Michael A. Rogawski - US grants

DESCRIPTION (provided by applicant): The rodenticide tetramethylenedisulfotetramine (TETS) and the organophosphorus (OP) pesticide parathion are considered credible terrorist threat agents. Current medical countermeasures for acute TETS or OP intoxication can prevent mortality but do not sufficiently protect the CNS from persistent seizures and/or permanent injury. Therefore, new and more effective countermeasures must be developed to facilitate better medical treatment of civilians, first responders and military personnel following exposure to acutely toxic levels of TETS, parathion and similar chemical threat agents. The goals of the proposed research are to develop rodent models of TETS- and parathion-induced seizures and then use these models to identify potential therapeutic agents. We seek to identify agents that can protect against the development of seizures or treat seizures once they have begun and/or can prevent irreversible neuronal injury. We plan to evaluate a 2,3- benzodiazepine AMPA receptor antagonist and a soluble epoxide hydrolase inhibitor (sEHi). We have recently shown that AMPA receptor antagonists provide sustained seizure protection and are able to block seizures when administered at later times when there is refractoriness to diazepam, the benzodiazepine typically used to treat acute TETS and OP intoxication. We and others have data indicating that sEHi also exhibit anti- epileptic properties. Perhaps more important in light of recent evidence suggesting that the use of anti- inflammatory compounds in combination with standard antidote significantly decreases neuronal damage in acute OP intoxication, we have demonstrated that sEHi are also potent anti-inflammatory compounds. These observations suggest the potential for AMPA receptor antagonists and sEHi to significantly improve the clinical management of acute TETS and parathion intoxication by extending the therapeutic window and enhancing neuroprotection. To test this hypothesis, we will address two specific aims: (1) Develop rodent models of acute TETS and parathion intoxication;and 2) Determine whether AMPA receptor antagonist and/or sEHi are of therapeutic benefit in acute TETS or parathion intoxication when administered prior to or after exposure to the chemical threat agent. Endpoints that will be measured across all Aims include time to onset, frequency and duration of clonic and tonic seizures, EEG, histological measures of neuropathology and blood levels of TETS, AMPA receptor antagonist and sEHi. AMPA receptor antagonists and sEHi are currently undergoing human clinical trials and have demonstrated an excellent safety record;which will facilitate translation of positive findings in these rodent models to human studies. In summary, this project will develop rodent models of acute TETS and parathion intoxication that will be useful to the field beyond the proposed studies, and it will generate critical information on novel treatment approaches of practical value for two diverse classes of chemical threat agents. PUBLIC HEALTH RELEVANCE: The rodenticide tetramethylenedisulfotetramine (TETS) and the organophosphorus (OP) pesticide parathion are considered credible terrorist threat agents. Current medical countermeasures for acute TETS or OP intoxication can prevent mortality but do not sufficiently protect the CNS from persistent seizures and/or permanent injury. The goal of this research project is to test the hypothesis that AMPA receptor antagonists and/or inhibitors of soluble epoxide hydrolases will significantly improve outcome following exposure to acutely toxic levels of TETS and parathion by extending the therapeutic window for seizure protection and enhancing neuroprotection.

Summary: The overall objective of this project is to identify new, more effective medical countermeasures for the acute seizures produced by organophosphate (OP) anticholinesterase inhibitor and GABAA receptor antagonist chemical threat agents. By preventing or stopping the acute seizures, these countermeasures will improve the survival and long-term outcome of individuals exposed to seizure-inducing doses of these agents. We have selected two OP agents diisopropylfluorophosphate (DFP) and soman (0-pinacolyl methylphosphonofluoridate; GD) and one GABAA receptor antagonist tetramethylenedisulfotetramine (TETS) to represent the two broad classes of chemical threat agents. Soman, a representative of the OP neurotoxin chemical warfare agents (nerve agents), is classified as a Schedule 1 substance under the Chemical Weapons Convention of 1993. DFP, while less potent than the nerve agents in the Chemical Weapons Convention toxic chemicals list, has nearly identical effects. As a volatile agent that could rapidly spread inside closed areas, it could cause many injuries and deaths and is therefore a credible threat agent. TETS, a representative of the GABAA receptor antagonist convulsants is a highly lethal toxin that has been used as a rodenticide but is now banned in most countries of the world. Nevertheless, lethal poisoning with TETS occur regularly. On a mg basis, TETS is equal in lethality to the OP chemical warfare agents. Recently, we have had the opportunity to characterize the convulsant activity of TETS in mice and rats when administered by the i.p., i.v., oral and intracerebroventricular routes. As a result of this work, we have developed the first animal model in which to evaluate potential treatments for TETS when administered before and after exposure. In this project, DFP and soman are administered to rats and TETS is administered to mice to induce seizures. The animals are treated before or at various intervals following exposure with various potential therapeutic agents (or combinations) to assess the ability of the test agent to prevent or terminate behavioral or electrographic seizure activity. Testing in the OP models occurs in a step-wise fashion, first against DFP seizures because DFP has reduced safety concerns and can be used in a regular laboratory environment, and then with soman, which requires extreme safety precautions available at the US Army Medical Research Institute of Chemical Defense. All test agents are compared with diazepam, the current standard of care for the emergency treatment of chemical exposure seizures. Diazepam may be effective if administered early in the course of seizures but seizures may recur and the drug may be inactive if administered later in the course of seizures. In addition, diazepam can depress blood pressure and respiration. A further concern is that diazepam is erratically absorbed when administered by the i.m. route (as with the injector systems stockpiled for use in mass casualty situations). We are seeking agents (or combinations) that improve upon diazepam with respect to any of these liabilities. We have identified a series of potential therapies (Tier 1) that are either currently approved for sale in the U.S. or for which human data is available and can therefore be rapidly deployed. In addition, novel potential therapies that are new molecular entities (Tier 2) will be developed in Core B and Project 3. Therapies that exhibit superior activity to diazepam will be studied in Project 2 for their ability to mitigate seizure-induced brain damage. Finally, in order to advance the most promising therapies, we will complete various additional studies, including formulation development and safety, toxicology and ADME studies in conjunction with Cores A and B, to assist in making go/no-go decisions regarding further development.

