paired primary and metastatic tumors from patients with MB. We also propose additional SB based experiments to identify metastases genes in a novel MYCN-driven model for MB which arises largely independently of Shh signaling and which models both classic (60% of human MB) and large cell, anaplastic pathologies (10% of human MB). Because classic human MB can be driven either by myc or by loss of p53, we will also mobilize SB in p53 deficient mice. The use of 3 models minimizes biological effects due to background, characterizes a broad genetic subset of MB, and facilitates identification and prioritization of 1). Genes driving metastases in 3 distinct models for MB. 2). Candidates metastases genes altered in human MB. 3). Potential therapeutic targets. These data have profound implications for therapy, which assumes that metastases are biologically similar to the primary tumor. A.1.To recover transposon insertion site sequences from matched primary/metastatic GEM MB in two models in order to identify genes and pathways important for MB pathogenesis. A.2.To validate the functional importance of candidate metastases genes identified by SB insertion. A.3.To evaluate hierarchical structures, genes and pathways important for leptomeningeal dispersion using human genomic data, and paired tumors and metastases from human MB. Medulloblastoma (MB) is a common and frequently lethal tumor of children, for which current therapies are often ineffective. Locoregional dissemination (metastasis) of MB through the cerebrospinal fluid essentially defines incurable MB. Our long term objective is to mobilize the transposable element Sleeping Beauty (SB) in three different genetically engineered mouse models of MB to dissect the clonal organization and genetic bases of malignant progression and leptomeningeal metastasis, to inform human MB. Successful completion of this proposal identifies metastases genes and new therapeutic targets for children with metastatic MB
Keywords: 0-11 years old; Accounting; allelic variant; Anaplastic Cell; Automobile Driving; base; Biological; biological signal transduction; Blood Circulation; Bloodstream; Brain; Brain Neoplasia; Brain Neoplasms; Brain Tumors; Cancer Cause; cancer cell; Cancer Etiology; cancer metastasis; Cancers; Candidate Disease Gene; Candidate Gene; Cell Communication and Signaling; Cell Signaling; Cerebrospinal Fluid; Cessation of life; Child; Child Youth; Childhood Brain Neoplasm; Childhood Brain Tumor; children; Children (0-21); Chromosomal Breaks; Chromosome Break; Circulation; Data; Data Set; Dataset; Death; Disease; disease/disorder; Disorder; DISSEC; Dissection; DNA Transposable Elements; driving; Drivings, Automobile; Encephalon; Encephalons; Epigenetic; Epigenetic Change; Epigenetic Mechanism; Epigenetic Process; Event; Evolution; experiment; experimental research; experimental study; Gene Expression; Gene Organization; Gene Structure; Gene Structure/Organization; Gene variant; Genes; Genes, p53; Genetic; Genetic Alteration; Genetic analyses; genetic analysis; Genetic Change; Genetic defect; Genetic Diversity; Genetic Variation; Genetically Engineered Mouse; Genetics-Mutagenesis; genome mutation; Genomics; Goals; Hedgehog (Hh) signal transduction pathway; hedgehog signaling pathway; Hematogenous; hh signaling pathway; HHG1; HLP3; HPE3; Human; human data; Human, Child; Human, General; in vivo; Intracellular Communication and Signaling; Leptomeningeal Metastasis; Lymphatic; malignancy; Malignant; Malignant - descriptor; Malignant Cell; Malignant Neoplasms; Malignant Tumor; Mammals, Mice; Man (Taxonomy); Man, Modern; Maps; Medulla Spinalis; medulloblastoma; Membrane; membrane structure; Meningeal Carcinomatosis; meningeal metastases; Messenger RNA; Metastasis; Metastasis to the Leptomeninges; Metastasize; Metastatic Lesion; Metastatic Neoplasm; Metastatic Neoplasm to the Leptomeninges; Metastatic to; Metastatic Tumor; Metastatic Tumor to the Leptomeninges; Methylation; Mice; Micro RNA; MicroRNAs; miRNA; Modeling; Molecular