Tag-reporter strategy for facile oligosaccharide synthesis on polymer support [16]

Full text of DOI:10.1021/ja003856c

3848
J. Am. Chem. Soc. 2001, 123, 3848-3849
Scheme 1. Tag-Reporter Strategy
Tag-Reporter Strategy for Facile Oligosaccharide
Synthesis on Polymer Support
Hiromune Ando,† Shino Manabe,† Yoshiaki Nakahara,†,‡ and
Yukishige Ito*,†
RIKEN (The Institute of Physical and Chemical
Research) and CREST, Japan Science and
Technology Corporation (JST), 2-1 Hirosawa, Wako-shi
Saitama 351-0198, Japan
Department of Industrial Chemistry, Tokai UniVersity
Kitakaname 1117, Hiratuka-shi, Kanagawa, 259-1292, Japan
ReceiVed NoVember 2, 2000
Development of a general method for the rapid assembly of
oligosaccharides based on polymer support technology has been
the subject of intense effort.1 Considering the structural diversity
of glycoconjugate derived oligosaccharides, the advantages of
polymer support synthesis are obvious, in terms of speeding up
as well as potential for extension to combinatorial and automated
synthesis. However, to reach this goal, several fundamental
problems have yet to be overcome, the most serious of which
are (1) the reduced reactivity of substrates bound to a polymer
support, (2) the difficulty of monitoring such reactions, and (3)
limitations on the ability to scale up reactions. Taking account
of these shortcomings, it would be most advantageous to merge
polymer support technology with solution-phase chemistry, so
that the major drawbacks inherent to solid-phase reactions could
be kept to a minimum.2 Herein, we report a novel technology for
monitorable, high-yielding soluble polymer support oligosaccha-
ride synthesis3 based on the “tag-reporter” strategy. This exploits
low-molecular weight poly(ethylene)glycol (PEG)4 as a polymer
support tag in combination with temporary chloroacetyl (CAc)
protection, which also serves as a reporter group (Scheme 1).
With this combination, the chain elongation process (glycosyla-
tion) and chemoselective deprotection can be monitored by matrix-
associated laser desorption/ionization mass spectrometory (MALDI-
TOF MS)5 and a coloring reaction, respectively.
Starting with PEG-tagged acceptor, each glycosylation reaction
is monitored by MALDI-TOF MS, because PEG-bound materials
are easily distinguished from others by their mountain-like shape
which derives from statistical distribution of the PEG chain length
consisting of 8-20 ethylene glycol units. Retrieval of the coupled
product can be achieved simply by direct chromatography of the
reaction mixture on silica gel. With ethyl acetate (AcOEt), the
PEG-tagged component stays at the origin, while nonsupported
sugars move rapidly through the column. After AcOEt washing,
PEG-bound products can be readily eluted with AcOEt-MeOH.
The CAc group was adopted as a “latent chromophore”, relying
upon its reactivity with p-nitrobenzylpyridine (PNBP) and piper-
idine to form a strongly colored internal salt.6
As a demonstration of the power of our strategy, the assembly
of tetrasaccharide 17 was conducted. Preparation of polymer-
bound sugar primer 12 began with straightforward synthesis of
nitro-containing linker 37 from commercially available 1 via 2,
which was subsequently bound to glucosamine derivative 4 in a
manner similar to that for conventional benzylidene formation
(Scheme 2). Subsequent manipulation via 6, 7, 8, and 9 gave 10,
which was subsequently deblocked by removal of the tBu group
to liberate the carboxylic acid, which was used for coupling with
PEG (av MW 550) to afford 11. As we hoped, the dechloro-
acetylation of 11 was easily monitored by the coloring test which
was carried out by successive treatment with PNBP and piperi-
dine. Quantification of the color density was carried out on TLC
plates which were processed using a scanner and the NIH Image8
* To whom correspondence should be addressed.
† RIKEN.
‡ Tokai University.
(1) Recent reviews: (a) Ito, Y.; Manabe, S. Curr. Opin. Chem. Biol. 1998,
2, 701-708. (b) Osborn, H. M. I.; Khan, T. H. Tetrahedron 1999, 55, 1807-
1850. (c) St. Hilaire, P. M.; Meldal, M. Angew. Chem., Int. Ed. 2000, 39,
1162-1179. (d) Hummel, G.; Hindsgaul, O. Angew. Chem., Int. Ed. 1999,
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M. J. J. Org. Chem. 1999, 64, 5926-5929. (f) Doi, T.; Sugiki, M.; Yamada,
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Nakai, Y.; Egusa, K.; Porca, J. A.; Kusumoto, S. Synlett 1999, 1074-1078.
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M.; Schmidt, R. R. Org. Lett. 2000, 2, 3043-3046. (k) Egusa, K.; Fukase,
K.; Nakai, Y.; Kusumoto, S. Synlett 2000, 27-32. (l) Tolborg, J. F.; Jensen,
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J. Tetrahedron Lett. 2000, 41, 6969-6972. On chemo-enzymatic synthesis:
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3166-3169. (s) Yan, F.; Wakarchuk, W. W.; Gilbert, M.; Richards, C.;
Whitfield, D. M. Carbohydr. Res. 2000, 328, 3-16.
(5) Reports on monitoring of reactions performed on polymer support by
spectroscopic means. MALDI-TOF MS: (a) Egner, B. J.; Langley, G. J.;
Brandle, M. J. Org. Chem. 1995, 60, 2652-2653. (b) Eager, B. J.; Brandley,
M. Tetrahedron 1997, 53, 14021-14030. (c) Thu¨rmer, R.; Meisenbach, M.;
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53, 665-672. NMR: (e) Seeberger, P. H.; Beebe, X.; Sukenick, G. D.;
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(8) Developed at the U.S. National Institutes of health and available on
the Internet at http://rbs.info.nih.gov/nih-image/.
(2) Review for organic synthesis on soluble polymer support: Granvert,
D. J.; Janda, K. D. Chem. ReV. 1997, 97, 489-509.
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1995, 117, 2116-2117. (b) Ito, Y.; Kanie, O.; Ogawa, T. Angew. Chem., Int.
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(4) Leading report on oligosaccharide synthesis utilizing lightweight PEG
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1996, 2193-2194.
10.1021/ja003856c CCC: $20.00 © 2001 American Chemical Society
Published on Web 03/31/2001

