Organophosphate (OP) nerve agents have been deployed as weapons of mass destruction in warfare arenas and in recent terrorist attacks; posing significant risks to military personnel, civilians, first responders and healthcare provider. The classic OP nerve agents such as sarin, soman, tabun, and VX, along with related OP pesticides represent some of the most toxic materials known to humankind. Their toxicity is principally due to their ability to irreversibly bind to acetylcholinesterase (AChE) in the CNS and blood. This process deactivates AChE and renders it unable to process acetylcholine in the synapse; resulting in excitotoxic levels of acetylcholine and hypersensitization of muscarinic and nicotinic receptors. This overload of excitotoxic chemicals incapacitates the victim via cholinergic toxidrome, resulting in seizures, paralysis, mucous secretions, eye irritation, cardiorespiratory depression, GI distress, and eventually, death. Additionally, OP exposure initiates a sequelae of mechanisms leading to delayed neurodegeneration in the CNS; including heightened excitotoxicity at other receptors and transporters and progressive demyelination and degradation of nerve axons. Cases of exposure to OP agents, whether intentional (warfare or terrorism) or accidental (pesticide mishandling), number in excess of 2 million per year; resulting in the deaths of several hundred thousand victims per year worldwide. Currently available countermeasures for OP poisoning involve the simultaneous administration (via autoinjectors) of three different agents: (1) an anticholinergic agent, (2) an anticonvulsant agent, and (3) a pyridinium oxime-bearing AChE reactivator. While the anticholinergic and anticonvulsant agents work to block the effects of excess acetylcholine, the oxime-based reactivator plays the key role of cleaving the phosphorylated serine esteratic site to release the active AChE molecule. However, currently used agents possess poor ADME properties due to their charged nature. Thus, significant deficiencies in current OP countermeasures include: (1) the requirement for simultaneous multi-therapeutic modality treatments with complicated dosing regiments, (2) poor CNS-permeability of currently used pyridinium oximes, and (3) the complete lack of efficacy in any of the current treatments to counteract the long-term neurodegenerative sequelae that manifests after prolonged exposure to nerve agents. It has been established that inhibitors of poly(ADP-ribose)polymerase-1 (PARP-1) are useful neuroprotective agents especially when the neurodegeneration is caused by excitotoxic insult. Inhibitors of PARP-1 have been investigated pre-clinically and clinically for cardiac ischemia, hemorrhagic shock syndrome, stroke, traumatic CNS injury, diabetes, inflammation, and cancer. The prototype PARP-1 inhibitor, benzamide, has been shown to be neuroprotective against soman-induced seizure-related brain damage in the rat. It has been recently suggested that the inherent multiple-pathway complexity of OP-induced long-term neuropathy disorders should be countered with a similarly multi-faceted treatment approach. Hence, the design of small molecules possessing dual- pharmacophores to simultaneously modulate two discreet molecular targets is an important strategy in drug discovery. There is a clear and proven advantage to having a single agent that can modulate multiple targets simultaneously as compared to either a cocktail or a single pill formulated with multi-component agents. A dual acting single agent designed by overlapping pharmacophores would have better ligand efficiency (e.g. reduced MW, LogP) and would provide one dimensional metabolism, pharmacokinetics, and safety profiles as opposed to co-administering two drugs where development metrics are complicated due to possible drug-drug interactions and differential kinetic profiles. It is the aim of this project to discover novel and mechanisticaly unique dual- acting agents which would represent a breakthrough innovative countermeasure against OP chemical threats. Thus, the first innovative aspect of this application is to discover a novel PARP-1 inhibitor that also possesses dual activity as an AChE-reactivator; via merging two known pharmacophores in the same molecule, to provide multiple pharmacological effects and a superior overall pharmacological profile to treat OP-poisoning. A single agent with dual PARP-1 inhibition and AChE-reactivating activity would represent an innovative and novel treatment for OP poisoning because: (1) the PARP-1 inhibition activity would attenuate the excitotoxic cascade of free-radical formation by preserving mitochondrial function and reducing oxidative stress and energetic crisis, and (2) the oxime-bearing fragment would salvage the OP-bound inactivated AChE and thereby reduce excitotoxic acetylcholine levels. The second innovative aspect of our approach is the design and synthesis of dual agents with suitable pharmaceutical profiles, such that they will be able to cross the blood brain barrier, a desired feature that is lacking from current AChE reactivators. The successful identification of two in vitro optimized series of novel, dual-active PARP-1 inhibitor/AChE reactivators in this Phase 1 campaign would provide critical tool compounds for in vivo PK assessment and POC studies in rodent models of OP-intoxication and neurodegeneration. This would set the stage for a final lead optimization campaign focused on identifying pre- clinical leads with well-balanced in vivo PK parameters during an SBIR Phase 2 continuing grant.