Biology, Mutagenesis; Morbidity; Morbidity - disease rate; Mortality; Mortality Vital Statistics; mouse model; mRNA; Murine; Mus; Mutagenesis; Mutation; MYCN; MYCN gene; Neoplasm Metastasis; neoplasm/cancer; neoplastic meningitis; Nervous System, Brain; new therapeutic target; NMYC; NMYC Gene; novel; P53; Pathogenesis; Pathology; pathway; Pathway interactions; Patients; Pattern; Pediatric Neoplasm of the Brain; Pediatric Tumor of the Brain; Plant Embryos; Primary Lesion; Primary Neoplasm; Primary Tumor; Protein Methylation; public health relevance; Refractory; research study; resistance to therapy; resistant to therapy; RNA, Messenger; Role; Sampling; Secondary Neoplasm; Secondary Tumor; seed; Seeds; Selection (Genetics); SHH; SHH gene; Signal Pathway; Signal Transduction; Signal Transduction Systems; Signaling; Site; Sleeping Beauty; SMMCI; smoothened signaling pathway; social role; Solid; Solid Neoplasm; Solid Tumor; Spinal Cord; spinal fluid; Testing; therapeutic target; therapy resistant; TP53; TP53 gene; Transposable Elements; TRP53; tumor; Tumor Cell Migration; Tumor Protein p53 Gene; tumors in the brain; Variation (Genetics); youngster; Zygotes, Plant
Relevance: Narrative: Medulloblastoma (MB) is a common and frequently lethal tumor of children, for which current therapies are often ineffective. Locoregional dissemination (metastasis) of MB through the cerebrospinal fluid essentially defines incurable MB. Our long term objective is to mobilize the transposable element Sleeping Beauty (SB) in three different genetically engineered mouse models of MB to dissect the clonal organization and genetic bases of malignant progression and leptomeningeal metastasis, to inform human MB. Successful completion of this proposal identifies metastases genes and new therapeutic targets for children with metastatic MB
Project start date: 2010-05-04
Project end date: 2015-02-28
Budget start date: 1-MAR-2011
Budget end date: 29-FEB-2012
PFA/PA: PA-07-070
5R01CA148699-02 (2011): $484312
STRUCTURE AND FUNCTION OF E. HISTOLYTICA ADHERENCE LECTIN
A William, Wade Hampton Frost Professor
University Of Virginia Charlottesvillecity: Charlottesville country: United States (us)
Grant 5R01AI026649-22 from National Institute Of Allergy And Infectious Diseases
Abstract: Amebic colitis and liver abscess are due to infection with the enteric protozoan parasite Entamoeba histolytica. The World Health Organization estimates that approximately 50 million people worldwide suffer from invasive amebic infection each year. The adherence of E. histolytica trophozoites to intestinal mucins and epithelial cells by a galactose-binding (Gal/GalNAc) lectin is necessary for the parasite to kill host cells and invade. We have discovered that the intermediate subunit of this lectin is part of an E. histolytica gene family of more than 80 transmembrane kinases (TMK). The TMK kinase domains are novel and the number of TMKs is unprecedented in a unicellular organism. We hypothesize that the TMKs are a major receptor system for sensing the environment and controlling the invasive behavior of the parasite. We propose to test if the TMKs undergo antigenic variation, determine if TMK 96 is a dual specificity (tyr and ser/thr) kinase and identify its downstream effectors and regulators, and study the biologic activities regulated by TMK 96. We will focus on the role of TMK 96 in the adherence, killing and endocytosis of apoptotic corpses as it has been identified as part the phagosome proteome. Successful completion of these studies will provide fundamental knowledge of these novel receptor kinases and insight into the means by which the parasite controls its invasion into the human intestine
