Rheal A. Towner - US grants

ABSTRACT NOT PROVIDED

magnetic resonance imaging; biomedical facility; bioimaging /biomedical imaging; clinical research;

2-Hydroxy-N,N,N-trimethylethanaminium; Ablation; Angiogram; Angiography; Animals; Bioenergetic; Bioenergetics; Biomedical Research; Body Tissues; Brain; Brain Neoplasia; Brain Neoplasms; Brain Tumors; Breast; CRISP; Cancer of Brain; Cancers; Choline; Common Rat Strains; Computer Retrieval of Information on Scientific Projects Database; Contrast Agent; Contrast Drugs; Contrast Media; Creatine; Development; Disease; Disorder; Electromagnetic, Laser; Encephalon; Encephalons; Ethanaminium, 2-hydroxy-N,N,N-trimethyl-; Experimental Animal Model; Funding; Glycine, N-(aminoiminomethyl)-N-methyl-; Grant; Image; Institution; Investigation; Investigators; Lasers; Lesion; Lipids; Liquid substance; Localized; MR Imaging; MR Spectroscopy; MR Tomography; MRI; MRS; MRSI; Magnetic Resonance Imaging; Magnetic Resonance Imaging Scan; Magnetic Resonance Spectroscopy; Magnetism; Malignant Melanoma; Malignant Neoplasms; Malignant Tumor; Malignant Tumor of the Brain; Malignant neoplasm of brain; Mammals, Mice; Mammals, Rats; Mammals, Rodents; Medical Imaging, Magnetic Resonance / Nuclear Magnetic Resonance; Metabolic; Methods; Methods and Techniques; Methods, Other; Mice; Microscopic; Microscopy; Modeling; Molecular; Molecular Target; Monitor; Murine; Mus; NIH; NMR Imaging; NMR Tomography; National Institutes of Health; National Institutes of Health (U.S.); Nervous System, Brain; Nuclear Magnetic Resonance Imaging; Organ; Pathogenesis; Phase; Phosphatides; Phospholipids; Radiation, Laser; Radiopaque Media; Rat; Rattus; Research; Research Personnel; Research Resources; Researchers; Resolution; Resources; Rodent; Rodentia; Rodentias; Source; Spectroscopy; Spectrum Analyses; Spectrum Analysis; Techniques; Technology; Temperature; Therapeutic Agents; Tissues; United States National Institutes of Health; Vascular System; Zeugmatography; base; design; designing; disease/disorder; fluid; imaging; imaging probe; in vivo; instrument; liquid; magnetic; magnetic field; malignancy; melanoma; neoplasm/cancer; tumor; tumors in the brain

[unreadable] DESCRIPTION (provided by applicant): This project is focussed on developing and assessing novel tumor antigen-specific magnetic resonance imaging (MRI) molecular probes for the in vivo detection of tumor antigens, which may serve as diagnostic biomarkers for early detection of malignant human cancers. A transgenic mouse hepatocarcinogenesis model will be used to follow sequential changes during neoplastic nodule, adenoma and carcinoma development. This research will aid in the development of diagnostic methods for in vivo detection of malignant tumor formation. Specifically, we plan to test the in vivo specificity of tumor-specific antigen MRI molecular targeting agents to be able to detect and progressively follow the formation of tumors by targeting antigens associated with tumor malignancy during various stages of experimental carcinogenesis in a mouse liver tumor model. SPECIFIC AIM 1: To determine the specificity of novel MRI molecular targets for tumor antigens in mouse hepatoma cell culture models: Objective 1: Determine the degree of expression of malignant-specific tumor antigens in mouse hepatoma cell lines (BW1J, HEPA 1-6 or MH-224) using immunohistochemistry with antibodies specific for the malignant-specific tumor antigens. Objective 2: Develop tumor antigen specific MRI contrast agents (with gadolinium (Gd) or iron oxide parent compounds) tagged with an anti-nodule/tumor marker antibody (Ab) specific for malignant-specific tumor antigens. Objective 3: To use MRI to initially determine the effectiveness of the antigen-specific contrast agents for antigen-binding specificity in various mouse hepatocellular carcinoma cell lines in vitro. SPECIFIC AIM 2: Use antigen specific MRI contrast agents to detect tumor antigens in vivo within experimental mouse carcinogenesis models: Objective 4: Use antigen specific MRI contrast agents to detect and predict the formation of tumors in vivo in experimental rodent carcinogenesis models. Objective 5: To compare in vivo MRI detection of tumor-specific molecular markers with conventional immunohistoehemistry of mouse liver slices for malignant-specific tumor antigens, and tumor grading. Once we have established the optimal molecular-targeting agent in a mouse hepatocarcinogenesis model, then the groundwork is established for initiation of human cell and planning of clinical evaluations for patients with liver cancer. [unreadable] [unreadable] [unreadable]

