Nonpeptide neurotrophic agents useful in the treatment of neurodegenerative diseases such as Alzheimer's disease

Accepted Manuscript Nonpeptide neurotrophic agents useful in the treatment of neurodegenerative diseases such as Alzheimer’s disease Masaaki Akagi, Nobuaki Matsui, Haruka Akae, Nana Hirashima, Nobuyuki Fukuishi, Yoshiyasu Fukuyama, Reiko Akagi PII:

S1347-8613(15)00004-3

DOI:

10.1016/j.jphs.2014.12.015

Reference:

JPHS 36

To appear in:

Journal of Pharmacological Science

Received Date: 2 July 2014 Revised Date:

12 December 2014

Accepted Date: 24 December 2014

Please cite this article as: Akagi M, Matsui N, Akae H, Hirashima N, Fukuishi N, Fukuyama Y, Akagi R, Nonpeptide neurotrophic agents useful in the treatment of neurodegenerative diseases such as Alzheimer’s disease, Journal of Pharmacological Science (2015), doi: 10.1016/j.jphs.2014.12.015. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

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Critical Review

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Nonpeptide neurotrophic agents useful in the treatment of neurodegenerative diseases such as Alzheimer’s disease Masaaki Akagi1,*, Nobuaki Matsui1, Haruka Akae1, Nana Hirashima1, Nobuyuki Fukuishi1, Yoshiyasu Fukuyama1, and Reiko Akagi2 1

Faculty of Pharmaceutical Sciences, Tukushima Bunri University, Yamashiro-cho,

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Tokushima 770-8514, Japan 2

Faculty of Pharmacy, Yasuda Women’s University, Yasuhigashi, Asaminami-ku,

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Hiroshima 731-0153, Japan *

Corresponding author. [email protected]

Running title: Nonpeptide neurotrophic agents for AD Abstract

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Developed regions, including Japan, have become “aged societies,” and the number of adults with senile dementias, such as Alzheimer’s disease (AD), Parkinson’s disease, and Huntington’s disease, has also increased in such regions. Neurotrophins (NTs) may play a role in the treatment of AD because endogenous neurotrophic factors (NFs)

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prevent neuronal death. However, peptidyl compounds have been unable to cross the blood–brain barrier in clinical studies. Thus, small molecules, which can mimic the functions

of

NFs,

might

be

promising alternatives for the

treatment of

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neurodegenerative diseases. Natural products, such as or nutraceuticals or those used in traditional medicine, can potentially be used to develop new therapeutic agents against neurodegenerative diseases. In this review, we introduced the neurotrophic activities of polyphenols honokiol and magnolol, which are the main constituents of Magnolia obovata Thunb, and methanol extracts from Zingiber purpureum (BANGLE), which

may have potential therapeutic applications in various neurodegenerative disorders. Keywords: nonpeptide neurotrophic agents, natural products, senescence-accelerated mice, rat cortical neurons

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1. Introduction The percentage of older adults is increasing with the medical advances that have

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prolonged the lifespan of individuals. The extensively accepted standard indicates that numerous countries, including Japan and Germany, have become “aged societies”

(Table 1). In Japan, the population ratio of people aged more than 65 years is estimated

to reach 29.1% by 2020 and further increase to 38.8% by 2050 (according to data from World Population Prospects: The 2010 Revision, Japan).

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The number of adults with senile dementias, such as Alzheimer’s disease (AD), Parkinson’s disease (PD), and Huntington’s disease (HD), has increased owing to the

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advent of this rapidly aging society. In Japan, the elderly population with dementia will increase to 4,700,000 by 2025 unless current medical treatments are not significantly improved (Table 2). Dementia causes serious health problems to the elderly and burdens the overall society. Although AD is not a normal consequence of aging, the greatest risk factor for AD is aging. AD is the most prevalent dementia (accounting for 50%–80% of dementias, according to the Alzheimer’s Association, www.alz.org) and some of its curative therapies.

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causes have recently been discovered. However, we still lack effective preventative or The senile dementias (AD, PD, and HD), brain trauma, and amyotrophic lateral sclerosis are neurodegenerative diseases that are often associated with synaptic loss and neuronal death. In most cases, neuronal injuries occur after self-degeneration, toxic

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insults, or lack of nutrients (1). AD is characterized by neuronal loss, and the overall treatment strategy for AD is to prevent neuronal death or to produce new neuronal cells in order to replenish the cells in the degenerated regions. Neurotrophins (NTs) are

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expected to play a role in the treatment of AD because neuronal death is prevented by endogenous neurotrophic factors (NFs), such as nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) (2). Therapeutic applications of NFs by intracranial injections, transplantation of cells secreting NFs, or gene therapy have shown promising results in animal models of neuronal degeneration with learning and memory impairments as well as in clinical trials with AD patients. Thus, NFs are likely to be useful agents for ameliorating neurodegeneration (3). However, NFs were unable to cross the blood–brain barrier in clinical studies and were easily metabolized. Moreover, they caused immune responses because of their peptidyl properties.

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Therefore, small molecules, which can mimic the functions of NFs, might be promising alternatives for the treatment of neurodegenerative diseases (4). In vitro, NTs can also promote process outgrowth and survival. Thus, rat cortical neurons were cultured to

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investigate the small neurotrophic molecules in natural products, which resulted in the discovery of several active compounds (5). There are some reports on the elucidation of

the mechanisms of natural products, but the detailed mechanism is yet unclear. The

present review focuses the neurotrophic activity of polyphenols, honokiol and magnolol, the main constituents of Magnolia obovata Thunb, and methanol extracts from Zingiber

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purpureum (BANGLE), which may have potential therapeutic applications in various

2 AD: Pathology and therapeutics 2.1. Pathology of AD

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neurodegenerative disorders.

AD is a progressive neurodegenerative disorder characterized by accelerated neuronal

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cell death resulting to dementia. The hallmark pathological features are extracellular amyloid plaques and intraneuronal fibrillar structures. The latter includes neurofibrillary tangles (NFTs), neuropil threads, and dystrophic neuritis that invade amyloid plaques. The extracellular deposition of proteolytic fragments of amyloid precursor protein

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(APP) is termed amyloid β (Aβ), whereas the fibrillar pathologies are composed of the microtubule-associated protein, tau, assembled into polymeric filaments, forming amyloid plaques (6). APP is a large type-1 transmembrane multidomain protein that

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performs multiple cellular activities. The most frequently expressed isoforms in the brain are APP695, APP751, and APP770. Secretases (α, β, and γ) catabolize APP, forming non-amyloidal and/or amyloid-derived products (7). β-secretase, also known as

beta-site amyloid precursor protein cleaving enzyme 1 (BACE1), cleaves APP at Met 671 to produce soluble amyloid precursor protein β (sAPPβ) and a C-terminal 99 fragment. This is then cleaved by γ-secretase at Val 711 and Ala 713, resulting in the formation of APP intracellular domain (AICD), Aβ40, and Aβ42 peptides (8). Aβ fibrils are formed by the aggregation of these amyloid peptides. On the other hand, α-secretase cleaves APP at Lys 687 to form the soluble amyloid precursor protein α (sAPPα) and a

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C-terminal fragment of 83 amino acid residues. This fragment is cleaved by γ-secretase to produce non-amyloidogenic Aβ17–42 [also known as protein 3 (p3)] and AICD. γ-Secretase is a heterotetrameric membrane-embedded aspartyl protease consisting of

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four subunits: nicastrin, presenilin, anterior pharynx, and presenilin enhancer 2 (9).

When α- or β-secretase cannot cleave APP, it is cleaved by γ-secretase, forming the

soluble amyloid precursor protein γ (sAPPγ) and AICD (10). Some drugs that target APP processing are complex and challenging to design because of the multiple functional enzymes and substrates involved. Polyphenols are potential activators of

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α-secretase and inhibitors of β- and γ-secretase, resulting in a reduction in amyloid fibril deposition in the brain (11).