Keywords: 1-Phosphatidylinositol 3-Kinase; Active Sites; Adherence (attribute); Amebiasis; Amebic colitis; Antibodies; Antigenic Variation; Apoptotic; base; Behavior; Binding (Molecular Function); Biochemical; Biochemistry; Biology; cDNA Library; Cell Surface Receptors; Cells; Chimera organism; Code; Complex; Controlled Environment; Cytoplasmic Tail; Dominant-Negative Mutation; Endocytosis; Entamoeba histolytica; Enteral; Environment; Epithelial Cells; Escherichia coli; Extracellular Domain; Family; Gal-GalNAc; Galactose; Gene Family; Genome; Genomics; Human; Immune Sera; In Vitro; Infection; Ingestion; insight; Interleukin 2 Receptor; Intestines; Invaded; Killings; knock-down; Knowledge; laser capture microdissection; Lectin; Life; Liver Abscess; Measurement; Measures; member; Messenger RNA; Modeling; Mucins; Mus; Mutation; novel; novel strategies; Organism; Parasite Control; Parasites; Peptides; Phagosomes; Phosphotransferases; positional cloning; Prevention; Process; programs; Property; Protein Kinase; Proteins; Proteome; Proteomics; Reaction; receptor; Regulation; Research Personnel; Role; Signal Transduction; Specificity; Structure; System; Testing; World Health Organization; yeast two hybrid system; Yeasts
Project start date: 1989-08-01
Project end date: 2012-03-31
Budget start date: 1-APR-2011
Budget end date: 31-MAR-2012
5R01AI026649-22 (2011): $328344
THE ROLE OF ABCC6 IN CHRONIC AND ACUTE CARDIOVASCULAR MINERALIZATION
A William
University Of Hawaii At Manoacity: Honolulu country: United States (us)
Grant 1R01HL108249-01A1 from National Heart, Lung, And Blood Institute
Abstract: Abnormal mineralization occurs in the context of several common conditions, including advanced age, diabetes, hypercholesterolemia, chronic renal failure and certain genetic conditions. Loss-of-function mutations in the ABCC6 gene cause chronic or acute forms of dystrophic mineralization in diseases such as Pseudoxanthoma elasticum (PXE) and dystrophic cardiac calcification (DCC). These pathologies are characterized by mineralization of cardiovascular and/or dermal tissues. PXE is heritable while DCC is an acquired phenotype resulting from cardiovascular insults. We have obtained preliminary data that suggest that ABCC6 initiates and modulates a calcification inhibitor pathway with a crucial role in both acute and chronic dystrophic calcification. We hypothesize that the loss of ABCC6 function in liver produces an imbalance of modulators of calcification in cardiovascular tissues either directly or through increased levels or modification of a circulating factor. To test this hypothesis, we will use a mouse models to characterize the effects of ABCC6 mutations on the structure and function of the protein, determine the response of Abcc6-/- mice to acute ischemic cardiac injuries and determine the role of Abcc6-/- in the calcification of the atherosclerotic plaque. The overall goal of this proposal is to address the role of the ABCC6 gene in chronic and acute dystrophic calcification that affects cardiac and vascular tissues. To achieve the goals of this proposal, we will use mouse models to characterize the effects of ABCC6 mutations on the structure and function of the protein, determine the response of Abcc6-/- mice to acute cardiac ischemic injuries and determine the role of Abcc6 in the calcification of the atherosclerotic plaque
Keywords: Acute; Address; Affect; Aging; Arterial Fatty Streak; Atherosclerosis; Backcrossings; Binding (Molecular Function); Blood Circulation; Blood Vessels; calcification; calcification inhibitor; Cardiac; Cardiovascular system; Cell membrane; Cellular translocation; Chronic; Chronic Kidney Failure; Complement; Complex; Coronary artery; Data; Deposition; Dermal; Diabetes Mellitus; Digestion; Disease; disease-causing mutation; Distal; Dystrophic Calcification; Elderly; Gene Expression; Genes; Goals; Health; Healthcare; Hepatic; Hepatocyte; Hereditary Disease; Human; human disease; Hydrolysis; hypercholesterolemia; Hyperlipidemia; improved; In Vitro; in vivo; Inflammatory; Inherited; inhibitor/antagonist; Injection of therapeutic agent; Injury; Kidney Failure; Knock-out; Knockout Mice; Ligation; Link; Liver; Location; loss of function mutation; Mechanics; Mediating; Metabolic; mineralization; Minerals; Modification; Molecular; mouse model; Mouse Protein; Mus; mutant; Mutation; novel; Pathologic Processes; Pathology; Pathway interactions; Patients; Phenotype; Population; Predisposition; protein structure function; Proteins; Pseudoxanthoma Elasticum; Publishing; Regulation; response; restoration; Role; Tail; Testing; Thalassemia; Tissues; Trypsin; Vascular calcification; Veins; Wild Type Mouse; Work
