The first F-ring modified ciguatoxin analogue showing significant toxicity

Article abstract of DOI:10.1039/b924375e

Ciguatoxins, the principal causative toxins of ciguatera seafood poisoning, are potent neurotoxic polycyclic ethers. We report herein the total synthesis of a 10-membered F-ring analogue of 51-hydroxyCTX3C, which constitutes the first example of an F-ring modified ciguatoxin that exhibits potent cytotoxicity as well as mouse acute toxicity. The Royal Society of Chemistry 2010.

Full text of DOI:10.1039/b924375e

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The first F-ring modified ciguatoxin analogue showing significant
toxicityw
Yuuki Ishihara,a Nayoung Lee,a Naomasa Oshiro,b Shigeru Matsuoka,c Shuji Yamashita,*a
Masayuki Inouec and Masahiro Hirama*ad
Received (in Cambridge, UK) 20th November 2009, Accepted 12th February 2010
First published as an Advance Article on the web 5th March 2010
DOI: 10.1039/b924375e
Ciguatoxins, the principal causative toxins of ciguatera seafood
poisoning, are potent neurotoxic polycyclic ethers. We report
herein the total synthesis of a 10-membered F-ring analogue of
51-hydroxyCTX3C, which constitutes the first example of an
F-ring modified ciguatoxin that exhibits potent cytotoxicity as
well as mouse acute toxicity.
Ciguatoxins (e.g., 1, Fig. 1), the principal causative toxins of
ciguatera seafood poisoning,1–4 are large ladder-like polycyclic
ethers 3 nm in length with 13 rings ranging from 5- to
9-membered.5 These toxins are produced by an epiphytic
dinoflagellate, Gambierdiscus toxicus,6 and transferred to
herbivorous and carnivorous fish through the aquatic food
chain, and eventually to humans. Ingestion of affected fish
leads to gastrointestinal, cardiovascular and neurological
disorders which may last for weeks or even years. The lethal
potency of ciguatoxins (LD50 = 0.25–4 mg kgꢀ1 2–5
by
)
intraperitoneal injection into mice was reported to be greater
than that of the structurally related red-tide toxins, brevetoxins
(LD50 4 200 mg kgꢀ1).7–10
These marine natural products exhibit their toxicities by
binding to the voltage-sensitive sodium channels (VSSC) of
excitable membranes, causing a continuous depolarization of
membrane potential.11,12 Unfortunately, the very limited
supply of ciguatoxins from natural sources has prevented the
biological and structure–activity relationship (SAR) studies
that would aid in understanding the structural requirements
necessary for ciguatoxin–VSSC interactions.13–15
Fig.
1
Structures of ciguatoxin 51-hydroxyCTX3C (1) and its
analogues.
SAR studies. The 8-membered ring (2) and the open chain
O-linked analogue (3) suggested the critical importance of the
9-membered F-ring of 1 for potent biological activity.18 Here,
we report the total synthesis and biological activity of a new
analogue, the 10-membered F-ring 51-hydroxyCTX3C (4),
which shows more potent activity than 2 and 3.
Recently, we reported the unified total synthesis of
ciguatoxins, CTX1B, CTX3C, and 51-hydroxyCTX3C (1), in
quantities sufficient for testing.16 Pharmacological studies
revealed that the synthetic ciguatoxins exerted multimodal
effects on VSSC with simultaneous stimulatory and inhibitory
aspects.17 Moreover, this unified synthetic strategy allowed us
to prepare fully synthetic analogues of the ciguatoxins for
The 10-membered F-ring analogue (4) was synthesized from
the previously reported key intermediate 516d,18 in seven
steps (Scheme 1). DIBAL reduction of ester 5 followed by one-
carbon homologation of alcohol 6 afforded nitrile 7. DIBAL
reduction of 7 and subsequent Wittig olefination of aldehyde 8
gave pentaene 9 in 90% overall yield. Ring-closing olefin
metathesis using Grubbs’ first generation catalyst19 provided
the desired tridecacyclic system 10 quantitatively. Finally, the
removal of four 2-naphthylmethyl (NAP) groups20 of 10 was
realized by DDQ oxidation to generate the target molecule 4.z
Biological in vitro and in vivo activities of 4 were assayed
(Table 1). To our surprise, the EC50 value, which was
evaluated using mouse neuroblastoma Neuro-2A cells,21
indicated that 4 exhibited 400–700 times greater toxicity than
the other F-ring modified analogues 2 and 3, although it was
still 1/50 less cytotoxic than 1. Moreover, 4 was the first
synthetic analogue to exhibit some mouse acute toxicity.
a Department of Chemistry, Graduate School of Science,
Tohoku University, Sendai 980-8578, Japan.
E-mail: s-yamashita@mail.tains.tohoku.ac.jp,
hirama@mail.tains.tohoku.ac.jp; Fax: +82-22-795-6566
b Okinawa Prefectural Institute of Health and Environment,
Nanjo 901-1202, Japan
c Graduate School of Pharmaceutical Sciences, The University of
Tokyo, Tokyo 113-0033, Japan
d Research and Analytical Center for Giant Molecules, Graduate
