Proinflammatory cytokine overproduction by activated glia offers a pathophysiology progression mechanism that can be targeted in new therapeutic development strategies with potential for disease modification in multiple CNS disorders, including Alzheimer's disease (AD). Current FDA-approved drugs that modulate cytokine function are macromolecules, which have disadvantages for clinical use in chronic CNS disorders. There is a critical unmet need for small molecule, orally active, brain-penetrant compounds that can reduce excessive proinflammatory cytokine production by glia back towards basal levels. Recently, we described an integrative drug discovery platform for rapid discovery of novel small molecule compounds that suppress excessive CNS proinflammatory cytokine production, with resultant neuroprotective effects in animals. The availability of this platform allows exploration of hypotheses about the role of discrete biological processes in glia proinflammatory cytokine production and the potential for in vivo modulation by bioavailable small molecules of neurologic outcomes in neurodegenerative disease models. Use the integrated synthetic chemistry and hierarchal biology screening platform focused on excessive proinflammatory cytokine production to develop experimental therapeutics active in modifying disease progression in animal models of neurodegenerative disease. Chemical diversification of an inactive molecular scaffold and consideration of potential molecular properties were used. Screening for suppression of excessive glia activation was used in an unbiased, function-driven discovery approach. Structure-assisted pharmacophore models of the protein kinase p38 MAPK, which plays a critical role in regulation of proinflammatory cytokine up-regulation and is an established therapeutic target for peripheral inflammatory disorders, were used in a single molecular target approach. The unbiased, function-driven approach and follow-up medicinal chemistry refinement yielded the novel drug, Minozac, now in clinical development. The single molecular target approach produced a novel p38 MAPK inhibitor currently in refinement. Both compounds are efficacious in an AD-relevant animal model. The findings validate the integrative approach for rapid discovery of novel small molecules that are candidates for drug development in AD and related disorders. Studies are in progress with other animal models in which increased levels of proinflammatory cytokines contribute to the progression of pathophysiology.

We report the development of a novel, aqueous-soluble, safe, small molecule, experimental therapeutic that suppresses injury-induced, proinflammatory cytokine increases in the brain, with resultant attenuation of synaptic protein biomarker loss and improvement in hippocampus-dependent behavioral deficits. A GMP production scheme for the active pharmaceutical ingredient, compound 17, is presented. The development and large-scale availability of this novel compound allow exploration of new, potentially disease-modifying, therapeutic approaches to CNS disorders.

Overproduction of proinflammatory cytokines by glia contributes to progression of pathophysiology in diverse neurological disorders. Thus, development of safe and efficacious therapeutics that suppress glia up-regulated proinflammatory cytokine production towards basal levels could benefit multiple CNS diseases. We recently described (Wing et al., 2006, Curr Alzheimer Res 3:205; Ralay Ranaivo et al., 2006, J Neurosci 26:662) a de novo discovery platform that integrates a chemistry platform with hierarchal activity and in vivo function platforms. A lead compound, MW01-5-188WH, with in vivo function and promising safety and bioavailability profiles was discovered. Medicinal chemistry refinement of this lead emphasized molecular property improvement with retention of in vivo functions, followed by development of a GMP compatible production scheme (Hu et al., 2006, Bioorg Med Chem Lett, doi:10.1016/j.bmcl.2006.10.028). The clinical candidate, Minozac, is under clinical development (www.neuromedixinc.com). Our results demonstrate the feasibility of the process for developing innovative, potentially disease-modifying therapeutics within a highly accelerated timeline, limited budget and focused synthetic chemistry campaign. (Supported by NIH & NeuroMedix, Inc.)

There is immediate potential to enhance success and innovation in drug development by pairing newly emerging approaches in medicinal chemistry and computational biology with knowledge gained from the recent era of high throughput screens and the early years of modern drug discovery when in vivo efficacy was an early "Go/No Go" project management decision. Focused, in-parallel synthetic chemistry platforms, combined with computational analyses serving as decision aids in planning, minimize the total number of compounds synthesized while maximizing the probability of creating bioavailable compounds that sample diverse chemical space. Incorporating a hierarchal strategy that emphasizes early selection of synthesized compounds based on biological or biophysical endpoints presents fewer and more relevant compounds for secondary evaluation of in vivo efficacy using animal screens with disease relevant or clinically translatable endpoints. We summarize here an interdisciplinary approach at the chemistry-biology interface that is used for the rapid discovery of novel lead compounds for neurodegenerative disorders, such as Alzheimer's disease (AD). The chemistry platform uses established chemistries amenable to in-parallel strategies to create synthetic diversifications of the privileged pyridazine chemotype that sample a restricted chemical space. The hierarchal biology platform uses primary screens for in vitro activity and selectivity with the target cell type, and rapid secondary screens for in vivo efficacy and toxicity in animal models with good phenotypic penetrance for disease relevant pathophysiological endpoints or clinically translatable surrogate endpoints. For the AD case study, novel lead compounds were developed in less than two years by a small academic group, and corporate sponsored clinical trials are planned.

A corollary of the neuroinflammation hypothesis is that selective suppression of neurotoxic products produced by excessive glial activation will result in neuroprotection. We report here that daily oral administration to mice of the brain-penetrant compound 4,6-diphenyl-3-(4-(pyrimidin-2-yl)piperazin-1-yl)pyridazine (MW01-5-188WH), a selective inhibitor of proinflammatory cytokine production by activated glia, suppressed the human amyloid-beta(A beta) 1-42-induced upregulation of interleukin-1 beta, tumor necrosis factor-alpha, and S100B in the hippocampus. Suppression of neuroinflammation was accompanied by restoration of hippocampal synaptic dysfunction markers synaptophysin and postsynaptic density-95 back toward control levels. Consistent with the neuropathophysiological improvements, MW01-5-188WH therapy attenuated deficits in Y maze behavior, a hippocampal-linked task. Oral MW01-5-188WH therapy begun 3 weeks after initiation of intracerebroventricular infusion of human A beta decreased the numbers of activated astrocytes and microglia and the cytokine levels in the hippocampus without modifying amyloid plaque burden or altering peripheral tissue cytokine upregulation in response to an in vivo inflammatory challenge. The results provide a novel integrative chemical biology proof in support of the neuroinflammation hypothesis of disease progression, demonstrate that neurodegeneration can be attenuated independently of plaque modulation by targeting innate brain proinflammatory cytokine responses, and indicate the feasibility of developing efficacious, safe, and selective therapies for neurodegenerative disorders by targeting key glial activation pathways.