Relevance: The overall goal of this proposal is to address the role of the ABCC6 gene in chronic and acute dystrophic calcification that affects cardiac and vascular tissues. To achieve the goals of this proposal, we will use mouse models to characterize the effects of ABCC6 mutations on the structure and function of the protein, determine the response of Abcc6-/- mice to acute cardiac ischemic injuries and determine the role of Abcc6 in the calcification of the atherosclerotic plaque
Project start date: 2012-02-24
Project end date: 2016-01-31
Budget start date: 24-FEB-2012
Budget end date: 31-JAN-2013
1R01HL108249-01A1 (2012): $375000
MECHANISMS OF PERCHLORATE-INDUCED DISRUPTION OF SEXUAL DIFFERENTIATION
A William
University Of Alaska Anchoragecity: Anchorage country: United States (us)
Grant 5R01ES017039-03 from National Institute Of Environmental Health Sciences
Keywords: Affect; Animal Model; Animals; Antisense Oligonucleotides; Aromatase; Behavior; Behavioral; Courtship; design; Development; developmental disease/disorder; Disease; Dose; Endocrine Disruptors; Environment; Epidemic; experience; Family member; Female; Fishes; Food; Frequencies (time pattern); Functional disorder; gain of function; Gasterosteidae; Gene Expression; Gene Family; gene function; Genes; Genetic; Genetic Transcription; Genome; geographic difference; Goals; Gonadal structure; Gonadotropins; Health; Hormonal; Human; human disease; Human Milk; Hypertrophy; In Situ Hybridization; Induced Mutation; Ingestion; Iodides; knock-down; loss of function; Maintenance; male; Mediating; Messenger RNA; Milk; Molecular; Molecular Profiling; Morphology; Mutate; Mutation; nuclease; Outcome; paralogous gene; Pathway interactions; Pattern; Perchlorates; Phenotype; Physiological; Physiology; Production; Proteins; public health relevance; receptor; reproductive; Reproductive Health; Research; research study; response; Risk; Role; Sex Differentiation; Sexual Development; SLC5A5 gene; sodium-iodide symporter; Supplementation; Testing; Testis; Thyroid Diseases; Thyroid Function Tests; Thyroid Gland; Thyroid Hormones; Tissues; toxicant; Transcript; Translating; United States; uptake; Water; Work; Zinc Fingers
Relevance: Perchlorate is a persistent, water-soluble contaminant that is pervasive in the United States and poses a major risk to human health through ingestion of contaminated water, food and milk. Perchlorate not only inhibits thyroid activity, but also alters sexual development in stickleback fish, a commonly used model organism in genetic studies. The recent dramatic increase and geographic differences in frequency of human reproductive disorders are likely due to changes in the environment, including perchlorate exposure. Proposed experiments will identify genes and gene functions that, under the insult of perchlorate contamination, disrupt normal development of male and female gonads. Experiments will also determine the hormonal mechanisms that translate genetic changes into developmental disorders of the gonads. This research will advance our understanding of human thyroid diseases and the recent epidemic of impaired human reproductive health
Project start date: 2010-03-01
Project end date: 2014-11-30
Budget start date: 1-DEC-2011
Budget end date: 30-NOV-2012
5R01ES017039-03 (2012): $486206
MITOCHONDRIAL CARBONIC ANHYDRASES AND DIABETIC BLOOD-BRAIN BARRIER DISRUPTION
A William
Saint Louis Universitycity: Saint Louis country: United States (us)
Grant 5R01DK083485-02 from National Institute Of Diabetes And Digestive And Kidney Diseases