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In AD, the tau that is present in NFLs is aberrantly phosphorylated and often proteolytically truncated at its C terminus (12). Cleavage of tau at Glu 391 by a yet-to-be-identified protease is one such proteolytic event. Tau truncated at this site is observed in the NFTs in brains of AD patients (12). Such post-translational modifications are believed to impair the ability of tau to bind/stabilize microtubules. The AD brain exhibits evidence of oxygen radical-mediated damage. However, the ability to directly implicate this mechanism in AD has been a difficult task because of

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the evanescent nature of oxygen radicals and the use of non-specific analytical approaches. Oxidative stress increases in the AD brain. The central nervous system (CNS) is particularly vulnerable to oxidative damage because it has a high energy requirement, high oxygen consumption rate, and relative deficit of antioxidant defense

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systems compared with that of other organs (13). Brain insulin/insulin-like growth factor (IGF) resistance and deficiency are

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accompanied by impairments in glucose utilization and disruption of energy metabolism, with attendant increases in the mitochondrial dysfunction, oxidative stress, production of reactive oxygen species (ROS), and DNA damage. All of these situations drive pro-apoptosis, pro-inflammatory, and pro-AβPP-Aβ cascades. Insulin stimulates the expression of glucose transporter protein 4 (GLUT4) and protein trafficking from the cytosol to the plasma membrane to modulate glucose uptake and utilization. The stimulation of GLUT4 with insulin is critical for regulating neuronal metabolism and energy production, which are needed for memory and cognition (14). Deficiencies in energy metabolism that are caused by the inhibition of insulin/IGF signaling promote

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oxidative stress, mitochondrial dysfunction, and pro-inflammatory cytokine activation. Oxidative stress results in the generation and accumulation of reactive oxygen and nitrogen species, which attack subcellular organelles and cause chemical modifications

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of lipids, proteins, DNA, and RNA. For example, adducts formed with macromolecules

can compromise the structural and functional integrity of neurons due to the loss of membrane functions, disruption of the cytoskeleton, dystrophy of synaptic terminals,

deficits in neurotransmitter functions and plasticity, and perturbation of signaling

pathways for energy metabolism, homeostasis, and cell survival. The expression and

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phosphorylation of tau are regulated by insulin and IGF. In AD, brain insulin/IGF resistance reduces signaling through phosphoinositol-3-kinase (PI3K) and Akt and

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increases activation of glycogen synthase kinase 3β (GSK-3β) (15). Overactivation of GSK-3β is partly responsible for the hyperphosphorylation of tau, thus promotes tau misfolding and fibril aggregation. The consequences include failure to generate sufficient quantities of normal soluble tau in relation to accumulation of hyperphosphorylated insoluble fibrillar tau and attendant exacerbation of cytoskeletal collapse, neurite retraction, and synaptic disconnection. Finally, intracellular AβPP-Aβ directly interferes with PI3K activation of Akt, impairing neuronal survival and

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increasing GSK-3β activation and hyperphosphorylation of tau. Consequently, hyperphosphorylated tau is prone to misfolding, aggregation, and ubiquitination, prompting the formation of dementia-associated paired-helical filament-containing

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neuronal cytoskeletal lesions.

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2.2. Therapeutics for AD

Alzheimer’s disease is a process in which toxic Aβ fragments are generated (7) due to

uncontrolled cleavage of APP by unknown inducing factors. Alzheimer’s disease is also characterized by amyloid fibril and phosphorylated tau aggregates and tangles. At present, there are two major problems with AD drug discovery. (1) There is no available animal model that reflects all the pathological events observed in the human AD brain.

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(2) There is a lack of reliable biomarkers to detect and understand the progression of AD.

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Scientists are trying to develop drugs that can simultaneously perform multiple tasks such as reducing inflammation, inhibiting β-secretase, activating α-secretase, preventing Aβ and tau aggregation, and driving the disintegration of preformed fibrils. However,

no perfect drug or treatment of AD has been discovered so far, and several drugs have

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failed in recent clinical trials.

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2.2.1 Experimental models to screen neurotrophic compounds

The brain contains many types of neurons having different degrees of selectivity for NTs. Neurotrophic activity is experimentally model-dependent; for example, septal cholinergic neurons in cultures were extremely sensitive to NGF, whereas septal dopaminergic neurons were not (16). Small molecular weight neurotrophic compounds

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also have different degrees of selectivity for neuronal cells. Therefore, a compound may be significantly sensitive to NTs in one experimental model and extremely insensitive in another model. Currently, several in vivo and in vitro screening models have been 3).

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designed to examine the different degrees of selectivity in neurotrophic studies (Table A small molecule can mimic NTs by directly activating or upregulating the NT receptors. Furthermore, a small molecule can upregulate the expression of NTs, enhance

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the intracellular NT signaling pathway, and/or inhibit the degradation of the trophic signal. The physiological structure and function of the neurons are maintained by the NTs when the neurons completely differentiate into their neuronal phenotypes. Therefore, a lack of NTs in the body or withdrawal of NTs from the culture medium results in deleterious signals, which cause neuronal apoptosis.

2.2.2 Small molecular weight compounds with neurotrophic activity

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Compounds such as 1-{3-[2-(1-benzothiophen-5-yl)ethoxy]propy-l}azetidin-3-ol

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maleate (T-817-MA), xaliproden, and DMXB-A have neurotrophic activities and are in the clinical testing stage. The first of these has been developed as a therapeutic agent to treat neurodegenerative disorders such as AD by modulating endogenous antioxidative

mechanisms. Phase II clinical trials are being conducted in the United States to assess

Xaliproden

{SR57746A;

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the efficacy of T-817-MA in treating AD (31).

1-[2-(naphth-2-yl)ethyl]-4-(3-trifluoromethylphenyl)]

-1,2,5,6 -tetrahydropyridine hydrochloride} is a selective and potent 5-HT1A receptor

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agonist with low nanomolar affinity for the 5-HT1A receptors. Studies on xaliproden have focused on the pro-neurotrophic properties of this compound in cellular models, including rat pheochromocytoma PC12 cells, primary neuronal cells, and animal models of neurodegeneration (32).

DMXB-A {GTS-21; 3-[(2,4-dimethoxy)benzylidene]-anabaseine dihydrochloride}is a derivative of the naturally occurring alkaloid, anabaseine (33). It is a partial agonist of

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human α7 nicotinic receptors and, at higher concentrations, a weak antagonist of α4β2 receptors and serotonin 5-HT3 receptors (33). Approximately 40% of the oral dose is absorbed within 1 h of administration (34). Metabolites with hydroxyl substituents at positions 2 and 4 are more efficacious agonists when they are bound to the receptor, but

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the extent of their production in the human brain in unknown (34). DMXB-A improves memory in several animal models, and it normalizes the inhibition of auditory responses in rodents with significantly less tachyphylaxis than that with nicotine. Mecamylamine,

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an α4β2 antagonist, has a deleterious effect on conditioned learning in young rabbits. The eyeblink classical conditioning acquisition is improved in young rabbits administered with mecamylamine and DMXB-A (35). In aged rats, DMXB-A improves one-way active avoidance and the performance in Lashley III maze testing and 17-arm radial maze test (35). Passive avoidance deficits are normalized in rats and ischemia-induced hippocampal cell death and impaired passive avoidance performance in gerbils are prevented by treatment with DMXB-A (36). DMXB-A also improves performance on the Morris water maze (35). Positive neurocognitive effects, particularly on attention, were observed in healthy volunteers administered with

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DMXB-A (37). All subjects were evaluated for performance on a computerized test battery to measure the effect of treatment on cognitive functions, including changes in attention [simple reaction time (RT), choice RT, and digit vigilance], numeric and

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spatial working memory, secondary episodic recognition memory (word and picture

recognition and immediate and delayed word recall), and visual tracking. DMXB-A

significantly improved performance on simple RT, choice RT, correct detection during

digit vigilance, both word and picture recognition memory, and both immediate and delayed word recall. DMXB-A improved subject performance speed on numeric and

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spatial working memory tasks. Improvement was generally observed with a 25-mg dose, with further improvement at a 75-mg dose, and an equivalent effect at a 150-mg dose

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(37).