DESCRIPTION (provided by applicant): This project will develop and implement a training program in neurotherapeutics discovery and development for faculty members and advanced postdoctoral fellows, centered around a 3-day short course that will provide the trainees with the various knowledge elements required to discover and advance a neurotherapeutic agent to IND. Following the short course, the training program will continue for a two-year period in which students will have individualized mentoring and assessment. The training, which is designed to be applicable to diverse diseases of the nervous system, will equip students with a broad understanding of the various component steps in the neurotherapeutics drug discovery and development process. Students will learn how to identify a good drug discovery target; how to construct an assay; the elements of medicinal chemistry; how to conduct animal efficacy testing; the principles of ADME studies, safety testing, and formulation; the principles of experimental medicine and biomarkers; the steps required to prepare an IND document and the principles for interacting with the FDA; the principles of intellectual property as they relate to neurotherapeutics discovery and development; and how to seek funding for academic drug discovery research. They will also receive training in responsible conduct of research. Students will be equipped with the skills to develop and coordinate an entire drug discovery and development effort, and to work collaboratively with experts in each of the component areas. The training will combine didactic lectures with active engagement activities in which the students will be challenged to utilize the lecture material to work through their own drug discovery project plan with the guidance of the area experts. The 3-day short course (followed by two-years of mentorship and assessment) will be offered annually, a total of five times. PUBLIC HEALTH RELEVANCE: This project will train researchers to discover and develop new treatments for disorders of the brain and nervous system so as to reduce disability and lead to improved quality of life.

Project Summary/Abstract This project will develop and implement a training program in neurotherapeutics discovery and development for faculty members and advanced postdoctoral fellows, centered around a 3½-day short course that will provide the trainees with the various knowledge elements required to discover and advance a neurotherapeutic agent to IND. Following the short course, the training program will continue for a two-year period in which students will have individualized mentoring and assessment. The training, which is designed to be applicable to diverse diseases of the nervous system, will equip students with a broad understanding of the various component steps in the neurotherapeutics drug discovery and development process. Students will learn how to identify a good drug discovery target; how to construct an assay; the elements of medicinal chemistry; how to conduct animal efficacy testing; the principles of ADME studies, safety testing, and formulation; the principles of experimental medicine and biomarkers; the steps required to prepare an IND document and the principles for interacting with the FDA; the principles of intellectual property as they relate to neurotherapeutics discovery and development; and how to seek funding for academic drug discovery research. They will also receive training in responsible conduct of research. Students will be equipped with the skills to develop and coordinate an entire drug discovery and development effort, and to work collaboratively with experts in each of the component areas. The training will combine didactic lectures with active engagement activities in which the students will be challenged to utilize the lecture material to work through their own drug discovery project plan with the guidance of the area experts. The 3½-day short course (followed by two-years of mentorship and assessment) will be offered annually, a total of five times.

Project Summary/Abstract This project seeks to advance the development of (2S,3S)-sec-butylpropylacetamide [(2S,3S)-SPD] for the treatment of chemical threat agent seizures. A priority of the NIH CounterACT Program is the development of medical countermeasures to treat seizures induced by anticholinesterase nerve agents such as soman and sarin or GABA-inhibiting agents such as tetramethylenedisulfotetramine (TETS). Benzodiazepines, the current standard-of-care therapies, fail to stop seizures induced by such agents when there is more than a brief delay between onset of seizures and administration of the treatment agent. In rodents, (2S,3S)-SPD is a broad- spectrum antiseizure agent that effectively stops benzodiazepine-resistant behavioral and electrographic status epilepticus in the lithium-pilocarpine model. Moreover, studies conducted at the U.S. Army Medical Research Institute of Chemical Defense indicate that (2S,3S)-SPD rapidly stops soman-induced benzodiazepine- resistant SE in rats at delayed time points, an effect not shared by benzodiazepines or other antiseizure drugs. Therefore, (2S,3S)-SPD has promise to provide a major improvement to the current standard-of-care for the treatment of nerve agent seizures. The objective of the proposed research is to generate the data required to advance the development of an intramuscular formulation (2S,3S)-SPD so that it is available as a medical countermeasure for nerve agent seizures in the setting of a civilian exposure. A scalable synthesis route for (2S,3S)-SPD will be developed. The API will be formulated in a solution designed for intramuscular administration, a ?mass-casualty-friendly? route of delivery that offers fast onset of action. Initial toxicology and safety studies will be conducted in animals, including studies to assess safety in both males and females and in special populations such as children. Additional proof-of-concept and dose-ranging studies in relevant animal models will be conducted, including in a model of TETS-induced status epilepticus. The proposed research will assess the potential of (2S,3S)-SPD as an improved medical countermeasure to treat seizures induced by diverse chemical threat agents. On completion of the project, a comprehensive set of data will be available to advance the development of (2S,3S)-SPD as an improved treatment for chemical threat agent seizures.