Keywords: Acetyl Coenzyme A; aerobic glycolysis; Affect; Apoptosis; Astrocytes; Bicarbonates; Binding (Molecular Function); Blood; Blood - brain barrier anatomy; Blood Vessels; Blood-Retinal Barrier; Brain; carbonate dehydratase; Carbonic Anhydrase Inhibitors; Catabolism; Cell Respiration; cell type; Cells; Citric Acid; Citric Acid Cycle; Coculture Techniques; Communication; Complications of Diabetes Mellitus; Cytoplasm; Deterioration; Development; Diabetes Mellitus; diabetic; Drug usage; Electron Transport; Endothelial Cells; Extravasation; Functional disorder; Generations; Glucose; glucose metabolism; Hexosamines; Hyperglycemia; In Vitro; in vitro Model; in vivo; in vivo Model; inhibitor/antagonist; Insulin; Insulin Resistance; Knock-out; Laboratories; Lead; Marketing; Measures; Membrane; Metabolism; Methods; Mitochondria; mitochondrial membrane; Modeling; Mus; Nervous System Physiology; Neuraxis; neurovascular unit; overexpression; Oxaloacetates; Oxidative Stress; Pathway interactions; Pericytes; Pharmaceutical Preparations; Plasma Proteins; polyol; prevent; Process; Production; Property; public health relevance; Pyruvate; Reactive Oxygen Species; research study; Resistance; Retinal; Role; Serum Proteins; Source; Streptozocin; System; Testing; Toxic effect; Translational Research; Wild Type Mouse; Work
Relevance: We will investigate a mechanism that could explain why diabetics develop problems with the small vessels of the brain and drugs that could reverse those problems. If successful, a major complication of diabetes might be treatable with a class of drugs already on the market
Project start date: 2011-02-01
Project end date: 2015-01-31
Budget start date: 1-FEB-2012
Budget end date: 31-JAN-2013
5R01DK083485-02 (2012): $320992
CONTROL OF DRUG DELIVERY TO THE CNS: MODULATING P-GLYCOPROTEIN ACTIVITY
A William
St. Louis Va Medical Centercity: St. Louis country: United States (us)
Relevance: Potential Impact on Veterans´ Health Care: If all or part of this cascade is active in vivo, then P-gp activity is likely modified in a host of diseases that would result in altered CNS drug activities. These diseases would include not only bacterial sepsis, but diseases such as rheumatoid arthritis, diabetes, stroke, CNS cancers, and even obesity and Alzheimer´s disease in which proinflammatory and neuroinflammatory conditions exist. The proposed work will have 3 major impacts on health care: 1) it will help to identify drugs and diseases in which a modified P-gp activity results in altered CNS drug profiles; 2) it will identify stategies and agents which can reverse those altered CNS drug profiles in those disease states; 3) it will lead to a new strategy, that of manipulation of P-gp activity, in the development of drugs both for targeting CNS diseases and for developing drugs with fewer CNS side effects
Project start date: 2009-04-01
Project end date: 2013-03-31
Budget start date: 1-APR-2010
Budget end date: 31-MAR-2011
PFA/PA: RFA-BX-08-001
5I01BX000457-02 (2011): $0
BEYOND PECAM: MECHANISMS OF TRANSENDOTHELIAL MIGRATION
A William, Magerstadt Professor And Chair
Northwestern Universitycity: Chicago country: United States (us)
Grant 5R37HL064774-11 from National Heart, Lung, And Blood Institute
Keywords: Asthma; Atherosclerosis; Autoimmune Diseases; Award; base; Blood Circulation; Cell Line; Chronic; Complement; Disease; Endothelial Cells; fighting; Funding; Goals; Graft Rejection; Grant; Immune response; In Vitro; in vivo; In-Migration; Inflammation; Inflammatory; Inflammatory Response; Instruction; Lead; Leukocytes; Membrane; Membrane Protein Traffic; microorganism; Microtubules; migration; Modeling; Molecular; Mus; Play; postcapillary venule; Principal Investigator; Process; programs; Recruitment Activity; Recycling; Research; response; Rheumatoid Arthritis; Role; Septic Shock; Signal Transduction; Site; Therapeutic; Time; Tissues; Work; Wound Healing
Project start date: 2000-04-01
Project end date: 2015-04-30
Budget start date: 1-MAY-2011
Budget end date: 30-APR-2012
5R37HL064774-11 (2011): $404052
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