2.2.3 Neurotrophic natural products

Natural products that are used in traditional medicine or as nutraceuticals could

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potentially be used to develop new therapeutic agents against neurodegeneration. Illicium, the sole genus of the family Illiciaceae, is distributed in the southern parts of China (38). Majority of these plants are considered toxic; however, compounds from these plants have demonstrated neuroactivity and are characterized according to their

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neurotrophic profiles. To characterize the plants according to their neurotrophic profiles, rat pheochromocytoma PC12 cells (17), primary cultured rat cortical/hippocampal

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neurons (21), and human neuroblastoma SH-SY5Y cells (18) were used (Table 3). The rat PC12 cells are easy to culture and have been extensively used as a neuronal model because they allow the screening of several compounds. Primary cultured neurons bear majority of the in vivo neuronal properties, whereas the SH-SY5Y cells contain human

biochemical characteristics. These three cellular models (PC12 cells, primary cultured neurons, and SH-SY5Y cells) were utilized, and the biological characteristics of the neurotrophic compounds discovered in the screening system were classified. As a result, several neurotrophic compounds (i.e., alkaloids, terpenes, and phenols) were revealed. However, it is unknown whether the neurotrophic activities that occurred in vitro would

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also transpire in vivo. Thus, it would be important to determine the amount of neurotrophic compounds that would be required for systemic administration. Table 4

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shows the compounds that have been investigated in our laboratory.

2.2.3.1 Polyphenolic compounds

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Recent research is focusing on natural products, such as the water extract of the leaves

of Caesalpinia crista and Centella asiatica, Ginkgo biloba extract, and extracts prepared from the medicinal herb, Paeonia suffruticosa. These products are alternatives

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for the treatment of AD (55). The water extract of the leaves of C. crista prevents Aβ aggregation from monomers and disintegrates preformed Aβ fibrils. C. asiatica prevents synuclein aggregation (56), G. biloba extract inhibits the formation of oligomers (55), and extracts prepared from the medicinal herb P. suffruticosa inhibit Aβ fibril formation and also destabilize the preformed amyloid fibrils (57). The anti-amyloidogenic properties observed are attributed to the polyphenolic compounds present in these

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extracts. The chemical structure of polyphenols includes two or more phenol rings with hydroxyl groups in ortho or para positions. The phenol rings are essential for redox reactions (58). There is a direct positive correlation between the antioxidant capacity and number of hydroxyl groups present in the polyphenols’ structure. Thus, an increase

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in the amount of hydroxyl groups in the polyphenol chemical structure is associated with an increase in redox potential and antioxidant activity (59). are

powerful

anti-amyloidogenic

compounds

owing

to

their

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Polyphenols

physicochemical features such as the presence of aromatic rings, molecular planarity, capacity to form hydrogen bonds, presence of an internal double bond, and molecular weights below 500 g/mol. The molecular weight allows for potential inhibition of APP pathways that, in turn, reduces amyloid load (60). Several members of the disintegrin and metalloproteinase (ADAM) family (ADAM9, 10, and 17) have been proposed to be physiologically active α-secretases. It has been

demonstrated that ADAM10 has the highest α-secretase activity in vivo. Moreover, it has been suggested that the upregulation of ADAM10 could be a potential therapeutic

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target for the treatment of AD. ADAM10 has a potential neuroprotective role because it promotes the non-amyloidogenic pathway (61). This enzyme is activated by the removal of the prodomain, which is probably promoted by the action of proprotein

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convertases. Phlorotannins, curcumin, and epigallocatechin-3-gallate (EGCG) have

been shown to increase the overexpression of sAPPα through activation of α-secretase, favoring neuroprotection (62).

The amino acids, Asp 32 and Asp 228, and two water molecules are essential for the

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catalytic activity of β-secretase, and all these components are located in the catalytic binding site. One of these water molecules participates in enzymatic proteolysis,

whereas the second water molecule is important for stabilizing a tetrahedral

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intermediate that is essential for protein cleavage (63). Computational modeling studies have clarified that Asp 32, not Asp 228, is protonated (63). Proteolytic β-secretase activity is initiated through a nucleophilic attack on the carbonyl group of the peptide bond by a water molecule (63). The inhibition of β-secretase represents a potential therapeutic target in AD treatment because it decreases the Aβ load. Isophthalamides have shown greater inhibitory effects on β-secretase (63), and myricetin is a potential

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β-secretase inhibitor (64). The proposed mechanism is that the displacement of a water molecule inhibits the polyphenol, and the water molecule then participates in a hydrogen-bonding network with Asp 32 and Asp 228, which is essential for β-secretase proteolytic activity (63).

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Presenilin I (PS1) protein is a member of the aspartic protease family, which is implicated in the regulation of intramembrane proteolysis (9, 65). Presenilin I has been

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identified as the catalytic subunit of the γ-secretase complex. This protein is composed of nine transmembrane domains, and domains 6 and 7 form the catalytic site. Mutations in PS1 have been linked to familial AD (FAD). Therefore, PS1 is a potential target in the design of drugs against AD (66). The catalytic activity of PS1 requires two opposing aspartyl groups (Asp 257 and Asp 385) in the active site. One of these aspartates is deprotonated and acts as a base, which activates a water molecule present in the catalytic site. The other aspartate donates a proton to the carbonyl group of the substrate (66). Therefore, it is suggested that polyphenols that can occupy the active site of

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γ-secretase and displace the water molecule required by the enzyme for catalysis, may disable the enzyme and reduce Aβ formation (66).

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Aggregation of Aβ42 is the central point of AD pathology. Consequently, the inhibition of Aβ42 aggregation is a key focus in drug discovery. There are multiple steps involved

in the biology of Aβ42 aggregation, i.e., monomers form oligomers, protofibrils, and

mature fibrils. Drug discovery efforts are focused on preventing the formation of either oligomers or fibrils. However, studies that explore the binding of drugs to Aβ42 and

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prevent fibril formation are restricted because more than 50% of the amino acids in this

peptide are hydrophobic residues. It has been suggested that the hydrophobic core is essential for fibrillogenesis (67). The fibrillization of Aβ, which begins with the

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formation of Aβ oligomers that are constituted by 24 monomers, is a multistep process (68). Amyloid fibril formation depends on concentration of Aβ42 peptide, low pH, the time of incubation, and the length of the carboxyl chain (69). The Aβ structure is stabilized by hydrophobic forces, aromatic stacking, and electrostatic interactions (70). The main physicochemical properties of molecules with the potential to inhibit amyloidal fibril formation might be due to the presence of aromatic rings in their

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chemical structure and the ability to form noncovalent interactions with amino acid residues of the Aβ peptide sequence (71). Moreover, the planarity of the inhibitor is essential for increasing surface contact with the Aβ peptides (72). Most polyphenols have more than two aromatic rings, which are essential for π–π stacking interactions

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with the hydrophobic amino acid residues of Aβ (Tyr and Phe), and at least three hydroxyl groups that form hydrogen bonds with the hydrophilic amino acid residues of Aβ (His6, Ser8, Tyr10, His14, and Lys16) (73). The resonance structure of polyphenols

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provides enough planarity to penetrate the Aβ fibril hydrophobic groove, thus disturbing

the fibril structure (74). Polyphenolic compounds, such as resveratrol, curcumin, and myricetin, have

demonstrated anti-Aβ aggregation properties (75, 76). Furthermore, flavanols possess more anti-amyloidogenic activity than isoflavonoids because they contain more hydroxyl groups that are able to form hydrogen bonds with Aβ peptides. Curcuminoids are more hydrophobic than flavonoids. This physicochemical property might enhance their affinity for binding with the hydrophobic core of the Aβ fibril, resulting in an

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increased anti-amyloidal activity (77). Tannins are complex polyphenols that have the highest number of hydroxyl groups of all the polyphenolic compounds; therefore, they have the strongest anti-aggregation activity (78). However, their large molecular weight

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reduces their suitability as a therapeutic drug (79).

Our understanding of the bioavailability and health benefits of polyphenols remains

unclear. However, it has been shown that some polyphenols can cross the blood–brain

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barrier and contribute to neuroprotection (80).

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2.2.3.2 Neurotrophic activity of honokiol and magnolol

2.2.3.2.1 Neurite outgrowth promotes activities in cortical neurons Honokiol and magnolol are bioactive compounds found in the stem bark of the M. obovata Thunb (Wakoboku) and Magnolia officinalis Rhed (Houpu) (81). Honokiol and magnolol are herbal drugs (called “Kouboku”) used in China and Japan for the treatment of digestive and nervous complaints.

