Carbohydrate-based VEGF inhibitors

FULL PAPER

DOI: 10.1002/ejoc.200700698

Carbohydrate-Based VEGF Inhibitors

Tobias Haag,^[a]Richard A. Hughes,^[b]Gerd Ritter,^[c]and Richard R. Schmidt*^[a]

Keywords: Carbohydrates / Amino acids / Mimetics / Peptides, cyclic / Angiogenesis, inhibition

Cyclic peptide–carbohydrates (compounds 1a–c, 2, 33, 34)

were designed and synthesized to act as mimetics of loop 2

of the proangiogenic molecule vascular endothelial growth

factor D (VEGF-D). The mimetics were designed to inhibit

dimerization of the receptors (VEGFR-2 and VEGFR-3) by

VEGF-D, and thus have the potential to inhibit angiogenesis.

To this end, in the previously described cyclic octapeptide

CNEESLIC and the cyclic nonapeptide CGNEESLIC inhibi-

tors derived from VEGF-D loop 2, the NEES tetrapeptide res-

idue was replaced by a carbohydrate scaffold having the

amino acid side chain mimics in positions proposed by mod-

eling studies. Attachment of the additional amino acids using

the Fmoc technology, then formation of the cyclic disulfides,

and finally total deprotection afforded the target molecules

of which 2 and 34 showed an ability to inhibit the biological

activity of VEGF-D through VEGFR-2 in cell-based assays,

albeit at high mimetic concentration.

Germany, 2007)

mers in the dimer provides two poles for binding to the

VEGF receptors (VEGFRs) of which the most important

are VEGFR-1, -2 and -3.^[8,11]They are transmembrane pro-

teins and, when dimerized by VEGF dimers, they undergo

a conformational change leading to autophosphorylation

of tyrosine side chains of the tyrosine kinase domain in the

intracellular part. This way, a signalling cascade leading to

angiogenesis is induced.

In the pathologic angiogenesis, VEGFR-1 and VEGFR-

2 seem to play a particularly important role,^[12,13]therefore

the search for therapeutic pro- and particularly anti-

angiogenic compounds has concentrated on these recep-

tors.^[8]Success in in vivo and in vitro studies strongly pro-

moted this endeavour,^{[2,8,14–16]}and various clinical studies

have been performed.^[17]Because of the complexity of angi-

ogenesis various possibilities exist to fight tumour

growth.^[18]We concentrated on the inhibition of the VEGF

homodimer binding to VEGFR.

It was previously found that monocyclic peptides pos-

sessing the sequence and the conformation, for instance of

loops 1–3 of VEGF-D by constructing the loop sequence

between two cysteine residues and cyclising this construct

by a disulfide bridge, leads to excellent inhibitors of VEGF-

D binding to VEGFR-2.^[9]The therapeutic importance of

such compounds is generally compromized by their lability

towards proteases.^[19]Therefore, mimicking peptides by dif-

ferent scaffolds has become an important goal. In this con-

text, carbohydrate scaffolds have been shown to be use-

ful:^[20–25]they are rigid, stereochemically defined, polyfunc-

tional, and generally the starting materials are readily avail-

able. This way, peptide mimetic molecules can be designed

with the required functionalities in the required positions

in space. In this paper carbohydrate-based mimetics of

Introduction

As far back as in 1971 it was proposed that angiogenesis

plays an important role in tumour growth.^[1]Since then,

this proposal has been confirmed and widely accepted^[2,3]

and the mechanism of angiogenesis has been extensively

studied.^[4–6]Various factors stimulating or inhibiting angi-

ogenesis have been found.^[6,7]Important stimulators of an-

giogenesis and tumour angiogenesis are the vascular endo-

thelial growth factors (VEGFs), a family consisting of

VEGF-A, -B, -C, -D, and -E.^[8]Their three-dimensional

structure has been determined by X-ray analysis^[10]A three-

dimensional model of the VEGF-D dimer was deduced

from the VEGF-A X-ray structure^[10]by applying protein-

homology modeling techniques.^[9]The VEGF monomer

consists essentially of a central four-stranded antiparallel β-

sheet, three additional β-sheet segments, and two short α-

helices. The stabilization of the tertiary structure of the

monomer is based on a cystine knot consisting of three in-

tramolecular disulfide bridges. There are three loops con-

necting the central β-sheets, which are located at the tips of

the monomers. The amino acids of these loops are responsi-

ble for the interaction with the receptors.^[8,10]The monomer

forms as quaternary structure a side-by-side homodimer, in

which the monomers are covalently linked through two di-

sulfide bridges. The head-to-tail orientation of the mono-

[a] Fachbereich Chemie, Universität Konstanz, Fach M 725,

78457 Konstanz, Germany

Fax: +49-7531-883135

E-mail: Richard.Schmidt@uni-konstanz.de

[b] Department of Pharmacology, University of Melbourne,

Victoria 3010 Australia

E-mail: rahughes@unimelb.edu.au

[c] Ludwig Institute for Cancer Research,

New York, NY 10158, USA

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Carbohydrate-Based VEGF Inhibitors

Figure 1. Structure of target molecules 1a–c and 2.

VEGF-D loop 2 mimetic CNEESLIC (A in Figure 1) are pound 4, which on O-benzylation with benzyl bromide and

presented, which were designed based on molecular model- sodium hydride in DMF furnished the fully protected glu-

ing studies.^[9]

cose derivative 5, which had at 3-O and 4-O the desired

Results and Discussion

Synthesis of Target Molecules 1a–1c and 2

The modeling studies exhibited that the NEES tetrapep-

tide moiety of cyclic octapeptide A can be relatively well

accommodated by an appropriately functionalized Glcβ(1–

4)GlcNH₂disaccharide residue. Hence, the mimetic 1a was

designed as target molecule (Figure 1). These studies also

permitted replacement of the leucine residue by isoleucine

or valine, therefore also the mimetics 1b and 1c, respec-

tively, were target molecules. Additionally, ring expansion

by one glycine residue between the first cysteine and the

asparagine residues gave a good fit with A, therefore also

compound 2 was considered as target molecule.

For the synthesis of the Glcβ(1–4)GlcNH₂disaccharide

moiety, the b residue (see Figure 1) requires temporary pro-

tection at 1-O, 2-O and 6-O. To this end, glucose was trans-

formed, according to a known procedure,^[26]into 1,2-O-

ethylidene derivative 3 (Scheme 1). Regioselective 6-O-tri-

Scheme 1. Synthesis of glycosyl donor 8. Reagents and conditions:

(a) Trt-Cl, Pyr (62%); (b) BnBr, NaH, DMF (78%); (c) HOAc,

H₂O; Ac₂O, Pyr (90%); (d) N₂H₄·HOAc (90%); (e) CCl₃CN, DBU,

tylation with trityl chloride in pyridine^[27]afforded com- CH₂Cl₂(99%).

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T. Haag, R. A. Hughes, G. Ritter, R. R. Schmidt

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“permanent” protection. Acid-catalyzed cleavage of the tri-

tyl and the 1,2-O-ethylidene groups and then O-acetylation

afforded compound 6. Treatment with hydrazinium acetate

led to regioselective 1-O-deacetylation (Ǟ 7); reaction with

trichloroacetonitrile in the presence of 1,8-diazabicy-

clo[5.4.0]undec-7-ene (DBU)^[28]afforded the O-glucosyl

trichloroacetimidate 8 in good overall yield.

For the synthesis of the a residue (see Figure 1) requiring

selective access to 1-O, 2-N, 3-O and 4-O, -glucosamine

was transformed into N-dimethylmaleoyl (N-DMM) pro-

tected thexyldimethylsilyl (TDS) 2-amino-4,6-O-benzyl-

idene-2-deoxy-glucopyranoside 9 according to a known

procedure (Scheme 2).^[29]3-O-Acetylation with acetic anhy-

dride in pyridine afforded compound 10 which on treatment

with sodium cyanoborohydride in the presence of hydro-

chloric acid in diethyl ether^[30]furnished the 6-O-benzyl-

protected derivative 11 which had the required protecting-

group pattern for selectively accessing the functional groups

in positions 1–4. Glycosylation of 11 with glycosyl donor 8

and trimethylsilyl trifluoromethanesulfonate (TMSOTf) as

catalyst (0.1 equiv.) in dichloromethane at –5 °C afforded

the desired β-linked disaccharide 12 (J_1b,2b= 8.0 Hz) in

high yield (Scheme 3). Treatment of 12 with tetrabutyl-

ammonium fluoride (TBAF) in THF led to the 1-O-desil-

ylated compound 13 which gave with trichloroacetonitrile

in the presence of DBU the trichloroacetimidate 14 as gly-

cosyl donor.

Scheme 3. Synthesis of disaccharide donor 14. Reagents and condi-

tions: (a) TMSOTf, –5 °C, CH₂Cl₂(95%); (b) TBAF, HOAc, THF

(95%); (c) CCl₃CN, DBU, CH₂Cl₂(92%).

Scheme 4. Synthesis of acceptor 17. Reagents and conditions: (a)

(1) NH₃, MeOH; Ac₂O, Pyr; (2) Trt-OH, H₂SO₄, Ac₂O (42%); (b)

NaOMe, MeOH (93%).

replaced by a Cbz group because the DMM group is not

stable under the following oxidation conditions. To this end,

19 was first treated with sodium hydroxide, and thereafter

the pH was adjusted to 4.5 with hydrochloric acid, this way

liberating the amino group;^[29,32]following treatment with

benzyloxycarbonyl (Cbz) chloride in the presence of potas-

sium carbonate furnished the Cbz-protected disaccharide

20. Chemoselective oxidation of the primary hydroxymethyl

Scheme 2. Synthesis of acceptor 11. Reagents and conditions: (a)

Ac₂O, Pyr (98%); (b) NaCHBH₃, HCl, Et₂O (93%).

For the attachment of the asparagine side-chain mimic,

ethyl glycolate (Scheme 4, 15)^[31]was treated with ammonia

in methanol; O-acetylation with acetic anhydride in pyr-

idine and following N-tritylation with triphenylcarbinol in

acetic anhydride/sulfuric acid furnished the intermediate 16.

O-Deacetylation with sodium methoxide in methanol gave

the desired acceptor 17.

Glycosylation of 17 with the disaccharide donor 14 and

TMSOTf as catalyst (0.08 equiv.) in dichloromethane at

–15 °C furnished the β-linked glycoside 18 (J_1a,2a= 8.5 Hz)

in high yield (Scheme 5). For the regioselective introduction

of the other amino acid side-chain mimics the O-acetyl

groups of 18 were removed by treatment with sodium meth-

Scheme 5. Synthesis of disaccharide 23. Reagents and conditions:

(a) TMSOTf, CH₂Cl₂(65 %); (b) NaOMe, MeOH (95 %); (c)

NaOH; HCl; Cbz-Cl, K₂CO₃(73%); (d) TEMPO, NaOCl; BnBr,

CsF (93%); (e) BrCH₂CO₂tBu, Ag₂O (82%); (f) Pd/C, H₂; Fmoc-

oxide in methanol (Ǟ 19) and then the DMM group was ONSu (68%).

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Carbohydrate-Based VEGF Inhibitors

group with tetramethylpiperidine N-oxide (TEMPO) and

NaOCl^[33–36]and then benzylation with benzyl bromide and

cesium fluoride^[37]transformed the b residue into the de-

sired glucuronic acid leading to compound 21. The gluta-

mate side-chain mimics were introduced with tert-butyl bro-

moacetate in the presence of silver oxide to afford com-

pound 22. Hydrogenolytic cleavage of the benzyl groups

with Pd/C as catalyst led to an amino acid intermediate

which on reaction with fluorenylmethoxycarbonyloxy–suc-

cinimide (Fmoc-ONSu)^[38]led to N-Fmoc protection fur-

nishing NEES tetrapeptide mimic 23.

The required leucinyl–isoleucinyl–cysteine tripeptide 27a

(Scheme 6) was obtained according to standard Fmoc stra-

tegies. The commercially available cysteine building block

24 was treated with piperidine and then with Fmoc-pro-

tected isoleucine in the presence of (benzotriazol-1-yloxy)tri-

pyrrolidinophosophonium hexafluorophosphate (PyBOP)^[39]

and Hünig’s base (N-ethyldiisopropylamine, DIPEA) to af-

ford the dipeptide 25. Similarly, with Fmoc-protected leu-

cine the tripeptide 26a was obtained. Removal of the Fmoc

group with piperidine furnished the derivative 27a which

was coupled to 23 under the same conditions to afford the

protected heptapeptide mimic 28a (Scheme 7). Attachment

of the cysteine residue 29 to the 2a-amino group of this

construct required again cleavage of the Fmoc group with

piperidine and then PyBOP/DIPEA-supported condensa-

tion to furnish the ring-open octapeptide mimic 30a in 64%

yield. Treatment of 30a with iodine in methanol led to loss

of the S-trityl groups and to disulfide bond formation.^[40]

Following cleavage of the acid-labile protecting groups with

trifluoroacetic acid (TFA) and addition of triethylsilane

(TES) to scavenge carbenium ion intermediates^[41]led to the

crude target molecule 1a. Purification by RP-18 flash

chromatography led to the separation of salts and most of

the byproducts. Final purification of 1a was performed by

RP-18 HPLC with acetonitrile/water/TFA as eluent to give

the target molecule in 80% yield. Similarly, from 25 the ad-

ditional leucine- or valine-containing tripeptides 26b and

26c, respectively, were obtained (Scheme 8); removal of the

Fmoc protecting group with piperidine afforded 27b and

27c. Condensation with 23 and with 29 as described above

and following ring closure under disulfide bond formation,

protecting group cleavage and then purification afforded

the target molecules 1b and 1c.

Scheme 7. Synthesis of target molecule 1a.

The target molecule 2 was obtained by the same pro-

cedure (Scheme 9). Reaction of 29 with glycine methyl ester

and PyBOP/DIPEA as condensing agent afforded dipeptide

Scheme 8. Synthesis of target molecules 1b and 1c.

Scheme 6. Synthesis of LIC tripeptide 27a.

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ester 31 in practically quantitative yield. Ester hydrolysis clic octapeptide mimic 33 and the cyclic nonapeptide mimic

with sodium hydroxide in aqueous methanol led to the de- 34 became the new target molecules (Figure 2).

sired dipeptide 32. Attachment of 32 to 28a required first

For the synthesis of allo-configured fragment a, com-

cleavage of the Fmoc residue from 28a with piperidine and pound 9 was transformed into the 3-O-levulinoyl (Lev) de-

then condensation with PyBOP/DIPEA. Treatment of this rivative 35 by treatment with levulinic acid and dicyclohexyl

intermediate with iodine in methanol led as described above carbodimide (DCC) and Steglich’s reagent (DMAP)^[43]as

to ring closure under disulfide bond formation; acid-cata- condensing agents (Scheme 10). Reductive opening of the

lyzed deprotection and then purification gave the target 4,6-O-benzylidene group with triethylsilane (TES) in the

molecule 2 in very good yield.

presence of trifluoroacetic anhydride and TFA at 0 °C^[44]

furnished the 4-O-unprotected 6-O-benzyl-protected glu-

cose derivative 36. Glycosylation with 8 as glycosyl donor

and TMSOTf as catalyst (0.08 equiv.) gave the β-linked di-

saccharide 37 (J_1b,2b= 8.0 Hz) in 68% yield. The levulinoyl

group was selectively cleaved by treatment with hydrazin-

ium acetate in pyridine^[45]to furnish 3a-O-unprotected de-

rivative 38. The inversion of the 3a-hydroxy group was per-

formed by oxidation with Dess–Martin periodinane to the

ketone, which upon reduction with sodium borohyd-

ride^[46,47]gave mainly the allo-configured compound 39. Re-

action with acetic anhydride in pyridine afforded com-

pound 40 which was selectively deprotected at 1-O by treat-

ment with TBAF in THF to afford compound 41. Reaction

of 41 with trichloroacetonitrile in the presence of DBU as

base led to trichloroacetimidate 42 as disaccharide donor.

Glycosylation of acceptor 17 with 42 as donor and tin(II)

triflate as catalyst^[48](0.005 equiv.) in dichloromethane at

room temperature gave the desired β-linked glycoside 43

(J_1a,2a= 8.7 Hz) in 64% yield. O-Deacetylation with so-

dium methoxide in methanol (Ǟ 44), cleavage of the DMM

group by treatment with sodium hydroxide, adjustment of

the pH to 4.5 with hydrochloric acid and finally introduc-

tion of the Cbz group to the liberated amino group with

Cbz-Cl in the presence of potassium carbonate led to the

desired 2a-O-, 2b-O- and 6b-O-unprotected intermediate

Scheme 9. Synthesis of target molecule 2.

In a cell-based assay^[9]compound 2 was found in prelimi-

nary studies to exhibit some inhibition (ca. 20%) of VEGF-

D-mediated survival activity through VEGFR-2, albeit at

high concentration of compound 2 (10^–4).^[42]The cyclic

octapeptide mimics 1a–c had only a marginal effect in this

assay.

Synthesis of Target Molecules 33 and 34

45.

The low inhibition of compounds 1a–c and 2 was the

Oxidation of the primary hydroxymethyl group of 45

reason to consider a configurational change for the attach- with TEMPO/NaOCl to a carboxy group and then treat-

ment of one of the glutamate side-chain mimics because an ment with benzyl bromide in the presence of cesium fluo-

even better fit with A (Figure 1) was proposed by the mod- ride as base in DMF afforded the disaccharide 46 contain-

eling studies for this structural modification. Hence, the cy- ing the desired benzyl ester at the glucuronic acid moiety

Figure 2. Structure of target molecules 33 and 34.

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Carbohydrate-Based VEGF Inhibitors

and 51 were deprotected and ring-closed to compounds 33

and 34, respectively, as described for 1a, thus providing

these target molecules in high yields.

Scheme 10. Synthesis of disaccharide 45. Reagents and conditions:

(a) Lev-OH, DCC, DMAP (89 %); (b) TES, TFA (78 %); (c)

TMSOTf, CH₂Cl₂(68 %); (d) N₂H₄·HOAc, Pyr (98 %); (e)

periodinane; NaBH₄(68%); (f) Ac₂O, Pyr (97%); (g) TBAF, THF

(93%); (h) CCl₃CN, DBU, CH₂Cl₂(98%); (i) 17, Sn(OTf)₂, CH₂Cl₂

(64%); (j) NaOMe, MeOH (98%); (k) NaOH; HCl; Cbz-Cl, K₂CO₃

(80%).

(Scheme 11). The glutamate side-chain mimics were again

introduced with tert-butyl bromoacetate and silver oxide to

afford compound 47. Palladium/carbon-catalyzed hydro-

genolysis of all O-benzyl groups led to an amino acid inter-

mediate which on reaction with Fmoc-ONSu led to N-

Fmoc protection and furnished the new NEES tetrapeptide

mimetic 48. Coupling of LIC-tripeptide 27a to 48 with

PyBOP/DIPEA as condensing agent afforded heptapeptide

mimetic 49 in 64% yield. Attachment of the cysteine residue

29 required first cleavage of the Fmoc group of 49 with

piperidine and then PyBOP/DIPEA-supported condensa-

Scheme 11. Scheme 7. Synthesis of target molecules 33 and 34.

Reagents and conditions: a) TEMPO, NaOCl; BnBr, CsF (78%);

b) Br-CH₂–CO₂tBu, Ag₂O (70 %); c) Pd/C, H₂; Fmoc-ONSu

(50%); d) PyBOP, DIPEA (64%); e) Piperidine; PyBOP, DIPEA

(50: 91%, 51: 91%); f) I₂, MeOH; TES, TFA, H₂O; RP-18 column;

RP-18 HPLC (33: 78%, 34: 75%).

Conclusion

In preliminary cell-based assays for VEGF-D-mediated

tion to give the octapeptide mimetic 50 in 91% yield. Simi- survival activity through VEGFR-2, compound 34 showed

larly, from 49 and CG-dipeptide 32 the nonapeptide mi- better inhibition values (inhibition ca. 35% at 10^–4 of 34)

metic 51 was obtained in the same yield. Compounds 50 than for compound 2,^[42]although it should be noted that

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T. Haag, R. A. Hughes, G. Ritter, R. R. Schmidt

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0.85 (h), 0.75 (t) (toluene/ethyl acetate, 4:1). [α]²_D⁰= –46.2 (c = 0.5,

CHCl₃) (h), [α]²_D⁰= –30.2 (c = 0.5, CHCl₃) (t).

