1998 — 2002 |
Prather, Paul L |
R29Activity Code Description: Undocumented code - click on the grant title for more information. |
Opioid Control Mechanisms of Signal Transduction @ University of Arkansas Med Scis Ltl Rock
DESCRIPTION (Applicant's Abstract): The long term objective of this project is to determine how activation of mu and delta-opioid receptors leads to distinct intracellular signals. Although mu- and delta-opioid receptors may relay intracellular messages in a similar fashion, a clear distinction in the abuse and dependence producing potential of opioids suggests significant differences are yet to be discovered. For this project, a cellular model has been developed in which cloned mu and delta-opioid receptors in stably transfected GH3 cells interact differently with two intracellular effectors, adenylyl cyclase and Ca++ channels. This unique and important model will be used to identify fundamental differences between mu- and delta-opioid signal transduction cascades. This will be accomplished by the systematic analysis of the interactions between opioid receptors, G proteins and effectors in these unique clones. First, the effect of receptor density on mu- and delta-coupling to effectors will be determined. Second, signal transduction in clones expressing only mu-, only delta, or both mu and delta-opioid receptors will be examined. Third, the association of opioid receptors with G proteins will be determined by purification of agonist-stimulated receptor-G protein complexes. Fourth, the activation of G proteins by opioid receptors will be studied using agonist-induced incorporation of [32P]azidoanilido-GTP into G-alpha subunits. Fifth, the composition of the heterotrimeric G proteins (G-alpha, G-beta- and G-gamma subunits) responsible for coupling mu and delta- receptors to effectors will be confirmed by the use of antisense oligonucleotides targeting specific G protein subunits. Finally, the association of Ca2+ channels with G alpha and/or G-beta,gamma subunits will be assessed after the immunoprecipitation of G protein/Ca2+ channel complexes. Understanding basic differences in the way mu and delta-opioid receptors relay information intracellularly could lead to the development of new methods for the treatment of opioid abuse and pain management.
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0.97 |
2001 — 2004 |
Prather, Paul L |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Cannabinoid Agonist Regulation of Signal Transduction @ University of Arkansas Med Scis Ltl Rock
DESCRIPTION(Adapted from applicant's abstract): This is a new application from an investigator who currently holds an R29 Award, but has never had an RO1 award. He proposes three specific aims to establish the multiplicity of coupling of cannabinoid (CB) receptors with G proteins and to determine which G proteins mediate downstream signaling in brain. In specific aim #1, he will investigate the interaction of CB1 receptors in rat brain with specific G-alpha subunits in response to full, partial and inverse cannabinoid agonists derived from four structural classes. He will use a newly developed elegant technique in which receptor-activated G protein autoradiography is performed using brain sections with a GTP photoaffinity ligand. The studies in the brain slices will be augmented by isolating key areas from each slice, solubilizing the sections and determining which G proteins are specifically activated by high resolution gel techniques. In some cases, the gel techniques will be augmented by immunoprecipitations with specific G protein antibodies. Full concentration-effect curves will be employed using membranes prepared from those specific brain regions. The second specific aim is to correlate CB ligand-selective G protein regulation with specific effects or coupling in cerebellar granular cells. He will use cerebellar granular cells as primary neuronal cultures to compare the potency and efficacy of selected full, partial and inverse cannabinoid agonists to activate specific G proteins in the membrane preparations. He will next determine which pertussis-toxin sensitive intracellular effectors are regulated by CB1 receptors in those cells. The effectors to be evaluated will include the inhibition of adenylyl cyclase activity, inhibition of voltage-dependent calcium currents, activation of MAPK activity and finally, activation of inwardly rectifying potassium channels. The third portion of this aim involves identifying the specific G protein alpha subunits responsible for the coupling to those effectors by employing antisense oligonucleotides directed against specific G protein alpha subunits. He will also determine the correlation between the potency and efficacy of different CB ligands to coupled individual G proteins and regulates the distinct intracellular effectors. The third specific aim is to investigate potential mechanisms underlying CB agonist specific trafficking of responses in CB1 receptor transfected C6 glioma cells. The hypothesis to be tested here is that the ratio of CB1 receptors to G proteins within cells will contribute to the ability of different agonists to selectively traffic responses. He will transfect C6 glioma cells with the cDNA's encoding CB1 receptors to generate clones expressing a wide range of receptor densities. He will then compare the potency and efficacy of selected full, partial and inverse CB agonist to activate specific G proteins and effectors as a function of receptor density.
