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Chemistry and Neurotrophic Activities of (–)-Talaumidin and Its Derivatives

(–)-Talaumidin (1), a 2,5-biaryl-3,4-dimethyltetrahydrofuran lignan isolated from Aristolochia arcuata Masters, exhibits significant neurite-outgrowth promotion and neuroprotection in primary cultured rat cortical neurons and in NGF-differentiated PC12 cells. The first enantioselective total synthesis of 1 was achieved by a flexible and reliable synthetic pathway involving an Evans asymmetric aldol reaction, as well as a stereocontrolled hydroboration and Friedel–Crafts arylation, to construct the four contiguous chiral centers on the tetrahydrofuran (THF) ring of 1. In order to investigate the stereochemistry–activity relationship of 1, a systematic synthesis of all diastereomers of 1 was accomplished by applying the synthetic strategy used for natural product 1. The evaluation of neurite-outgrowth promotion by all of the synthesized diastereomers indicated that the (–)-(1S,2R,3S,4R)-isomer 1e was significantly more active than naturally occurring 1. Additionally, we established a synthetic methodology for talaumidin derivatives that could be used to prepare a variety of analogs in a few steps and on a large scale. The synthesized racemic analog rac-1e (56a) exhibited neurite-outgrowth promoting activity in NGF-differentiated PC12 cells to the same degree as the optically active (–)-1e, revealing that a relative configuration bearing all-cis- substituents is important for potent neurotrophic activity, whilst the absolute configuration does not affect activity. Fourteen analogs based on (±)-56a were prepared via the same synthetic methodology. Among them, 56b with a methylenedioxy group on both benzene rings was found to exhibit the most significant neurite outgrowth promotion. In addition, 56a and 56b induced regeneration of the mouse optic nerve in vivo, and their activity was higher than that of talaumidin, as well as their in vitro measured activity. Furthermore, the structure–activity relationship of 56b indicated that the two benzene rings were essential structures, and that the methyl groups on the THF ring could enhance the neurotrophic activity. This result suggests that the two benzene rings of the talaumidin derivatives are essential structures for neurotrophic activity, while the two methyl groups on the THF ring can enhance neurite-outgrowth activity. Finally, it was observed that 1 and derivatives 56a and 56b exhibited potent regenerative activity in the injured mouse optic nerve in vivo.