Communications to the Editor
J. Am. Chem. Soc., Vol. 123, No. 16, 2001 3849
Scheme 2. Synthesis of PEG-Tagged Sugara
Figure 2. MALDI-TOF MS spectra showing the course of coupling
reaction of 12 and 13. (a) 0 min; (b) 1 min; (c) 3 min; (d) 20 min.
Monosaccharide (12 + Na)+ and (12 + K)+ are appearing as a
mountaneous shape on left side (m/z ) 1104 to 1648) and disaccharide
(14 + Na)+ and (14 + K)+ are appearing on right side (m/z ) 1652 to
2064).
a Reagents and yields: 1) allyl bromide, K2CO3/CH3CN, 60 °C, quant.;
2) CH(OCH3)3, CSA/Drierite, CH3CN, 40 °C; 3) 4, CSA/Drierite, CH3CN,
55 °C, 95%; 4) BnBr, NaH/DMF, 93%; 5) 4.0 M HCl in dioxane,
NaBH3CN/THF, quant.; 6) Pd(PPh3)4, Et3SiH, AcOH/toluene, 92%; 7)
Scheme 4. Tetrasaccharide Assembly on Polymer Support and
Its Releasea
t
BrCH2CO2 Bu, K2CO3/CH3CN, 60 °C, 92%; 8) (ClCH2CO)2O, Pyr/
CH2Cl2, 0 °C, 97%; 9) TFA/CH2Cl2; 10) PEG monomethyl ether, DCC,
DMAP/CH2Cl2, 89% (over 2 steps); 11) DMAP, 50% aq Pyr (pH 8-9),
95%.
Figure 1. Monitoring of dechloroacetylation of 11 by coloring with
PNBP and piperidine. (a) TLC profile (lane 1: 0 h, lane 2: 0.5 h, lane
3: 3.0 h, lane 4: 3.5 h, lane 5: 6 h), (b) plot profile and density values
analyzed by NIH Image program (numbers are corresponding to theirs
a
in (a)). Density percentage to that of lane 1.
Scheme 3. Coupling Reaction on Polymer Support
by color and MALDI-TOF MS, to afford tetrasaccharide 16
(Scheme 4). After the CAc group was removed, liberation of the
tetrasaccharide from the polymer support was performed under
previously reported conditions for reductive cyclorelease by the
action of Sn(SPh)2, PhSH, and Et3N in benzene.7 The resulting
hydroxamic acid 17a was treated with CSA in MeOH at 45 °C7
to give 17b (74% from 16; overall yield from 14, 59%).
In summary, we have developed a new method for monitoring
polymer-supported glycan chain assembly. The process is rapid,
efficient, and high-yielding, and in addition, the course of each
reaction can be monitored in real-time and to very sensitive
degree. Research to further refine this technology is ongoing as
part of work on developing a general method for automated
oligosaccharide synthesis.
program. As shown in Figure 1, dechloroacetylation was ac-
companied by almost complete disappearance of color. The
resultant 12 was determined to be quantitatively deprotected by
1H NMR, which revealed the complete disappearance of the low-
field H-4 signal that originally appeared at δ 5.25.
The coupling reaction was performed using fluoride 13 under
Suzuki conditions9 and was monitored by MALDI-TOF MS
(Scheme 3, Figure 2). Complete consumption of 12 as well as
near quantitative formation of the disaccharide was observed as
a shift of the broad peak. The PEG-bound fractions were separated
using a silica gel column, and formation of disaccharide 14 was
Acknowledgment. This work was partly supported by New Energy
and Industrial Technology Development Organization (NEDO). We thank
Ms. A. Takahashi for technical assistance, Dr. T. Chihara and his staff
for elemental analysis, and Dr. R. Walton for valuable comments.
1
confirmed by H NMR. The coupling yield was estimated to be
greater than 98% judging from the relative intensity of the H-1
signals of the disaccharide (δ 5.41 and 5.26) and the unreacted
monosaccharide (δ 5.63). This nearly quantitative coupling was
reproducible on gram scales.
Thus obtained disaccharide 14 was subjected to two dechlo-
roacetylation (89-96%)/coupling (96-97%) cycles, monitored
Supporting Information Available: Experimental details, 1H NMR
1
spectra of compounds 2, 3, 5, 6, 7, 8, 9, 10, 11, and 17b, H- and 13C
NMR (DEPT) of 12, 14, 15, and 16, and MALDI-TOF MS profiles of
coupling reactions (PDF). This material is available free of charge via
the Internet at http://pubs.acs.org.
(9) Suzuki, K.; Maeta, H.; Matsumoto, T. Tetrahedron Lett. 1989, 30,
4853-4856.
JA003856C