5h: ¹H NMR (250 MHz, CDCl₃): δ = 1.37 (d, ³J = 4.9 Hz, 1 H,

neither 2 nor 34 were as effective as the parent cyclic octa-

peptide A as previously reported.^[9]Therefore, further struc-

tural optimization of these VEGF-D loop mimetics is in

progress. Detailed studies on the in vivo stability, potential

unspecific interactions, and toxicities of 1a–c, 2, 33 and 34

will be reported in due course.

2

3

CH₃), 3.22 (dd, J_gem= 10.1, J_vic= 2.9 Hz, 1 H, 6-H), 3.50 (dd,

²J_gem= 10.0, J_vic= 1.4 Hz, 1 H, 6Ј-H), 3.78 (m, 3 H, 3-H, 4-H, 5-

3

H), 4.31 (d, J = 10.7 Hz, 1 H, CHHPh), 4.36 (dd, J_1,2= J_2,3

=

5 Hz, 1 H, 2-H), 4.64 (m, 2 H, CH₂Ph), 4.82 (d, ³J = 11.5 Hz, 1 H,

CHHPh), 4.52 (q, ³J = 4.9 Hz, 1 H, CH₃), 5.74 (d, ³J_1,2= 4.9 Hz, 1

H, 1-H), 7.13–7.45 (m, 25 H, phenyl) ppm.

Experimental Section

5t: ¹H NMR (250 MHz, CDCl₃): δ = 1.44 (d, ³J = 4.9 Hz, 1 H,

2

3

CH₃), 3.18 (dd, J_gem= 10.1, J_vic= 3.5 Hz, 1 H, 6-H), 3.50 (dd,

General: Solvents were purified by standard procedures. NMR

spectra were recorded at 22 °C with a Bruker AC 250 Cryospec or

a Bruker DRX 600 spectrometer. Tetramethylsilane (TMS) or the

resonance of residual undeuterated solvent was used as internal

standard: CDCl₃(δ = 7.24 ppm), D₂O (δ = 4.63 ppm), [D₆]DMSO

(δ = 2.49 ppm). MALDI mass spectra were recorded with a Kratos

Kompact Maldi 2 spectrometer, and 2,5-dihydroxybenzoic acid

(DHB) or 6-aza-2-thiothymine (ATT) was used as matrix. FAB

mass spectra were obtained with a Finnigan MAT 312/AMD 5000

instrument; +6 kV for positive ions, –4 kV for negative ions. Thin-

layer chromatography was performed on Merck 60 F₂₅₄silica gel

plastic plates or Merck RP-18 F₂₅₄glass plates; compounds were

visualized by treatment with a solution of [(NH₄)₆Mo₇O₂₄·4H₂O]

(20 g) and Ce(SO₄)₂(0.4 g) in 10% sulfuric acid (400 mL) and then

heating to 120 °C. Flash chromatography was performed on J. T.

Baker silica gel 60 (40–63 µm) at a pressure of 0.3 bar. Preparative

RP-18 HPLC was carried out with a Eurospher 100 C18 column

(Fa. Knauer) with a Shimadzu LC-8A pump and a Rainin Dyna-

max UV-1 detector at a flow rate of 10 mL/min. Optical rotations

were measured at 20 °C with a Polar-Monitor, Fa. Büchi, at the

sodium D line.

²J_gem= 10.0, J_vic= 1.7 Hz, 1 H, 6Ј-H), 3.85–3.92 (m, 3 H, 3-H, 4-

H, 5-H), 4.14 (dd, 1 H, 2-H), 4.23 (d, J = 11.2 Hz, 1 H, CHHPh),

3

4.43 (d, J = 10.8 Hz, 1 H, CHHPh), 4.62 (2d, 2 H, CH₂Ph), 5.11

(q, ³J = 4.9 Hz, 1 H, CH₃), 5.68 (d, ³J_1,2= 4.9 Hz, 1 H, 1-H), 7.12–

7.46 (m, 25 H, phenyl) ppm. C₄₁H₄₀O₆(628.7): calcd. C 78.32, H

6.41; found C 78.27, H 6.81.

Acetyl 2,6-Di-O-acetyl-3,4-di-O-benzyl-α,β-D-glucopyranoside (6):

Compound 5 (7.4 g, 11.76 mmol) was dissolved in acetic acid

(200 mL), H₂O (40 mL) and the mixture stirred in an oil bath at

125 °C for 8 h; the reaction mixture was concentrated in vacuo and

coevaporated four times with toluene. The residue was diluted in

pyridine (200 mL), and acetic acid anhydride (100 mL) was slowly

added while cooling in an ice bath. After 16 h, the reaction mixture

was concentrated in vacuo and coevaporated twice with toluene.

Purification of the residue by flash chromatography (silica gel, tolu-

ene/ethyl acetate, 7:1 Ǟ 5:1) furnished the anomeric mixture of

compound 6 (5.2 g, 10.6 mmol, 90%) as colorless oil. TLC: R_f=

0.65 (toluene/ethyl acetate, 3:1). [α]²_D⁰= +47.2 (c = 1, CHCl₃). H

1

NMR (250 MHz, CDCl₃): δ = 1.92, 1.94, 1.95, 2.01, 2.02, 2.06 (6

s, 9 H, acetyl), 3.62–3.73 (m, 2 H, 3-H, 4-H), 3.98–4.02 (m, 1 H,

5-H), 4.21–4.32 (m, 1 H, 2-H), 4.52–4.60 (m, 2 H, CH₂Ph), 4.69

(d, 1 H, CH₂Ph), 4.78–4.89 (m, 3 H, 6-H, CH₂Ph), 5.02–5.10 (m,

1,2-O-Ethylidene-6-O-(triphenylmethyl)-α-D-glucopyranose

(4):

Compound 3^[26](6.0 g, 29.1 mmol) was dissolved in dry pyridine

(50 mL), and TrtCl (6.74 g, 26.2 mmol) was added. After stirring

overnight, the reaction mixture was diluted with ethyl acetate and

washed three times with saturated NaCl solution. The organic

phase was dried with MgSO₄, filtered, purified and coevaporated

twice with toluene. Purification of the residue by flash chromatog-

3

1 H, 6Ј-H), 5.60 (d, J_1,2= 8.2 Hz, 0.5 H, 1-H), 6.22 (d, J_1,2

=

3.6 Hz, 0.5 H, 1-H), 7.21–7.35 (m, 10 H, phenyl) ppm. C₂₆H₃₀O₉

(486.5): calcd. C 64.19, H 6.22; found C 64.28, H 6.36.

2,6-Di-O-acetyl-3,4-di-O-benzyl-α,β-D-glucopyranose (7): Com-

pound 6 (5.0 g, 10.3 mmol) was dissolved in dry DMF (25 mL),

and N₂H₄·HOAc (1.13 g, 1.2 equiv.) was added at room tempera-

ture. After 1 h, the reaction mixture was diluted with ethyl acetate

and washed three times with cold saturated NaCl solution. The

organic phase was dried with MgSO₄and concentrated in vacuo.

The residue was purified by column chromatography (silica gel,

toluene/ethyl acetate, 5:1) which furnished the anomeric mixture of

compound 7 (4.1 g, 9.2 mmol, 90%) as colorless oil. TLC: R_f=

raphy (silica gel, toluene/acetone, 3:1) furnished

4 (8.0 g,

17.8 mmol, 62%) as slightly yellow foam. TLC: R_f= 0.48 (toluene/

acetone, 2:1). [α]²_D⁰= +9.6 (c = 1, CHCl₃). ¹H NMR (250 MHz,

CDCl₃): δ = 1.31 (d, ³J = 4.9 Hz, 1.5 H, CH₃), 1.41 (d, ³J = 4.9 Hz,

1.5 H, CH₃), 2.5 (br. s, 2 H, OH), 3.26–3.41 (m, 2 H, 6-H), 3.61–

3

3.89 (m, 3 H, 3-H, 4-H, 5-H), 4.02 (dd, J_1,2= J_2,3= 4.9 Hz, 0.5

3

H, 2-H), 4.10 (dd, J_1,2= 4.9, J_2,3= 6.4 Hz, 0.5 H, 2-H), 5.06 [q,

³J = 4.9 Hz, 0.5 H, CH(CH₃)₂], 5.40 [q, ³J = 4.9 Hz, 0.5 H,

CH(CH₃)₂], 5.54 (d, J_1,2= 4.6 Hz, 0.5 H, 1-H), 5.55 (d, J_1,2

0.25 (toluene/ethyl acetate, 3:1). [α]²_D⁰= +70.0 (c = 1, CHCl₃). H

1

3

=

NMR (250 MHz, CDCl₃): δ = 1.99, 2.01 (2 s, 6 H, acetyl), 3.14

(br. s, 1 H, OH), 3.54–3.62 (m, 2 H, 3-H, 4-H), 4.20–4.35 (m, 3 H,

5-H, CH₂Ph), 4.61 (d, 1 H, CH₂Ph), 4.72–4.82 (m, 4 H, 2-H, 6-H,

CH₂Ph), 5.41 (br. s, 1 H, 1-H), 7.21–7.39 (m, 10 H, phenyl) ppm.

C₂₄H₂₈O₈(444.5): calcd. C 64.85, H 6.35; found C 64.67, H 6.70.

5.1 Hz, 0.5 H, 1-H), 7.11–7.39 (m, 15 H, phenyl) ppm. MALDI-

MS (pos. mode, matrix DHB, THF): m/z = 471 [M + Na]⁺, 487

[M + K]⁺.

3,4-Di-O-benzyl-1,2-O-ethylidene-6-O-(triphenylmethyl)-α-D-gluco-

pyranose (5): Compound 4 (6.5 g, 14.5 mmol) was dissolved in dry

DMF (120 mL), and NaH (1.4 g, 58.3 mmol) was added at 0 °C

under cooling in an ice bath. After 10 min, benzyl bromide (10.3 g,

60 mmol) was added, and the solution was then stirred at room

temperature for 4 h. The reaction mixture was then diluted with

MeOH (10 mL), concentrated under reduced pressure, dissolved in

ethyl acetate and washed twice with saturated NaCl solution. The

organic phase was dried with MgSO₄and concentrated. Purifica-

tion by flash chromatography (silica gel, petroleum ether/ethyl ace-

tate, 10:1 Ǟ 5:1) furnished the two epimers (h: 4.45 g, 7.08 mmol;

t: 2.62 g, 4.17 mmol, 78%) as slightly yellowish foam. TLC: R_f=

O-2,6-Di-O-acetyl-3,4-di-O-benzyl-α-D-glucopyranosyl Trichlo-

roacetimidate (8): Compound 7 (4.5 g, 10.1 mmol) was dissolved in

dry CH₂Cl₂(40 mL), and trichloroacetonitrile (5.81 mL,

57.9 mmol) was added. After the addition of 5 drops of DBU, the

reaction mixture was stirred for 1 h. The dark reaction mixture was

concentrated to 3/4 of its volume, and the residue was purified by

column chromatography (silica gel, petroleum ether/ethyl acetate,

2.5:1 + 1% NEt₃). This furnished imidate 8 (5.8 g, 9.8 mmol, 97%)

which was immediately used for the next step. TLC: R_f= 0.45 (pe-

troleum ether/ethyl acetate, 1:1). ¹H NMR (250 MHz, CDCl₃): δ =

3

1.93 (s, 3 H, acetyl), 2.02 (s, 3 H, acetyl), 3.69 (dd, J_2,3≈ J_3,4

≈

6022

www.eurjoc.org

Eur. J. Org. Chem. 2007, 6016–6033

Carbohydrate-Based VEGF Inhibitors

3

9.6 Hz, 1 H, 3-H), 4.01–4.07 (m, 1 H, 5-H), 4.12 (dd, J_3,4≈ J_4,5

= J_4,5= 9.4 Hz, 1 H, 4a-H), 3.93 (dd, J_1,2= 8.1, J_2,3= 10.9 Hz,

≈ 9.2 Hz, 1 H, 4-H), 4.20–4.33 (m, 2 H, 6-H), 4.59 (d, ³J = 10.7 Hz, 2a-H), 4.21 (m, 2 H, 6b-H), 4.45 (d, J_1,2= 8.0 Hz, 1 H, 1b-H),

3

1 H, CH₂Ph), 4.75–4.89 (m, 3 H, CH₂Ph), 5.06 (dd, J_1,2= 3.6, 4.50, 4.53, 4.63, 4.72, 4.74, 4.77 (6 d, 6 H, 3ϫ CH₂Ph), 4.88 (m, 1

³J_2,3= 10.1 Hz, 1 H, 2-H), 6.45 (d, ³J_1,2= 3.4 Hz, 1 H, 1-H), 7.24–

H, 2b-H), 5.35 (d, ³J_1,2= 8.1 Hz, 1 H, 1a-H), 5.52 (dd, ³J_2,3= 10.5,

7.31 (m, 10 H, phenyl), 8.58 (s, 1 H, NH). C₂₆H₂₈Cl₃NO₈(588.9) ³J_3,4= 9.2 Hz, 1 H, 3a-H), 7.23–7.37 (m, 15 H, phenyl) ppm. ¹³C

ppm.

NMR (150.8 MHz, CDCl₃, selected data): δ = 56.9 (1 C, 2a-C),

63.0 (1 C, 6b-C), 67.7 (1 C, 6a-C), 70.9 (1 C, 3a-C), 72.8 (1 C, 5b-

C), 73.0 (1 C, 2b-C), 74.7 (1 C, 5a-C), 75.9 (1 C, 4a-C), 77.2 (1 C,

3b-C), 83.2 (1 C, 4b-C), 93.1 (1 C, 1a-C), 100.4 (1 C, 1b-C) ppm.

MALDI-MS (pos. mode, matrix DHB, THF): m/z = 1010 [M +

Na]⁺, 1026 [M + K]⁺. C₅₃H₆₉NO₁₅Si (988.2): calcd. C 64.42, H

7.04, N 1.42; found C 64.32, H 7.39, N 1.41.

Dimethyl(thexyl)silyl 3-O-Acetyl-4,6-O-benzylidene-2-deoxy-2-(di-

methylmaleimido)-β-D

-glucopyranoside (10): To a solution of 9^[29]

(25.5 g, 49.2 mmol) in pyridine (200 mL) acetic acid anhydride

(100 mL) was slowly added dropwise at 0 °C. After 3 h, the reaction

mixture was concentrated in vacuo and coevaporated four times

with toluene. The residue was purified by flash chromatography

(silica gel, petroleum ether/ethyl acetate, 8:1 Ǟ 5:1) which furnished

acetylated compound 10 (27.1 g, 48.4 mmol, 98%) as colorless

foam. TLC: R_f= 0.25 (toluene/ethyl acetate, 3:1). [α]²_D⁰= –34.8 (c

= 1, CHCl₃). H NMR (250 MHz, CDCl₃): δ = 0.01, 0.05 (2 s, 6

H, 2 ϫ SiCH₃), 0.72 (m, 12 H, 4 ϫ CH₃), 1.48 [m, 1 H, CH-

(CH₃)₂], 1.92 (s, 9 H, acetyl, DMM-H), 3.63–3.72 (m, 2 H, 4-H, 5-

H), 3.79 (dd, J_1,2

10.4, J_vic= 8.0 Hz, 1 H, 6-H), 4.30 (dd, J_gem= 10.4, J_vic

4.3 Hz, 1 H, 6Ј-H), 5.46 (d, J_1,2= 9.4 Hz, 1 H, 1-H), 5.48 (s, 1 H,

CHphenyl), 5.70 (dd, J_2,3= 9.0, J_3,4= 10.2 Hz, 1 H, 3-H), 7.31–

7.44 (m, 5 H, phenyl) ppm. C₂₉H₄₁NO₈Si (559.7): calcd. C 62.23,

H 7.38, N 2.50; found C 62.23, H 7.13, N 2.67.

(2,6-Di-O-acetyl-3,4-di-O-benzyl-β-

acetyl-6-O-benzyl-2-deoxy-2-(dimethylmaleimido)-α,β-

D

-glucopyranosyl)(1Ǟ4)-3-O-

-gluco-

D

pyranose (13): Acetic acid (0.48 mL, 1 equiv.) was added to a solu-

tion of 12 (7.8 g, 7.83 mmol) in THF (130 mL), and the mixture

was cooled to 0 °C. After addition of TBAF (1  in THF, 9.3 mL,

9.3 mmol), the reaction mixture was stirred at 0 °C for 2 h, then

diluted with diethyl ether and washed twice with saturated NaCl

solution. The aqueous phase was reextracted with diethyl ether, and

the combined organic phases were dried with MgSO₄, filtered, and

concentrated in vacuo. Purification by flash chromatography fur-

nished the anomeric mixture of compound 13 (6.45 g, 7.62 mmol,

95%) as colorless foam. TLC: R_f= 0.35 (toluene/ethyl acetate, 6:4).

[α]²_D⁰= +18.7 (c = 1, CHCl₃). ¹H NMR (600 MHz, CDCl₃): δ =

1.86–2.04 (8 s, 15 H, 3ϫ acetyl, DMM-H), 3.39 (m, 1 H, 5b-H),

3.50 (m, 1 H, 3b-H), 3.57 (m, 1 H, 4b-H), 3.64 [m, 0.5 H, 5a-H(β)],

4.68 [m, 0.5 H, 6a-H(α)], 3.74 [m, 0.5 H, 6a-H(β)], 3.79 [m, 0.5 H,

6aЈ-H(β)], 3.86 [m, 0.5 H, 6aЈ-H(α)], 3.94–3.96 [m, 1 H, 2a-H(β),

4a-H(β)], 4.00 [m, 0.5 H, 4a-H(α)], 4.17 [m, 0.5 H, 5a-H(α)], 4.24

(m, 2 H, 6b-H), 4.38 [m, 0.5 H, 2a-H(α)], 4.41 (m, 1 H, 1b-H),

4.47–4.54 (m, 2 H, CH₂Ph), 4.64 (m, 1 H, CHHPh), 4.75–4.80 (m,

3 H, CH₂Ph, CHHPh), 4.88 (m, 1 H, 2b-H), 5.30 [d, ³J_1,2= 3.5 Hz,

1

3

2

=

³J_3,4= 9.5 Hz, 1 H, 2-H), 3.98 (dd, J_gem

=

3

2

3

Dimethyl(thexyl)silyl 3-O-Acetyl-6-O-benzyl-2-deoxy-2-(dimethyl-

maleimido)-β-D-glucopyranoside (11): Compound 10 (6.0 g,

10.7 mmol) was dissolved in dry THF (100 mL), cooled down to

0 °C, NaCNBH₃(6.72 g, 100 mmol) was added, and then a satu-

rated solution of HCl in diethyl ether (20 mL) was slowly added

in the presence of freshly heated molecular sieves (4 Å). After gas

development had ceased, the reaction mixture was immediately

neutralized with solid NaHCO₃, diluted with diethyl ether and

washed three times with saturated NaHCO₃solution. The organic

phase was dried with MgSO₄, filtered, concentrated and the residue

purified by column chromatography (silica gel, toluene/ethyl ace-

tate, 6:1) to furnish compound 11 (5.6 g, 10.0 mmol, 93 %) as

amorphous solid. TLC: R_f= 0.25 (toluene/ethyl acetate, 3:1). [α]²_D⁰

= –16.6 (c = 1, CHCl₃). ¹H NMR (250 MHz, CDCl₃): δ = 0.01,

0.10 (2 s, 6 H, 2ϫ SiCH₃), 0.72–0.75 (m, 12 H, 4ϫ CH₃), 1.47

3

0.5 H, 1a-H(α)], 5.44 [d, J_1,2= 8.5 Hz, 0.5 H, 1a-H(β)], 5.56 [dd,

3

³J_2,3= 9.1, J_3,4= 10.6 Hz, 0.5 H, 3a-H(β)], 5.71 [dd, J_2,3= 8.9,

³J_3,4= 11.3 Hz, 0.5 H, 3a-H(α)], 7.26–7.40 (m, 15 H, phenyl) ppm.