(-)-Talaumidin (1), a 2,5-biaryl-3,4-dimethyltetrahydrofuran lignan isolated from Aristolochia arcuata Masters, exhibits significant neurite-outgrowth promotion and neuroprotection in primary cultured rat cortical neurons and in NGF-differentiated PC12 cells. The first enantioselective total synthesis of 1 was achieved by a flexible and reliable synthetic pathway involving an Evans asymmetric aldol reaction, as well as a stereocontrolled hydroboration and Friedel-Crafts arylation, to construct the four contiguous chiral centers on the tetrahydrofuran (THF) ring of 1. In order to investigate the stereochemistry-activity relationship of 1, a systematic synthesis of all diastereomers of 1 was accomplished by applying the synthetic strategy used for natural product 1. The evaluation of neurite-outgrowth promotion by all of the synthesized diastereomers indicated that the (-)-(1S,2R,3S,4R)-isomer 1e was significantly more active than naturally occurring 1. Additionally, we established a synthetic methodology for talaumidin derivatives that could be used to prepare a variety of analogs in a few steps and on a large scale. The synthesized racemic analog rac-1e (56a) exhibited neurite-outgrowth promoting activity in NGF-differentiated PC12 cells to the same degree as the optically active (-)-1e, revealing that a relative configuration bearing all-cis-substituents is important for potent neurotrophic activity, whilst the absolute configuration does not affect activity. Fourteen analogs based on (±)-56a were prepared via the same synthetic methodology. Among them, 56b with a methylenedioxy group on both benzene rings was found to exhibit the most significant neurite outgrowth promotion. In addition, 56a and 56b induced regeneration of the mouse optic nerve in vivo, and their activity was higher than that of talaumidin, as well as their in vitro measured activity. Furthermore, the structure-activity relationship of 56b indicated that the two benzene rings were essential structures, and that the methyl groups on the THF ring could enhance the neurotrophic activity. This result suggests that the two benzene rings of the talaumidin derivatives are essential structures for neurotrophic activity, while the two methyl groups on the THF ring can enhance neurite-outgrowth activity. Finally, it was observed that 1 and derivatives 56a and 56b exhibited potent regenerative activity in the injured mouse optic nerve in vivo.
Keywords: talaumidin, neurotrophic activities, PC12, regenerative activity, primary cultured rat cortical neurons INTRODUCTION Neurotrophins (NGF, BDNF, NT3, and NT4/5) are known to play essential roles in neuron survival, process outgrowth, and synaptic connectivity during development and nervous system plasticity in adults. Hence, they have a potential to become useful agents for neurodegeneration (Pardridge, 2002). Although, these polypeptide cannot cross the brain-blood barrier because of their high molecular weight and easily metabolize by peptidases under physiological conditions (Pardridge, 2002;Thoenen and Sendtner, 2002). Therefore, small molecules that can mimic the functions of neurotrophic factors might be promising alternatives for the treatment of neurodegenerative diseases (Xie and Longo, 2000;Massa et al., 2002). Neurotrophins also are able to promote process outgrowth and survival neuronal cells in vitro. Thus, we have been investigating neurotrophin-mimic small molecules from natural products based on rat cortical neuron cultures and PC12 cells, resulting in the discovery of interesting neurotrophic compounds (Huang et al., 2000(Huang et al., , 2001Fukuyama et al., 2002;Yokoyama et al., 2002;Kubo et al., 2009Kubo et al., , 2010Kubo et al., , 2012Kubo et al., , 2013Kubo et al., , 2015Matsui et al., 2012).
In our continuing studies on neurotrophic compounds, we isolated 1 from Blazilian plant Aristolochia arcuata. In addition to significant neurite-outgrowth promotion in primary cultured rat cortical neurons, we found that 1 and its analogs also exhibited neuroprotections against cell death induced by several insults (Zhai et al., 2004(Zhai et al., , 2005. Furthermore, 1, belonging to a diaryltetrahydrofuran-type lignan, possesses a tetrahydrofuran ring bearing four contiguous stereogenic centers. These promising biological activities and the prospective selective preparation of the possible stereoisomers with regard to the four stereogenic centers of 1 make it an attractive synthetic target. In this review, we focus and summarize neuriteoutgrowth promotion activities in primary cultured rat cortical neurons (Zhai et al., 2004) and in NGF-differentiated PC12 cells. Additionally, we describe the synthesis of 1 and all stereoisomers of 1 (Esumi et al., 2006;Fukuyama et al., 2008), and discuss structure-activity relationships between 1 and its analogs on PC12 cells . Furthermore, we report their regenerative activity toward mouse optic nerves as a neurotrophic activity in vivo, reinforcing their potential as therapeutic agents for neurodegenerative disease .

Neurite-Outgrowth Promoting Activity of Talaumidin in PC12 Cells
Rat pheochromocytoma PC12 cells have been widely used as a model cells of neurons (Vaudry et al., 2002). When PC12 cells are stimulated with NGF, they cease growth and begin to grow neurites, eventually differentiating into a neuron-like phenotype. In the absence of NGF, talaumidin had no morphological effects on PC12 cells. In the presence of 20 ng/mL NGF, however, talaumidin promoted neurite outgrowth dose-dependently at concentrations of 1-30 µM, inducing longer average neurite length, as well as a higher percentage of neurite-bearing cells (Figure 2). These effects were validated through morphological observations (Figures 2C vs. 2B) and quantitative analysis of neurites ( Figure 2D).