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0.97 |
2008 — 2009 |
Prather, Paul L |
R21Activity Code Description: To encourage the development of new research activities in categorical program areas. (Support generally is restricted in level of support and in time.) |
Selective Cb2 Cannabinoid Agonists as Candidate Therapeutics For Als @ Univ of Arkansas For Med Scis
DESCRIPTION (provided by applicant): Project Summary: Amyotrophic Lateral Sclerosis (ALS) is a neurodegenerative disease characterized by progressive motor neuron loss, paralysis and death within 2 to 5 years of diagnosis. Currently, no effective pharmacological agents exist for the treatment of this devastating disease. Neuroinflammation may greatly influence the progression of motor neuron loss during ALS. Cannabinoids produce anti-inflammatory actions via CB1 and CB2 receptors and delay the progression of pathologic conditions characterized by neuroinflammation. In G93A-SOD1 (G93A) mutant mice, the most well-characterized animal model of ALS, we demonstrate that mRNA, receptor binding and function of CB2, but not CB1, receptors are dramatically and selectively upregulated in the spinal cords of G93A mice in a temporal pattern closely paralleling disease progression. More importantly, daily injections of two structurally diverse selective CB2 agonists (AM-1241 and L- 759,633) initiated at symptom onset, markedly maintain motor function and increase the survival interval after disease onset. Therefore, we propose that selective CB2 agonists may represent a novel therapeutic modality for ALS. While CB2 agonists may prove useful for this devastating neurodegenerative disease, their development as pharmaceutical agents has been fundamentally hindered by their relative insolubility in aqueous, or other biocompatible, vehicles. Indeed, several lines of evidence indicate that the actual maximal efficacy of AM-1241 might have been underestimated due to less than optimal drug delivery by the vehicle employed in our preliminary studies. As such, we propose that pharmacokinetic studies are needed to determine the most efficient vehicle, route of administration and/or dose required to produce the maximal efficacy of AM-1241 in G93A mice. The current A1 revision of this R21 application will "identify candidate therapeutics" and "obtain preliminary data on the efficacy of candidate therapeutics" for ALS by conducting the following two Specific Aims: Specific Aim 1 will identify novel CB2 agonists as candidate therapeutics for ALS by screening a series of indole and classic cannabinoid-based CB2 agonists provided by Dr. John W. Huffman (Clemson University, SC) for their ability to slow disease progression and prolong survival of G93A mice. Specific Aim 2 will optimize the therapeutic potential of AM-1241, a CB2 agonist with proven efficacy in the G93A mouse model of ALS. This will be accomplished by employing the vehicle and route of administration demonstrated by pharmacokinetic studies to produce the greatest delivery of AM-1241 to serum and spinal cords. Selective CB2 agonists identified by this project could be the first efficacious drugs for the management of ALS. Most importantly, our long-term goal is to translate results from these studies into a future clinical trial in ALS patients. Project Narrative: Amyotrophic Lateral Sclerosis (ALS) is a neurodegenerative disease characterized by progressive motor neuron loss, paralysis and death within 2 to 5 years of diagnosis. Currently, no effective drugs exist for the treatment of this devastating disease. Based on some exciting preliminary evidence, this project will seek to discover new drugs called "CB2 agonists" that could be the first class of effective drugs to prolong the lives of ALS patients.