¹³C NMR (150.8 MHz, CDCl₃, selected data): δ = 54.7 [0.5 C, 2a-

C(α)], 56.3 [0.5 C, 2a-C(β)], 62.8 (1 C, 6b-C), 67.1 [0.5 C, 6a-C(α)],

67.4 [0.5 C, 6a-C(β)], 68.5 [0.5 C, 3a-C(α)], 69.4 [0.5 C, 5a-C(α)],

70.7 [0.5 C, 3a-C(β)], 72.8 (1 C, 5b-C), 72.9 (1 C, 2b-C), 75.4 [0.5

C, 4a-C(β)], 75.5 [0.5 C, 4a-C(α)], 77.1 (1 C, 4b-C), 83.2 (1 C, 3b-

C), 92.6 (1 C, 1a-C), 100.4 (1 C, 1b-C) ppm. C₄₅H₅₁NO₁₅(845.9):

calcd. C 63.90, H 6.08, N 1.66; found C 63.55, H 6.24, N 2.38.

3

[sept, J = 6.8 Hz, 1 H, CH(CH₃)₂], 1.91 (s, 6 H, DMM-H), 1.97

(s, 3 H, acetyl), 2.96 (br. s, 1 H, OH), 3.61–3.78 (m, 4 H, 2-H, 4-

2

3

H, 5-H, 6-H), 3.93 (dd, J_gem= 10.8, J_vic= 8.0 Hz, 1 H, 6Ј-H),

4.59 (s, 2 H, CH₂Ph), 5.36 (d, ³J = 8.1 Hz, 1 H, 1-H), 5.50 (dd,

³J_2,3= 8.3, J_3,4= 10.8 Hz, 1 H), 7.13–7.33 (m, 5 H, phenyl) ppm.

3

O-[(2,6-Di-O-acetyl-3,4-di-O-benzyl-β-

O-acetyl-6-O-benzyl-2-deoxy-2-(dimethylmaleimido)-β-

pyranosyl] Trichloroacetimidate (14): Trichloroacetonitrile

D-glucopy- (4.25 mL, 6.12 g, 42.4 mmol) was added to a solution of 13 (6.25 g,

D

-glucopyranosyl)(1Ǟ4)-3-

C₂₉H₄₃NO₈Si (561.7): calcd. C 62.01, H 7.72, N 2.49; found C

62.00, H 7.89, N 2.42.

D

-gluco-

Dimethyl(thexyl)silyl (2,6-Di-O-acetyl-3,4-di-O-benzyl-β-

ranosyl)(1Ǟ4)-3-O-acetyl-6-O-benzyl-2-deoxy-2-(dimethylmale-

imido)-β- -glucopyranoside (12): TMSOTf (0.1  in dry CH₂Cl₂,

7.38 mmol) in dry CH₂Cl₂(35 mL), then DBU (5 drops), and the

solution was stirred at room temperature for 1 h. The dark reaction

mixture was concentrated in vacuo to 3/4 of its volume and the

residue purified by column chromatography (silica gel, petroleum

ether/ethyl acetate, 3:2 + 1 % NEt₃) to furnish 14 (7.43 g,

6.80 mmol, 92%) as slightly brownish foam which was immediately

used in the next step. TLC: R_f= 0.69 (petroleum ether/ethyl acetate,

1:1). [α]²_D⁰= +20.1 (c = 1, CHCl₃). ¹H NMR (250 MHz, CDCl₃): δ

= 1.87, 1.89, 2.01 (3 s, 15 H, 3ϫ acetyl, DMM-H), 3.34–3.40 (m,

1 H, 5b-H), 3.41–3.52 (m, 2 H, 3b-H, 4b-H), 3.42–3.53 (m, 3 H,

5a-H, 6a-H), 4.02 (dd, ³J_3,4≈ ³J_4,5≈ 9.4 Hz, 1 H, 4a-H), 4.07–4.20

D

8 mL, 0.1 equiv.) was added to a solution of 8 (5.06 g, 8.6 mmol)

and 11 (5.4 g, 9.62 mmol) in dry CH₂Cl₂at –5 °C; the reaction

mixture was stirred at this temperature for 1 h and then neutralized

with NEt₃. The reaction mixture was concentrated in vacuo and

the residue purified by flash chromatography (silica gel, petroleum

ether/ethyl acetate, 5:1) to furnish compound 12 (8.11 g, 8.2 mmol,

95%) as colorless foam. TLC: R_f= 0.48 (petroleum ether/ethyl ace-

tate, 2:1). [α]²_D⁰= +3.6 (c = 1, CHCl₃). ¹H NMR (600 MHz,

CDCl₃): δ = 0.02, 0.13 (2 s, 6 H, 2ϫ SiCH₃), 0.70–0.76 (m, 12 H,

4ϫ CH₃), 1.49 [sept, ³J = 6.8 Hz, 1 H CH(CH₃)₂], 1.86 (d, 1 H, (m, 2 H, 6b-H), 4.27 (dd, J_1,2= 9.1, J_2,3= 10.5 Hz, 1 H, 2a-H),

DMM-H), 1.88, 1.90, 2.01 (3 s, 12 H, acetyl), 3.41 (m, 1 H, 5b-H),

3.54 (m, 2 H, 3b-H, 4b-H), 3.64 (dd, J_gem= 10.8 Hz, 1 H, 6a-H), 4.59 (d, 1 H, CHHPh), 4.72–4.78 (m, 3 H, CH₂Ph, CHHPh), 4.84

3.77 (dd, 2J_gem= 10.9, J_vic= 2.9 Hz, 1 H, 6aЈ-H), 3.90 (dd, J_3,4(dd, J_1,2

3

4.39 (d, J_1,2= 8.0 Hz, 1 H, 1b-H), 4.44, 4.47 (2 d, 2 H, CH₂Ph),

2

³J_2,3≈ 8.4 Hz, 1 H, 2b-H), 4.84 (dd, J_2,3

≈

³J_3,4

≈

3

≈

Eur. J. Org. Chem. 2007, 6016–6033

www.eurjoc.org

6023

T. Haag, R. A. Hughes, G. Ritter, R. R. Schmidt

FULL PAPER

3

9.8 Hz, 1 H, 3a-H), 6.40 (d, J_1,2= 8.9 Hz, 1 H, 1a-H), 7.22–7.34 [N-(Triphenylmethyl)carbamoyl]methyl (3,4-Di-O-benzyl-β-

D-gluco-

(m, 15 H, phenyl), 8.62 (s, 1 H, NH). C₄₇H₅₁Cl₃N₂O₁₅(990.3) ppm. pyranosyl)(1Ǟ4)-6-O-benzyl-2-deoxy-2-(dimethylmaleimido)-β-

D-glu-

copyranoside (19): Sodium methoxide (20 mg, 0.37 mmol) was

added to a solution of 18 (5.0 g, 4.4 mmol) in dry MeOH and the

mixture stirred at room temperature overnight. The reaction mix-

ture was neutralized with ion exchange resin IR 120 (H⁺), filtered,

and concentrated in vacuo to furnish 19 (4.3 g, 4.2 mmol, 95%) as

slightly yellowish foam. TLC: R_f= 0.35 (toluene/ethyl acetate, 1:1).

N-(Triphenylmethyl)methoxyacetamide (16): A solution of 15 (5.2 g,

50 mmol, 4.82 mL) was combined with a 2  methanolic NH₃solu-

tion (15 mL) and the mixture stirred at room temperature over-

night. The solvents were removed under reduced pressure to furnish

a white, amorphous solid (3.75 g, 49.9 mmol, quant.). The interme-

diate was dissolved in pyridine (50 mL), and acetic anhydride

(25 mL) was added under ice-bath cooling. After stirring overnight,

the reaction mixture was concentrated and coevaporated four times

with toluene. The acetylated amide (5.0 g, 42.1 mmol) was sus-

pended with triphenylmethanol (22 g, 84.2 mmol) in acetic acid

(120 mL) and acetic anhydride (8 mL) and after addition of sulfuric

acid (2.56 mL, concentrated) stirred at 55 °C for 4 h. The dark red

solution was then slowly poured into ice-cold water (400 mL) while

stirring to furnish a white amorphous solid. Filtration and drying

in vacuo furnished 16 (7.54 g, 21 mmol, 42% from 33) as colorless

powder. TLC: R_f= 0.60 (toluene/ethyl acetate, 6:4). ¹H NMR

(250 MHz, CDCl₃): δ = 2.14 (s, 3 H, acetyl), 4.57 (s, 2 H,

CH₂C=O), 7.16–7.31 (m, 15 H, Trityl) ppm. C₂₃H₂₁NO₃(359.4):

calcd. C 76.86, H 5.89, N 3.90; found C 76.27, H 6.02, N 3.91.

[α]²_D⁰= –14.9 (c = 1, CHCl₃). H NMR (600 MHz, [D₆]DMSO): δ

1

3

= 1.80 (s, 6 H, DMM-H), 3.26 (dd, J_1,2≈ J_2,3≈ 8.0 Hz, 1 H, 2b-

3

H), 3.31 (m, 1 H, 5b-H), 3.34 (dd, J_3,4≈ J_4,5≈ 9.0 Hz, 1 H, 4b-

3

H), 3.44 (m, 2 H, 3b-H, 6b-H), 3.52 (dd, J_3,4= J_4,5= 8.8 Hz, 1

H, 4a-H), 3.65 (m, 3 H, 2a-H, 5a-H, 6bЈ-H), 3.77 (m, ²J_gem= 10.4,

2

³J_vic= 4.4 Hz, 1 H, 6a-H), 3.82 (dd, J_gem= 10.4 Hz, 1 H, 6aЈ-H),

3

4.05–4.15 (m, 3 H, 3a-H, CH₂Ph), 4.31 (d, J_1,2= 7.8 Hz, 1 H, 1b-

H), 4.38 (2 d, 2 H, OCH₂C=O), 4.54, 4.67, 4.73 (3 d, 3 H, CH₂Ph,

CHHPh), 4.74 (m, 1 H, 6b-OH), 4.82 (m, 1 H, 3a-OH), 4.90 (d, 1

3

H, CHHPh), 5.03 (d, 1 H, J_1,2= 8.6 Hz, 1a-H), 5.69 (d, 1 H, J

= 5.7 Hz, 2b-OH), 7.08–7.35 (m, 30 H, phenyl), 7.99 (s, 1 H, NH)

ppm. ¹³C NMR (150.8 MHz, [D₆]DMSO, selected data): δ = 55.7

(1 C, 2a-C), 60.1 (1 C, 6b-C), 68.2 (1 C, 6a-C), 68.5 (1 C, 3a-C),

72.3 (1 C, OCH₂C=O), 73.5 (1 C, 2b-C), 74.0 (1 C, 5a-C), 75.1 (1

C, 5b-C), 77.1 (1 C, 4b-C), 80.4 (1 C, 4a-C), 84.3 (1 C, 3b-C), 98.2

(1 C, 1a-C), 103.0 (1 C, 1b-C) ppm. C₆₆H₆₂N₂O₁₃(1019.1): calcd.

C 70.71, H 6.13, N 2.75; found C 70.43, H 6.14, N 3.00.

2-Hydroxy-N-(triphenylmethyl)acetamide (17): Sodium methoxide

(0.1  in dry MeOH, 5 mL) was added to a solution of 16 (7.37 g,

20.5 mmol) in dry MeOH (60 mL). The solution was then stirred

at room temperature for 2 h and neutralized with ion exchange

resin IR 120 (H⁺), filtered, and concentrated in vacuo. The residue

was dissolved in ethyl acetate and heated to reflux; then n-hexane

was added until the mixture became slightly turbid. After cooling,

the residue was filtered and dried under reduced pressure which

furnished 17 (6.1 g, 19 mmol, 93%) as colorless powder. TLC: R_f

[N-(Triphenylmethyl)carbamoyl]methyl (3,4-Di-O-benzyl-β-

pyranosyl)(1Ǟ4)-6-O-benzyl-2-[(benzyloxycarbonyl)amino]-2-de-

oxy-β- -glucopyranoside (20): NaOH (790 mg, 19.8 mmol) was

D-gluco-

D

added to a solution of 19 (1.4 g, 1.37 mmol) in dioxane (80 mL)

and water (20 mL) at room temperature, and the reaction mixture

was stirred at room temperature overnight. Then the pH was ad-

justed to 4.5 with 1  HCl, monitored every 30 min for the next

4 h and if necessary adjusted to 4.5 with 1  HCl. After stirring

overnight at pH = 4.5, the reaction mixture was neutralized with

ethanolamine (110 µL) and NaOH. Thereafter K₂CO₃(600 mg,

4.4 mmol) was added; after addition of benzyl chloroformate

(0.56 mL, 669 mg, 3.92 mmol), the mixture was stirred at room

temperature for 1.5 h. The reaction mixture was concentrated to 1/2

of its volume and extracted three times with CH₂Cl₂. The combined

organic phases were dried with MgSO₄, filtered and concentrated.

Purification by flash chromatography (silica gel, toluene/acetone,

4:1) furnished Z-protected 20 (1.05 g, 1.0 mmol, 73%) as colorless,

1

= 0.25 (toluene/ethyl acetate, 6:4). H NMR (250 MHz, CDCl₃): δ

= 3.99 (s, 2 H, CH₂–CO), 7.16–7.26 (m, 15 H, Trityl), 7.53 (br. s,

1 H, NH) ppm. C₂₁H₁₉NO₂(317.4): calcd. C 79.47, H 6.03, N 4.41;

found C 78.89, H 6.13, N 4.38.

[N-(Triphenylmethyl)carbamoyl]methyl (2,6-Di-O-acetyl-3,4-di-O-

benzyl-β-D-glucopyranosyl)(1Ǟ4)-3-O-acetyl-6-O-benzyl-2-deoxy-2-

(dimethylmaleimido)-β-

D

-glucopyranoside (18): A solution of 14

(5.84 g, 5.90 mmol) and 17 (2.15 g, 6.77 mmol, 1.15 equiv.) in dry

CH₂Cl₂(30 mL) was cooled to –5 °C; a TMSOTf solution (0.1 

in dry CH₂Cl₂, 5.0 mL, 0.08 equiv.) was added and the mixture

stirred at –5 °C for 1 h. After neutralising the reaction mixture with

NEt₃, it was concentrated in vacuo and the residue purified twice

by column chromatography (silica gel, toluene/ethyl acetate, 4:1 Ǟ

3:1) to furnish 18 (4.2 g, 3.67 mmol, 65%) as colorless foam. TLC:

R_f= 0.30 (toluene/ethyl acetate, 6:4). [α]²_D⁰= –4.9 (c = 1, CHCl₃).

¹H NMR (600 MHz, CDCl₃): δ = 1.83 (br. d, 6 H, DMM-H), 1.85,

1.86, 2.00 (3 s, 12 H, 4ϫ acetyl), 3.34 (m, 1 H, 5b-H), 3.41 (dd,

hygroscopic foam. TLC: R_f= 0.57 (toluene/acetone, 1:1). [α]²_D⁰

=

1

–16.3 (c = 1, CHCl₃). H NMR (250 MHz, CDCl₃): δ = 2.32 (m,

1 H, OH), 3.32–3.52 (m, 5 H, 2b-H, 3b-H, 4b-H, 5b-H, 6b-H),

3.53–3.69 (m, 3 H, 2a-H, 6a-H, 6bЈ-H), 3.81 (dd, ²J_gem= 12.5, ³J_vic

3

= 3.5 Hz, 6aЈ-H), 4.12 (d, 1 H, CHHPh), 4.26 (dd, J_2,3≈ J_3,4

≈

7.8 Hz, 3a-H), 4.29 (s, 2 H, CH₂Ph), 4.51 (m, 2 H, CH₂–CO), 4.59

(d, ³J_1,2= 8.9 Hz, 1 H, 1b-H), 4.81–4.90 (m, 4 H, 2ϫ CH₂Ph), 5.03

(d, ³J_1,2= 9.5 Hz, 1 H, 1a-H), 7.13–7.35 (m, 35 H, phenyl), 7.91 (s,

1 H, NH) ppm. MALDI-MS (pos. mode, matrix DHB, THF): m/z

= 1068 [M + Na]⁺, 1084 [M + K]⁺.

3

³J_2,3= J_3,4= 9.1 Hz, 1 H, 3b-H), 3.52 (dd, J_3,4= J_4,5= 9.2 Hz,

1 H, 4b-H), 3.56 (m, 2 H, 5a-H, 6a-H), 3.68 (dd, ²J_gem= 10.8, ³J_vic

= 2.4 Hz, 6aЈ-H), 3.96 (dd, J_3,4= J_4,5= 9.1 Hz, 1 H, 4a-H), 4.03

(dd, 1 H, 2a-H), 4.04, 4.08, 4.21 (3 d, 3 H, CH₂Ph, CHHPh), 4.22

(m, 2 H, 6b-H), 4.34 (d, ³J_1,2= 8.0 Hz, 1 H, 1b-H), 4.42, 4.49, 4.58

3

[N-(Triphenylmethyl)carbamoyl]methyl (Benzyl 3,4-di-O-benzyl-β-D-

(3 d, 3 H, CH₂Ph, CHHPh), 4.71, 4.76 (2 d, CH₂C=O), 5.27 (d, glucopyranosyluronate)(1Ǟ4)-6-O-benzyl-2-[(benzyloxycarbonyl)-

³J_1,2= 8.5 Hz, 1 H, 1a-H), 5.46 (dd, ³J_2,3≈ ³J_3,4≈ 9.8 Hz, 1 H, 3a-

amino]-2-deoxy-β- -glucopyranoside (21): A mixture of an NaOCl

H), 7.14–7.33 (m, 30 H, phenyl), 7.73 (s, 1 H, NH) ppm. ¹³C NMR solution (10 %, 1.2 mL), H₂O (1 mL) and an NaHCO₃solution

D

(150.8 MHz, CDCl₃, selected data): δ = 54.7 (1 C, 2a-C), 62.8 (1

(saturated, 1.7 mL) was added dropwise to a solution of 20

C, 6b-C), 66.7 (1 C, 6a-C), 70.7 (1 C, 3a-C), 72.7 (2 C, 2b-C, 5b-

(300 mg, 0.29 mmol), NaBr (5 mg), TBAB (5 mg), and TEMPO

C), 74.6 (1 C, 5a-C), 74.7 (1 C, 4a-C), 76.9 (1 C, 4b-C), 83.0 (1 C, (4 mg, 0.025 mmol) in CH₂Cl₂(5.2 mL) and H₂O (0.85mL) at 0 °C;

3b-C), 99.0 (1 C, 1a-C), 100.1 (1 C, 1b-C) ppm. MALDI-MS (pos. then the mixture was stirred at 0 °C for 20 min, then methanol

mode, matrix DHB, THF): m/z = 1069 [M + Na]⁺. C₆₆H₆₈N₂O₁₆(1 mL) was added. The reaction mixture was diluted with H₂O and

(1145.3): calcd. C 69.22, H 5.98, N 2.45; found C 69.18, H 6.25, N

2.51.

extracted with CH₂Cl₂. The aqueous phase was acidified with a

KHSO₄solution (5%) and extracted three times with CH₂Cl₂. The

6024

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Eur. J. Org. Chem. 2007, 6016–6033

Carbohydrate-Based VEGF Inhibitors

combined organic phases were dried with MgSO₄, filtered, and

concentrated in vacuo to furnish the free acid (290 mg, 95%) as

intermediate which was dissolved in dry DMF (5 mL); CsF (90 mg,

0.59 mmol) and benzyl bromide (75 µL, 107.9 mg, 0.63 mmol) were

added, and the mixture was stirred at room temperature overnight.

The reaction mixture was then diluted with ethyl acetate and

washed 3ϫ with saturated NaCl solution. The organic phase was

dried with MgSO₄, filtered, and the solvents were evaporated. After

coevaporating twice with toluene, the residue was purified by col-

umn chromatography (silica gel, toluene/acetone, 4.5:1) to furnish

21 as colorless foam (305 mg, 0.27 mmol, 93%). TLC: R_f= 0.57

(toluene/acetone, 1:1). [α]²_D⁰= –11.4 (c = 1, CHCl₃). ¹H NMR

(250 MHz, CDCl₃): δ = 2.66 (s, 1 H, OH), 3.39–3.51 (m, 3 H, 5a-

H, 3b-H, 2b-H), 3.53–3.68 (m, 5 H, 2a-H, 3a-H, 6a-H, 5b-H), 3.88

(d, 1 H, CHHPh), 4.02 (dd, 1 H, 4b-H), 4.14 (dd, 1 H, 4a-H), 4.23

the mixture was stirred at room temperature overnight. Then the

reaction mixture was concentrated in vacuo until it became turbid,

then diluted with H₂O and extracted four times with CHCl₃. The

combined organic phases were dried with MgSO₄, concentrated,

and the residue was purified by column chromatography (silica gel,

ethyl acetate/MeOH, 10:3 Ǟ 3:1). Not all byproducts could be sep-

arated. Compound 23 (82 mg, 0.074 mmol, 68%) was used in the

next step without further purification. MALDI-MS (pos. mode,

matrix DHB, THF): m/z = 1128 [M + Na]⁺, 1144 [M + K]⁺.