[unreadable] DESCRIPTION (provided by applicant): [unreadable] [unreadable] This project is focused on establishing the mechanism of alpha-phenyl-tert-butyl nitrone (PBN), in its ability to inhibit glioma formation in experimental rodent models that vary in tumor grade. We have promising preliminary data that clearly demonstrates that PBN can potentially be an effective anti-tumor agent in vivo, prior to tumor implantation, or once a tumor has formed. Our hypothesis is that c-Met, a tyrosine kinase receptor for the hepatocyte growth factor (scatter factor) plays an integral role in the formation of malignant gliomas, and that altered c-Met levels from PBN treatment suppresses glioma formation. Important hallmarks of malignant gliomas include their invasive behavior and angiogenesis. c-Met is thought to be a mediator in many of the processes of malignant brain tumor progression, including cell proliferation, cell invasion, apoptosis and angiogenesis. Approximately 15,000 patients in the U.S.A. die with glioblastomas per year. Due to the infiltrative nature of gliomas, surgery is rarely effective. Despite modern diagnostics and treatments the median survival time for patients with glioblastomas does not exceed 15 months. A potential chemo-preventative agent that can suppress glioma growth or even eradicate gliomas entirely would be of tremendous benefit to patients that develop malignant tumors. In vivo magnetic resonance (MR) methods are ideally suited to follow changes in tumor growth, angiogenesis, as well as the assessment of c- Met levels which can be used as a marker for predicting the prognosis of gliomas. We currently use non-invasive in vivo MR imaging (MRI) methods to detect tumor morphology, tumor vasculature or angiogenesis (using MR angiography (MRA)), and have preliminary data on c-Met expression (using molecular-targeted MRI). In addition we use molecular biology methods (Western blots, immunofluorescence histology) and histological tumor grading, to correlate with MR imaging, angiography and molecular targeted data. This project will assess PBN in its ability to prevent malignant glioma formation and development, use c-Met knockdown glioma cell clones, and incorporate novel in vivo molecular-targeted MRI techniques for in vivo detection of c-Met in intracranial rat gliomas varying in tumor grades. Intracerebral implantation of rat C6 (forming grade II or III tumors) or RG2 (forming more aggressive grade III or IV tumors) glioma cells will be used in Fisher 344 rats as the glioma models. Specific aim 1 will be to assess the levels of c-Met in the 2 glioma models (C6 and RG2); specific aim 2 will be to establish the effectiveness of PBN as a c-Met suppressor in preventing glioma growth and formation, and to use c-Met knockdown glioma models to further elucidate the role of c-Met in glioma formation; and specific aim 3 will be to develop and validate improved MRI methods for in vivo detection of c-Met expression. There are very few existing agents that are currently applicable for the intervention of malignant gliomas. Other promising characteristics of PBN, in addition to c-Met suppression, are that it is orally bio-available, and it is able to cross the blood-brain-barrier (BBB). PBN seems to be a promising candidate in its ability to be an effective agent against glioma formation, and may be useful as a chemo-preventative agent against glioblastoma multiforme in humans. We have preliminary data that PBN (alpha-phenyl-tert-butyl nitrone) seems to be a promising candidate in its ability to be an effective agent against glioma formation, and may be useful as a chemo-preventative agent against glioblastoma multiforme in humans. Our hypothesis is that c-Met, a tyrosine kinase receptor, plays an integral role in the formation of malignant gliomas, and that PBN is able to decrease c-Met levels, resulting in the suppression of glioma formation. This project will assess PBN in its ability to prevent malignant glioma formation and development, use c-Met knockdown glioma cell clones to elucidate the role of c-Met, and incorporate novel in vivo moleculartargeted MRI (magnetic resonance imaging) techniques for in vivo detection of c-Met in intracranial rat gliomas varying in tumor grades. [unreadable] [unreadable] [unreadable]