Public Health Relevance Statement: Public Health Relevance: Multi-targeted Countermeasures against Acute and Delayed Effects of OP Exposure NARRATIVE Organophosphate (OP) nerve agents have been stockpiled and deployed as weapons of mass destruction. While originally developed as pesticides in the 1930's, their potent effects on the central nervous system (CNS) led to their optimization / large-scale production and their addition to the chemical arsenals of military organizations and extremist groups throughout the world. The classic OP nerve agents such as sarin, soman, tabun, and VX, along with related OP pesticides, represent some of the most toxic materials known to humankind. Their principal mechanism of action involves inhibition of acetylcholinesterase (AChE) which normally hydrolyses acetylcholine in the synapse. This synaptic "flooding" of acetylcholine results in an excitotoxic sequelae including seizures, muscle paralysis, mucous secretions, irritation, blurry vision, cardiorespiratory depression, loss of consciousness, and either immediate or eventual death as a result of progressive neurodegeneration. Intentional or accidental misuse of these agents leads to approximately 2 million cases of OP exposure per year worldwide; resulting in several hundred thousand deaths per year. Currently available countermeasures for OP poisoning are woefully inadequate and involve the administration of a "triple cocktail" including an anticholinergic agent, a benzodiazepine anticonvulsant agent, and an oxime agent which can re-activate the inhibited AChE. However, the well-documented long-term secondary neuronal damage that results from initial exposure to OP poisons is not addressed by the current countermeasures. Therefore, it is the aim of this project to discover novel and mechanistically unique dual-acting agents which could represent a breakthrough innovative countermeasure against OP chemical threats that includes treatment of acute OP-poisoning and mitigation of long-term neurodegenerative effects.
Project Terms: Acetylcholine; Acetylcholinesterase; Acetylcholinesterase Inhibitors; Acute; Address; Anti-Cholinergics; Anticonvulsants; Anxiety; Attenuated; Axon; base; Benzamides; Benzodiazepines; Binding (Molecular Function); Blood; Blood - brain barrier anatomy; Brain Injuries; Cardiac; central nervous system injury; Cessation of life; Charge; Chemicals; cholinergic; Cleaved cell; Clinical; Cognitive deficits; Demyelinations; design; Diabetes Mellitus; Disease; Distress; Dose; drug development; drug discovery; Drug Interactions; Drug Kinetics; drug market; emergency service/first responder; Emotional Disturbance; Equilibrium; excitotoxicity; experience; Exposure to; Eye; Floods; Free Radical Formation; Freedom; Generations; Goals; Grant; Health Personnel; Hemorrhagic Shock; Hydrolysis; improved; In Vitro; in vivo; Inflammation; inhibitor/antagonist; innovation; Intellectual Property; Intoxication; irritation; Ischemia; Kinetics; knowledge base; large scale production; late disease onset; Late-Onset Disorder; Lead; Learning Disabilities; Libraries; Ligands; Link; Malignant Neoplasms; Measures; Mental Depression; Metabolism; Metric; Military Personnel; Mind; Mitochondria; Modality; Molecular Target; Mucous body substance; Muscarinic Acetylcholine Receptor; Muscle; National Institute of Neurological Disorders and Stroke; Nature; Nerve; nerve agent; Nerve Degeneration; nervous system disorder; Neuraxis; Neurons; Neuropathy; neuroprotection; Neuroprotective Agents; neuropsychiatry; Niacinamide; Nicotinic Receptors; novel; organophosphate poisoning; Organophosphates; Oxidative Stress; Oximes; Paralysed; Parents; Pathway interactions; Penetration; Permeability; Pesticides; Pharmaceutical Chemistry; Pharmaceutical Preparations; Pharmacologic Substance; pharmacophore; Phase; pill (pharmacologic); Play; Poly(ADP-ribose) Polymerases; Post-Traumatic Stress Disorders; pre-clinical; Process; progressive neurodegeneration; Property; prototype; public health relevance; Rattus; receptor; Reporting; response; Risk; Rodent Model; Safety; Sarin; scaffold; screening; Seizures; Series; Serine; Site; Small Business Innovation Research Grant; Small Business Technology Transfer Research; small molecule; Soman; Staging; Status Epilepticus; stroke; Synapses; Syndrome; tabun; Terrorism; Therapeutic; tool; Toxic effect; toxic organophosphate insecticide exposure; Toxin; Treatment outcome; Unconscious State; United States National Institutes of Health; Vision; weapons of mass destruction; Work