N-[(9-Fluorenyl)methoxycarbonyl]-S-(triphenylmethyl)-L-cysteine

tert-Butyl Ester (24): TBTA (874 mg, 4 mmol) in dry cyclohexane

(4 mL) was added to a solution of Fmoc-Cys(Trt)-OH (1.17 g,

2.0 mmol) in CH₂Cl₂(2 mL). After addition of BF₃·Et₂O (40 µL),

the reaction mixture was stirred at room temperature for 3 h and

neutralized with NEt₃. The reaction mixture was then concentrated

and the residue purified by column chromatography (silica gel, pe-

troleum ether/ethyl acetate, 8:1) to furnish 24 (1.12 g, 1.75 mmol,

87%) as colorless foam. TLC: R_f= 0.70 (petroleum ether/ethyl ace-

tate, 2:1). [α]²_D⁰= +10.0 (c = 1, CHCl₃). ¹H NMR (250 MHz,

CDCl₃): δ = 1.41 (s, 9 H, tert-butyl), 2.54 (m, 2 H, Cys-β-H), 4.25

3

(d, 2 H, CH₂Ph), 4.42 (d, 2 H, CH₂Ph), 4.60 (bd, J_1,2≈ 8.0 Hz, 1

3

H, 1a-H), 4.64 (d, 1 H, CHHPh), 4.72 (s, 1 H, OH), 4.86 (d, J_1,2

= 8.4 Hz, 1 H, 1b-H), 4.89 (d, 2 H, CH₂Ph), 5.12 (s, 1 H, OH),

7.13–7.35 (m, 40 H, phenyl) ppm. MALDI-MS (pos. mode, matrix

DHB, THF): m/z = 1183 [M + Na]⁺, 1199 [M + K]⁺. C₆₉H₆₈N₂O₁₄

(1149.3): calcd. C 72.11, H 5.96, N 2.44; found C 71.87, H 6.01, N

2.46.

3

(m, 1 H, Cys-α-H), 5.31 (d, J = 7.8 Hz, 1 H, NH), 7.16–7.29 (m,

17 H, phenyl, Fmoc), 7.36–7.43 (m, 4 H, Fmoc) ppm. C₄₁H₃₉NO₄S

(641.8): calcd. C 76.73, H 6.12, N 2.18; found C 76.41, H 6.54, N

2.10.

[N-(Triphenylmethyl)carbamoyl]methyl [Benzyl 3,4-di-O-benzyl-2-

O-(tert-butyloxycarbonylmethyl)-β-

6-O-benzyl-2-[(benzyloxycarbonyl)amino]-3-O-[(tert-butyloxy-

carbonyl)methyl]-2-deoxy-β- -glucopyranoside (22): tert-Butyl bro-

D-glucopyranosyluronate](1Ǟ4)-

N-[(9-Fluorenyl)methoxycarbonyl]-

L-isoleucyl-S-(triphenylmethyl)-

D

L-cysteine tert-Butyl Ester (25): Piperidine (4 mL) was added to a

moacetate (880 µL, 1.16 g, 5.96 mmol) and molecular sieves (4 Å)

were added to a solution of 21 (960 mg, 0.82 mmol) in CH₂Cl₂

(25 mL). After stirring at room temperature for 10 min, silver(I)

oxide (2.85 g, 12.3 mmol) and TBAI (700 mg, 1.89 mmol) were

added to the reaction mixture which was then stirred in the dark

overnight. The reaction mixture was filtered through Celite and

purified by flash chromatography (silica gel, petroleum ether/ethyl

acetate, 3:1 Ǟ 2.5:1 Ǟ 2:2) to furnish 22 (936 mg, 0.68 mmol, 82%)

as colorless foam. TLC: R_f= 0.30 (toluene/ethyl acetate, 4:1).

[α]²_D⁰= –20.0 (c = 1, CHCl₃). ¹H NMR (600 MHz, CDCl₃): δ =

1.46, 1.47 (2 s, 18 H, 2ϫ tert-butyl), 3.23 (dd, ³J_1,2= ³J_2,3= 8.4 Hz,

solution of 24 (3.0 g, 4.67 mmol) in CH₂Cl₂(25 mL) and the mix-

ture stirred at room temperature for 3 h. The reaction mixture was

concentrated in vacuo and coevaporated with toluene. After purifi-

cation of the residue by column chromatography (silica gel, tolu-

ene/ethyl acetate, 20:1 Ǟ 4:1 Ǟ 2.5:1) the resulting unprotected

amino acid (1.8 g, 4.29 mmol, 92%) was dissolved in dry CH₂Cl₂

(25 mL), and Fmoc-Ile-OH (1.8 g, 5.15 mmol, 1.2 equiv.) and

PyBOP (2.6 g, 4.99 mmol) were added. The pH was adjusted to ca.

8 with DIPEA (ca. 1.1 g, 2 equiv., 1.46 mL). After stirring at room

temperature for 1 h, the reaction mixture was diluted with ethyl

acetate and washed with KHSO₄and NaHCO₃solution. The or-

ganic phase was dried with MgSO₄, filtered, concentrated and the

residue purified by flash chromatography (silica gel, toluene/ethyl

acetate, 15:1) to furnish dipeptide 25 (3.1 g, 4.11 mmol, 96%) as

3

1 H, 2b-H), 3.39 (m, 1 H, 5a-H), 3.49 (dd, J_2,3= 8.4 Hz, 1 H, 3b-

2

H), 3.57 (m, 1 H, 3a-H), 3.58 (m, 1 H, 2a-H), 3.64 (dd, J_gem

=

3

10.8 Hz, 1 H, 6a-H), 3.69 (d, J = 9.8 Hz, 1 H, 5b-H), 3.73 (dd, 1

3

2

3

H, J_3,4

≈ =

³J_4,5≈ 4b-H), 3.85 (dd, 1 H, J_gem= 11.0 Hz, J_vic

colorless foam. TLC: R_f= 0.65 (toluene/ethyl acetate, 5:1). [α]²_D⁰

=

2.7 Hz, 6aЈ-H), 4.02 (m, 1 H, 4a-H), 4.03–4.08 (m, 2 H, CH₂–CO),

4.12–4.20 (m, 2 H, CH₂–CO), 4.27–4.34 (m, 3 H, CH₂–CO,

1

+2.5 (c = 1, CHCl₃). H NMR (250 MHz, CDCl₃): δ = 0.92 (m, 6

H, 2ϫ Ile-CH₃), 1.20 (m, 1 H, Ile-CHH), 1.42 (s, 9 H, tert-butyl),

1.44 (m, 1 H, Ile-CHH), 1.84 (m, 1 H, Ile-β-H), 2.49–2.62 (m, 2 H,

Cys-β-H), 4.04 (m, 1 H, Cys-α-H), 4.21 (m, 1 H, Ile-α-H), 4.35–

4.51 (m, 1 H, Fmoc-CH₂), 5.38 (d, ³J = 8.5 Hz, 1 H, NH), 6.18 (d,

³J = 7.6 Hz, 1 H, NH), 7.11–7.39 (m, 19 H, phenyl, Fmoc), 7.85,

7.75 (2 m, 4 H, Fmoc) ppm. C₄₇H₅₀N₂O₅S (754.98): calcd. C 74.77,

H 6.68, N 3.71; found C 74.98, H 6.95, N 3.90.

3

CHHPh), 4.40 (d, 1 H, J_1,2= 7.5 Hz, 1a-H), 4.41–4.46 (m, 3 H,

CH₂Ph, 1b-H), 4.68–4.76 (m, 3 H, CH₂Ph, CHHPh), 4.92 (d, 1 H,

CHHPh), 4.95 (s, 1 H, CHHPh), 5.13, 5.16 (2 d, 2 H, CH₂Ph), 5.30

(s, 1 H, CHHPh), 6.74 (s, 1 H, 2a-NH), 7.13–7.31 (m, 40 H,

phenyl), 8.13 (s, 1 H, NH) ppm. ¹³C NMR (150.8 MHz, CDCl₃,

selected data): δ = 28.0 (3 C, tert-butyl), 28.1 (3 C, tert-butyl), 56.1

(1 C, 2a-C), 67.2 (1 C, 6a-C), 73.9 (1 C, 5b-C), 74.8 (1 C, 5a-C),

77.6 (1 C, 4a-C), 78.6 (1 C, 3a-C), 79.2 (1 C, 4b-C), 82.8 (1 C, 2b-

C), 83.2 (1 C, 3b-C), 102.2 (1 C, 1b-C), 103.5 (1 C, 1a-C), 168.2 (1

C, 6b-C) ppm. MALDI-MS (pos. mode, matrix DHB, THF): m/z

= 1400 [M + Na]⁺, 1416 [M + K]⁺.

N-[(9-Fluorenyl)methoxycarbonyl]-

L-leucyl-L-isoleucyl-S-(triphenyl-

methyl)- -cysteine tert-Butyl Ester (26a): Piperidine (3 mL) was

L

added dropwise to a solution of 25 (1.88 g, 2.5 mmol) in CH₂Cl₂

(20 mL). After stirring at room temperature for 1 h, the reaction

mixture was concentrated, coevaporated with toluene and the resi-

due purified by column chromatography (silica gel, toluene/ethyl

[N-(Triphenylmethyl)carbamoyl]methyl {2-O-[(tert-Butyloxycar-

bonyl)methyl]-β-D-glucopyranosyluronate}-(1Ǟ4)-3-O-[(tert-butyl-

oxycarbonyl)methyl]-2-deoxy-2-{[(9-fluorenyl)methoxycarbonyl]- acetate, 10:1 Ǟ 5:1 Ǟ 2:1). The resulting unprotected dipeptide

amino}-β-

D

-glucopyranoside (23): Pd/C (100 mg) was added to a

(1.1 g, 2.0 mmol, 80%) was dissolved in dry CH₂Cl₂(15 mL), and

Fmoc-Leu-OH (0.85 g, 2.4 mmol) and PyBOP (1.25 g, 2.4 mmol)

were added. The pH was adjusted to 8 with DIPEA (ca. 0.68 mL)

and the reaction mixture stirred at room temperature. After 1 h, it

was diluted with CHCl₃and washed with KHSO₄, NaHCO₃and

NaCl solution. The organic phase was dried with MgSO₄, filtered

solution of 22 (150 mg, 0.11 mmol) in THF/H₂O (5:1, 20 mL). Af-

ter stirring under hydrogen for 48 h, the catalyst was filtered off

through Celite, the filtrate concentrated to 1/2 of its volume, diluted

with CH₃CN/H₂O (2:1, 20 mL), and then Fmoc-ONSu (90 mg,

0.27 mmol) and NaHCO₃(225 mg, 2.67 mmol) were added, and

Eur. J. Org. Chem. 2007, 6016–6033

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6025

T. Haag, R. A. Hughes, G. Ritter, R. R. Schmidt

FULL PAPER

and concentrated. Purification by column chromatography (silica

gel, toluene/ethyl acetate, 10:1) furnished tripeptide 26a (1.68 g,

1.93 mmol, 97%) as colorless foam. TLC: R_f= 0.59 (toluene/ethyl

(pos. mode, matrix DHB, THF): m/z = 1756 [M + Na]⁺, 1772 [M

+ K]⁺.

[N-(Triphenylmethyl)carbamoyl]methyl {2-O-[(tert-Butyloxycar-

acetate, 1:1). [α]²_D⁰= +3.6 (c = 0.5, CHCl₃). H NMR (600 MHz,

1

bonyl)methyl]-β-

phenylmethyl)-

tyloxycarbonyl)methyl]-2-{[N-(tert-butyloxycarbonyl)-S-(triphenyl-

methyl)- -cysteinyl]amido}-2-deoxy-β- -glucopyranoside (30a):

D

-glucopyranosyluronyl[

L-leucyl-L-isoleucyl-S-(tri-

[D₆]DMSO): δ = 0.75 (m, 3 H, Ile-CH₃), 0.79 (m, 3 H, Ile-CH₃),

0.83 (m, 6 H, Leu-CH₃), 1.03 (m, 1 H, Ile-CHH), 1.27 (s, 9 H, tert-

butyl), 1.39 (m, 1 H, Ile-CHH), 1.45 (m, 2 H, Leu-β-H), 1.57 [m,

1 H, Leu-CH(CH₃)₂], 1.67 (m, 1 H, Ile-β-H), 2.30, 2.34 (2 m, Cys-

β-H), 3.92 (m, 1 H, Cys-α-H), 4.09 (m, 1 H, Leu-α-H), 4.19–4.29

(m, 3 H, Fmoc-CH₂, Ile-α-H), 7.21–7.32 (m, 19 H, phenyl, Fmoc),

7.39, 7.67 (2 m, 4 H, Fmoc), 7.54 (m, 1 H, Leu-NH), 7.70 (m, 1

H, Ile-NH), 8.37 (m, 1 H, Cys-NH) ppm. ¹³C NMR (150.8 MHz,

CDCl₃, selected data): δ = 10.8 (1 C, Ile-CH₃), 14.9 (1 C, Ile-CH₃),

21.1 (1 C, Leu-CH₃), 22.8 (1 C, Leu-CH₃), 23.8 (1 C, Ile-CH₂),

23.9 [1 C, Leu-CH(CH₃)₂], 32.5 (1 C, Cys-β-C), 36.9 (1 C, Ile-β-

C), 40.5 (1 C, Leu-β-C), 46.5 (1 C, Fmoc), 52.0 (1 C, Cys-α-C),

52.9 (1 C, Leu-α-C), 55.9 (1 C, Ile-α-C) ppm. C₅₃H₆₁N₃O₆S (868.1):

calcd. C 73.33, H 7.08, N 4.84; found C 73.26, H 7.37, N 4.80.

L

-cysteine tert-butyl ester]}-(1Ǟ4)-3-O-[(tert-bu-

L

D

Piperidine (1 mL) was added to a solution of 28a (50 mg,

0.029 mmol) in CH₂Cl₂(5 mL). After 3 h, the reaction mixture was

concentrated in vacuo, coevaporated twice with toluene and the

residue purified by column chromatography (silica gel, CHCl₃/

MeOH, 200:1 Ǟ 50:1 Ǟ 25:1). The NH-unprotected compound

(30 mg, 0.020 mmol, 70 %) was dissolved with 29 (11 mg,

0.023 mmol) and PyBOP (14 mg, 0.027 mmol) in dry DMF (1 mL).

The pH was adjusted to 8 with DIPEA (ca 8 µL), and the reaction

mixture stirred at room temperature for 1 h. After dilution with

ethyl acetate, the mixture was washed with NH₄Cl solution and

NaCl solution. The organic phase was dried with MgSO₄, filtered

and concentrated. Purification by flash chromatography (silica gel

toluene/acetone, 4.5:1 Ǟ 3.25:1) furnished 30a (36 mg, 0.018 mmol,

93 %, 64 % over 2 steps) as colorless lyophilisate from dioxane.

L-Leucyl-L-isoleucyl-S-(triphenylmethyl)-L-cysteine tert-Butyl Ester

(27a): The protected tripeptide 26a (867 mg, 1.0 mmol) was dis-

solved in CH₂Cl₂(15 mL), and piperidine (3 mL) was added. After

3 h of stirring at room temperature, the reaction mixture was con-

centrated and coevaporated twice with toluene. The residue was

purified by flash chromatography (silica gel, toluene/ethyl acetate,

10:1 Ǟ 5:1) to furnish deprotected tripeptide 27a (640 mg,

0.99 mmol, 99%) which was immediately used in the next step.

1

TLC: R_f= 0.62 (toluene/acetone, 1:1). H NMR (600 MHz, [D₆]-

DMSO): δ = 0.78 (m, 13 H, 2ϫ Leu-CH₃, 2ϫ Ile-CH₃, Ile-β-H),

1.02 (m, 1 H, Ile-CHH), 1.28–1.40 (m, 37 H, 3ϫ tert-butyl, Boc,

Ile-CHH), 1.44 (m, 2 H, Leu-β-H), 1.63 [m, 2 H, Ile-β-H, Leu

CH(CH₃)₂], 2.34 (m, 2 H, Cys-β-H), 2.44 (m, 2 H, Cys-β-H), 2.99

(m, 1 H, 2b-H), 3.30–3.34 (m, 3 H, 5a-H, 3b-H, 4b-H), 3.48 (dd, 1

H, 2a-H), 3.54 (m, 1 H, 6a-H), 3.65–3.71 (m, 4 H, 3a-H, 4a-H, 6aЈ-

H, 5b-H), 3.88–3.90 (m, 3 H, CHHC=ONHTrt, Cys-α-H, Cys-α-

[N-(Triphenylmethyl)carbamoyl]methyl {2-O-[(tert-Butyloxycar-

bonyl)methyl]-β-

phenylmethyl)-

butyloxycarbonyl)methyl]-2-deoxy-2-{[(9-fluorenyl)methoxycarbon-

yl]amino}-β- -glucopyranoside (28a): A solution of 23 (55 mg,

D

-glucopyranosyluronyl[

L-leucyl-L-isoleucyl-S-(tri-

2

H), 4.02 (d, J_gem= 15 Hz, 1 H, CHHC=ONHTrt), 4.14 (m, 2 H,

L

-cysteine tert-butyl ester]}-(1Ǟ4)-3-O-[(tert-

CH₂C=O), 4.19 (m, 2 H, CH₂C=O), 4.26 (dd, ³J = 8.2 Hz, 1 H,

3

Ile-α-H), 4.36 (m, 1 H, Leu-α-H), 4.53 (d, J_1,2≈ 7.3 Hz, 2 H, 1a-

D

H, 1b-H), 4.63 (m, 1 H, OH), 5.24 (m, 1 H, OH), 5.40 (m, 1 H,

0.049 mmol), 27a (38 mg, 0.058 mmol) and PyBOP (34 mg,

0.065 mmol) in DMF (3 mL) was treated with DIPEA (ca. 20 µL)

to adjust the pH to ca. 8. After stirring at room temperature for

2 h, the reaction mixture was diluted with ethyl acetate and washed

with KHSO₄solution (5%), NaHCO₃solution and saturated NaCl

solution. The organic phase was dried with MgSO₄, filtered, and

concentrated in vacuo. Purification by column chromatography

(silica gel, toluene/acetone, 4:1 Ǟ 3.5:1) furnished 28a (70 mg,

0.040 mmol, 82%) as colorless lyophilisate from dioxane. TLC: R_f

= 0.75 (toluene/acetone, 1:1). [α]²_D⁰= –16.7 (c = 0.3, CHCl₃). ¹H

NMR (600 MHz, CDCl₃): δ = 0.84–0.88 (m, 12 H, CH₃), 1.09 (m,

1 H, Ile-CHH), 1.38 (s, 9 H, tert-butyl), 1.42 (m, 1 H, Ile-CHH),

OH), 6.49 (d, ³J = 8.1 Hz, 1 H, Cys-NH), 7.15–7.33 (m, 45 H, Trt),

3

7.72 (d, J = 7.4 Hz, 1 H, 2a-NH), 7.91 (d, J = 9.0 Hz, 1 H, Ile-

NH), 8.03 (s, 1 H, C=ONHTrt), 8.05 (d, ³J = 7.3 Hz, 1 H, Leu-

NH), 8.47 (d, ³J = 7.2 Hz, 1 H, Cys-NH) ppm. ¹³C NMR

(150.8 MHz, [D₆]DMSO, selected data): δ = 50.9 (1 C, Leu-α-C),

52.0 (1 C, Cys-α-C), 53.2 (1 C, Cys-α-C), 53.9 (1 C, 2a-C), 55.9 (1

C, Ile-α-C), 58.9 (1 C, 6a-C), 68.2 (1 C, CH₂C=O), 68.5 (1 C,

CH₂C=O), 69.0 (1 C, CH₂C=O), 74.6 (1 C, 5b-C), 76.5 (1 C, 4a-

C), 78.9 (1 C, 3a-C), 81.7 (1 C, 2b-C), 101.1 (1 C, 1b-C), 101.4 (1

C, 1a-C) ppm. MALDI-MS (pos. mode, matrix DHB, THF): m/z

= 1978 [M + Na]⁺, 1994 [M + K]⁺.