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Neurons were placed in cultures that contained honokiol or magnolol. The neurons that were in honokiol-containing cultures had extremely long neurites and more prominent, darker-stained neuronal soma compared with those of the neurons in the control culture. In addition, one of the processes was much longer and displayed more

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neurite branches. If bFGF were added to this cell culture, the neurite extension could be significantly enhanced. The neurons that were in magnolol-containing cultures also had

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long neurites. The length of neurites increased in the cultures that were treated with honokiol or magnolol in a dose-dependent manner and the neurites reached their maximum length at 10 µM. Magnolol had a weaker effect on neurite extension

compared with that of honokiol. This difference in the trophic effects caused by honokiol and magnolol presumably results from the different positions of an aryl–aryl bond that is formed between two units of allylphenol (Fig. 1) (82). These results are consistent with previous studies that have shown that honokiol is more effective than magnolol at producing various CNS-related effects (83).

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2.2.3.2.2 Prevention

of

learning

and

memory

impairment

in

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senescence-accelerated-prone mice (SAMP)

Senescence-accelerated mice (SAM) are selected by brother–sister mating of AKR/J

mice and are characterized by a rapid accumulation of senile features and a shortened lifespan (1–1.5 years compared with that of 2–2.5 years in control animals) (84). SAM

are composed of nine inbred strains of senescence-accelerated-prone mice (SAMP), and

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three strains of senescence-accelerated-resistant mice (SAMR), the latter of which show normal aging. Neither SAMP1 nor SAMR1 mice are behaviorally or morphologically

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different until they are 4 months old. At that age, the SAMP1 mice begin to rapidly accumulate senile changes. These mice have decreases in orientation ability and learning capacity and the appearance of age-specific morphological changes such as lordokyphosis, hair loss, frequent corneal cataracts, and mucosal inflammation (28). Among the various SAMP substrains, SAMP8 mice exhibit early-onset impairment of learning and memory (85). Recent studies have reported that aged SAMP8 mice had cholinergic neuron deficits in the septal region of the forebrain (86). These findings

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suggest that the SAMP8 mouse might be a useful animal model for investigating the cholinergic hypothesis of human dementia in advanced aging and AD. Magnolol (5 and 10 mg/kg) was orally administered once a day for 14 days to 2- or 4-month-old mice. The mice were examined at 4 and 6 months of age. In the SAMP1

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mice, the density of the neurofibrils in the stratum radiatum of the CA1 region in the hippocampus decreased with age. This result was not observed in the SAMR1 mice. When 2-month-old mice were treated with magnolol, the decrease in neurofibrils in the

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CA1 region was significantly prevented. However, magnolol administration at 4 months of age did not result in a protective effect. These findings suggest that magnolol must be administered before neuronal loss begins in order to have a protective effect on the hippocampus (28).

Magnolol or honokiol was orally administered once a day for 14 days to 2-month-old

SAMP8 mice. Passive avoidance tests and object recognition tests were used to evaluate learning and memory at 4 and 6 months of age. Learning and memory in the SAMP8 mice were markedly impaired at 4 and 6 months of age. However, this age-related learning and memory impairment was prevented by the pretreatment with magnolol (10

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mg/kg)

or

honokiol

(1

mg/kg).

Immunohistochemical

analysis

of

choline

acetyltransferase (ChAT) was used to evaluate the forebrain cholinergic neuron densities in the medial septum and vertical limb of the diagonal band. Significant

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cholinergic deficits were observed in 6-month-old SAMP8 mice. However, this age-related cholinergic deficit was prevented by the pretreatment with magnolol (10 mg/kg) or honokiol (1 mg/kg) (29).

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2.2.3.2.3 Neurotrophic mechanisms of honokiol and magnolol

Figure 2 illustrates the signal pathways of the NTs in neuronal cells (87).

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Neurotransmitters, such as NGF, BDF, and NT-3, bind to the extracellular domain of the tyrosine kinase receptors, TrkA, TrkB, and TrkC, respectively, and activate tyrosine kinase in the intracellular domain. The autophosphorylation of multiple tyrosine residues within the intracellular segments produces several binding sites for intracellular signaling proteins. When the target signal proteins bind to tyrosine kinase, they are phosphorylated to adopt active forms and then activate downstream products (88). The NT-activated signal molecules {Ca2+, MAPK [Extracellular signal-regulated kinases the nuclei.

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(ERK)], and Akt} in Figure 2 are indispensable for transferring intracellular signals to The effects of honokiol and magnolol have been explained by their effects on GABAA receptors (89). In primary cultured rat cortical neurons and human SH-SY5Y

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cells, honokiol and magnolol increased cytoplasmic free Ca2+ with a characteristic lag phase. The increase in cytoplasmic free Ca2+ was independent of extracellular Ca2+, but dependent on phospholipase C (PLC) and inositol 1,4,5-trisphosphate (IP3) receptor

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activation. These results suggest that honokiol and magnolol increase cytoplasmic free Ca2+ through a PLC-mediated pathway. Furthermore, the release of Ca2+ from

intracellular stores mainly contributes to an increase in cytoplasmic free Ca2+. Thus, honokiol and magnolol may be involved in the activation mechanism associated with intracellular Ca2+ mobilization (18). ERK1/2 and Akt might be related to the honokiol-induced neurite outgrowth and intracellular Ca2+ mobilization. This is suggested because the outgrowth in the cultured rat cortical neurons was markedly reduced by PD98059, a mitogen-activated protein kinase kinase (MAPKK, MAPK/ERK kinase MEK, directly upstream from ERK1/2) inhibitor but not by

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LY294002, a PI3K (upstream from Akt) inhibitor. Honokiol also highly enhanced the phosphorylation of ERK1/2 in a concentration-dependent manner. The phosphorylation of ERK1/2 was enhanced by honokiol and inhibited by KN93, PD98059,

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Ca2+/calmodulin-dependent kinase II (CAMK II) inhibitor, and MAPKK inhibitor. Products of the phosphoinositide specific PLC-derived IP3 and 1,2-diacylglycerol

(DAG) were measured after honokiol treatment (48). These results suggest that the

signal transduction from PLC, IP3, Ca2+, and CAM II to ERK1/2 is involved in the honokiol-induced neurite outgrowth.

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The Akt pathway is a prominent regulator of NT-mediated survival responses in neurons. Activated Akt inhibits apoptosis through multiple mechanisms and negatively

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regulates the phosphorylation and inactivation of the forkhead transcription factors. Akt activity was decreased in the forebrain of SAMP8 mice at 2 months of age compared with that of SAMR1 mice. However, treatment with either magnolol or honokiol increased Akt phosphorylation in the forebrain of SAMP8 mice (29). Regulation of oxidative stress may also explain the protective effects of magnolol and honokiol in preventing age-related impairment in SAMP8 mice. Magnolol and honokiol are known to have antioxidant effects (90, 91). The onset of increased lipid

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hydroperoxide in the brain coincides with the onset of cognitive impairment in SAMP8 mice that are 2 months old. Thus, oxidative stress may be important in the development of impaired learning and memory (92). Evaluation

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2.2.3.3 BANGLE:

of

hippocampal

neurogenesis

in

olfactory

bulbectomized (OBX) mice, an experimental depression and dementia

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animal model

In the early stages of AD, the olfactory system is significantly impaired (93); it is

particularly rich in acetylcholine and other neurotransmitters and has decreased ChAT activity (94). These results indicate that degeneration of cholinergic neurons may occur in the olfactory system. Patients with AD have increased numbers of senile plaques containing Aβ and NFTs in the olfactory bulbs (OBs) and the anterior olfactory nuclei

(94). Previous studies have shown that when the OBs are removed in animals (i.e., OBX mice), the result is impaired learning and memory (95) with increased exploratory behaviors (96). These results suggest that OBX mice are an animal model for the

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investigation

of

experimental

depression

and

dementia.

In

mice,

the

intracerebroventricular application of fibroblast growth factor-2 (FGF-2) ameliorates the reduction in hippocampal neurogenesis and reverses depressive behaviors induced

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by OB removal (97). The olfactory bulbectomization procedure was performed

according to a previously described method by Hozumi et al. (98). Two butenoid dimmers, trans-3-(3′,4′-dimethoxyphenyl)-4-[(E)-3″,4″-dimethoxystyryl]cyclohex-1-ene (compound

1)

and

cis-3-(3′,4′-dimethoxyphenyl)-4-[(E)-3′′,4′′-dimethoxystyryl]

cyclohex-1-ene (compound 2) (Table 4) were isolated as neurotrophic molecules from

SC

an Indonesian medicinal plant, Z. purpureum (Bangle). These two compounds were administered once a day on days 15–28 after. Chronic treatment with compounds 1 and

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2 significantly increased the number of BrdU/NeuN double-labeled cells (30). These results indicate that compounds 1 and 2 have neurotrophic effects and have the potential to modify depression and dementia.