Neurotrophic Effects of Talaumidin in Primary Cultured Rat Cortical Neurons
In tissue sections and cell cultures, the anti-MAP2 antibody can stain neuronal cell bodies and dendrites but cannot use for efficient recognition of axons (Kaufmann et al., 1997). Alternatively, the anti-tau antibody reacts with tau proteins, which are distributed over the entire neuron surface, thus staining the cell body, dendrites as well as axons of neurons (Dotti et al., 1987). First, the morphological effects of talaumidin on cultured rat cortical neurons were evaluated by the anti-MAP2 staining method (Figures 3A,B). Talaumidin has been found to exhibit a significant promoting neurite outgrowth in the primary cultures of rat cortical neurons at concentration of 10 µmol/L. Measurements of each neuron stained by anti-MAP2  *P < 0.05, **P < 0.01, ***P < 0.001 compared with control (Ctrl) (Zhai et al., 2004).
was performed by morphological analysis of process outgrowth brought on by talaumidin, and the quantitative results are shown in Figure 3C. It is obvious that talaumidin promotes process elongation dose-dependently at concentrations ranging from 3 to 30 µM. The longest processes stained with the anti-tau method are referred to as axon-like neurites, while others are referred to as dendrite-like neurites for clarity in the description of effects of 1 on neurite outgrowth. The morphological evaluation was carried out by anti-tau staining method (Figures 3D,E). According to expectations, 1 was observed to significantly promote dendrite-like processes, as well as axon-like processes at 10 µM. Quantitative analysis indicated that 1 enhanced process elongation in a dose-dependent manner at concentrations ranging from 1 to 30 µM ( Figure 3F). Additionally, 1 also showed neuroprotective effects against serum deprivationinduced cell death in rat cortical neurons (Zhai et al., 2004).

Evaluation of Neurite-Outgrowth Promoting Activity of Talaumidin and Other 2,5-diaryl-3,4-dimethyltetrahydrofuran Neolignans
The neurotrophic effects of 2,5-diaryl-3,4dimethyltetrahydrofuran neolignans (2-7) were compared with talaumidin (1) (Zhai et al., 2005). Comparison of the effects of compounds 1-7 in the neurite-outgrowth assay indicated that 5, 6, and 7 were similar to 1, but 2 and 4 had fewer effects than 1 (Figure 4). Especially, all-cis-substituted -type 6 showed the most potent neurite-outgrowth promoting activity at 30 µM. Furthermore, compounds 5 and 7, whose stereochemistry are trans-cis-trans (4,5-trans, 3,4-cis, and 2,3-trans), presented similar activity of 1. Curiously, trans-cis-trans 3 could not be attributed to effect. The above preliminary structure-activity analysis assumes that the stereochemistry of tetrahydrofuran ring and substituents on two benzene groups would make an important contribution to the enhancement of activity. It's very interesting challenge to investigate the relationship between stereochemistry and substituent and the neurotrophic activity of talaumidin.