1.45 (s, 9 H, tert-butyl), 1.50 (s, 9 H, tert-butyl), 1.52, 1.68 (2 m, 2 Carbamoylmethyl [2-O-(Carboxymethyl)-β-

D

-glucopyranosyluron-

-hemicystine)]-(1Ǟ4)-3-O-(carboxymethyl)-

-hemicystinyl)amido]-β- -glucopyranoside (1a): A

H, Leu-β-H), 1.58 [m, 1 H, Leu-CH(CH₃)₂], 1.78 (m, 1 H, Ile-β-

H), 2.46, 2.64 (2 m, 2 H, Cys-β-H), 3.09 (m, 1 H, 2b-H), 3.35 (m,

1 H, 4a-H), 3.61 (m, 1 H, 2a-H), 3.63–3.70 (m, 3 H, 3b-H, 4b-H,

5b-H), 3.72–3.76 (m, 4 H, 5-H, 6-H, 6Ј-H, Fmoc), 3.97 (m, 1 H,

yl(

L-leucyl-L-isoleucyl-L

2-deoxy-2-[(

L

D

solution of 30a (35 mg, 0.018 mmol) in CH₂Cl₂/MeOH (7:1,

18 mL) was slowly added dropwise to a solution of iodine (23 mg,

3a-H), 4.04–4.10 (m, 3 H, OCH₂C=O, Fmoc), 4.10–4.18 (m, 3 H, 0.091 mmol) in CH₂Cl₂/MeOH (7:1, 18 mL) at 0 °C . After stirring

Fmoc, OCH₂C=O), 4.19–4.22 (m, 3 H, Ile-α-H, OCH₂C=O), 4.23–

4.41 (m, 5 H, Cys-α-H, Leu-α-H, 1b-H, OCH₂C=O), 4.61 (m, 1 H,

1a-H), 6.01 (br. s, 1 H, Leu-NH), 6.68 (d, 1 H, Ile-NH), 6.83 (br.

at 0 °C for 30 min, 0.01  sodium thiosulfate solution was added

until decolorization and the reaction mixture extracted three times

with CHCl₃. The combined organic phases were dried with MgSO₄,

s, 1 H, Cys-NH), 7.12–7.73 (m, 39 H, Fmoc, phenyl, 2a-NH), 8.10 filtered, and concentrated in vacuo. The intermediate was then dis-

(br. s, 1 H, NHTrt) ppm. ¹³C NMR (150.8 MHz, CDCl₃, selected

data): δ = 11.5, 15.1, 21.6, 22.8 (4 C, 4ϫ CH₃), 24.7 [2 C, Leu-

CH(CH₃)₂, Ile-CH₂], 27.8, 28.1, 28.2 (9 C, 3ϫ tert-butyl), 33.7 (1

C, Cys-β-C), 37.8 (1 C, Ile-β-C), 40.9 (1 C, Leu-β-C), 46.4 (1 C,

Fmoc), 51.8 (1 C, Leu-α-C), 52.0 (1 C, Cys-α-C), 57.4 (1 C, Ile-α-

C), 60.0 (1 C, 6a-C), 66.8 (1 C, Fmoc), 69.2 (2 C, OCH₂C=O), 70.1

(1 C, OCH₂C=O), 72.3 (1 C, 4b-C), 72.7 (1 C, 5a-C), 74.8 (1 C,

3b-C), 75.2 (1 C, 4a-C), 77.7 (1 C, 3a-C), 78.6 (1 C, 2a-C), 83.4 (1

C, 2b-C), 102.1 (1 C, 1a-C), 103.6 (1 C, 1b-C) ppm. MALDI-MS

solved in TFA (8 mL), Et₃SiH (0.4 mL) and H₂O (0.2 mL) and the

mixture stirred at 0 °C for 4 h. After removal of TFA in vacuo, the

reaction mixture was dried in vacuo and prepurified by chromatog-

raphy (RP-18 silica gel, CH₃CN/H₂O, 1:2.5). RP-18 HPLC fur-

nished 1a (14 mg, 0.015 mmol, 80 %) as colorless lyophilisate.

HPLC (prep. RP-18: 0–5 min isocratic 5% CH₃CN + 0.1% TFA,

5–60 min linear gradient 5–50% CH₃CN + 0.1% TFA, flow 10 mL/

1

min): t_R= 30.0 min. H NMR (600 MHz, [D₆]DMSO): δ = 0.82–

0.85 (m, 12 H, 4ϫ CH₃), 1.08 (m, 1 H, Ile-CHH), 1.43–1.56 (m, 2

6026

www.eurjoc.org

Eur. J. Org. Chem. 2007, 6016–6033

Carbohydrate-Based VEGF Inhibitors

H, Ile-CHH, Leu-β-H), 1.52 (m, 1 H, Leu-β-H), 1.60 (m, 1 H, Ile-

β-H), 1.68 [m, 1 H, Leu-CH(CH₃)₂], 2.92 (m, 1 H, Cys-β-H), 3.07

chromatography (silica gel, toluene/ethyl acetate, 10:1 Ǟ 5:1). To

the reaction mixture was added Fmoc-Val-OH (165 mg,

(m, 2 H, Cys-β-H, 2b-H), 3.17 (m, 1 H, Cys-β-H), 3.35–3.42 (m, 3 0.49 mmol), PyBOP (240 mg, 0.46 mmol) and DIPEA (ca. 150 µL)

H, 3b-H, 4b-H, Cys-β-H), 3.48 (m, 1 H, 5a-H), 3.63–3.67 (m, 5 H, in CH₂Cl₂(3 mL) at pH = 8. After stirring at room temperature

3

2a-H, 3a-H, 6a-H, 5b-H), 3.73 (dd, J_3,4= ³J_4,5= 7.4 Hz, 1 H, 4a-

for 2 h, it was diluted with ethyl acetate and washed with saturated

2

H), 3.89 (d, J_gem= 15 Hz, 1 H, CHHC=ONHTrt), 4.06–4.09 (m, NaCl solution. The organic phase was dried with MgSO₄, filtered,

3 H, CHHC=ONHTrt, Cys-α-H, Ile-α-H), 4.23–4.42 (m, 6 H, Leu-

concentrated and purified by flash chromatography to furnish tri-

peptide 26c (307 mg, 0.36 mmol, 85%) as colorless foam. TLC: R_f

= 0.60 (toluene/ethyl acetate, 3:1). [α]²_D⁰= –12.6 (c = 1, CHCl₃). ¹H

NMR (250 MHz, CDCl₃): δ = 0.87–0.91 (m, 12 H, 2ϫ Ile-CH₃,

2ϫ Val-CH₃), 1.21 (m, 1 H, Ile-CHH), 1.41 (s, 9 H, tert-butyl),

α-H, 2b-H, 2ϫ CH₂–CO), 4.47 (d, ³J_1,2= 7.2 Hz, 1 H, 1a-H), 4.58

3

(m, 1 H, Cys-α-H), 4.63 (d, J_1,2= 7.7 Hz, 1 H, 1b-H), 4.77, 5.20,

5.66 (3 br.s, 3 H, OH), 7.68 (s, 1 H, Ile-NH), 7.90 (s, 1 H, Leu-

NH), 8.36 (s, 1 H, Cys-NH), 8.45 (s, 1 H, Cys-NH), 8.58 (s, 1 H,

1 3

2 a - N H ) p p m .

C N M R ( 1 5 0 . 8 M H z , [ D ] - 1.42 (m, 1 H, Ile-CHH), 1.82 (m, 1 H, Ile-β-H), 2.09 (m, 1 H, Val-

6

DMSO, selected data): δ = 10.8, 15.1, 21.3, 23.1 (4 C, 4ϫ CH₃),

β-H), 2.49–2.63 (m, 2 H, Cys-β-H), 4.00 (m, 1 H, Cys-α-H), 4.12–

3

24.0 (1 C, Ile-β-C), 24.1 (1 C, Ile-CH₂), 36.4 [1 C, Leu-CH(CH₃)₂], 4.53 (m, 5 H, Val-α-H, Ile-α-H, Fmoc), 5.39 (d, J = 8.6 Hz, 1 H,

41.3 (1 C, Leu-β-C), 51.3 (1 C, Leu-α-C), 51.6 (1 C, Cys-α-C), 52.8 NH), 6.07 (d, ³J = 8.4 Hz, 1 H, NH), 6.39 (d, ³J = 8.4 Hz, 1 H,

(1 C, Ile-α-C), 54.3 (1 C, 2a-C), 56.8 (1 C, Cys-α-C), 59.7 (1 C, 6a-

C), 67.1, 67.9 (2 C, 2ϫ CH₂C=O), 68.7 (1 C, 2b-C), 72.4 (1 C, 3b-

NH), 7.15–7.45 (m, 19 H, phenyl, Fmoc), 7.68, 7.77 (2d, 4 H,

Fmoc) ppm. C₅₂H₅₉N₃O₆S (854.1): calcd. C 73.12, H 6.96, N 4.92;

C), 74.9 (2 C, 5b-C, 4b-C), 76.3 (1 C, 5a-C), 77.0 (1 C, 4a-C), 78.9 found C 72.74, H 7.12, N 5.54.

(1 C, 3a-C), 81.9 (1 C, 2b-C), 100.0 (1 C, 1a-C), 102.0 (1 C, 1b-C)

L

-Valyl-L-isoleucyl-S-(triphenylmethyl)-L-cysteine tert-Butyl Ester

ppm. MALDI-MS (pos. mode, matrix DHB, THF): m/z = 982 [M

(27c): Piperidine (2 mL) was added to a solution of 26c (300 mg,

0.35 mmol) in DMF (10 mL). After stirring at room temperature

for 2 h, the reaction mixture was concentrated in vacuo and puri-

fied by flash chromatography (silica gel, toluene/ethyl acetate, 5:1

Ǟ 2:1) to furnish deprotected tripeptide 27c (202 mg, 0.32 mmol

91%) as colorless foam. TLC: R_f= 0.10 (toluene/ethyl acetate, 3:1).

[α]²_D⁰= –15.1 (c = 1, CHCl₃). ¹H NMR (250 MHz, CDCl₃): δ =

0.78–0.99 (m, 12 H, 2ϫ Val-CH₃, 2ϫ Ile-CH₃), 1.12 (m, 1 H, Ile-

CHH), 1.41 (s, 9 H, tert-butyl), 1.43 (m, 1 H, Ile-CHH), 1.91 (m,

1 H, Ile-β-H), 2.32 (m, 1 H, Val-β-H), 2.56 (m, 2 H, Cys-β-H), 3.22

(d, 1 H, Val-α-H), 4.27 (dd, 1 H, Ile-α-H), 4.42 (m, 1 H, Cys-α-H),

6.32 (d, 1 H, NH), 7.14–7.39 (m, 15 H, phenyl), 7.86 (d, 1 H, NH)

ppm. C₃₇H₄₉N₃O₄S (631.9): calcd. C 70.33, H 7.82, N 6.65; found

C 70.45, H 7.68, N 6.78.

+ Na]⁺, 998 [M + K]⁺.

N-[(9-Fluorenyl)methoxycarbonyl]-

L-isoleucyl-L-isoleucyl-S-(tri-

phenylmethyl)- -cysteine tert-Butyl Ester (26b): Compound 25

L

(317 mg, 0.42 mmol) was deprotected with piperidine (1.5 mL) in

DMF (7 mL), the mixture concentrated in vacuo and purified by

column chromatography (silica gel, toluene/ethyl acetate, 10:1 Ǟ

5:1). To the reaction mixture were added Fmoc-Ile-OH (171 mg,

0.49 mmol), PyBOP (240 mg, 0.46 mmol) and DIPEA (ca. 150 µL)

in CH₂Cl₂(3 mL) at pH = 8. After stirring at room temperature

for 2 h, the mixture was diluted with ethyl acetate and washed with

saturated NaCl solution. The organic phase was dried with MgSO₄,

filtered, concentrated and purified by flash chromatography to fur-

nish tripeptide 26b (326 mg, 0.37 mmol, 89%) as fine colorless

foam. TLC: R_f= 0.60 (toluene/ethyl acetate, 3:1). [α]²_D⁰= –16.8 (c

Carbamoylmethyl [2-O-(Carboxymethyl)-β-

yl( -isoleucyl- -isoleucyl- -hemicystine)]-(1Ǟ4)-3-O-(carboxy-

methyl)-2-deoxy-2-[( -hemicystinyl)amido]-β- -glucopyranoside

(1b): As in the synthesis of 1a, for the preparation of 1b the hepta-

peptide mimetic was synthesized as intermediate from 27b and 23,

which after Fmoc deprotection, a new peptide coupling, cyclization

and final deprotection gave 1b. HPLC (prep. RP-18: 0–5 min iso-

cratic 5% CH₃CN + 0.1% TFA, 5–50 min linear gradient 5–50%

D-glucopyranosyluron-

1

= 0.5, CHCl₃). H NMR (250 MHz, CDCl₃): δ = 0.85–0.90 (m, 12

L

H, 4ϫ CH₃), 1.10, 1.15 (2 m, 2 H, Ile-CH₂), 1.41 (s, 9 H, tert-

butyl), 1.45 (m, 2 H, Ile-CH₂), 1.60 (m, 1 H, Ile-β-H), 1.83 (m, 1

H, Ile-β-H), 2.54, 2.60 (2 dd, 2 H, Cys-β-H), 4.04 (m, 1 H, Cys-α-

H), 4.18–4.46 (m, 5 H, Ile-α-H, Ile-α-H, Fmoc), 5.38 (d, ³J =

8.1 Hz, 1 H, NH), 6.07 (d, ³J = 7.3 Hz, 1 H, NH), 6.40 (d, ³J =

7.9 Hz, 1 H, NH), 7.15–7.42 (m, 19 H, phenyl, Fmoc), 7.57, 7.75

(2 d, 4 H, Fmoc) ppm. C₅₃H₆₁N₃O₆S (868.1): calcd. C 73.33, H

7.08, N 4.84; found C 73.00, H 6.99, N 5.17.

L

D

1

CH₃CN + 0.1% TFA, flow 10 mL/min): t_R= 33.6 min. H NMR

(600 MHz, [D₆]DMSO): δ = 0.77 (m, 3 H, Ile-CH₃), 0.82 (m, 6 H,

2ϫ Ile-CH₃), 0.87 (m, 3 H, Ile-CH₃), 0.99 (m, 1 H, Ile-CHH), 1.11

(m, 1 H, Ile-CHH), 1.38 (m, 1 H, Ile-CHH), 1.48 (m, 1 H, Ile-

CHH), 1.70 (m, 1 H, Ile-β-H), 1.75 (m, 1 H, Ile-β-H), 3.03 (m, 2

H, Cys-β-H), 3.10 (dd, ³J_1,2= ³J_2,3= 8.3 Hz, 1 H, 2b-H), 3.16 (dd,

L-Isoleucyl-L-isoleucyl-S-(triphenylmethyl)-L-cysteine tert-Butyl Es-

ter (27b): Piperidine (2 mL) was added to a solution of 26b

(300 mg, 0.34 mmol) in DMF (10 mL). After stirring at room tem-

perature for 2 h, the reaction mixture was concentrated in vacuo

and purified by column chromatography (silica gel, toluene/ethyl

acetate, 5:1 Ǟ 2:1) to furnish deprotected tripeptide 27b (212 mg,

0.33 mmol 95%) as colorless foam. TLC: R_f= 0.10 (toluene/ethyl

acetate, 3:1). [α]²_D⁰= –15.6 (c = 1, CHCl₃). ¹H NMR (250 MHz,

CDCl₃): δ = 0.85–0.95 (m, 12 H, 4ϫ Ile-CH₃), 1.02 (m, 2 H, Ile-

CH₂), 1.36, 1.47 (2 m, 2 H, Ile-CH₂), 1.40 (s, 9 H, tert-butyl), 1.97

(m, 2 H, 2ϫ Ile-β-H), 2.46 (m, 2 H, Cys-β-H), 3.26 (d, 1 H, Ile-α-

H), 4.26 (dd, 1 H, Ile-α-H), 4.39 (m, 1 H, Cys-α-H), 6.35 (d, 1

H, NH), 7.17–7.38 (m, 15 H, phenyl), 7.87 (d, 1 H, NH) ppm.

C₃₈H₅₁N₃O₄S (645.9): calcd. C 70.66, H 7.96, N 6.51; found C

69.94, H 7.81, N 6.92.

²J_gem= 14.1, J_vic= 4.1 Hz, 1 H, Cys-β-H), 3.30 (m, J_gem

14.1 Hz, 1 H, Cys-βЈ-H), 3.33–3.40 (m, 2 H, 3b-H, 4b-H), 3.47 (m,

=

3

2

3

1 H, 3a-H), 3.64 (m, 3 H, 2a-H, 5a-H, 6a-H), 3.68 (d, J_4,5

=

9.4 Hz, 1 H, 5b-H), 3.72 (dd, 1 H, 4a-H), 3.88 (d, ²J_gem= 15.1 Hz,

1 H, CHHC=ONH₂), 3.99–4.08 (m, 3 H, Cys-α-H, Ile-α-H,

CHHC=ONH₂), 4.27 (m, 3 H, Ile-α-H, CH₂C=O), 4.45 (m, 2 H,

3

CH₂C=O), 4.48 (br. d, J_1,2≈ 7.1 Hz, 1 H, 1a-H), 4.54 (dd, 1 H,

Cys-α-H), 4.62 (d, 1 H, J_1,2= 7.7 Hz, 1b-H), 6.88, 7.35 (2 br. s, 2

3

H, OH), 7.58 (br. s, 1 H, Ile-NH), 7.83 (br. s, 1 H, Ile-NH), 8.26

(br. s, 1 H, Cys-NH), 8.37 (br. s, 1 H, Cys-NH), 8.58 (br. s, 2a-NH)

ppm. ¹³C NMR (150.8 MHz, [D₆]DMSO, selected data): δ = 10.9,

11.1, 15.2, 15.4 (4 C, 4ϫ CH₃), 24.0, 24.4 (2 C, CH₂CH₃), 36.0,

37.3 (2 C, 2ϫ Ile-β-C), 40.1 (1 C, Cys-β-C), 40.6 (1 C, Cys-β-C),

51.7 (1 C, Cys-α-C), 52.9 (1 C, Cys-α-C), 54.3 (1 C, 2a-C), 56.5 (1

N-[(9-Fluorenyl)methoxycarbonyl]-

L-valyl-L-isoleucyl-S-(triphenyl-

methyl)- -cysteine tert-Butyl Ester (26c): Compound 25 (317 mg,

L

0.42 mmol) was deprotected with piperidine (1.5 mL) in DMF C, Ile-α-C), 57.3 (1 C, Ile-α-C), 59.4 (1 C, 6a-C), 67.0 (1 C,

(7 mL), the solution concentrated in vacuo and purified by column CH₂C=O), 67.6 (1 C, CH₂C=O), 68.7 (1 C, CH₂C=O), 72.7 (1 C,

Eur. J. Org. Chem. 2007, 6016–6033

www.eurjoc.org

6027

T. Haag, R. A. Hughes, G. Ritter, R. R. Schmidt

FULL PAPER

4b-C), 74.4 (1 C, 5b-C), 74.9 (1 C, 3b-C), 75.9 (1 C, 3a-C), 77.4 (1

βЈ-H), 3.47 (d, 2 H, Gly-α-H), 3.89 (m, 1 H, Cys-α-H), 7.07 (d,

C, 4a-C), 78.4 (1 C, 5a-C), 82.0 (1 C, 2b-C), 99.9 (1 C, 1a-C), 101.8 1 H, NH), 7.20 (m, 15 H, phenyl), 7.55 (br. s, 1 H, NH) ppm.

(1 C, 1b-C) ppm. MALDI-MS (pos. mode, matrix DHB, THF):

C₂₉H₃₁N₂NaO₅S·H₂O (560.6): calcd. C 62.13, H 5.72, N 5.00;

found C 62.29, H 5.72, N 5.04.

m/z = 960 [M + H]⁺, 982 [M + Na]⁺, 998 [M + K]⁺.