2.2.3.4 Structure-activity relationships of natural products

Understanding the structure–activity relationship of NFs is important. However,

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using natural products to search for NF-like molecules with similar structures is difficult. Thus, these molecules should be synthesized. Although this type of research has not yet progressed, we have introduced it below.

The morphological effects of honokiol and its analogs on the cultured rat cortical

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neurons were evaluated by the anti-MAP2 staining method. The 4′-hydroxy group and the 5′-allyl group were deemed responsible for the honokiol-mediated neurite outgrowth-promoting activity. However, the 3′-allyl group was not considered essential

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for neurite growth (Fig. 1) (99). Illicinin A (A) and its derivatives (Fig. 3) were synthesized for the structure–activity

relationship studies. Sequential Stille reactions introduced a prenyl and allyl group to the benzene ring of A and its derivatives. Thus, an allylphenyl moiety in the A molecule

is essential for its neurotrophic properties (42). To further understand the structure–activity relationship, the neurotrophic properties of Aa–Ad were evaluated by measuring the longest neurites among the rat cortical

neurons cultured in the presence of each analogue. Among the screened compounds, Ac decreased neurite outgrowth, whereas Aa, Ab, and Ad were comparable to A in terms

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of neurite outgrowth. These results indicate that an allyl function in A is essential for preserving neurotrophic activity. Among the active analogues, the activities of Aa and Ab, which possess a 1,2-free diol group, slightly increased. This implies that the

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presence of two vicinal oxy groups is important for neurotrophic activity. However, the

methylenedioxy forms of Aa and Ab did not enhance their neurotrophic activities. In addition, the presence and position of the prenyl group in A did not affect its neurotrophic activity (42). Conclusions

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3

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Several small molecules, which can mimic the functions of NFs, have been discovered in natural products. However, the neurotrophic activity has only been evaluated in vivo for a few compounds. Magnolol, honokiol, and BANGLE extracts can supply the amount of neurotrophic compounds that is required for adequate systemic administration. Promising compounds with neurotrophic activity may eventually be used for systemic administration. The rapid discovery and production of a synthetic method for producing an effective compound in large quantities would be highly

Conflicts of Interest

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desirable for the treatment of AD.

References

Friedlander RM. Apoptosis and caspases in neurodegenerative diseases. N Engl J

AC C

1

EP

The authors declare no conflict of interest.

Med. 2003;348:1365-1375.

2

Counts SE, Mufson EJ. The role of nerve growth factor receptors in cholinergic

basal forebrain degeneration in prodromal Alzheimer’s disease. J Neuropathol Exp Neurol. 2005;64:263-272.

3

Tuszynski MH, Thal L, Pay M, Salmon DP, U HS, Bakay R, et al. A phase 1 clinical trial of nerve growth factor gene therapy for Alzheimer disease. Nat Med. 2005;11:551-555.

4

Uwabe K, Iwakawa T, Matsumoto M, Takahashi K, Nagata K. HU0622: a small

ACCEPTED MANUSCRIPT

molecule promoting GAP-43 activation and neurotrophic effects. Neuropharmacol. 2006;51:727-736. 5

Kubo M, Ishii R, Ishino Y, Harada K, Matsui N, Akagi M, et al. Evaluation of

RI PT

constituents of Piper retrofractum fruits on neurotrophic activity. J Nat Prod. 2013;76:769-773.

6 Nelson PT, Alafuzoff I, Bigio EH, Bouras C, Braak H, Cairns N, Davies P, Tredici KD, Duyckaerts C, Frosch MP, Hof PR, Hulette C, Hyman BT, Iwatsubo T,

Jellinger KA, Jicha GA, Kovari E, Kukull WA, Leverenz JB, Love S, Mackenzie IR,

SC

Mann DM, Masliah E, Mc- Kee A, Montine TJ, Morris JC, Schneider JA, Sonnen JA, Thal DR, Trojanowski JQ, Troncoso JC, Wisniewski T, Woltjer RL, Beach TG:

M AN U

Correlation of Alzheimer’s disease neuropathologic changes with cognitive status: a review of the literature. J Neuropath Exp Neurol. 2012;71:362–381. 7 Sambamurti K, Greig NH, Utsuki T, Barnwell EL, Sharma E, Mazell C, Bhat NR, Kindy MS, Lahiri DK, Pappolla MA. Targets for AD treatment: conflicting messages from

-secretase inhibitors. J Neurochem. 2011;117:359–374.

TE D

8 Rajendran L, Schneider A, Schlechtingen G, Weidlich S, Ries J, Braxmeier T, Schwille P, Schulz JB, Schroeder C, Simons M, Jennings G, Knölker H-J, Simons K. Efficient inhibition of the Alzheimer’s disease

-secretase by membrane

targeting. Science. 2008;320:520–523.

EP

9 Tiedt H, Lueschow A, Winter P, Müller U. Previously not recognized deletion in presenilin-1 (p.Leu174del.) in a patient with early-onset familial Alzheimer’s

AC C

disease. Neurosci Lett. 2013;544:115–118. 10 Utsuki T, Shoaib M, Holloway H, Ingram D, Wallace W, Haroutunian V, Sambamurti K, Lahiri D, Greig N. (2002) Nicotine lowers the secretion of the Alzheimer’s amyloid beta-protein precursor that contains amyloid beta-peptide in rat. J Alzheimers Dis. 2002;4:405.

11 Guerrero E, Padmaraju V, Hegde ML, Britton GB, Rao KS. Recent advances in α-synuclein functions, advanced glycation, and toxicity: implications for Parkinson’s disease. Mol Neurobiol. 2013;47:525– 536. 12 Novak M, Kabal J, Wischik CM. Molecular characterization of the minimal

ACCEPTED MANUSCRIPT

protease resistant tau unit of the Alzheimer’s disease paired helical filament. EMBO J. 1993;12:365-370. 13 Smith MA, Rottkamp CA, Nunomura A, Raina AK, Perry G. Oxidative stress in

RI PT

Alzheimerís disease. Biochim Biophys Acta. 2000;1502:139-144.

14 Gonzalez-Sanchez JL, Serrano-Rios M. Molecular basis of insulin action. Drug News Perspect. 2007;20:527-531.

15 Schubert M, Gautam D, Surjo D, Ueki K, Bauder S, Schubert D, Kondo T, Alber J,

Galldiks N, Kustermann E, Amdt S, Jacobs AH, Krone W, Kahn CR, Bruning JC.

SC

Role for neuronal insulin resistance in neurodegenerative diseases. Proc Natl Acad Sci USA. 2004;101:3100-3105.

M AN U

16 Knusel B, Michel PP, Schwaber JS, Hefti F. Selective and nonselective stimulation of central cholinergic and dopaminergic development in vitro by nerve growth factor, basic fibroblast growth factor, epidermal growth factor, insulin and insulin-like growth factors I and II. J Neurosci. 1990;10:558-570. 17 Mei JM, Niu CS. Effects of CDNF on 6-OHDA-induced apoptosis in PC12 cells via modulation of Bcl-2/Bax and caspase-3 activation. Neurol Sci. 2014;Mar 15. 18 Zhai H, Nakade K, Mitsumoto Y, Fukuyama Y. Honokiol and magnolol induce Ca2+

TE D

mobilization in rat cortical neurons and human neuroblastoma SH-SY5Y cells. Eur J Pharmacol. 2003;474:199-204.

19 Tadtong S, Meksuriyen D, Tanasupawat S, Isobe M, Suwanborirux K. Geldanamycin derivatives and neuroprotective effect on cultured P19-derived

EP

neurons. Bioorg Med Chem Lett. 2007 May 15;17(19:2939-2943. 20 Oh-Hashi K, Hirata Y, Kiuchi K. Transcriptional regulation of mouse mesencephalic astrocyte-derived neurotrophic factor in Neuro2a cells. Cell Mol

AC C

Biol Lett. 2013 Sep;18(3):398-415.