Total Syntheses of (-)-Talaumidin
Due to its unique structure and potent neurotrophic activities, talaumidin has been the target of extensive synthetic efforts over the years. To date, six syntheses have been reported. In 2006, we reported the first total synthesis of (2S,3S,4S,5S)talaumidin via a flexible and stereo-controlled synthetic strategy (Esumi et al., 2006;Fukuyama et al., 2008). In addition, we determined the absolute configuration of (-)-talaumidin to be (2S,3S,4S,5S) during the course of the synthetic study. In 2007, Hanessian et al. attained the synthesis of unnatural (+)talaumidin along with four tetrahydrofuran analogs (Hanessian and Reddy, 2007). In the same year, Kim et al. disclosed the stereoselective reductive deoxygenation/epimerization of cyclic hemiacetals and the synthesis of (+)-talaumidin (Kim et al., 2007). Alternative syntheses of talaumidin have additionally been FIGURE 4 | Comparison of the effects of compounds 1-7 on neurite outgrowth in primary cultured rat cortical neurons. # P < 0.001 vs. control (Zhai et al., 2005). SCHEME 1 | First synthesis of (-)-talaumidin by Fukuyama et al. reported by Matcha and Ghosh (2008), Rye andBarker (2009), andXue et al. (2009). Synthetic studies of tetrahydrofuran-type lignans are actively continuing worldwide.
Synthesis of (+)-Talaumidin by Hanessian et al. In 2007, Hanessian et al. reported a second synthesis of talaumidin, which was performed as part of a synthetic study on tetrahydrofuran-type lignan compounds (Scheme 2) (Hanessian and Reddy, 2007). The synthesis began with a chiral cyanohydrin 18, in turn prepared by a catalytic asymmetric reaction of 17, according to Belokon's protocol (Belokon et al., 2000). After Wittig olefination, 1,4-addition of 19 with dimethyllithium cuprate and TMSCl afforded anti-configuration in 12:1 dr. Subsequent α-alkylation of the ester with MeI via an enolate provided 20 with high diastereoselectivity. Then, a Grignard reaction with the aldehyde in the presence of CeCl 3 provided 21 having the two (R)hydroxy moieties. The key cycloetherification of 21 gave rise to the talaumidin skeleton in 90% yield through a quinone methide intermediate. Following deprotection, the total synthesis of (+)-talaumidin was attained in an overall yield of 12.7% over 16 steps. SCHEME 2 | Total syntheses of talaumidin.
Frontiers in Chemistry | www.frontiersin.org Synthesis of (+)-Talaumidin by Kim et al. Kim et al. investigated synthetic methodologies for the assembly of tetrahydrofuran lignans and the total synthesis of (+)talaumidin was accomplished as a part of these studies (Kim et al., 2007). Talaumidin was constructed utilizing the Evans aldol, anti-selective dimethylation, and Friedel-Crafts reactions as key transformations. In addition, they established a synthetic strategy featuring Lewis acid promoted deoxygenation followed by epimerization of a hemiacetal, and accomplished the synthesis of five tetrahydrofuran-type lignans. The efficiency of Kim's synthesis was showed by the short step and high overall yield of 31%.

Synthesis of (±)-Talaumidin by Rye et al.
In 2009, Rye et al. reported a straightforward synthetic methodology for the preparation of tetra-substituted tetrahydrofuran lignans such as (±)-talaumidin and (±)fragransin A 2 . The synthetic pathway began with an acyl-Claisen rearrangement to construct two successive tertiary stereocenters. After the introduction of an aryl group, the intramolecular cyclization of monoprotected 1,4-diol 34 gave the talaumidin skeleton. Removal of the MOM group completed the total synthesis of racemic talaumidin in an overall yield of 13.8%. Its analog, racemic franransin A 2, was synthesized in the same manner, in an overall yield of 5.8%. Although it was a racemic synthesis, the economic synthesis of 1 was accomplished from inexpensive starting materials in 10 steps.

Stereoselective Construction of Tetrahydrofuran-Type Lignan Skeleton
In these syntheses, some common procedures were established in order to construct the four chiralities of talaumidin. Evans aldol reaction has been utilized by three research groups and proven to be an optimal procedure forming successive chiral centers at C2 and C3 of 1. On the other hand, the third chiral center at C4 was constructed by α-substitution of carbonyl group, except for Fukuyama's synthesis. Finally, Friedel-Crafts reaction or intramolecular etherification have been adopted for the control of chirality at C5. The both reactions are found to be appropriate to control the 4,5-trans-configuration. These procedures would be a standard strategy for the synthesis of tetrahydrofuran-type lignans.