Carbamoylmethyl [2-O-(Carboxymethyl)-β-

yl( -valyl- -isoleucyl- -hemicystine)]-(1Ǟ4)-3-O-(carboxymethyl)-

2-deoxy-2-[( -hemicystinyl)amido]-β- -glucopyranoside (1c): As in

D

-glucopyranosyluron-

Carbamoylmethyl [2-O-(Carboxymethyl)-β-D-glucopyranosyluron-

L

yl(

L

-leucyl-

L

-isoleucyl-

L

-hemicysteine)]-(1Ǟ4)-3-O-(carboxymeth-

-glucopyranoside (2):

L

D

yl)-2-deoxy-[(

L

-hemicystinylglycyl)amido]-β-D

the synthesis of 1a, for the preparation of 1c the heptapeptide mi-

metic was synthesized as intermediate from 27c and 23, which after

Fmoc deprotection, a new peptide coupling, cyclization and final

deprotection gave 1c. HPLC (prep. RP-18: 0–5 min isocratic 6%

As in the synthesis of 1a, for the preparation of 2 the nonapeptide

mimetic was synthesized as intermediate from 32 and 28a, which

after cyclization and final deprotection gave 2 (45%). HPLC (prep.

RP-18: 0–5 min isocratic 6% CH₃CN + 0.1% TFA, 5–55 min linear

CH₃CN + 0.1% TFA, 5–55 min linear gradient 6–50% CH₃CN +

0.1% TFA, flow 10 mL/min): t_R= 29.6 min. H NMR (600 MHz, 31.1 min. H NMR (600 MHz, [D₆]DMSO): δ = 0.80 (m, 3 H, Ile-

[D₆]DMSO): δ = 0.77 (m, 3 H, Val-CH₃), 0.81 (m, 3 H, Ile-CH₃), CH₃), 0.83–0.87 (m, 9 H, Ile-CH₃, 2ϫ Leu-CH₃), 1.13 (m, 1 H,

gradient 6–50 % CH₃CN + 0.1 % TFA, flow 10 mL/min): t_R=

1

0.86 (m, 6 H, Val-CH₃, Ile-CH₃), 1.12 (m, 1 H, Ile-CHH), 1.48 (m,

Ile-CHH), 1.31 (m, 1 H, Leu-β-H), 1.44 (m, 2 H, Leu-βЈ-H, Ile-

1 H, Ile-CHH), 1.70 (m, 1 H, Ile-β-H), 2.02 (m, 1 H, Val-β-H),

3.03 (m, 2 H, Cys-β-H), 3.10 (dd, J_1,2= J_2,3= 8.9 Hz, 1 H, 2b-

H), 3.15 (dd, 1 H, Cys-β-H), 3.30 (dd, 1 H, Cys-β-H), 3.35–3.38

CHH), 1.54 [m, 1 H, Leu-CH(CH₃)₂], 1.72 (m, 1 H, Ile-β-H), 3.00

3

(m, 2 H, Cys-β-H), 3.07 (dd, J_1,2

=

³J_2,3= 8.2 Hz, 1 H, 2b-H),

3.14 (m, 1 H, Cys-β-H), 3.31–3.39 (m, 4 H, 5a-H, 3b-H, 4b-H, Cys-

(m, 2 H, 3b-H, 4b-H), 3.47 (dd, 1 H, 3a-H), 3.65–3.83 (m, 6 H, 2a- β-H), 3.65–3.77 (m, 7 H, 2a-H, 3a-H, 4a-H, 6a-H, 5b-H, Gly-α-H),

H, 4a-H, 5a-H, 6a-H, 5b-H), 3.88 (d, ³J = 15.4 Hz, 1 H,

CHHC=ONH₂), 3.99–4.08 (m, 3 H, Ile-α-H, Cys-α-H,

CHHC=ONH₂), 4.27 (m, 3 H, Val-α-H, CH₂C=O), 4.47–4.52 (m,

3.88 (d, 1 H, CHHC=O), 3.99–4.01 (m, 2 H, Cys-α-H, Gly-α-H),

4.04–4.07 (m, 2 H, Ile-α-H, CHHC=O), 4.12 (d, 1 H, CHHC=O),

4.24 (m, 2 H, CH₂C=O), 4.26 (d, 1 H, CHHC=O), 4.45 (m, 2 H,

3

3 H, CH₂C=O, 1a-H), 4.55 (dd, 1 H, Cys-α-H), 4.62 (d, ³J = Cys-α-H, 1a-H), 4.55 (m, 1 H, Leu-α-H), 4.66 (d, J_1,2= 7.7 Hz, 1

7.7 Hz, 1 H, 1b-H), 6.87, 7.36 (2 s, 2 H, OH), 7.55 (br. s, 1 H, Val- H, 1b-H), 7.25 (d, ³J = 7.0 Hz, 1 H, Leu-NH), 7.83 (d, ³J = 7.2 Hz,

3

NH), 7.83 (br. s, 1 H, Ile-NH), 8.24 (br. s, 1 H, Cys-NH), 8.37 (br.

1 H, 2a-NH), 8.15 (d, J = 6.8 Hz, 1 H, Ile-NH), 8.39 (br. s, 1 H,

s, 3 H, Cys-NH, C=ONH₂), 8.57 (br. s, 1 H, 2a-NH) ppm. ¹³C Cys-NH), 8.57 (d, J = 7.6 Hz, 1 H, Cys-NH), 8.72 (m, 1 H, Gly-

3

NMR (150.8 MHz, [D₆]DMSO, selected data): δ = 10.9, 15.2 (2 C,

2ϫ Ile-CH₃), 17.8, 19.2 (2 C, 2ϫ Val-CH₃), 24.3 (1 C, CH₂CH₃),

31.2 (1 C, Val-β-C), 35.8 (1 C, Ile-β-C), 40.1 (1 C, Cys-β-C), 40.4

(1 C, Cys-β-C), 51.8 (1 C, Cys-α-C), 52.9 (1 C, Cys-α-C), 54.4 (1

C, 2a-C), 57.0 (1 C, Val-α-C), 57.3 (1 C, Ile-α-C), 59.4 (1 C, 6a-C),

NH) ppm. ¹³C NMR (150.8 MHz, [D₆]DMSO, selected data): δ =

10.9, 15.2 (2 C, 2ϫ Ile-CH₃), 21.8, 22.9 (2 C, 2ϫ Leu-CH₃), 24.3

[2 C, CH₂CH₃, CH(CH₃)₂], 36.0 (1 C, Ile-β-C), 38.9 (1 C, Cys-β-

C), 40.1 (1 C, Cys-β-C), 41.6 (1 C, Leu-β-C), 42.2 (1 C, Gly-α-C),

49.8 (1 C, Leu-α-C), 50.9 (1 C, Cys-α-C), 51.2 (1 C, Cys-α-C), 53.7

67.6, 68.8, 70.1 (3 C, 3ϫ CH₂C=O), 72.8 (1 C, 4b-C), 74.6 (1 C, (1 C, 2a-C), 57.2 (1 C, Ile-α-C), 59.4 (1 C, 6a-C), 67.3, 68.1, 68.9

5b-C), 74.8 (1 C, 3b-C), 75.8 (1 C, 3a-C), 77.5 (1 C, 4a-C), 78.2 (1

C, 5a-C), 81.9 (1 C, 2b-C), 100.1 (1 C, 1a-C), 102.0 (1 C, 1b-C)

ppm. MALDI-MS (pos. mode, matrix DHB, THF): m/z = 967 [M

+ Na]⁺, 984 [M + K]⁺.

(3 C, 3ϫ CH₂C=O), 72.3 (1 C, 4b-C), 73.1 (1 C, 5b-C), 74.9 (2 C,

5a-C, 3b-C), 76.5 (1 C, 4a-C), 80.2 (1 C, 3a-C), 82.3 (1 C, 2b-C),

100.4 (1 C, 1a-C), 101.2 (1 C, 1b-C) ppm. MALDI-MS (pos. mode,

matrix DHB, THF): m/z = 1039 [M + Na]⁺, 1055 [M + K]⁺.

N-(tert-Butyloxycarbonyl)-S-(triphenylmethyl)-

Methyl Ester (31): DIPEA (ca. 0.2 mL) was added to a solution

of Boc-Cys(Trt)-OH (230 mg, 0.50 mmol), NH₂-Gly-OMe (69 mg, (3.0 g, 25.8 mmol) and dicyclohexylcarbodiimide (5.3 g,

L

-cysteinylglycine

Dimethyl(thexyl)silyl 4,6-O-Benzylidene-2-deoxy-2-(dimethylmale-

imido)-3-O-levulinoyl-β-D-glucopyranoside (35): Levulinic acid

0.55 mmol) and PyBOP (301 mg, 0.58 mmol) in dry CH₂Cl₂until

pH = 8 was reached. The reaction mixture was stirred at room

temperature for 2 h, diluted with ethyl acetate and washed twice

with saturated NaCl solution. The organic phase was dried with

MgSO₄, filtered and concentrated. Purification by column

chromatography (silica gel, toluene/ethyl acetate, 3.5:1) furnished

31 (265 mg, 49 mmol, 98%) as colorless powder. TLC: R_f= 0.25

(toluene/ethyl acetate, 3:1). [α]²_D⁰= –15.3 (c = 0.5, CHCl₃). ¹H NMR

25.8 mmol) were added to a solution of 9^[29](6.0 g, 11.5 mmol) in

dry CH₂Cl₂(60 mL). After the addition of catalytic amounts of

DMAP, the turbid solution was stirred at room temperature for

2 h. The precipitated urea was filtered off, and the filtrate washed

with saturated NaHCO₃solution. The organic phase was dried

with MgSO₄, filtered and concentrated. Purification by flash

chromatography (silica gel, petroleum ether/ethyl acetate, 4.5:1 Ǟ

3:1) furnished 35 (6.3 g, 10.2 mmol 89%) as colorless powder. TLC:

R_f= 0.48 (petroleum ether/ethyl acetate, 2:1). [α]²_D⁰= –2.5 (c = 1,

CHCl₃). ¹H NMR (250 MHz, CDCl₃): δ = 0.01 (s, 3 H, SiCH₃),

0.08 (s, 3 H, SiCH₃), 0.68–0.75 (m, 12 H, 4ϫ CH₃), 1.47 [m, 1 H,

2

(250 MHz, CDCl₃): δ = 1.37 (s, 9 H, Boc), 2.50 (dd, J_gem= 12.9,

³J_vic= 5.1 Hz, 1 H, Cys-β-H), 2.67 (dd, 1 H, Cys-βЈ-H), 3.67 (s, 3

H, OCH₃), 3.83 (m, 1 H, Cys-α-H), 3.92 (m, 2 H, Gly-α-H), 4.72

(br. d, 1 H, NH), 6.49 (br. s, 1 H, NH), 7.13–7.40 (m, 15 H, phenyl) CH(CH₃)₂], 1.93 (s, 6 H, DMM-H), 2.04 (s, 3 H, Lev-C=OCH₃),

ppm. C₃₀H₃₄N₂O₅S (534.7): calcd. C 67.39, H 6.41, N 5.24; found

C 67.09, H 6.16, N 5.16.

2.39–2.65 (m, 4 H, 2ϫ Lev-CH₂), 3.62–3.69 (m, 3 H, 3-H, 4-H, 5-

H), 3.78 (dd, ³J_1,2≈ ³J_2,3≈ 9.9 Hz, 1 H, 2-H), 3.98 (dd, ²J_gem= 10.5,

2

3

³J_vic= 8.0 Hz, 1 H, 6-H), 4.30 (dd, J_gem= 10.4, J_vic= 4.3 Hz, 1

N-(tert-Butyloxycarbonyl)-S-(triphenylmethyl)-L-cysteinylglycine

3

H, 6Ј-H), 5.43 (d, J_1,2= 8.0 Hz, 1 H, 1-H), 5.47 (s, 1 H,

(32): NaOH (100 mg, 2.5 mmol) was added to a solution of 31

(250 mg, 0.46 mmol), dioxane (20 mL), MeOH (7 mL) and H₂O

(7 mL), stirred overnight and then neutralized with ion exchange

resin IR 120 (H⁺), filtered and concentrated to furnish 32 (235 mg,

0.45 mmol, 99%) which was immediately used in the next step with-

out further purification. TLC: R_f= 0.04 (toluene/ethyl acetate, 3:1).

3

CHphenyl), 4.24 (dd, J_2,3= 10.5, J_3,4= 8.9 Hz, 1 H, 3-H), 7.32

(m, 3 H, phenyl), 7.41 (m, 2 H, phenyl) ppm. C₃₂H₄₅NO₉Si (615.8):

calcd. C 62.41, H 7.37, N 2.27; found C 62.36, H 7.43, N 2.47.

Dimethyl(thexyl)silyl 6-O-Benzyl-2-deoxy-2-(dimethylmaleimido)-3-

O-levulinoyl-β-D-glucopyranoside (36): Triethylsilane (13 mL) was

added to a solution of 35 (6.0 g, 9.74 mmol) in dry CH₂Cl₂

(50 mL). The reaction mixture was then cooled to 0 °C, trifluoro-

1

[α]²_D⁰= +10.5 (c = 0.5, CHCl₃). H NMR (250 MHz, [D₆]DMSO):

δ = 1.37 (s, 9 H, Boc), 2.36 (dd, 1 H, Cys-β-H), 2.48 (m, 1 H, Cys-

6028

www.eurjoc.org

Eur. J. Org. Chem. 2007, 6016–6033

Carbohydrate-Based VEGF Inhibitors

2

acetic anhydride (3.5 mL) and trifluoroacetic acid (3.2 mL) were

added, and after stirring at 0 °C for 4 h, the mixture was poured

into a cold saturated NaHCO₃solution. The mixture was then di-

luted with ethyl acetate and washed twice with NaHCO₃solution.

The organic phase was dried with MgSO₄, filtered, and concen-

trated in vacuo. Purification by flash chromatography (silica gel,

toluene/acetone, 8:1) furnished 36 (4.7 g, 7.61 mmol, 78%) as col-

orless oil. TLC: R_f= 0.39 (toluene/acetone, 8:1). [α]²_D⁰= –8.9 (c =

1, CHCl₃). ¹H NMR (250 MHz, CDCl₃): δ = 0.01 (s, 3 H, SiCH₃),

0.11 (s, 3 H, SiCH₃), 0.67–0.74 (m, 12 H, 4ϫ CH₃), 1.46 [m, 1 H,

CH(CH₃)₂], 1.91 (s, 6 H, DMM-H), 2.09 (s, 3 H, Lev-CH₃), 2.44

= 6.2 Hz, 1 H, 6aЈ-H), 4.25 (m, J_gem= 11.9 Hz, 2 H, 6b-H), 4.39

(d, ³J_1,2= 8.1 Hz, 1 H, 1b-H), 4.49–4.80 (m, 6 H, 3ϫ CH₂Ph), 4.95

(dd, ³J_1,2≈ ³J_2,3≈ 8.4 Hz, 1 H, 2b-H), 5.21 (d, J_1,2= 8.1 Hz, 1 H,

3

1a-H), 7.20–7.25 (m, 15 H, phenyl) ppm. C₅₁H₆₇NO₁₄Si (946.16):

calcd. C 64.74, H 7.14, N 1.48; found C 64.66, H 7.17, N 1.47.

Dimethyl(thexyl)silyl 2,6-Di-O-acetyl-3,4-di-O-benzyl-β-

anosyl-(1Ǟ4)-6-O-benzyl-2-deoxy-2-(dimethylmaleimido)-β-

D-glucopyr-

D

-allo-

pyranoside (39): Dess–Martin periodinane reagent (1.40 g,

3.30 mmol) was added to a solution of 38 (1.10 g, 1.16 mmol) in

CH₂Cl₂(10 mL), the reaction mixture stirred at room temperature

for 3 h, then sodium sulfate solution (8 mL) and NaHCO₃solution

(8 mL) were added, and the mixture was stirred for 30 min. The

phases were separated, the aqueous phase was extracted twice with

CHCl₃, and the combined organic phases were dried with MgSO₄,

filtered, and concentrated in vacuo. The residue (1.15 g, quant.),

the oxidized intermediate, was diluted in CH₂Cl₂(9 mL) and

MeOH (9 mL) and the mixture cooled to –10 °C. Under vigorous

stirring, NaBH₄(37.4 mg, 0.99 mmol) was added in one portion

D-gluco- and the reaction mixture stirred for 210 s. Then acetone (2 mL) and

3

(m, 2 H, Lev-CH₂), 2.68 (m, 2 H, Lev-CH₂), 3.12 (d, J = 2.4 Hz,

1 H, OH), 3.61–3.69 (m, 2 H, 4-H, 5-H), 3.70–3.77 (m, 2 H, 6-H),

3

3.94 (dd, J_1,2= 8.0, J_2,3= 10.9 Hz, 1 H, 2-H), 4.59 (s, 2 H,

3

CH₂Ph), 5.33 (d, J_1,2= 8.0 Hz, 1 H, 1-H), 5.54 (dd, J_2,3= 10.9,

³J_3,4= 8.3 Hz, 1 H, 3-H), 7.26–7.33 (m, 5 H, phenyl) ppm.

C

₃₂H₄₁NO₉Si (617.8): calcd. C 62.21, H 7.67, N 2.27; found C

61.78, H 7.49, N 2.51.

Dimethyl(thexyl)silyl 2,6-Di-O-acetyl-3,4-di-O-benzyl-β-

pyranosyl-(1Ǟ4)-6-O-benzyl-2-deoxy-2-(dimethylmaleimido)-3-O-le- NH₄Cl solution were added, and the mixture was stirred for

vulinoyl-β-

D

-glucopyranoside (37): TMSOTf (0.1  in CH₂Cl₂,

10 min. After dilution with H₂O, the reaction mixture was ex-

tracted three times with CH₂Cl₂, and the combined organic phases

4 mL) was added to a mixture of 8 (2.90 g, 4.94 mmol) and 36

(2.78 g, 4.5 mmol) in dry CH₂Cl₂(30 mL) at room temperature. were dried with MgSO₄, filtered and concentrated. Purification by

After 1.5 h, the reaction mixture was neutralized with NEt₃and

concentrated. Purification by column chromatography (silica gel,

toluene/ethyl acetate, 7:1) furnished 37 (3.21 g, 3.07 mmol, 68%) as

column chromatography (silica gel, petroleum ether/ethyl acetate,

3.5:1) furnished, beside some reactant 38 (105 mg, 0.11 mmol, 9%),

the epimerized allose building block 39 (750 mg, 0.79 mmol, 68%)

as white amorphous solid. TLC: R_f= 0.41 (petroleum ether/ethyl

acetate, 2:1). [α]²_D⁰= –6.1 (c = 1, CHCl₃). ¹H NMR (600 MHz,

CDCl₃): δ = 0.08 (s, 3 H, SiCH₃), 0.16 (s, 3 H, SiCH₃), 0.71–0.78

(m, 12 H, 4ϫ CH₃), 1.50 [m, 1 H, CH(CH₃)₂], 1.89 (s, 3 H, acetyl),

colorless foam. TLC: R_f= 0.45 (toluene/ethyl acetate, 3:1). [α]²_D⁰

=

1

+6.3 (c = 1, CHCl₃). H NMR (600 MHz, CDCl₃): δ = 0.02 (s, 3

H, SiCH₃), 0.12 (s, 3 H, SiCH₃), 0.70–0.76 (m, 12 H, 4ϫ CH₃),

1.49 [m, 1 H, CH(CH₃)₂], 1.89–1.95 (m, 9 H, DMM-H, acetyl),

2.05, 2.07 (2 s, 6 H, acetyl, Lev-C=OCH₃), 2.35, 2.40, 2.50, 2.60 (4

m, 4 H, Lev-CH₂), 3.43 (m, 1 H, 5b-H), 3.53–3.56 (m, 3 H, 5a-H,

3b-H, 4b-H), 3.65 (dd, ²J_gem= 10.5 Hz, 1 H, 6a-H), 3.76 (dd, ²J_gem

1.94 (s, 3 H, acetyl), 1.95 (s, 6 H, DMM-H), 3.52–3.55 (m, 2 H,

2

4b-H, 5b-H), 3.60–3.68 (m, 2 H, 3b-H, 6a-H), 3.67 (dd, J_gem

=

3

11.2, J_vic= 4.9 Hz, 1 H, 6aЈ-H), 3.86 (dd, J_3,4= 2.9, J_4,5

3

= 11.1, J_vic= 3 Hz, 1 H, 6aЈ-H), 3.89 (dd, 1 H, J_3,4

≈

³J_4,5

≈

9.8 Hz, 1 H, 4a-H), 3.92 (dd, J_1,2= 8.2, J_2,3= 2.8 Hz, 1 H, 2a-

H), 3.99 (m, 1 H, 5a-H), 4.05 (dd, J_gem= 11.8, J_vic= 5.2 Hz, 1

3

2

3

9.5 Hz, 4a-H), 3.93 (dd, 1 H, J_1,2= 8.1, J_2,3= 10.8 Hz, 2a-H),

4.19–4.22 (m, 2 H, 6b-H), 4.46 (d, 1 H, ³J_1,2= 8.0 Hz, 1b-H), 4.50,

4.52, 4.62 (3 d, 3 H, CH₂Ph, CHHPh), 4.71–4.78 (m, 3 H, CH₂Ph,

3

H, 6b-H), 4.21 (dd, J_2,3= 2.8, J_3,4= 2.9 Hz, 3a-H), 4.27 (dd,

²J_gem= 11.8, J_vic= 1.7 Hz, 1 H, 6bЈ-H), 4.48 (d, J_1,2= 8.2 Hz, 1

3

CHHPh), 4.87 (dd, J_1,2≈ J_2,3≈ 8.5 Hz, 2b-H), 5.35 (d, 1 H, J_1,2

H, 1b-H), 4.54. 4.55, 4.65, 4.66, 4.79, 4.81 (6 d, 6 H, 3ϫ CH₂Ph),

3

= 8.1 Hz, 1a-H), 5.57 (dd, 1 H, J_2,3= 10.7, J_3,4= 9.2 Hz, 3a-H), 4.99 (dd, J_1,2= 7.9, J_2,3= 9.7 Hz, 1 H, 2b-H), 5.86 (d, J_1,2

=

7.23–7.37 (m, 15 H, phenyl) ppm. ¹³C NMR (150.8 MHz, CDCl₃,

selected data): δ = 56.5 (1 C, 2a-C), 63.0 (1 C, 6b-C), 67.6 (1 C,

6a-C), 70.5 (1 C, 3a-C), 72.8 (1 C, 5b-C), 72.9 (1 C, 2b-C), 74.6 (1

8.2 Hz, 1 H, 1a-H), 7.24–7.35 (m, 15 H, phenyl) ppm. ¹³C NMR

(150.8 MHz, CDCl₃, selected data): δ = 57.3 (1 C, 2a-C), 63.0 (1

C, 6b-C), 68.9 (1 C, 6a-C), 70.3 (1 C, 3a-C), 72.1 (1 C, 5a-C), 72.9

C, 5a-C), 75.7 (1 C, 4a-C), 77.2 (1 C, 3b-C), 83.2 (1 C, 4b-C), 93.1 (1 C, 2b-C), 73.2 (1 C, 4b-C), 77.2 (1 C, 4a-C), 77.5 (1 C, 5b-C),

(1 C, 1a-C), 100.3 (1 C, 1b-C) ppm. C₅₆H₇₉NO₁₆Si (1044.3): calcd.