21 Kellner Y, Godecke N, Dierkes T, Thieme N, Zagrebeisky M, Korte M. The BDNF effects on dendric spines of mature hippocampal neurons depend on neuronal activity. Front Synaptic Neurosci. 2014 Mar 20;6:5.

22 Qiao LY, Yu SJ, Kay JC, Xia CM. In vivo regulation of brain-derived neurotrophic factor in dorsal root ganglia is mediated by nerve growth factor-triggered Akt activation during ctstitis. PLoS One. 2013 Nov 26;8(11):e81547.

23 Gransee HM, Zhan WZ, Sieck GC, Mantilla CB. Targeted delivery of TrkB receptor to phrenic motoneurons enhances functional recovery of rhythmic phrenic

ACCEPTED MANUSCRIPT

activity after cervical spinal hemisection. PLoS One. 2013 May 28;8(5):e64755. 24 Lazo OM, Mauna JC, Pissani CA, Inestrosa NC, Bronfman FC. Axotomy-induced neurotrophic withdrawal causes the loss of phenotypic differentiation and

RI PT

downregulation of NGF signaling, but not death of septal cholinergic neurons. Mol Neurodegener. 2010 Jan 19;5:5.

25 Yamada Y, Prosser RA. Copper chelation and exogenous copper affect circadian

clock phase resetting in the suprachiasmatic nucleus in vitro. Neuroscience. 2014 Jan 3;256:252-261.

SC

26 Elmann A, Teleman A, Erlank H, Mordechay S, Rindner M, Ofir R, et al. Protective and antioxidant effects of a chalconoid from Pulicaria incisa on brain astrocytes.

M AN U

Oxid Med Cell Longev. 2013;2013:694398.

27 Yang J, Harte-Hargrove LC, Siao CJ, Marinic T, Clarke R, Ma Q, et al. proBDNF negatively regulates neuronal remodeling, synaptic transmission, and synaptic plasticity in hippocampus. Cell Report. 2014 May 8;7(3)796-806. 28 Matsui N, Nakashima H, Ushio Y, Tada T, Shirono A, Fukuyama Y, et al. Neurotrophic effect of magnolol in the hippocampal CA1 region of senescence -accelerated mice (SAMP1). Biol Pharm Bull. 2005;28:1762-1765.

TE D

29 Matsui N, Takahashi K, Takeichi M, Kuroshita T, Noguchi K, Yamazaki K, et al. Magnolol and honokiol prevent learning and memory impairment and cholinergic deficit in SAMP8 mice. Brain Res. 2009;1305:108-117. 30 Matsui N, Kido Y, Okada H, Kubo M, Nakai M, Fukuishi N, et al. Phenylbutanoid

EP

dimers isolated from Zingiber purpureum exert neurotrophic effects on cultured neurons and enhanced hippocampal neurogenesis in olfactory bulbectomized mice. Neurosci Lett. 2012;513:72-77.

AC C

31 Maekawa H, Matsunobu T, Satoh Y, Kurioka T, Nakamura A, Iwakami N, et al. Protective effect of neurotrophic agent T-817MA against inner ear barotrauma in the guinea pig. J Pharmacol Sci. 2011;117:67-70.

32 Lemaire L, Fournier J, Ponthus C, Le Fur Y, Confort-Gouny S, Vion-Dury J, et al. Magnetic resonance imaging of the neuroprotective effect of xaliproden in rats. Invest Radiol. 2002;37:321– 327. 33 Kem WR, Mahnir VM, Prokai L, Papke RL, Cao X, LeFrancois S, Wildeboer K, Prokai-Tatrai K, Porter-Papke J, Soti F. Hydroxy metabolites of the Alzheimer’s drug

candidate

3-[(2,4-

dimethoxy)benzylidene]-anabaseine

dihydrochloride

ACCEPTED MANUSCRIPT

(GTS-21): their molecular properties, interactions with brain nicotinic receptors, and brain penetration. Mol Pharmacol. 2004;65:56–67.

RI PT

34 Mahnir VM, Lin B, Prokai-Tatrai K, Kem WR. Pharmacokinetics and urinary excretion of DMXBA (GTS-21), a compound enhancing cognition. Biopharm Drug Dispos. 1998;19:147–151.

35 Meyer EM, Tay EE, Papke RL, Meyers C, Huang G, deFiebre CM. 3-[2,47

SC

Dimethoxybenzylidene]anabaseine (DMXB) selectively activates rats

receptors and improves memory-related behaviors in a mecamylamine-sensitive

M AN U

manner. Brain Res. 1997;768:49–56.

36 Nanri M, Yamamoto J, Miyake H, Watanabe H. Protective effect of GTS-21, a novel nicotinic receptor agonist, on delayed neuronal death induced by ischemia in gerbils. Jpn. J. Pharmacol. 1998;76:23–29.

37 Kitagawa H, Takenouchi T, Azuma R, Wesnes KA, Kramer WG, Clody DE. Safety, pharmacokinetics, and effects on cognitive function of multiple doses of GTS-21 in

TE D

healthy, male volunteers. Neuropsychopharmacology. 2003;28:542–551. 38 Fukuyama Y, Huang J-M. Studies in Natural Products Chemistry. Vol 32, Attaur-Rahman, editors. Amsterdam: Elsevier; 2005. p. 395–429.

EP

39 Inoue M, Zhai H, Sakazaki H, Furuyama H, Fukuyama Y, Hirama M. TMC-95A, a reversible proteasome inhibitor, induces neurite outgrowth in PC12 cells. Bioorg

AC C

Med Chem Lett. 2004;14:663-665. 40 Zhai H, Nakatsuka M, Mitsumoto Y, Fukuyama Y. Neurotrophic effects of talaumidin, a neolignan from Aristolochia arcuata, in primary cultured rat cortical neurons. Planta Med. 2004;70:598-602.

41 Fukuyama Y, Kuwayama A, Minami H. Garsubellin A, a novel polyprenylated phloroglucin derivative, increasing acetyltransferase (ChAT) activity in postnatal rat septal neuron cultures. Chem Pharm Bull. 1997;45:947-949.

ACCEPTED MANUSCRIPT

42 Takaoka S, Takaoka N, Minoshima Y, Huang J-M, Kubo M, Harada K, Hioki H, Fukuyama Y. Isolation, synthesis, and neurite outgrowth- promoting activity of

RI PT

illicinin A from the flowers of Illicium anisatum. Tetrahedron 2009;65:8354-8361. 43 Moriyama M, Huang J-M, Yang C-S, Hioki H, Kubo M, Harada K, Fukuyama Y. Structure and neurotrophic activity aof novel sesqui- neolignans from the pericarps of Illicium fargesii. Tetrahedron 2007;63:4243-4249.

SC

44 Kubo M, Okada C, Huang J-M, Harada K, Hioki H, Fukuyama Y. Novel pentacyclic seco-prezizaane-type sesquiterpenoids with neurotrophic properties from Illicium

M AN U

jiadifengpi. Org Lett. 2009;11:5190-5193.

45 Huang J-m, Yokoyama R, Yang C-s, Fukuyama Y. Merrilactone A, a novel neurotrophic sesquiterpene dilactone from Illicium merrillianum. Tetrahedron Lett. 2000;41:6111-6114.

46 Fukuyama Y, Hata Y, Kodama M. Bicycloillicinone asarone acetal: A novel prenylated C6-C3 compound increasing choline acetyltransferase (ChAT) activity

TE D

from Illicium tashiroi. Planta Med. 1997;63:275-277.

47 Liu Y, Kubo M, Fukuyama Y. Spirocyclic nortriterpenoids with NGF-potentiating activity from the fruits of Leonurus heterophyllus. J Nat Prod. 2012;75:1353-1358.

EP

48 Zhai H, Nakade K, Oda M, Mitsumoto Y, Akagi M, Sakurai J, et al. Honokiol-induced neurite outgrowth promotion depends on activation of extracellular

signal-regulated

kinases

(ERK1/2).

Eur

J

Pharmacol.

AC C

2005;516:112-117.

49 Fukuyama

Y,

Asakawa

Y.

Novel neurotrophic

isocuparane-type

dimers,

mastigophorenes A, B, C and D, isolated from the liverwort Mastigophora diclados. J Chem Soc Perkin Trans. 1991;1:2737-2741.