The Relationship Between Stereochemistry and Neurite-Outgrowth Activity of Talaumidin (1)
Following our asymmetric total synthesis of (-)-talaumidin, we embarked on structure-activity relationship (SAR) studies with the aim of potential drug discovery based on talaumidin. Initially, the relationship between stereochemistry and the neurotrophic activity of talaumidin was investigated. As 1 possesses four successive asymmetric carbons on the THF structure, seven diastereomers 1a-1g are possible, in addition to the enantiomer of each. The successful control of the four contiguous stereocenters in an asymmetric synthesis of all seven diastereomers would be crucial for the elucidation of the relationship between stereochemistry and neurotrophic activity, and likewise be an important achievement from the viewpoint of organic synthetic chemistry. In 2015, we published the systematic synthesis of talaumidin diastereomers and their evaluation of neurotrophic activity (Scheme 3) .
Diastereomers 1a-1c having (2S,3S)-configuration were synthesized from 13, which was an intermediate in the synthesis of 1 (Esumi et al., 2006;Fukuyama et al., 2008). After the introduction of the aryl group, the resulting ketone 38 was converted to dihydrofuran 39. The stereoselective hydrogenation of dihydrofuran 39 provided (2S,3S,4S,5R)-1a as a sole stereoisomer. Next, treatment of hemiacetal 40 with BF 3 ·OEt 2 followed by reduction with NaBH 3 CN furnished (2S,3S,4R,5S)-1b. In this reductive sequence, the hydride nucleophile approaches from the opposite face to the adjacent methyl group. On the other hand, the synthesis of (2S,3S,4R,5R)-1c was attained by applying a Mitsunobu-type reaction with 1,4-diol 41 using DEAD and PPh 3 . Noteworthy, the stereoselective cyclization proceeded with net retention of original configurations at C1 and C4. This surprising stereoselectivity was rationalized by preferential elimination of the PPh 3 -activated hydroxy group over the normal substitution pathway, giving rise to quinone methide intermediates (Harada et al., 2011b). The steric hindrance between a methyl group and the adjacent aryl group forces the product to adopt the desired conformations, resulting in the construction of (2S,3S,4R,5R)-configuration.
Subsequently, the synthesis of stereoisomers 1d-1g with (2S,3R)-configuration was achieved via common intermediate 44. The (2S,3R)-configuration of 44 was constructed by synselective Evans aldol reaction between 8 and 9 in 80% yield with >99% de. According to the same synthetic procedure  for (-)-1, the key intermediate 44 was derived from 43 by hydroboration with 9-BBN. With 44 in hand, the synthesis of (2S,3R,4S,5S)-1d was firstly attained by a cyclization of diol 13 under conditions of Mitsunobu-type reaction (Harada et al., 2011a). In this case, the benzyl group was converted to a tosylate prior to cyclization, in order to enhance the selectivity eliminating the hydroxy group at C4. Next, hydrogenation of dihydrofuran 11b gave all-cis-substituted (2S,3R,4S,5R)-1e with high diastereoselectivity. In addition, the C4 position of lactone 21 was epimerized, and then led to 23 by reduction of the lactone, followed by sulfonation. In accordance with Ley's method (Brown et al., 1989;Kim et al., 2007), a Grignard reaction with zinc bromide introduces the methylenedioxy benzene moiety from the β-face to avoid steric hindrance between the aryl groups (Harada et al., 2011a). Then, (2S,3R,4R,5S)-1f was synthesized by removal of the benzyl group. Finally (2S,3R,4R,5R)-1g was synthesized by the reduction of 27 with NaBH 3 CN/BF 3 ·OEt 2 conditions. Regarding this stereochemistry, the epimerization at C3 proceeded spontaneously in order to decrease the steric hindrance from the methyl group.
These synthetic studies provided useful information for the analysis of stereochemistries of tetrahydrofuran lignans. The characteristic 1 H and 13 C NMR data of 1 and 1a-1g are summarized in Tables 1, 2. Although coupling constants are indecisive, chemical shifts play a role in identifying relative stereochemistries on the THF ring. In the case of 2,3-transand/or 4,5-trans-configurations, the signal of methyl group appears at 0.99-1.04 ppm. In contrast, 2,3-cis-and/or 4,5-cisoriented methyl groups are shielded by the adjacent aromatic ring to result in upfield shift of the signal at 0.59-0.69 ppm. Moreover, the relative 2,3-stereochemistry is also able to be confirmed by 13 C NMR. The chemical shifts of benzylic carbon are at 85.7-88.5 ppm for 2,3-trans and at 82.7-84.8 ppm for 2,3-cis, respectively. On the other hand, the relative configuration of 3,4-dimethyl groups can be distinguished by the chemical shifts of 13 C NMR. The signal of 3,4-trans-dimethyl groups appears at 13.8-15.1 ppm, whereas 3,4-cis-dimethyl groups have lower chemical shifts around 9.4-12.9 ppm. These results of NMR experiments are consistent with those of natural products 2-7 and useful for the determination of relative stereochemistries of tetrahydrofuran type lignans.
Once the synthesis of all diastereomers was complete, their neurotrophic activity was compared with that of natural talaumidin . Talaumidin (1) and isomers 1a-1g were assessed their neurite-outgrowth promoting activity together with enantiomer of (-)-talaumidin. Consequently, the enantiomer of (-)-talaumidin exhibited activity similar to the natural product, and all synthesized compounds induced neurite-outgrowth promotion. Particularly, 1e having all-cisconfiguration was found to show more potent activity than naturally occurring talaumidin ( Figure 5). Furthermore, their neurite-outgrowth promoting activity of stereoisomers in primary cultured rat cortical neurons was evaluated at 0.01 µM.
The results indicated that all-cis-substituted 1e also exhibited the most significant neurite-outgrowth promotion among all of the stereoisomers (Figure 5).