DESCRIPTION (provided by applicant): The scope of this project is to establish whether magnetite or iron oxide (IO) nanoprobes, specific for tumor markers such as c-Met (tyrosine kinase receptor for the hepatocyte growth/scatter factor) or VEGF-R2 (vascular endothelial growth factor receptor), can be used therapeutically to inhibit or diminish glioma tumor growth in experimental rodent models. We plan to develop and synthesize IO nanoprobes, or characteristic tumor antigen-specific antibody tagged IO-based MRI contrast agents, for the in vivo detection of molecular events associated with tumor malignancy or angiogenesis in rat glioma models. We will assess the ability of these IO nanoprobes to detect malignant tumor markers such as c-MET, over-expressed in tumors that are invasive in nature, and VEGF-R2, which is associated with angiogenesis in malignant tumors, in an experimental C6 rat glioma model. It has also been recently established that IO nanoparticles can induce hyperthermia by being subjected to an alternating magnetic field (AMF), and that this may be used as a possible therapeutic modality against tumors. In a novel approach, we wanted to combine the therapeutic nature of the IO nanoparticles, and the specificity of tumor marker specific nanoprobes to direct hyperthermia therapy to malignant tumors targeted by anti-c-Met or anti-VEGF-R2 nanoprobes. Molecular magnetic resonance imaging (mMRI) will be used to assess the nanoprobe specificity for either c-Met or VEGF-R2, and morphological MRI will be used to assess the therapeutic efficacy of the nanoprobes on glioma growth in a rat glioma model. It is important that validation steps are initially studied in experimental animal models prior to the development of methods for clinical diagnosis and/or treatment. The advantages of MRI as a molecular imaging modality are a higher spatial resolution (5m) and the ability to obtain morphological, physiological and metabolic information in a single imaging session. The specific aims that we will use to assess tumor marker specific nanoprobes as potential diagnostic and therapeutic agents are as follows: (1) Assess tumor marker specific magnetite nanoprobes in their ability to detect markers associated with tumor malignancy and angiogenesis, and (2) assess anti-tumor therapeutic effect of nanoprobes when used with an alternating magnetic field. For specific aim 1, we plan to covalently bind the IO nanoparticles to cross-linked IO (CLIO) in conjunction with anti-VEGF-R2 Ab or anti-c-Met Ab, so that they can be used as vascular nanoprobes. Dextran-based CLIO contrast agents have been previously used as 'blood pool'agents. The IO nanoprobes will be tested in a C6 rat glioma model extensively used in our laboratory. For a non-specific control, a normal non-immune rat IgG will be coupled to the CLIO moiety. For specific aim 2, the focus will be to combine the tumor marker specificity of the nanoprobes to target malignant tumors, and to utilize the IO component on these compounds as a means of generating hyperthermia via an alternating magnetic field (100 kHz to 500 kHz frequency range) as a possible anti-tumor therapeutic treatment. We plan to subject C6 glioma-bearing rats to alternating magnetic fields (AMF), following administration of IO nanoprobes targeting either c-Met or VEGF-R2, and monitor tumor growth via MRI. Controls will be treated with non-specific IO nanoparticles (containing normal non-immune rat IgG) and subjected to an AMF. The proposed research is highly innovative and involves the research and development of molecular targeting agents, magnetic nanoprobes, and therapeutic options associated with these nanoprobes, that can be used to inhibit the growth and possibly eradicate malignant gliomas. Our plan is to incorporate the potential hyperthermia therapeutic effect of the nanoparticles in combination with the specificity of targeting tumor markers on malignant tumors. This combination of concepts has never been attempted before by targeting c- MET or VEGF-R2 within a glioma model. The development of in vivo biomedical research tools to assess the detection of potential tumor biomarkers may have a significant impact on future diagnostic approaches to be used in the early detection of human gliomas. In addition, this project will also assess the chemo-preventative capabilities of potential agents in their ability to prevent malignant glioma formation. To accomplish the goals of the project we have compiled a multi-institutional team from the Oklahoma Medical Research Foundation (OMRF), Oklahoma State University (OSU), and the University of Central Oklahoma (UCO), with expertise in mMRI, nanoparticles, radiofrequency bio-engineering, and cancer therapeutics. PUBLIC HEALTH RELEVANCE: This project is focused on developing and assessing tumor marker specific magnetite or iron oxide (IO) nanoprobes in their ability to detect tumor markers in vivo, and in addition be used as anti-cancer therapeutic agents. IO nanoprobes, such as ultrasmall superparamagnetic iron oxides (USPIOs) have recently become popular as molecular reporting probes with the use of molecular magnetic resonance imaging (mMRI). We propose to develop and assess IO nanoprobes with the use of molecular magnetic resonance imaging (mMRI) in a rodent glioma model to follow molecular markers associated with tumor malignancy and angiogenesis, and to utilize the nanoprobes themselves as potential therapeutic agents. Studying a potentially effective anti- tumor therapeutic agent, as well as developing novel in vivo diagnostic and predictive procedures may significantly impact patient prognosis and survivability in the future.