50 Liu Y, Kubo M, Fukuyama Y. Nerve growth factor-potentiating benzofuran derivatives

from

2012;75:2152-2157.

the

medicinal

fungus

Phellinus

ribis.

J

Nat

Prod.

ACCEPTED MANUSCRIPT

51 Takahashi H, Yanagi K, Ueda M, Nakade K, Fukuyama Y. Structures of 1,4-benzodioxane derivatives from the seeds of Phytolacca americana

and

their

2003;51:1377-1381.

RI PT

neuritogenic activity in primary cultured rat cortical neurons. Chem Pharm Bull.

52 Tang W, Hioki H, Harada K, Kubo M, Fukuyama Y. Clerodane diterpenoids with NGF-potentiating

activity

from

Ptychopetalum

J

Nat

Prod.

SC

2008;71:1760-1763.

olacoides.

53 Fukuyama Y, Kubo M, Esumi T, Harada K, Hioki H. Chemistry and biological

M AN U

activities of vibsane-type diterpenoids. Heterocycles 2010;81:1571-1602. 54 Kubo M, Kishimoto Y, Harada K, Hioki H, Fukuyama Y. NGF-potentiating vibsane-type diterpenoids from Viburnum sieboldii. Bioorg Med Chem Lett. 2010;20:2566-2571.

55 Kumar GP, Khanum F. Neuroprotective potential of phytochemicals. Pharmacogn Rev. 2012;6:81–90.

TE D

56 Berrocal R, Vasudevaraju P, Indi SS, Sambasiva Rao KR, Rao KS. In vitro evidence that an aqueous extract of Centella asiatica modulates

-synuclein

aggregation dynamics. J Alzheimers Dis. 2014;39(2):457–465.

EP

57 Fujiwara H, Tabuchi M, Yamaguchi T, Iwasaki K, Furukawa K, Sekiguchi K, Ikarashi Y, Kudo Y, Higuchi M, Saido T, Maeda S, Takashima A, Hara M, Yaegashi N, Kase Y, Arai H. A traditional medicinal herb Paeonia suffruticosa and

AC C

its active constituent 1,2,3,4,6-penta-O-galloyl-beta-D-glucopyranose have potent anti-aggregation effects on Alzheimer’s amyloid beta proteins in vitro and in vivo. J Neurochem. 2009;169:1648–1657.

58 Cieslik E, Gre da A, Adamus W. Contents of polyphenols in fruit and vegetables. Food Chem. 2006;94:135–142.

59 Zettersten C, Co M, Wende S, Turner C, Nyholm L, Sjöberg PJR. Identification and characterization of polyphenolic antioxidants using on-line liquid chromatography,

ACCEPTED MANUSCRIPT

electrochemistry, and electrospray ionization tandem mass spectrometry. Anal Chem. 2009;81:8968–8977.

RI PT

60 Ge JF, Qiao JP, Qi CC, Wang CW, Zhou JN. The binding of resveratrol to monomer and fibril amyloid beta. Neurochem Int. 2012;61:1192–1201.

61 Endres K, Fahrenholz F. Regulation of alpha-secretase ADAM10 expression and activity. Exp Brain Res. 2012;217:343–352.

SC

62 Kang IJ, Jang BG, In S, Choi B, Kim M, Kim MJ. Phlorotannin-rich Ecklonia cava

reduces the production of beta- amyloid by modulating alpha- and gamma-secretase

M AN U

expression and activity. Neurotoxicology. 2013;34:16–24.

63 Mancini F, De Simone A, Andrisano V. Beta-secretase as a target for Alzheimer’s disease drug discovery: an overview of in vitro methods for characterization of inhibitors. Anal Bioanal Chem. 2011;400:1979–1996.

64 Chakraborty S, Kumar S, Basu S. Conformational transition in the substrate binding domain of

-secretase exploited by NMA and its implication in inhibitor

TE D

recognition: BACE1-myricetin a case study. Neurochem Int. 2011;58:914–923. 65 Zhou Y, Suram A, Venugopal C, Prakasam A, Lin S, Su Y, Li B, Paul SM, Sambamurti K. Geranylgeranyl pyrophosphate stimulates

and APP-CTF . Fed Am Soc Exp Biol J. 2008;22:47–54.

EP

the generation of A

-secretase to increase

66 Haass C, De Strooper B. The presenilins in Alzheimer’s disease—proteolysis holds

AC C

the key. Science. 1999;286:916–919. 67 Geng J, Li M, Wu L, Ren J, Qu X. Liberation of copper from amyloid plaques: making a risk factor useful for Alzheimer’s dis- ease treatment. J Med Chem. 2012;55:9146–9155.

68 Gupta VB, Rao KS. Anti-amyloidogenic activity of S-allyl- L-cysteine and its activity to destabilize Alzheimer’s 2007;429:75–80.

-amyloid fibrils in vitro. Neurosci Lett.

ACCEPTED MANUSCRIPT

69 Chaudhary N, Singh S, Nagaraj R. Aggregation properties of a short peptide that mediates amyloid fibril formation in model proteins unrelated to disease. J Biosci.

RI PT

2011;36:679–689. 70 Sinha S, Lopes DHJ, Bitan G. A key role for lysine residues in amyloid

-protein

folding, assembly, and toxicity. ACS Chem Neurosci. 2012;3:473–481.

71 Ge JF, Qiao JP, Qi CC, Wang CW, Zhou JN. The binding of resveratrol to

SC

monomer and fibril amyloid beta. Neurochem Int. 2012;61:1192–1201.

72 Porat Y, Abramowitz A, Gazit E. Inhibition of amyloid fibril formation by

M AN U

polyphenols: structural similarity and aromatic inter- actions as a common inhibition mechanism. Chem Biol Drug Des. 2006;67:27–37. 73 Dolai S, Shi W, Corbo C, Sun C, Averick S, Obeysekera D, Farid M, Alonso A, Banerjee P, Raja K. “Clicked” sugar–curcumin conjugate: modulator of amyloidand tau peptide aggregation at ultralow concentrations. ACS Chem Neurosci. 2011;2:694–699.

hinders beta

TE D

74 Convertino M, Pellarin R, Catto M, Carotti A, Caflisch A. 9, 10-Anthraquinone -aggregation: how does a small molecule interfere with A -peptide

amyloid fibrillation? Protein Sci. 2009;18:792–800.

EP

75 Yang F, Lim GP, Begum AN, Ubeda OJ, Simmons MR, Ambegaokar SS, Chen P, Kayed R, Glabe CG, Frautschy SA, Cole GM. Curcumin inhibits formation of amyloid

oligo- mers and fibrils, binds plaques, and reduces amyloid in vivo. J

AC C

Biol Chem. 2005;280:5892–5901.

76 Marin E, Briceño MI, Caballero-George C. Critical evalua- tion of biodegradable polymers used in nanodrugs. Int J Nanomedicine. 2013;8:3071–3091.

77 Rivière C, Richard T, Vitrac X, Mérillon JM, Valls J, Monti JP. New polyphenols active on

-amyloid aggregation. Bioorg Med Chem Lett. 2008;18:828–831.

ACCEPTED MANUSCRIPT

78 Ono K, Hamaguchi T, Naiki H, Yamada M. Anti- amyloidogenic effects of antioxidants: implications for the prevention and therapeutics of Alzheimer’s

RI PT

disease. Biochim Biophys Acta. 2006;1762:575–586. 79 Jung HA, Oh SH, Choi JS. Molecular docking studies of phlorotannins from Eisenia bicyclis with BACE1 inhibitory activity. Bioorg Med Chem Lett. 2010;20:3211– 3215.

SC

80 D’Archivio M, Filesi C, Varì R, Scazzocchio B, Masella R. Bioavailability of the polyphenols: status and controversies. Int J Mol Sci . 2010;11:1321–1342.

M AN U

81 Fujita M, Itokawa H, Sashida Y. Studies on the components of Magnolia obovata Thunb. 3. Occurrence of magnolol and honokiol in M. obovata and other allied plants. Yakugaku Zasshi 1973;93:429-434.

82 Fukuyama Y, Nakade K, Minoshima Y, Yokoyama R, Zhai H, Mitsumoto Y. Neurotrophic activity of honokiol on the cultures of fetal rat cortical neurons. Bioorg Med Chem Lett. 2002;12:1163-1166.