School of Science, Tohoku University, Sendai 980-8578, Japan
w Electronic supplementary information (ESI) available: Detailed
experimental procedures and associated spectrometric data. See
DOI: 10.1039/b924375e
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Scheme 1 Total synthesis of 10-membered F-ring analogue (4) of 51-hydroxyCTX3C. Cy = cyclohexyl, DDQ = 2,3-dichloro-5,6-dicyano-1,4-
benzoquinone, DIBAL = diisobutylaluminium hydride, DMSO = dimethylsulfoxide, Ts = p-toluenesulfonyl.
satisfied the NOE experiment of the EF0GH-ring model 11
Table 1 Biological assays of 51-hydroxyCTX3C (1), F-ring modified
analogues 2, 3 and 4
(Fig. 2 and 3A). Although 4 shows a bent form in the most
stable state, a relatively straight rod-like structure similar to 1
Compounds Cytotoxicity EC50/nMa Acute toxicity LD50/mg kgꢀ1b
was found at a higher level (2.74 kcal molꢀ1 more) than the
1
0.0049
103
170
0.31
most stable conformer (Fig. 3B). The root mean square
2c
3c
4
4667
4667
600
0.24
a
Cytotoxicity was determined as EC50 on mouse neuroblastoma cells
b
Neuro-2A. Acute toxicity was determined as LD50 on i.p. injected
c
mice. Ref. 18.
Modeling studies of 4 (MM2* in MacroModel Ver. 8.1)22
suggested that the assay data could be related to the overall
shapes of the molecules.18 The most stable conformer of 4 was
chosen from the pool of energy-minimized structures that
Fig. 3 (A) Energy-minimized structures of 10-membered F-ring
analogue (4) of 51-hydroxyCTX3C. (B) Energy-minimized structures
of 51-hydroxyCTX3C (1) (green) and 4 (yellow) and the straight
Fig. 2 Energy-minimized structure (MM2*, Macro Model Ver. 8.1)
and NOE data for the model compound 11. The phenyl groups were
not included in the energy calculation.
conformer of
4
(pink) with ABCDE-rings superimposed
(MM2*, Macro Model Ver. 8.1).
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deviation (RMSD) of 17 oxygen atoms between the most
stable structure of 1 and the straight conformer of 4 was
calculated to be 0.65 A. In contrast, the minimum RMSD
between the most stable conformer of 1 and the 8-membered
F-ring analogue 2 was found to be much larger (1.69 A).
The corresponding straight structure of 2 was not found
within the 12.0 kcal molꢀ1 range from the most stable state.
In conclusion, we have synthesized the 10-membered F-ring
analogue (4) of 51-hydroxyCTX3C and found an apparent
relationship between the biological activities and the
overall shapes of the polycyclic molecules (2–4). While a
conformational change of 4 to the straight form might be
induced by the interaction with VSSC, it is more likely that the
minor straight rod-like conformer of 4 fits into and best
interacts with VSSC using a hydrogen-bond network similar
to natural product 1. Further SAR studies of ciguatoxins are
under active investigation in our laboratory.
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This work was supported financially by a Grant-in-Aid for
Specially Promoted Research from the Ministry of Education,
Culture, Sports, Science and Technology (MEXT) in Japan.
Fellowships for Young Scientists to Y. I. from the Japan
Society for the Promotion of Science (JSPS) are gratefully
acknowledged.
Notes and references
z Selected analytical data: 4, colorless amorphous; 1H NMR
(500 MHz, C6D5N, 25 1C) d 7.28 (1H, d, J = 3.5 Hz, OH7), 6.72
(1H, d, J = 3.5 Hz, OH44), 6.52 (1H, d, J = 4.0 Hz, OH51), 6.07 (1H,
m, H19), 5.89 (1H, m, H13), 5.85 (1H, m, H23), 5.83 (1H, m, H18),
5.81 (1H, m, H2), 5.72 (1H, m, H3), 5.69 (1H, m, H14), 5.57 (1H, m,
H24), 4.98 (1H, m, OH29), 4.84 (1H, m, H51), 4.46 (1H, ddd,
J = 10.5, 10.5, 4.5 Hz, H41), 4.30 (1H, dd, J = 10.5, 5.5 Hz, H1),
4.22 (1H, m, H29), 4.21 (1H, m, H44), 4.19 (1H, m, H12), 4.16 (2H, m,
H52 ꢂ 2), 4.16 (1H, m, H15), 4.05 (1H, m, H20), 4.05 (1H, m, H45),
4.02 (1H, m, H1), 3.99 (1H, m, H7), 3.95 (1H, dd, J = 10.5, 10.5 Hz,
H46), 3.89 (1H, m, H27), 3.71 (1H, m, H16), 3.68 (1H, m, H26), 3.62
(1H, m, H6), 3.55 (1H, m, H5), 3.49 (1H, dd, J = 8.5, 8.5 Hz, H11),
3.44 (1H, ddd, J = 9.5, 9.5, 3.5 Hz, H21), 3.40 (1H, m, H34), 3.38 (1H,
m, H9), 3.37 (1H, m, H8), 3.32 (1H, m, H39), 3.26 (1H, ddd, J = 12.5,
10.5, 4.5 Hz, H33), 3.20 (1H, m, H31), 3.19 (1H, m, H38), 3.17 (1H, m,
H22), 3.15 (1H, m, H42), 2.87 (1H, m, H17), 2.66 (1H, m, H4), 2.62
(1H, m, H28), 2.60 (1H, m. H47), 2.58 (1H, m, H40), 2.57 (1H, m,
H22), 2.50 (1H, ddd, J = 11.0, 4.5, 4.5 Hz, H10), 2.43 (1H, m, H4),
2.32 (1H, dd, J = 13.5, 3.5 Hz, H50), 2.29 (1H, m, H250), 2.28 (1H, m,
H28), 2.27 (1H, m, H17), 2.25 (1H, m, H32), 2.04 (1H, m, H25), 2.02
(1H, m, H250), 2.01 (1H, m, H37), 1.99 (1H, m, H43), 1.91 (1H, m,
H32), 1.90 (1H, m, H36), 1.83 (1H, m, H35), 1.78 (1H, m, H40), 1.74
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This journal is The Royal Society of Chemistry 2010
2970 | Chem. Commun., 2010, 46, 2968–2970