Abstract
1,1′-Methylenebis{4-[(hydroxyimino) methyl]-pyridinium} dimethanesulfonate (MMB4 DMS), a member of the bisquaternary pyridinium aldoximes group, is currently being developed by the Department of Defense as adjunct therapy in the case of organophosphorus (OP) nerve agent exposure. This issue of International Journal of Toxicology is dedicated to the MMB4 DMS preclinical advanced drug development activities that comprised the investigational new drug (IND) application so as to initiate a first-in-man phase 1 clinical study.
The OP nerve agents are among the most lethal chemical weapons. They are chemically stable, easily dispersed, highly toxic, and have rapid effects both when absorbed through the skin and via respiration. The pathophysiology of nerve agent intoxication is well understood. The OP nerve agents irreversibly bind to acetylcholinesterase (AChE), causing the phosphorylation and deactivation of AChE. The clinical effects are secondary to acetylcholine (ACh) excess at cholinergic junctions (muscarinic effects) in the central nervous system (CNS) and at the skeletal nerve–muscle junctions and autonomic ganglia (nicotinic effects). Medical counter measures (MCMs) against nerve agents include a combination of up to 4 drug classes: (1) a carbamate (pyridostigmine bromide [PB]) which when orally administered prior to exposure to a nerve agent serves to sequester a pool of AChE by reversibly inhibiting the enzyme so that it cannot be irreversibly bound by nerve agent; (2) an antimuscarinic (atropine) that antagonizes the effects of ACh at postsynaptic muscarinic receptors in peripheral tissues and CNS and prevents early life-threatening symptoms such as bronchoconstriction, excessive bronchosecretion, and impaired respiratory drive; (3) an oxime that reactivates nerve agent-inhibited AChE in peripheral tissues and restores the ability of the enzyme to hydrolyze ACh; and (4) a benzodiazepine (diazepam) for postexposure treatment to prevent or minimize seizure activity though activation of centrally mediated γ-aminobutyric acid pathways.
The only oxime licensed in the United States for the treatment of nerve agent exposure is 2-pyridine aldoxime methyl chloride, 2-pralidoxime (2-PAM). The antidote treatment nerve agent autoinjector (ATNAA) that contains 2-PAM and atropine is the current MCM for nerve agent poisoning. Although 2-PAM has acceptable efficacy against certain nerve agents (e.g., GB [Sarin] and VX {O-ethyl S-[2-(diisopropylamino)ethyl] methylpho-sphonothioate}), it lacks the desired level of efficacy against other agents (e.g., GA [Tabun], GD [Soman], GF [Cyclosarin], and VR N, N-diethyl-2-(methyl-(2-methylpropoxy)phosphoryl)sulfanyl-ethanamine), even when combined with PB pretreatment and atropine and diazepam postexposure. This lack of efficacy toward a broad scope of nerve agents has been a driving factor in the search for more efficacious and safer alternatives to 2-PAM in order to alleviate a serious unmet medical need and, as a result of these efforts, the advanced drug development of MMB4 DMS was initiated. Once realized, the ATNAA will be replaced by the Improved Nerve Agent Treatment System that contains MMB4 DMS and atropine.
The preclinical development program conducted in support of the IND submission to the US Food and Drug Administration (FDA) was composed of a series of pharmacology, pharmacokinetic, and toxicology studies. Pivotal IND-enabling studies were all conducted in compliance with the FDA’s Good Laboratory Practice (GLP) Regulations and 21 Code of Federal Regulations Part 58 for the conduct of nonclinical laboratory studies. The intramuscular (IM) route of administration was used in the nonclinical studies, because this is the intended route of administration in man. Of the toxicology species tested in the repeated dose studies used to calculate the maximum recommended starting dose and appropriate safety factors, rabbits were the most sensitive species to the effects of MMB4 DMS at equivalent doses. In support of the toxicology studies, full toxicokinetic evaluations were established, and comparative pharmacokinetic studies were conducted as well. Pharmacology studies explored the efficacy of MMB4 DMS against a host of conventional nerve agents, and these data were used to establish a minimum effective dose that could be translated to man. In addition, as prescribed by the regulations, safety pharmacology studies that evaluated potential effects on the cardiovascular, pulmonary, and CNS systems were conducted as well. Additionally, optimization of the formulation that would provide sufficient stability characteristics under field conditions was imperative to the success of the program.