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. OK-INBRE undergraduate partner institutions is now charged with considering how to "reinvent" Oklahoma's undergraduate science curriculum along the lines of the Bio2010 model (Bio2010: Transforming Undergraduate Education for Future Research Biologists, by the National Research Council of the National Academies, 2003;National Academies of Science, Washington, DC). It is our expectation that these curriculum changes will be implemented incrementally during the next round of OK-INBRE funding. To begin this new type of multidisciplinary teaching, in the next funding period, we propose to provide to PUI students two different week-long course modules developed by the Bioinformatics and accessory MRI Core facility directors. Briefly, the course provided by Dr. Dyer and the Bioinformatics Core has the goal of introducing the students to phylogenetic analyses using free bioinformatics resources available from the web. The students will be exposed to the principles of evolution, sequence databases, sequence comparisons and analyses, sequence similarity searching, multiple sequence alignments, and phylogenetic tree building. Towards the end of this module, the students should understand the applications of bioinformatics resources towards elucidating important and relevant evolutionary information from a sequence dataset. The module will focus on using phylogenetics to make functional assignments, identify homologous relationships between genes, and construct phylogenies between organisms. This course will be taught in upper division molecular biology, genetics, microbiology, or evolution courses and will bridge the disciplines of biology, chemistry, mathematics and computer science. The module will be offered twice in the academic calendar year at various PUIs, as well as during the summer research program on the OUHSC campus. The other multidisciplinary course to be offered to the OK-INBRE PUIs will be taught by Dr. Towner from the accessory MRI core facility located at the OMRF. Biomedical imaging incorporates multiple disciplines. Physics and chemistry are used to understand at the atomic level how protons in MRI and X-rays in CT scans can generate the signals used to generate anatomical images that can be applied for medical diagnosis. Image display and processing involves the use of mathematical algorithms, such as Fourier Transformation in MRI. Biophysical processes, such as selective contrast agent uptake, water diffusion and perfusion, involve combined biology, physics and chemistry concepts. The extension of MRI to obtain metabolic information associated with pathological processes involves a strong reliance on chemistry and biochemistry. Molecular imaging incorporates a molecular-specific affinity component as well as a reporting or signaling component and draws molecular biology (immunology, cell biology) and chemistry (synthesis of molecular probes, pharmacokinetics). Imaging analysis involves the use of mathematical and computer programming for post-processing and quantitative assessment of morphological, molecular, functional and metabolic information that can be obtained from the data. This Bio2010 module will be targeted towards upper division undergraduate students in biochemistry, chemistry, immunology, cell biology, and physics. The module will be offered twice in the academic calendar year, as well as during the summer research program at the OMRF (see Appendix for detailed outlines and syllabi for both of these new Bio2010 multidisciplinary module courses).

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The Small Animal Imaging Core (Core 2) will provide support for in vivo imaging of rodent models for the Proposed Projects. Dr. Towner, the Director of the Advanced Magnetic Resonance Center, and of Core 2, will oversee all aspects of in vivo rodent imaging involving the use of the 7 Tesla small animal MRI that will be utilized in the Proposed Projects. For vascular imaging, magnetic resonance (MR) imaging on large (angiography (MRA)) and small vessels (DCE (dynamic contrast-enhanced)-MRI and perfusion MRI) can be used to assess angiogenesis for Projects 1 and 5. Towner has extensive experience in developing molecular MRI nanoprobes that can be used to assess levels of inflammatory (e.g. iNOS) and angiogenesis (e.g. VEGF-R2) markers, which can be used in Project 4 to assess intracapsular iNOS levels, and the same approach for VEGF-R3 in Projects 2 and 5 to assess in vivo VEGF-R3 levels. These compounds also have the capability of being monitored by other imaging modalities such as microscopic fluorescence imaging via a biotin tag on the molecular MRI probes in collaboration with Core 1. Dr. Towner is also developing iron oxide-based nanoparticles, which can be used to detect macrophages in vivo in atherosclerosis for Project 3. Recently we have used iron oxide-based nanoprobes to specifically detect VEGF-R2 levels in vivo.