TE D

83 Maruyama Y, Kuribara H, Morita M, Yuzurihara M, Weintraub ST. Identification of magnolol and honokiol as anxiolytic agents in extracts of saiboku-to, an oriental herbal medicine. J Nat Prod. 1998;61:135-138.

EP

84 Takeda T, Hosokawa M, Higuchi K. Senescence-accerelated mouse SAM): a novel murin model of senescence. Exp Gerontol. 1997;32:105-109.

AC C

85 Nomura Y, Okuma Y. Age-related defects in lifespan and learning ability in SAMP8 mice. Neurobiol Aging. 1999;20:111-115.

86 Strong R, Reddy V, Morley JE. Cholinergic deficits in the septal-hippocampal pathway

of

the

SAM-P/8

senescence

accelerated

mouse.

Brain

Res.

2003;966:150-156.

87 Pollack SJ, Harper SJ. Small molecule Trk receptor agonists and other neurotrophic factor mimetics. Curr Drug Targets CNS Disord. 2002;1:59-80. 88 Potapoutian A, Reichardt LF. Trk receptors: mediators of neurotrophin action. Curr Opin Neurobiol. 2001;11:272-280.

ACCEPTED MANUSCRIPT

89 Ai J, Wang X, Nielsen M. Honokiol and magnolol selectively interact with GABAA receptor subtypes in vitro. Pharmacology 2001;63:34-41. 90 Haraguchi H, Ishikawa H, Shirataki N, Fukuda A. Antiperoxidative activity of

RI PT

neolignans from Magnolia obovata. J Pharm Pharmacol. 1997;49:209-212.

91 Lee J, Jung E, Park J, Lung K, Lee S, Hong S, et al. Anti-inflammatory effects of

magnolol and honokiol are mediated through inhibition of the downstream pathway of MEK-1 in NF-kappaB activation signaling. Planta Med. 2005;71:338-343.

92 Yasui F, Ishibashi M, Matsugo S, Kojo S, Oomura Y, Sasaki K. Brain lipid

SC

hydroperoxide level increase in senescence-accelerated mice at an early age. Neurosci Lett. 2003;350:66-68.

M AN U

93 Kovacs T, Cairms NJ, Lantos PL. Olfactory centre in Alzheimer’s disease: olfactory bulb is involved in early Braak’s stage. Neuroreport. 2001;12:285-288. 94 Esiri MM, Wilcock GK. The olfactory bulbs in Alzheimer’s disease. J Neurol Neurosurg Psychiatry. 1984;47:58-60.

95 Thorme BM, Aaron M, Latham EE. Effect of olfactory bulb abiation upon emotionality and muricidal behavior in four rat strains. J Comp Physiol Psychol. 1973;34:339-344.

TE D

96 Sieck MH. The role of the olfactory system in avoidance learning and activity. Physiol Behav. 1972;8:705-710.

97 Jarosik J, Legutko B, Werner S, Unsicker K, von Bohlen Und Halbach O. Roles of exogenous and endogenous FGF-2 in animal models of depression. Restor Neurol

EP

Neurosci. 2011;29:153–165.

98 Hozumi S, Nakagawasai O, Tan-No K, Niijima F, Yamadera F, Murata A, et al.

AC C

Characteristics of changes in cholinergic function and impairment of learning and memory-related behavior induced by olfactory bulbectomy. Behav Brain Res. 2003;138:9–15.

99 Esumi T, Makado G, Zhai H, Shimizu Y, Mitsumoto Y, Fukuyama Y. Efficient synthesis

and structure-activity

relationship

of honokiol, a

biphenyl-type neolignan. Bioorg Med Chem Lett. 2004;14: 2621-2625.

neurotrophic

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Table 1 The increase of people over 65-years old in various countries Ratio of aged 65+ (%) 2030

2040

2050

World

9.4

11.7

14.3

16.2

More developed regions

19.0

22.4

24.5

25.7

Less developed regions

7.5

9.8

Japan

29.1

31.6

China

12.0

16.5

India

6.3

8.3

United States of America

16.2

19.9

United Kingdom

18.7

21.1

Germany

23.0

28.0

France

20.3

RI PT

2020

23.1

12.5

14.7

36.1

38.8

23.3

25.6

10.5

13.5

20.9

21.2

23.0

23.6

31.0

30.9

24.9

24.9

SC

M AN U

Year

Note: data from "World Population Prospects : The 2010 Revision" by ¨United Nations. More developed regions: Europe, Northern America, Australia/New Zealand and Japan

The data for China don not include Hong Kong and Macao, Special Administrative

D

Regions (SAR) of China.

Japan: "2010 Population Census " by the Ministry of Internal Affairs and Communi-

TE

cations. Future population "Population & Household Projection (mediummortality assumption) by the National Institute of Population and Social

AC C

EP

Security Research.

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Table 2 Patients with dementia (x 10,000) and the increasing trend 2010

2015

2020

2025

Patients with Dementia

280

345

410

470

9.5

10.2

11.3

12.8

% of people above 65 years old

RI PT

Year

Note: data from National Institute of Populstion and Social Security Research,

AC C

EP

TE

D

M AN U

SC

"Japan's Future Estimated Population: 2006 median Estimate"

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Table 3 Experimental models to screen neurotrophic compounds PC12 cells (17), SH-SY5Y cells (18), P19 cells (19),

Cell lines

Neuro2A cells (20) et al.

RI PT

cortical neurons (21), hippocampal neurons (21), dorsal root ganglion neurons (22), spinal ventral horn neurons Primary cultured cells

(23),

medial septal neurons (24), retinal ganglion cells (25), asrtrocytes (26) et al. hippocampal slices (27)

et al.

SC

Tissue slices

senescence-accelerated mice 1 (SAMP1) (28), SAMP8 (29),

Whole animals

AC C

EP

TE

D

M AN U

olfactory bulbectomized mice (30) et al.

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Table 4 Representative small-molecular weight natural compounds with neurotrophic activity identified in our group Compound

References

Apiospora montagnei

TMC-95A

39

Aristolochia arcuata

talaumidin, nectandrin A, isonectandrin B, nectandrin B,

MASTERS

galgravin, aristolignin

Garcinia subelliptica

garsubellin A

RI PT

Sourse

41

illicinin A,

Illicium anisatum

SC

4-allyl-2-methoxy-6-(2-methylbut-3-en-2-yl)phenol

Illicium fargesii

isodunnianol

M AN U

jiadifenoxolane A, jiadifenolide,

Illicium jiadifengpi

(2R)-hydroxy-norneomajucin

Illicium merrillianum

merrilactone A

isodunnianin, tricycloillicinone, bicycloillicinone

Illicium tashiroi

40

asarone acetal, tricycloillicinone

42 43 44 45 46

leonurusoleanolide A, B, C, D

Magnolia ovobata

honokiol, magnolol

Mastigophora diclados

mastigophorenes A, B, D

49

Phellinus ribis

ribisin A, B, C, D

50

Phytolacca americana

americanoic acid A methyl ester

51

Plagiochila fruticosa

47 28, 29. 48

piperodione

5

plagiochilal B

27

AC C

Ptychopetalum

TE

EP

Piper retrofractum

D

Leonurus heterophyllus

ptychonal hemiaccetal, ptychonal

52

Viburnum awabuki

neovibsanin, neovibsanins A, B

53

Viburnum sieboldii

neovibsanins A, B

54

olaacoides

trans-3-(3',4'-dimethoxyphenyl)-4-[(E)-3",4"-dimethoxy

Zingiber purpureum

styryl]cyclohex-1-ene,

(BANGLE)

cis-3-(3',4'-dimethoxyphenyl)-4-[(E)-3",4"-dimethoxyst yryl]cyclohex-1-ene

30

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OH 2

1 1' 4' 3'

OH

honokiol OH

M AN U

SC

OH

RI PT

5

magnolol

AC C

EP

TE

D

Fig. 1 Structures of honokiol and magnolol

D

M AN U

SC

RI PT

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AC C

EP

TE

Fig. 2 Neurotrophic pathways of neurotrophins (NTs)

M AN U

SC

RI PT

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AC C

EP

TE

D

Fig. 3 Structures of illicinin A (A), and Aa - d