C 64.41, H 7.05, N 1.34; found C 64.33, H 7.12, N 1.44.

82.9 (1 C, 3b-C), 91.2 (1 C, 1a-C), 101.3 (1 C, 1b-C) ppm. MALDI-

MS (pos. mode, matrix DHB, THF): m/z = 969 [M + Na]⁺.

C₅₁H₆₇NO₁₄Si (946.2): calcd. C 64.74, H 7.14, N 1.48; found C

64.52, H 6.92, N 1.33.

Dimethyl(thexyl)silyl 2,6-Di-O-acetyl-3,4-di-O-benzyl-β-

D

-gluco-

-glu-

pyranosyl-(1Ǟ4)-6-O-benzyl-2-deoxy-2-(dimethylmaleimido)-β-

D

copyranoside (38): Acetic acid (29 mL) was added to a solution of

37 (4.29 g, 4.11 mmol) in pyridine (45 mL), the mixture cooled to

0 °C and hydrazine hydrate (3.25 mL, 3.35 g, 66.9 mmol) added

dropwise. After stirring at 0 °C for 9 min, the reaction mixture was

diluted with CH₂Cl₂and the organic phase washed with saturated

NaHCO₃solution and NaCl solution. The organic phase was dried

with MgSO₄, filtered, concentrated and coevaporated twice with

toluene. Purification by column chromatography (silica gel, tolu-

ene/acetone, 7:1) furnished 38 (3.83 g, 4.04 mmol, 98%) as colorless

Dimethyl(thexyl)silyl 2,6-Di-O-acetyl-3,4-di-O-benzyl-β-

pyranosyl-(1Ǟ4)-3-O-acetyl-6-O-benzyl-2-deoxy-2-(dimethylmale-

imido)-β- -allopyranoside (40): Acetic acid anhydride (10 mL) was

D-gluco-

D

added to a solution of 39 (1.00 g, 1.06 mmol) in pyridine (20 mL)

at 0 °C. After stirring at room temperature overnight, the reaction

mixture was concentrated in vacuo and coevaporated three times

with toluene. Purification by flash chromatography furnished acet-

ylated compound 40 (1.01 g, 1.02 mmol, 97%) as colorless foam.

TLC: R_f= 0.49 (petroleum ether/ethyl acetate, 2:1). [α]²_D⁰= +22.3

foam. TLC: R_f= 0.68 (petroleum ether/ethyl acetate, 1:1). [α]²_D⁰

=

(c = 0.5, CHCl₃). H NMR (600 MHz, CDCl₃): δ = 0.04 (s, 3 H,

1

+12.0 (c = 1, CHCl₃). H NMR (250 MHz, CDCl₃): δ = 0.01 (s, 3

H, SiCH₃), 0.11 (s, 3 H, SiCH₃), 0.69–0.75 (m, 12 H, 4ϫ CH₃),

1.48 [m, 1 H, CH(CH₃)₂], 1.88 (s, 3 H, acetyl), 1.89 (s, 6 H, DMM-

SiCH₃), 0.12 (s, 3 H, SiCH₃), 0.66–0.72 (m, 12 H, 4ϫ CH₃), 1.45

[m, 1 H, CH(CH₃)₂], 1.82 (d, 6 H, DMM-H), 1.88, 1.94, 1.97 (3 s,

3

9 H, 3ϫ acetyl), 3.42 (m, 1 H, 5b-H), 3.49 (dd, J_3,4

=

³J_4,5

=

2

H), 1.96 (s, 3 H, acetyl), 3.50–3.64 (m, 7 H, 3a-H, 4a-H, 5a-H, 6a- 9.3 Hz, 1 H, 4b-H), 3.58 (m, 2 H, 3b-H, 6a-H), 3.63 (dd, J_gem

3

H, 3b-H, 4b-H, 5b-H), 3.84 (dd, J_1,2= 8.2, J_2,3= 10.9 Hz, 1 H,

11.2, J_vic= 3.6 Hz, 1 H, 6aЈ-H), 3.87 (dd, J_3,4= 3.0, J_4,5

9.8 Hz, 1 H, 4a-H), 3.93 (dd, J_2,3= 2.2 Hz, 1 H, 2a-H), 3.94 (m,

3

2

3

2a-H), 3.91 (d, J = 1.2 Hz, 1 H, OH), 4.04 (dd, J_gem= 11.9, J_vic

Eur. J. Org. Chem. 2007, 6016–6033

www.eurjoc.org

6029

T. Haag, R. A. Hughes, G. Ritter, R. R. Schmidt

1 H, 5a-H), 4.09 (dd, ²J_gem= 11.7, J_vic= 4.8 Hz, 1 H, 6b-H), 4.18 (dimethylmaleimido)-β-

-allopyranoside (43): A solution of 42

FULL PAPER

3

D

2

3

(dd, J_gem= 11.7, J_vic= 1.3 Hz, 1 H, 6bЈ-H), 4.39 (d, J_1,2

=

(2.00 g, 2.02 mmol) and 17 (0.80 g, 2.52 mmol) in CH₂Cl₂(25 mL)

was cooled to –5 °C, then tin triflate (5 mg, 0.01 mmol) was added

and the mixture stirred at room temperature for 1 h. The reaction

7.9 Hz, 1 H, 1b-H), 4.48, 4.52, 4.61, 4.63, 4.73, 4.74 (6 d, 6 H, 3ϫ

CH₂Ph), 4.89 (dd, ³J_1,2= 8.2, J_2,3= 9.2 Hz, 1 H, 2b-H), 5.47 (dd,

3

1 H, 3a-H), 5.24 (d, J_1,2= 8.2 Hz, 1 H, 1a-H), 7.21–7.31 (m, 15 mixture was neutralized with NEt₃and concentrated. Purification

H, phenyl) ppm. ¹³C NMR (150.8 MHz, CDCl₃, selected data): δ

= 56.0 (1 C, 2a-C), 62.7 (1 C, 6b-C), 68.6 (1 C, 6a-C), 70.7 (1 C,

3a-C), 72.8 (1 C, 2b-C), 72.9 (1 C, 5b-C), 73.1 (1 C, 5a-C), 74.4 (1

by column chromatography (toluene/ethyl acetate, 15:1 Ǟ 14:1)

furnished, beside some elimination product 43Ј, compound 43

(1.47 g, 1.28 mmol, 64%) as colorless foam. TLC: R_f= 0.68 (petro-

C, 4a-C), 77.3 (1 C, 4b-C), 82.8 (1 C, 3b-C), 91.4 (1 C, 1a-C), 101.2 leum ether/ethyl acetate, 1:1). [α]²_D⁰= +21.5 (c = 0.5, CHCl₃). ¹H

(1 C, 1b-C) ppm. C₅₃H₆₉NO₁₅Si (988.2): calcd. C 64.42, H 7.04, N NMR (250 MHz, CDCl₃): δ = 1.81 (s, 6 H, DMM-H), 1.96 (s, 3

1.42; found C 64.28, H 7.10, N 1.39.

2,6-Di-O-acetyl-3,4-di-O-benzyl-β- -glucopyranosyl-(1Ǟ4)-3-O-ace-

tyl-6-O-benzyl-2-deoxy-2-(dimethylmaleimido)-α,β- -allopyranose

H, acetyl), 2.03 (s, 3 H, acetyl), 3.35–3.56 (m, 6 H, 5a-H, 6a-H, 6aЈ-

H, 3b-H, 4b-H, 5b-H), 3.98–4.12 (m, 3 H, 2a-H, CHHC=ONHTrt,

C H H P h ) , 4 . 2 0 – 4 . 2 6 ( m , 4 H , 4 a - H , 6 b - H , 6 b Ј - H ,

D

3

CHHC=ONHTrt), 4.35 (d, 1 H, CHHPh), 4.42 (d, J_1,2= 7.9 Hz,

(41): Acetic acid (135 µL, 141 mg, 2.36 mmol) was added to a solu-

tion of 40 (2.20 g, 2.22 mmol) in tetrahydrofuran (40 mL) and the

reaction mixture cooled to 0 °C. After addition of a TBAF solution

(1  in THF, 2.64 mL, 2.64 mmol) and stirring at 0 °C for 3 h, the

reaction mixture was diluted with diethyl ether and washed with

NaCl solution. The organic phase was dried with MgSO₄, filtered

and concentrated. Purification by flash chromatography (silica gel,

petroleum ether/ethyl acetate, 3:2) furnished anomerically depro-

tected 41 (1.75 g, 2.07 mmol, 93%) with an anomer ratio of α/β Ͼ

10:1 as colorless foam. TLC: R_f= 0.25 (petroleum ether/ethyl ace-

tate, 1:1). [α]²_D⁰= +21.6 (c = 1, CHCl₃). ¹H NMR (600 MHz,

CDCl₃): δ = 1.77 (s, 3 H, acetyl), 1.85 (s, 6 H, DMM-H), 1.93 (s,

1 H, 1b-H), 4.51 (d, 1 H, CHHPh), 4.62 (d, 1 H, CHHPh), 4.78

3

(m, 2 H, CH₂Ph), 4.84 (dd, J_1,2≈ J_2,3≈ 8.8 Hz, 2b-H), 5.53 (dd,

3

1 H, 3a-H), 5.73 (d, J_1,2= 8.7 Hz, 1 H, 1a-H), 7.09–7.36 (m, 15

H, phenyl), 7.71 (s, 1 H, C=ONHTrt) ppm. MALDI-MS (pos.

mode, matrix DHB, THF): m/z = 1168 [M + Na]⁺, 1184

[M + K]⁺. C₆₆H₆₈N₂O₁₆(1145.3): calcd. C 69.22, H 5.98, N 2.45;

found C 69.25, H 5.86, N 2.68.

2,6-Di-O-acetyl-3,4-di-O-benzyl-β-

D-glucopyranosyl-(1Ǟ4)-3-O-

acetyl-1,5-anhydro-6-O-benzyl-2-deoxy-2-(dimethylmaleimido)-

D

-

arabino-hex-1-enitol (43Ј): Colourless foam. TLC: R_f= 0.56 (tolu-

ene/ethyl acetate, 3:2). [α]²_D⁰= +122.4 (c = 1, CHCl₃). ¹H NMR

(600 MHz, CDCl₃): δ = 1.85 (s, 3 H, acetyl), 1.95 (s, 6 H, DMM-

H), 2.02 (s, 3 H, acetyl), 3.49 (m, 1 H, 5b-H), 3.52 (dd, ³J_3,4≈ ³J_4,5

3

3 H, acetyl), 1.99 (s, 3 H, acetyl), 3.37 (d, J = 6.0 Hz, 1 H, OH),

3

3.41 (m, 1 H, 5b-H), 3.49 (dd, J_3,4= J_4,5= 9.2 Hz, 1 H, 4b-H),

3

=

³J_3,4= 9.1 Hz, 1 H, 3b-H), 3.60 (m, 2 H, 6a-H,

3

3.54 (dd, J_2,3

≈ 9.1 Hz, 4b-H), 3.61 (dd, J_2,3≈ J_3,4≈ 9.0 Hz, 3b-H), 3.77 (m, 2

3

2

3

6aЈ-H), 3.83 (dd, J_3,4= 3.0, J_4,5= 9.8 Hz, 1 H, 4a-H), 3.93 (dd,

H, 6a-H), 4.11 (dd, J_gem= 11.7, J_vic= 4.7 Hz, 6b-H), 4.21 (dd,

²J_gem= 11.7 Hz, 6bЈ-H), 4.25 (m, 2 H, 4a-H, 5a-H), 4.53 (m, 3 H,

1b-H, CH₂Ph), 4.64 (m, 2 H, CH₂Ph), 4.78 (m, 2 H, CH₂Ph), 4.96

³J_1,2= 8.6, J_2,3= 2.2 Hz, 1 H, 2a-H), 4.02 (m, 1 H, 5a-H), 4.05

3

(dd, ²J_gem= 11.8, ³J_vic= 5.0 Hz, 1 H, 6b-H), 4.18 (dd, ²J_gem= 11.8,

³J_vic= 1.1 Hz, 1 H, 6aЈ-H), 4.35 (d, J_1,2= 7.9 Hz, 1 H, 1b-H),

3

(dd, J_1,2≈ J_2,3≈ 8.6 Hz, 2b-H), 5.25 (d, J_3,4= 2.2 Hz, 1 H, 3a-

H), 6.59 (s, 1 H, 1a-H), 7.25–7.36 (m, 15 H, phenyl) ppm. ¹³C

NMR (150.8 MHz, CDCl₃, selected data): δ = 62.9 (1 C, 6b-C),

65.5 (1 C, 3a-C), 68.0 (1 C, 6a-C), 72.9 (1 C, 5b-C), 73.0, 73.3 73.7

(3 C, 4a-C, 5a-C, 2b-C), 77.4 (1 C, 4b-C), 82.9 (1 C, 3b-C), 101.4

(1 C, 1b-C), 105.8 (1 C, 2a-C), 149.3 (1 C, 1a-C) ppm. MALDI-

MS (pos. mode, matrix DHB, THF): m/z = 850 [M + Na]⁺, 866

[M + K]⁺. C₄₅H₄₉NO₁₄·0.5H₂O (836.9): calcd. C 64.58, H 6.02, N

1.67; found C 64.47, H 6.12, N 1.71.

4.46, 4.48, 4.59, 4.61, 4.73, 4.74 (6d, 6 H, 3ϫ CH₂Ph), 4.85 (dd,

3

³J_1,2= 7.9, J_2,3= 9.1 Hz, 1 H, 2b-H), 5.49 (dd, 1 H, 3a-H), 5.91

3

(dd, J_1,2= 8.3, J_1,OH= 6.1 Hz, 1 H 1a-H), 7.21–7.30 (m, 15 H,

phenyl) ppm. ¹³C NMR (150.8 MHz, CDCl₃, selected data): δ =

55.5 (1 C, 2a-C), 62.8 (1 C, 6b-C), 68.8 (1 C, 6a-C), 70.8 (1 C, 3a-

C), 72.9 (1 C, 5b-C), 73.0 (1 C, 2b-C), 73.2 (1 C, 5a-C), 74.2 (1 C,

4a-C), 77.2 (1 C, 4b-C), 83.0 (1 C, 3b-C), 90.4 (1 C, 1a-C), 101.0

(1 C, 1b-C) ppm. C₄₅H₅₁NO₁₅(845.88): calcd. C 63.90, H 6.08, N

1.66; found C 63.95, H 6.19, N 1.58.

[N-(Triphenylmethyl)carbamoyl]methyl 3,4-Di-O-benzyl-β-

D-gluco-

Trichloro-O-[2,6-Di-O-acetyl-3,4-di-O-benzyl-β-

(1Ǟ4)-3-O-acetyl-6-O-benzyl-2-deoxy-2-(dimethylmaleimido)-β-

D

-glucopyranosyl-

-al-

pyranosyl-(1Ǟ4)-6-O-benzyl-2-deoxy-2-(dimethylmaleimido)-β-

D-al-

D

lopyranoside (44): Sodium methoxide (20 mg, 0.37 mmol) was

added to a solution of 43 (1.40 g, 1.22 mmol) in dry MeOH

(40 mL) and the mixture stirred overnight. The reaction mixture

was neutralized with ion exchange resin IR 120 (H⁺), filtered and

the filtrate concentrated in vacuo. Purification by column

chromatography (silica gel, toluene/acetone, 5:1) furnished 44

(1.22 g, 1.20 mmol, 98%) as colorless, amorphous solid. TLC: R_f

= 0.25 (petroleum ether/ethyl acetate, 1:1). [α]²_D⁰= +3.2 (c = 1,

CHCl₃). ¹H NMR (600 MHz, [D₆]DMSO): δ = 1.75 (br. s, 6 H,

DMM-H), 3.22 (m, 1 H, 5b-H), 3.26 (m, 1 H, 2b-H), 3.38 (m, 2

H, 3b-H, 4b-H), 3.54 (m, 1 H, 6b-H), 3.61 (m, 1 H, 6bЈ-H), 3.71–

lopyranosyl]acetimidate (42): Trichloroacetonitrile (1.35 mL, 1.94 g,

13.5 mmol) and then DBU (5 drops) were added to a solution of

41 (1.70 g, 2.01 mmol) in CH₂Cl₂(15 mL). The reaction mixture

was stirred at room temperature for 1 h, concentrated to 3/4 of its

volume and purified by column chromatography (silica gel, petro-

leum ether/ethyl acetate, 3:2 + 1% NEt₃) to furnish 42 as slightly

yellowish foam (1.95 g, 1.97 mmol, 98%) which was stored under

argon at –20 °C. TLC: R_f= 0.65 (petroleum ether/ethyl acetate,

1

1:1). H NMR (250 MHz, CDCl₃): δ = 1.58 (s, 3 H, acetyl), 1.59

(br. s, 6 H, DMM-H), 1.98, 2.03 (2 s, 6 H, 2ϫ acetyl), 3.47–3.62

(m, 3 H, 3b-H, 4b-H, 5b-H), 3.69–3.74 (m, 2 H, 6a-H), 4.04 (dd,

2

3.73 (m, 3 H, 2a-H, 4a-H, 6a-H), 3.78 (dd, J_gem= 10.4 Hz, 1 H,

3

³J_3,4= 2.9, J_4,5= 9.8 Hz, 1 H, 4a-H), 4.14 (m, 2 H, 2a-H, 5a-H),

2

6aЈ-H), 3.98 (d, J_gem= 15.3 Hz, 1 H, CHHC=ONHTrt), 4.00 (m,

2

3

2

4.21 (dd, J_gem= 9.1 Hz, 1 H, 6b-H), 4.29 (dd, J_gem= 9.1, J_vic

=

1 H, 5a-H), 4.13 (m, 1 H, 3a-H), 4.19 (d, J_gem= 15.2 Hz, 1 H,

3

2.4 Hz, 1 H, 6bЈ-H), 4.43 (d, J_1,2= 7.9 Hz, 1 H, 1b-H), 4.51 (2 d,

CHHC=ONHTrt), 4.35 (d, 1 H, CHHPh), 4.36 (d, 1 H, 1b-H),

4.40 (d, 1 H, CHHPh), 4.57 (d, 1 H, CHHPh), 4.61 (m, 1 H, 6b-

OH), 4.69 (d, 1 H, CHHPh), 4.73 (d, 1 H, CHHPh), 4.89 (d, 1 H,

2 H, CH₂Ph), 4.63, 4.67 (2 d, 2 H, CH₂Ph), 4.77 (d, 2 H, CH₂Ph),

3

4.89 (dd, J_1,2≈ J_2,3≈ 8.5 Hz, 1 H, 2b-H), 5.60 (dd, 1 H, J_2,3

≈

³J_3,4≈ 2.6 Hz, 3a-H), 6.89 (d, 1 H, ³J_1,2= 9.1 Hz, 1a-H), 7.22–7.33

3

CHHPh), 5.08 (d, J = 4.3 Hz, 1 H, 3a-OH), 5.62 (d, J = 5.4 Hz,

3

(m, 15 H, phenyl), 8.68 (s, 1 H, NH) ppm.