Synthesis of Racemic Compound 56a and Relationship Between Substituents on the Benzene Ring and Neurotrophic Activity
(-)-Talaumidin and the stereoisomer (-)-1e were found to possess potent neurotrophic activity, however, preparative procedures of optically active (-)-talaumidin and (-)-1e required long synthetic steps and high cost. Therefore, drug discovery based on talaumidin necessitated a simplification of the structure and synthetic methodology for talaumidin derivatives. Then, we focused on the efficient synthetic methodology of talaumidin derivatives and exploration of new compounds that could be supplied on a large scale . In section The Relationship Between Stereochemistry and Neurite-Outgrowth Activity of Talaumidin (1), it was revealed that there are few difference in neurotrophic activity between both enantiomers of 1. This result suggested that a racemic mixture of 1e could have activity similar to optically active 1e. Accordingly, racemic 1e (56a) was decided on as the next target compound. The developed step-economic synthesis of rac-1e (56a) is shown in Scheme 4. The synthesis began with a Grignard reaction onto the commercially available benzaldehyde 8, followed by a Dess-Martin oxidation. After bromination of 51, the obtained bromide 52 was coupled with 53, giving rise to a diketone 54 in 86% yield. Subsequently, Paal-Knorr furan synthesis of 54 under acidic conditions provided a furan compound 55 in good yield. At last, hydrogenation of the furan ring completed the synthesis of 56a bearing all-cis-configuration. Consequently, the synthesis of racemic 1e (56a) was accomplished in 6 steps with an overall SCHEME 4 | Step-economic synthesis of talaumidin derivatives. yield of 39%. In addition, five novel talaumidin derivatives were prepared by applying this synthetic methodology.
With six derivatives 56-ad, 56f, and 56g and their precursor furan compounds 55-ad, 55f, and 55g in hand, the neuriteoutgrowth activity was evaluated in NGF-differentiated PC12 cells at 30 µM. As results, tetrahydrofuran compounds tended to promote neurite-outgrowth to a higher degree than furan-type compounds (Figure 6). Among the tetrahydrofuran compounds, 56b having two methylenedioxyphenyl groups was found to exhibit the most significant activity. In addition, the stepeconomic synthesis of talaumidin derivatives allowed adequate quantities of samples to be prepared for in vivo experiments. Thus, we evaluated the optic nerve regenerating activity of talaumidin derivatives as an in vivo experiment. Remarkably, the all-cis-derivatives 56a and 56b showed high regenerative activity toward the injured optic nerve.