? DESCRIPTION (provided by applicant): The classical amyloid cascade hypothesis of Alzheimer's disease (AD) states that soluble amyloid beta (Aß) monomers aggregate into fibrillar plaques and lead to hyperphosphorylated tau protein filaments, neurofibrillary tangles, gliosis, neuronal loss and dementia. However, Aß plaques are not specific to AD, and are seen with other neurodegenerative disorders as well as cognitively normal patients, particularly in the elderly. The causes and clinical significance of Aß plaque formation in cognitively normal subjects is not fully understood although numerous studies support that increased fibrillar Aß on PET is a risk factor for future cognitive decline. Sepsis, a severe systemic inflammatory condition, results in short and long term neurocognitive dysfunction. Acutely, sepsis causes mitochondrial dysfunction and oxidative damage. Longer-term brain dysfunction following sepsis is poorly understood. We have shown a transient increase in cytokines and soluble Aß monomers in the rat brain with experimental sepsis (LPS) but progressive accumulation of Aß neuritic plaques throughout the interval of observation (7-9 d). Preliminary RNAseq analysis suggests increased levels of transcripts with LPS that may affect formation, stabilization or reduced clearance of neuritic Aß plaques. We hypothesize that sepsis and other systemic inflammatory conditions result in neuroinflammation, contribute to Aß neuritic plaque burden and increase the risk of cognitive dysfunction. We proposed to clarify the molecular basis, neurocognitive features, and long-term outcome of sepsis-induced brain dysfunction. Inherent in the success of this goal is the development of non-invasive imaging tools to track acute and chronic neuropathological manifestations of sepsis. The specific aims are: (1) To determine whether Aß neuritic plaques that accumulate in the rat sepsis model eventually resolve or whether they result in the same downstream neuropathological consequences as occur in AD. This will be compared, spatially and temporally, to evidence for mitochondrial dysfunction and oxidative damage; (2) to define molecular events resulting from experimental sepsis that mediate Aß plaque formation and neuronal damage based on our preliminary RNAseq data; (3) To perform longitudinal neurocognitive tests in rats to identify cognitive abnormalities resulting from experimental sepsis, and correlate these findings with neuropathology in regions implicated as abnormal; (4) To perform longitudinal studies with and without aggressive immune modulation to determine whether LPS-induced Aß plaque formation, associated neuropathological lesions, mitochondrial dysfunction and associated neurocognitive abnormalities can be pharmacologically mitigated or reversed; (5) to develop imaging strategies that best identify both acute and chronic brain pathology resulting from experimental sepsis, and correlate changes in these imaging findings with pharmacologic immune modulation (aim 4). These will include advanced microPET and MRI techniques. These imaging measures will be validated by relevant tissue correlates in the brain itself.

Project Summary We are proposing to update our current Bruker Biospin Advance I 70/30 USR pre-clinical MR imaging system (originally set up in 2004) to an Avance III HD (AV3) console upgrade with Paravision 6 software, not including the 7T (Tesla) 30 cm magnet, gradient and shim coils, gradient/shim cooling, RF amplifiers, shim amplifier, and Farady cage. For the proposed application, the major projects consist of 5 funded NIH R01 projects (Towner, Sun, Ahamed, Wu, and Deshmukh), a pending NIH R01 application (Srinivasan), a funded NIH R21 project (Plafker), and a funded American Cancer Association project (Woo), as well as 4 minor projects (Towner, Deak, Chen), primarily from two institutions (Oklahoma Medical Research Foundation (OMRF) and the University of Oklahoma Health Sciences Center ( OUHSC)), that will all directly benefit from the AV3 and Paravision 6 equipment upgrade. The proposed Advance III HD hardware and Paravision 6 upgrades to the 70/30 USR Bruker Biospin small animal MRI system will allow researchers in Oklahoma, particularly OMRF and OUHSC, to obtain faster, non-distorted imaging data for neurological, cardiac and tumor investigations. The projects range in biomedical research from neurological diseases (Towner (OMRF) ? septic encephalopathy; Plafker (OMRF) ? optic neuritis; Deak (OUHSC) ? Alzheimer's disease), cardiac diseases (Sun (OUHSC) ? cardiomyopathy; Ahamed (OMRF) ? cardiac fibrosis), cancer (Wu (OUHSC) ? lung cancer; Ramesh (OUHSC) ? nanoparticle-based gene delivery for cancer treatments; Woo (OUHSC) ? anti-angiogenic therapy for ovarian cancer; Chen (UCO) ? metastatic tumor therapy using combined laser and immunoadjuvant therapy), autoimmune diseases (Plafker (OMRF) ? autoimmune demyelinating optic neuritis; Deshmukh (OMRF) ? Sjogren's syndrome); vascular-related diseases (Srinivasan (OMRF) ? lymphatic system formation and lymphedema); to obesity (Towner ? molecular alterations).