This series of articles provides a synopsis of the drug development strategy used for the characterization of MMB4 DMS. More specifically, Dr Clark et al reviewed the development and validation of the GLP program that involved the methodology used to optimize the test article as a nanoparticle suspension, whereas Dixon et al focused on optimizing the EF in order to increase the stability and shelf life of MMB4 with the development of a formulation consisting of MMB4 micro/nanoparticles suspended in cottonseed oil, with the intent that this formulation would be used in a phase 1 clinical trial. Harvilchuck et al found that when evaluated for effectiveness in guinea pigs and Rhesus macaques as an antidote against GF, treatment with MMB4 DMS (in conjunction with atropine and diazepam) resulted in AChE reactivation, high percentages of survival, decreased severity of clinical signs, and improved quality-of-life scores. A further study by Harvilchuck et al investigated a dose of MMB4 DMS that would result in 80% survival following a single cyclosarin challenge, and found survival in the guinea pig was ≥50% at dose levels of 10 mg/kg or more MMB4 DMS, and in Rhesus macaques the survival was 100% at dose levels of ≥0.1 mg/kg MMB4 DMS. Safety pharmacology studies investigating the potential cardiovascular and pulmonary effects in beagle dogs or CNS effects in rats administered MMB4 DMS were investigated by Dr Roche et al. In dogs, 100 mg/kg MMB4 DMS resulted in increased blood pressure, slightly increased heart rate, slightly prolonged QTc, and moderately increased respiratory rate. There were no toxicological effects of MMB4 DMS on neurobehavioral function in rats administered up to 340 mg/kg MMB4 DMS. The results generated from an Absorption, Distribution, Metabolism and Excretion/metabolism study was reported by Dr Lusiak et al in which 14C-MMB4 DMS was administered intramuscularly to rats and rabbits followed by temporal collection of biological samples. The majority of the radioactivity was excreted in the urine of both species with no apparent species or dose differences in the urine excretion pattern. Two metabolites of MMB4 DMS were detected in rat and rabbit urine and were identified as 4-pyridine aldoxime (4-PA) and isonicotinic acid (pyridine-4-carboxylic acid). In addition, the in vitro metabolism of MMB4 DMS as well as drug–drug interactions and protein binding properties of the drug were investigated and reported by Hong et al. The incubation of 14C-MMB4 DMS in hepatocytes resulted in the formation of 4-PA in all species and the formation of isonicotinic acid in rat, rabbit, and monkey hepatocytes. In addition, MMB4 DMS exhibited reversible inhibition in a concentration-dependent manner toward cytochrome P450 (CYP)1A2, CYP2C9, CYP2C19, CYP2D6, and CYP3A4 in human liver microsomes showing the highest inhibition for CYP2D6. Finally, it was found that MMB4 was not extensively bound to plasma protein in any of the species evaluated. Hong et al investigated the pharmacokinetics of MMB4 DMS following a single intravenous administration in the rat, rabbit, and dog and found that the plasma MMB4 concentration versus time profiles were biphasic for all the species tested and fit a 2-compartment model with first-order elimination. There were no overt sex-related differences in MMB4 PK profiles, and the volume of distribution was conserved across the species while demonstrating species-related differences in the elimination of half-life and clearance of MMB4. Finally, Osheroff et al reported on the comparative toxicology following the repeated dose IM administration of MMB4 DMS to Sprague-Dawley rats, New Zealand White rabbits, and Rhesus monkeys. In all the species, the IM injection site displayed inflammation and necrosis following vehicle and test article administration, albeit the severity was increased with MMB4 DMS administration. Additionally, only in the rabbit were there other test article-related effects in the target organs identified as the heart, lungs, and kidneys with clinical pathology, biomarker evaluations, and light microscopic evidence to support these findings. The New Zealand White rabbit was identified as the most sensitive species, and the no-observed adverse effect level (NOAEL) was determined as 50 mg/kg/d; the NOAEL in the rat was 100 mg/kg/d; and the NOAEL in Rhesus monkey was >600 mg/kg/d. Hong et al supported these studies with an article reporting the comparative toxicokinetics of MMB4 DMS in rats, rabbits, dogs, and monkeys following single and repeated IM administration. Dr Hong found that in general, there were no significant gender effects for all the species tested in this study. After a single IM administration, average T max values ranged from 5 to 30 minutes, and that while C max values did not increase dose proportionally, area under the curve to infinity (AUC∞) values did increase in an approximate dose-proportional manner. The MMB4 DMS displayed greater than 80% absolute bioavailability values for rats, rabbits, and dogs. A comparison of exposure parameters revealed that C max and AUC values decreased in monkeys given 450 and 600 mg/kg MMB4 DMS following repeated administrations for 7 days.