The Role of Dimethyl and Diaryl Groups on THF Ring of Talaumidin Derivatives
Furthermore, the role of the phenyl and methyl groups on the THF ring of 56b was examined as part of the SAR study. In order to determine which moieties were necessary for neurotrophic properties, 57 lacking one benzene ring and 58 lacking two methyl groups were prepared, and their neurotrophic activity was assessed in NGF-differentiated PC12 cells (Figure 7). It was found that monophenyl analog 57 exhibited no activity   at all, whereas 58 had lower activity than 56b. These results indicate that the two benzene rings of the talaumidin derivatives are essential structures for neurotrophic activity while the two methyl groups at C3 and C4 positions can increase the neuriteoutgrowth activity.

Mechanistic Study on Neurite-Outgrowth Activity of Staurosporine-Differentiated RGC-5 Cells
In 2018, Koriyama et al. examined the neurite-outgrowth promotion of talaumidin in the neuroretinal cell line, RGC-5 (Koriyama et al., 2018). They assessed the neurite outgrowth effect and elucidated a mechanism of its neurotrophic action. At concentrations ranging from 1 to 10 µM, talaumidin promoted neurite outgrowth dose-dependently in differentiated RGC-5 cells. Its neurite-outgrowth promoting activity was not altered by PD98059, an extracellular signal-regulated kinase inhibitor. On the other hand, LY29002, a PI3K inhibitor, decreased the talaumidin-mediated neurite outgrowth. These results indicate that the PI3K-Akt signaling is involved in downstream pathway in talaumidin-induced neurite-outgrowth activity of RGC-5 cells.

CONCLUSION
In conclusion, the neurotrophic and protective activities of talaumidin have been found in several cellular models. Talaumidin can not only promote neurite outgrowth in NGFdifferentiated PC12 cells but also enhance cell survival after NGF withdrawal in differentiated PC12 cells. These are coincident with the fact that talaumidin has neurotrophic effects on primary cultured rat cortical neurons. In addition, the neurotrophic activities of talaumidin are extended to neuroprotection, which are deleterious factors in Alzheimer's disease. Due to its interesting structure and neurotrophic activity, talaumidin have attracted considerable attentions from synthetic chemists. We achieved the first enantioselective synthesis of (-)-(2S,3S,4S,5S)talaumidin using a Evans aldol reaction, hydroboration, and FriedelCrafts reaction. In addition, the systematic synthesis of all of the stereoisomers of (-)-talaumidin was accomplished, and their neurotrophic activity was evaluated. As results, the all-cis-substituted isomer 1e showed more potent neuriteoutgrowth promotion in NGF-differentiated PC12 cells than natural product talaumidin. Furthermore, we established a stepeconomic synthesis that could prepare a compound library based on talaumidin, and 14 derivatives were synthesized. As a result, compound 56b having two methylenedioxyphenyl groups was found to show the most potent neurite-outgrowth promoting activity in vitro. Moreover, it was found that derivatives 56a and 56b could induce the regeneration of mouse optic nerve in vivo. These consequences indicate that talaumidin derivatives can be an innovative agent for neurodegenerative diseases such as glaucoma, depression, and Alzheimer's disease. Further mechanistic and pharmacological investigations of neurotrophic activities of talaumidin derivatives are currently ongoing.

AUTHOR CONTRIBUTIONS
KH, MK, and YF wrote the manuscript. All authors discussed the results and commented on the manuscript.