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0.972 |
2016 — 2020 |
Prather, Paul L |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Synthetic Cannabinoid Toxicity: Role of Biotransformation @ Univ of Arkansas For Med Scis
? DESCRIPTION (provided by applicant): The terms K2 and Spice refer to any number of commercial products usually sold as legal marijuana. These products contain dangerous synthetic cannabinoid (SCBs) that are presumed to possess psychoactive properties similar to ?9-tetrahydrocannabinol (?9-THC), the natural cannabinoid found in marijuana. ?9-THC and SCBs both produce psychotropic actions by activating CB1 cannabinoid receptors (CB1Rs) in the CNS. However, SCBs are a chemically diverse group of compounds that are structurally distinct from ?9-THC, and thus detection of their use is difficult and has led to widespread abuse. Medical use of marijuana and ?9-THC has been shown to be safe. In marked contrast, no information is known concerning the safety or efficacy of any SCB found in K2, and reports suggest that many clinical effects of K2 products are distinct from those produced by marijuana and may present health risks. In this regard, our preliminary analysis of urine samples from SCB users by LC-MS/MS suggests that levels of SCB metabolites correlate with clinical symptoms that may be life threatening. Furthermore, we reported that several hydroxylated metabolites of SCBs retain high affinity and activity at CB1R and CB2Rs, and dramatically increase acute effects of parent SCBs. Therefore, in an individual user, the physiological effects of SCBs may represent an entourage effect caused 1) by the distinct blend of SCBs in a given product, and 2) further influenced by the individual's metabolic capacity to transform SCBs into distinct Phase I and II active metabolites. Thus, it is important to define the metabolic profile of SCBs in humans and their biological activity at CB1Rs and CB2Rs. The goal of this project is to elucidate the biodisposition, biotransformation, and biological activity of SCBs and their metabolites at CB1Rs and CB2Rs in humans, and correlate these findings with acute and chronic adverse effects in mice. We will test the hypothesis that in vivo hydroxylation of SCBs by cytochromes P450 (CYPs) and subsequent conjugation by UDP-glucuronosyltransferases (UGTs) and sulfotransferases (SULTs) produces a complex mixture of high affinity CB1R and CB2R agonists, antagonists, and inverse agonists. These metabolites acting in concert with parent SCBs produce the distinct pharmacologic effects and toxicity of SCBs in humans. Our interdisciplinary team will explore this hypothesis by four Specific Aims. Aim 1 will employ LC-MS/MS to identify in human urine the primary and secondary metabolites of 9 high priority SCBs abused in K2 products. Clinical symptom profiles will also be collected for each patient. Experiments in Aim 2 will characterize the human Phase I and II enzymes responsible for the in vitro metabolism of SCBs. In Aim 3, SCBs and their metabolites will be examined for the ability to bind to and activate human CB1Rs and CB2Rs. Finally, studies in Aim 4 will determine the pharmacokinetic profile of SCBs and determine if these compounds and their metabolites elicit cannabimimetic effects in mice. This collaborative translational project will provide information concerning the metabolism, pharmacology and toxicology of SCBs to identify likely health risks to the public.
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0.972 |
2020 — 2021 |
Prather, Paul L |
T32Activity Code Description: To enable institutions to make National Research Service Awards to individuals selected by them for predoctoral and postdoctoral research training in specified shortage areas. |
Systems Pharmacology and Toxicology Training Program @ Univ of Arkansas For Med Scis
Project Summary/Abstract: Over the past few decades, our knowledge of the mechanisms by which cells interact with drugs and toxins has exploded due to new molecular analysis techniques and the application of genomic methods. Accordingly, the emphasis of graduate education in the disciplines of pharmacology and toxicology needs to shift from a reductionist view to a systems approach in which doctoral students are comprehensively trained so they can formulate a strategy to solve important biological questions not only at the molecular, cellular, and tissue levels but also at the whole-animal level. Systems pharmacology and toxicology describes a field of study that considers the broad view of drug action. A systems approach using in vivo animal models is necessary to establish efficacy, safety and the pharmacodynamic/pharmacokinetic profile of candidate drugs but there is a shortage of students trained in this area. This Systems Pharmacology and Toxicology (SPaT) Program is designed for PhD students in their second year of graduate study pursuing dissertation research projects in the pharmacological sciences. Trainees (2/year for up to 2 years of support) and training faculty (30 mentors and 4 teachers) are drawn from the Graduate Program in Interdisciplinary Biomedical Sciences and the MD/PhD Program. The SPaT program will train students to use an in vivo approach to answering relevant questions in pharmacology and toxicology with emphasis on metabolism, drug design, pharmacodynamics, pharmacokinetics, and signaling. The rationale for SPaT is that this type of training provides students with a much broader perspective on pharmacology and toxicology that better prepares them to be leaders of multi-disciplinary research teams in the pharmacological sciences. We will integrate SPaT into PhD training programs already active at our graduate training sites in the Little Rock area that include faculty/scientists in the Colleges of Medicine, Pharmacy, and Public Health on the UAMS, the Arkansas Children's Hospital campus, and at the Food and Drug Administration-funded National Center for Toxicological Research. The unique focus of SPaT is training with in vivo systems pharmacological and toxicological approaches and concepts. The objective of SPaT is to provide in vivo pharmacology and toxicology training that complement the cellular and molecular training that students receive in their home programs. The training program consists of didactic training in pharmacology, toxicology, physiology, pharmacokinetics, metabolism, biostatistics, grant writing, and the Responsible Conduct of Research along with laboratory research using an in vivo model of human disease. The SPaT program will also provide strong mentoring, extensive networking, and teaching and leadership opportunities for its trainees through its programmatic activities.
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0.972 |