In previous studies, we have shown that mefloquine disrupts calcium homeostasis in neurons by depletion of endoplasmic reticulum (ER) stores, followed by an influx of external calcium across the plasma membrane. In this study, we explore two hypotheses concerning the mechanism(s) of action of mefloquine. First, we investigated the possibility that mefloquine activates non-N-methyl-D-aspartic acid receptors and the inositol phosphate 3 (IP3) signaling cascade leading to ER calcium release. Second, we compared the disruptive effects of mefloquine on calcium homeostasis to those of ionomycin in neuronal and nonneuronal cells. Ionomycin is known to discharge the ER calcium store (through an undefined mechanism), which induces capacitative calcium entry (CCE). In radioligand binding assays, mefloquine showed no affinity for the known binding sites of several glutamate receptor subtypes. The pattern of neuroprotection induced by a panel of glutamate receptor antagonists was dissimilar to that of mefloquine. Both mefloquine and ionomycin exhibited doserelated and qualitatively similar disruptions of calcium homeostasis in both neurons and macrophages. The influx of external calcium was blocked by the inhibitors of CCE in a dose-related fashion. Both mefloquine and ionomycin upregulated the IP3 pathway in a manner that we interpret to be secondary to CCE. Collectively, these data suggest that mefloquine does not activate glutamate receptors and that it disrupts calcium homeostasis in mammalian cells in a manner similar to that of ionomycin.Mefloquine is an antimalarial with utility for chemoprophylaxis and treatment. The drug has been associated with adverse central nervous system (CNS) effects in a dose-related manner (reviewed in reference 14). As a consequence, its continued use in an otherwise healthy population for prophylaxis is controversial. The precise etiology of mefloquine-induced adverse effects is unknown. There is clinical evidence that P glycoprotein polymorphisms are associated with adverse neurological outcomes (1). Numerous putative CNS targets have been proposed (reviewed in reference 12) in either in vitro or ex vivo contexts. Recent studies in our laboratory demonstrated mefloquine induction of brain stem histopathology consistent with direct cellular neurotoxicity in rats (12) and disruption of neuronal calcium homeostasis in vitro (14). The latter effect involves discharge of the endoplasmic reticulum (ER) calcium store and induction of a subsequent influx of calcium into the cell from the extracellular space (14). Initially we suspected that this effect might occur as a consequence of the inhibition of the thapsigargin-sensitive sarcoplasmic reticulum/ER Ca 2ϩ -ATPase (SERCA), followed by subsequent triggering of capacitative calcium entry (CCE). However, in subsequent gene expression studies, we observed that mefloquine failed to induce the downstream stress responses typical of thapsigargininduced ER calcium depletion (13). These observations suggested either that the interaction of mefloquine ...

The clinical potential of mefloquine has been compromised by reports of adverse neurological effects. A series of 4-quinolinecarbinolamines were compared in terms of neurotoxicity and antimalarial activity in an attempt to identify replacement drugs. Neurotoxicity (MTT [thiazolyl blue reduction] assay) was assessed by exposure of cultured embryonic rat neurons to graded concentrations of the drugs for 20 min. The 50% inhibitory concentration (IC 50 ) of mefloquine was 25 M, while those of the analogs were 19 to 200 M. The relative (to mefloquine) therapeutic indices of the analogs were determined after using the tritiated hypoxanthine assay for assessment of the antimalarial activity of the analogs against mefloquine-sensitive (W2) and -resistant (D6 and TM91C235) Plasmodium falciparum strains. Five analogs, WR157801, WR073892, WR007930, WR007333, and WR226253, were less neurotoxic than mefloquine and exhibited higher relative therapeutic indices (RTIs) against TM91C235 (2.9 to 12.2). Conventional quinoline antimalarials were generally less neurotoxic (IC 50 s of 400, 600, and 900 for amodiaquine, chloroquine, and quinine) or had higher RTIs (e.g., 30 for halofantrine against TM91C235). The neurotoxicity data for the 4-quinolinecarbinolamines were used to develop a three-dimensional (3D), function-based pharmacophore. The crucial molecular features correlated with neurotoxicity were a hydrogen bond acceptor (lipid) function, an aliphatic hydrophobic function, and a ring aromatic function specifically distributed in the 3D surface of the molecule. Mapping of the 3D structures of a series of structurally diverse quinolines to the pharmacophore allowed accurate qualitative predictions of neurotoxicity (or not) to be made. Extension of this in silico screening approach may aid in the identification of less-neurotoxic quinoline analogs.

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Mefloquine has been one of the more valuable antimalarial drugs but has never reached its full clinical potential due to concerns about its neurologic side effects, its greater expense than that of other antimalarials, and the emergence of resistance. The commercial development of mefloquine superseded that of another quinolinyl methanol, WR030090, which was used as an experimental antimalarial drug by the U.S. Army in the 1970s. We evaluated a series of related 2-phenyl-substituted alkylaminoquinolinyl methanols (AAQMs) for their potential as mefloquine replacement drugs based on a series of appropriate in vitro and in vivo efficacy and toxicology screens and the theoretical cost of goods. Generally, the AAQMs were less neurotoxic and exhibited greater antimalarial potency, and they are potentially cheaper than mefloquine, but they showed poorer metabolic stability and pharmacokinetics and the potential for phototoxicity. These differences in physiochemical and biological properties are attributable to the "opening" of the piperidine ring of the 4-position side chain. Modification of the most promising compound, WR069878, by substitution of an appropriate N functionality at the 4 position, optimization of quinoline ring substituents at the 6 and 7 positions, and deconjugation of quinoline and phenyl ring systems is anticipated to yield a valuable new antimalarial drug.In the late 1960s to early 1970s, Plasmodium falciparum malaria in Southeast Asia had begun to develop resistance to all of the available antimalarial drugs (6). Cure rates were 11 to 20% and 26 to 50% for chloroquine and quinine, respectively, and had declined to only 90% for the triple combination of quinine/pyrimethamine/dapsone (6). All of these regimens were associated with adverse side effects (6). As a consequence, the U.S. Army began routinely employing two experimental antimalarial drugs, WR030090 and WR033063, for the treatment of recrudescent malaria infections at the Walter Reed Army Medical Center (6). Subsequent field trials demonstrated that WR030090, a quinolinyl methanol, exhibited cure rates of at least 88% and was better tolerated than quinine (6, 21).Shortly thereafter, mefloquine was discovered and was developed commercially by Hoffman La Roche and the U.S. Army. Mefloquine exhibited a long half-life in humans, and this desirable property facilitated its administration as a single dose for malaria treatment and as a once-weekly dosing for prophylaxis (50). In contrast, WR030090 was only partially effective as a prophylactic agent, required a dosing regimen similar to that of quinine to effect cures, and was subsequently abandoned (6,9,21,30). However, it is important to recognize that this occurred because of unfavorable pharmacokinetic characteristics, not as a consequence of unacceptable toxicity.Mefloquine combined with artesunate constitutes one of the most effective combination agents for treatment of malaria (41). Mefloquine is also the only once-weekly drug approved for malaria chemoprophylaxis in the United States that...