1 H, 2b-OH), 5.75 (d, J_1,2= 8.6 Hz, 1 H, 1a-H), 7.06–7.34 (m, 30

[N-(Triphenylmethyl)carbamoyl]methyl 2,6-Di-O-acetyl-3,4-di-O-

H, phenyl, Trt), 7.84 (s, 1 H, C=ONHTrt) ppm. ¹³C NMR

benzyl-β-D-glucopyranosyl-(1Ǟ4)-3-O-acetyl-6-O-benzyl-2-deoxy-2-

(150.8 MHz, [D₆]DMSO, selected data): δ = 56.5 (1 C, 2a-C), 60.1

6030

www.eurjoc.org

Eur. J. Org. Chem. 2007, 6016–6033

Carbohydrate-Based VEGF Inhibitors

(1 C, 6b-C), 68.8 (1 C, 6a-C), 68.9 (1 C, 3a-C), 69.8 (1 C, OCH₂- purified by column chromatography (silica gel, toluene/acetone, 6:1

C=ONHTrt), 72.3 (1 C, 5a-C), 73.6 (1 C, 2b-C), 75.0 (1 C, 5b-C), Ǟ 5:1) to furnish 46 as colorless foam (420 mg, 0.37 mmol, 78%).

75.3 (1 C, 4a-C), 76.6 (1 C, 3b-C), 84.3 (1 C, 4b-C), 97.3 (1 C, 1a- TLC: R_f= 0.55 (toluene/acetone, 6:4). [α]²_D⁰= –15.7 (c = 1, CHCl₃).

C), 103.9 (1 C, 1b-C) ppm. C₆₀H₆₂N₂O₁₃(1019.1): calcd. C 70.71,

H 6.13, N 2.75; found C 70.30, H 6.22, N 2.88.

¹H NMR (600 MHz, CDCl₃): δ = 3.46 (m, 2 H, 2b-H, 3b-H), 3.57

(dd, ²J_gem= 9.9 Hz, 1 H, 6a-H), 3.60 (dd, ²J_gem= 9.9 Hz, 1 H, 6aЈ-

H), 3.75 (m, 1 H, 4b-H), 3.79 (m, 1 H, 4a-H), 3.84–3.87 (m, 2 H,

[N-(Triphenylmethyl)carbamoyl]methyl 3,4-Di-O-benzyl-β-

pyranosyl-(1Ǟ4)-6-O-benzyl-2-[(benzyloxycarbonyl)amino]-2-deoxy-

β- -allopyranoside (45): Solid NaOH (600 mg, 15.0 mmol) was

D-gluco-

3

2

2a-H, 4a-H), 3.90 (d, J_4,5= 9.3 Hz, 1 H, 5b-H), 4.07 (d, J_gem

=

15.2 Hz, 1 H, CHHC=ONHTrt), 4.20 (dd, 1 H, 3a-H), 4.25 (d, 1

H, CHHC=ONHTrt), 4.35 (m, 2 H, 1b-H, CHHPh), 4.48 (d, 1 H,

CHHPh), 4.60 (d, ³J = 8.3 Hz, 1 H, 1a-H), 4.70 (d, 1 H, CHHPh),

4.74–4.81 (m, 3 H, CH₂Ph, CHHPh), 4.94 (d, 1 H, CHHPh), 5.12

D

added to a solution of 44 (1.20 g, 1.17 mmol) in a mixture of diox-

ane (48 mL) and water (12 mL). The turbid two-phase solution was

stirred at room temperature overnight. The pH was adjusted to 4.5

with 1  HCl and monitored every 30 min for the next 5 h and if

necessary adjusted to 4.5 with 1  HCl. The reaction mixture was

stirred at room temperature overnight, ethanolamine (80 µL,

80.9 mg, 1.325 mmol) was added and the solution neutralized with

1  NaOH. K₂CO₃(540 mg, 3.91 mmol) was immediately added

and then benzyl chloroformate (480 µL, 0.57 g, 3.36 mmol). The

reaction mixture was stirred at room temperature for 1 h, concen-

trated to 1/2 of its volume, diluted with NaCl solution and ex-

tracted three times with CHCl₃. The combined organic phases were

dried with MgSO₄, filtered and concentrated. Purification by col-

umn chromatography (silica gel, toluene/acetone, 5:1) furnished 45

(0.98 g, 9.38 mmol, 80%) as colorless, hygroscopic foam. TLC: R_f

= 0.45 (toluene/acetone, 2:1). [α]²_D⁰= –9.6 (c = 1, CHCl₃). ¹H NMR

(600 MHz, CDCl₃): δ = 3.13 (br. s, 1 H, OH), 3.28 (m, 1 H, 5b-H),

3

(d, 1 H, CHHPh), 5.15 (d, 1 H, CHHPh), 5.42 (d, J = 9.5 Hz, 1

H, 2a-NH), 7.13–7.31 (m, 40 H, phenyl), 7.99 (s, 1 H, C=ONHTrt)

ppm. ¹³C NMR (150.8 MHz, CDCl₃, selected data): δ = 53.2 (1 C,

2a-C), 69.0 (1 C, 6a-C), 69.8 (1 C, 3a-C), 70.1 (1 C, OCH₂-

C=ONHTrt), 71.2 (1 C, 5a-C), 73.7 (1 C, 2b-C), 74.4 (1 C, 5b-C),

77.4 (1 C, 4a-C), 78.6 (1 C, 4b-C), 82.9 (1 C, 3b-C), 101.1 (1 C, 1a-

C), 103.8 (1 C, 1b-C) ppm. MALDI-MS (pos. mode, matrix DHB,

THF): m/z = 1172 [M + Na]⁺, 1188 [M + K]⁺. C₆₉H₆₈N₂O₁₄(1149.3):

calcd. C 72.11, H 5.96, N 2.44; found C 71.71, H 5.96, N 2.51.

[N-(Triphenylmethyl)carbamoyl]methyl {Benzyl 3,4-di-O-benzyl-2-

O-[(tert-butyloxycarbonyl)methyl]-β-

(1Ǟ4)-6-O-benzyl-2-[(benzyloxycarbonyl)amino]-3-O-[(tert-butyl-

oxycarbonyl)methyl]-2-deoxy-β- -allopyranoside (47): Molecular

D-glucopyranosyluronate}-

D

sieves (4 Å) and tert-butyl bromoacetate (0.36 mL, 0.47 g,

2.43 mmol) were added to a solution of 46 (400 mg, 0.34 mmol)

3

3.43 (dd, J_1,2≈ J_2,3≈ 8.2 Hz, 1 H, 2b-H), 3.48 (dd, J_2,3≈ J_3,4

≈

8.9 Hz, 1 H, 3b-H), 3.54–3.58 (m, 2 H, 4b-H, 6a-H), 3.63 (dd, ²J_gemwith TBAI (240 mg, 0.64 mmol) in dry CH₂Cl₂(8 mL); the mixture

= 9.6 Hz, 1 H, 6aЈ-H), 3.68 (dd, J_gem= 13.2 Hz, 1 H, 6b-H), 3.77

(dd, J_gem= 12.9 Hz, 1 H, 6bЈ-H), 3.80 (m, 1 H, 4a-H), 3.84–3.86 (800 mg, 3.43 mmol) was added. After stirring overnight, the reac-

( m, 2 H, 2 a- H, 5 a - H ) , 4 . 0 7 ( d , J_{g e m}= 1 5 . 1 Hz , 1 H, tion mixture was diluted with ethyl acetate and filtered through

2

was stirred at room temperature for 10 min, and then Ag₂O

2

CHHC=ONHTrt), 4. 22 (m, 1 H, 3a-H), 4. 24 (d, 1 H,

Celite. The filtrate was concentrated, coevaporated twice with tolu-

CHHC=ONHTrt), 4.27 (d, 1 H, CHHPh), 4.31 (d, J_1,2= 7.4 Hz, ene, and the residue purified by column chromatography (silica gel,

3

1 H, 1b-H), 4.36 (d, 1 H, CHHPh), 4.59 (d, J_1,2= 8.2 Hz, 1 H,

petroleum ether/ethyl acetate, 3.25:1) to furnish 47 (330 mg,

0.24 mmol, 70%) as colorless foam. TLC: R_f= 0.40 (toluene/ace-

1a-H), 4.64. 4.76 (2d, 2 H, CH₂Ph), 4.83 (m, 3 H, CH₂Ph,

CHHPh), 4.93 (d, 1 H, CHHPh), 5.44 (d, 1 H, 2a-NH), 7.20–7.36 tone, 6:4). [α]²_D⁰= –23.6 (c = 1, CHCl₃). ¹H NMR (600 MHz,

(m, 35 H, phenyl), 7.96 (s, 1 H, C=ONHTrt) ppm. ¹³C NMR CDCl₃): δ = 1.45, 1.46 (2 s, 18 H, 2ϫ tert-butyl), 3.25 (dd, J_1,2

=

3

(150.8 MHz, CDCl₃, selected data): δ = 53.3 (1 C, 2a-C), 61.4 (1

³J_2,3= 8.4 Hz, 1 H, 2b-H), 3.54 (dd, J_2,3

=

³J_3,4= 8.0 Hz, 1 H,

C, 6b-C), 69.0 (1 C, 6a-C), 69.7 (1 C, 3a-C), 70.1 (1 C, OCH₂- 3b-H), 3.65 (m, 2 H, 6a-H), 3.72 (m, 2 H, 3a-H, 4b-H), 3.82 (m, 1

C=ONHTrt), 71.4 (1 C, 5a-C), 74.3 (1 C, 2b-C), 75.5 (1 C, 5b-C), H, 2a-H), 3.84 (m, 1 H, 4a-H), 3.89 (m, 1 H, 5a-H), 3.92 (m, 1 H,

76.3 (1 C, 4b-C), 77.0 (1 C, 4a-C), 83.9 (1 C, 3b-C), 100.9 (1 C, 1a-

C), 103.7 (1 C, 1b-C) ppm. MALDI-MS (pos. mode, matrix DHB,

THF): m/z = 1068 [M + Na]⁺, 1084 [M + K]⁺. C₆₂H₆₄N₃O₁₃(1045.2):

calcd. C 71.25, H 6.17, N 2.68; found C 71.13, H 6.25, N 2.47.

5b-H), 4.06, 4.07 (2 d, 2 H, CH₂C=O), 4.12, 4.17 (2 d, 2 H,

CH₂C=O), 4.28 (m, 2 H, CH₂C=O), 4.23 ppm. 4.34 (2 d, 2 H,

CH₂Ph), 4.43 (m, 3 H, 1b-H, CH₂Ph), 4.59 (d, J_1,2= 8.2 Hz, 1 H,

3

1a-H), 4.72 (m, 4 H, 2ϫ CH₂Ph), 4.98 (m, 2 H, CH₂Ph), 5.14 (m,

2 H, CH₂Ph), 7.11–7.33 (m, 40 H, CH₂Ph), 8.03 (s, 1 H,

C=ONHTrt) ppm. ¹³C NMR (150.8 MHz, CDCl₃, selected data):

δ = 53.7 (1 C, 2a-C), 68.3 (1 C, 6a-C), 72.3 (1 C, 5a-C), 74.1 (1 C,

3a-C), 74.9 (1 C, 5b-C), 78.0 (1 C, 4b-C), 82.6 (1 C, 4a-C), 82.7 (1

C, 2b-C), 83.1 (1 C, 3b-C), 101.7 (1 C, 1a-C), 103.3 (1 C, 1b-C)

ppm. MALDI-MS (pos. mode, matrix DHB, THF): m/z = 1400

[M + Na]⁺, 1344 [M + Na – tBu]⁺.C₈₁H₈₈N₂O₁₈·0.5H₂O

(1386.6) calcd. C 70.16, H 6.47, N 2.02; found C 69.95, H 6.51, N

1.88.

[N-(Triphenylmethyl)carbamoyl]methyl (Benzyl 3,4-di-O-benzyl-β-

glucopyranosyluronate)-(1Ǟ4)-6-O-benzyl-2-[(benzyloxycarbonyl)-

amino]-2-deoxy-β- -allopyranoside (46): The pyranoside 45

D-

D

(500 mg, 0.47 mmol), NaBr (8 mg, 0.077 mmol) and TBAB (8 mg,

0.024 mmol) were dissolved in a two-phase mixture of CH₂Cl₂/H₂O

(6:1, 10 mL), and at 0 °C under ice-bath cooling TEMPO (5 mg,

0.032 mmol) was added. Then slowly a solution of NaOCl (5%,

3.6 mL) and a saturated NaHCO₃solution (2.8 mL) were added

dropwise, the reaction mixture was stirred at 0 °C for 30 min, di-

luted with H₂O and extracted with CH₂Cl₂. The aqueous phase

was acidified with KHSO₄solution (5%) and extracted three times

with CH₂Cl₂. The combined organic phases were dried with

MgSO₄, filtered, and concentrated in vacuo to furnish the free acid

(480 mg, 95%) as intermediate which was dissolved in dry DMF

(10 mL); CsF (140 mg, 0.59 mmol) and benzyl bromide (100 µL,

[N-(Triphenylmethyl)carbamoyl]methyl 2-O-[(tert-butyloxycarbon-

yl)methyl]-β-

carbonyl)methyl]-2-deoxy-2-{[(9-fluorenyl)methoxycarbonyl]amino}-

β- -allopyranoside (48): Pd/C (200 mg) was added to a solution of

D-glucopyranosyluronate-(1Ǟ4)-3-O-[(tert-butyloxy-

D

47 (300 mg, 0.22 mmol) in MeOH/H₂O (5:1, 30 mL) and the mix-

ture stirred under hydrogen for 48 h; the catalyst was filtered off

144 mg, 0.84 mmol) were added, and the mixture was stirred at through Celite, the filtrate concentrated to 1/2 of its volume, diluted

room temperature overnight. The reaction mixture was diluted with

ethyl acetate and washed three times with a saturated NaCl solu-

tion. The organic phase was dried with MgSO₄, filtered and con-

centrated. After two coevaporations with toluene, the residue was

with CH₃CN/H₂O (2:1, 40 mL), and Fmoc-ONSu (180 mg,

0.54 mmol) and NaHCO₃(600 mg, 7.12 mmol) were added. The

reaction mixture was stirred at room temperature overnight, con-

centrated until it became turbid, diluted with H₂O and then ex-

Eur. J. Org. Chem. 2007, 6016–6033

www.eurjoc.org

6031

T. Haag, R. A. Hughes, G. Ritter, R. R. Schmidt

FULL PAPER

tracted four times with CHCl₃. The combined organic phases were

dried with MgSO₄, concentrated, and the residue was purified by

column chromatography (silica gel, ethyl acetate/MeOH, 9:1 Ǟ

4:1). This way not all byproducts could be separated. Therefore,

compound 48 (120 mg, 0.108 mmol, 50%) was used in the next

step without further purification. MALDI-MS (pos. mode, matrix

DHB, THF): m/z = 1029 [M + Na]⁺.

MgSO₄, filtered and concentrated. Purification by flash chromatog-

raphy (silica gel toluene/acetone, 4.5:1 Ǟ 3.5:1) furnished 50

(50 mg, 0.026 mmol, 91%) as colorless lyophilisate from dioxane.

1

TLC: R_f= 0.62 (toluene/acetone, 1:1). H NMR (600 MHz, [D₆]-

DMSO): δ = 0.77–0.79 (m, 13 H, 2ϫ Leu-CH₃, 2ϫ Ile-CH₃, Ile-

β-H), 1.03 (m, 1 H, Ile-CHH), 1.27–1.41 (m, 37 H, 3ϫ tert-butyl,

Boc, Ile-CHH), 1.43 (m, 2 H, Leu-β-H), 1.62 [m, 2 H, Ile-β-H, Leu

CH(CH₃)₂], 2.35 (m, 2 H, Cys-β-H), 2.44 (m, 2 H, Cys-β-H), 2.98

(m, 1 H, 2b-H), 3.29–3.35 (m, 3 H, 5a-H, 3b-H, 4b-H), 3.49 (dd, 1

H, 2a-H), 3.55 (m, 1 H, 6a-H), 3.64–3.72 (m, 4 H, 3a-H, 4a-H, 6aЈ-

H, 5b-H), 3.87–3.91 (m, 3 H, CHH=ONHTrt, Cys-α-H, Cys-α-H),

4.02 (d, ²J_gem= 15 Hz, 1 H, CHHC=ONHTrt), 4.15 (m, 2 H, CH₂–

CO), 4.18 (m, 2 H, CH₂–CO), 4.27 (dd, Ile-α-H), 4.36 (m, 1 H,

[N-(Triphenylmethyl)carbamoyl]methyl 2-O-[(tert-Butyloxycarbon-

yl)methyl]-β-

phenylmethyl)-

carbonyl)methyl]-2-deoxy-2-{[(9-fluorenyl)methoxycarbonyl]amino}-

β- -allopyranoside (49): A solution of 48 (110 mg, 0.099 mmol), 27a

D

-glucopyranosyluronyl[

L-leucyl-L-isoleucyl-S-(tri-

L-cystine tert-butyl ester]-(1Ǟ4)-3-O-[(tert-butyloxy-

D

(80 mg, 0.124 mmol) and PyBOP (73 mg, 0.140 mmol) in dry DMF

(5 mL) was treated with DIPEA (ca. 40 µL) to adjust the pH to ca.

8. After stirring at room temperature for 2 h, the reaction mixture

was diluted with ethyl acetate and washed with a KHSO₄solution

(5%), a NaHCO₃solution and a saturated NaCl solution. The or-

ganic phase was dried with MgSO₄, filtered and concentrated. Puri-

fication by column chromatography (silica gel, toluene/acetone,

3.75:1 Ǟ 3.5:1) furnished 49 (110 mg, 0.063 mmol, 64%) as color-

less lyophilisate from dioxane. TLC: R_f= 0.68 (toluene/acetone,

3

Leu-α-H), 4.55 (d, J_1,2≈ 7.3 Hz, 2 H, 1a-H, 1b-H), 4.64 (m, 1 H,

3

OH), 5.25 (m, 1 H, OH), 5.41 (m, 1 H, OH), 6.49 (d, J = 8.1 Hz,

3

1 H, Cys-NH), 7.15–7.33 (m, 45 H, Trt), 7.72 (d, J = 7.4 Hz, 1 H,

2a-NH), 7.91 (d, ³J = 9.0 Hz, 1 H, Ile-NH), 8.03 (s, 1 H,

C=ONHTrt), 8.05 (d, ³J = 7.3 Hz, 1 H, Leu-NH), 8.47 (d, ³J =

7.2 Hz, 1 H, Cys-NH) ppm. ¹³C NMR (150.8 MHz, [D₆]DMSO,

selected data): δ = 51.0 (1 C, Leu-α-C), 52.1 (1 C, Cys-α-C), 53.4

(1 C, Cys-α-C), 54.1 (1 C, 2a-C), 56.0 (1 C, Ile-α-C), 58.9 (1 C, 6a-

C), 68.3 (1 C, CH₂–CO), 68.5 (1 C, CH₂–CO), 69.1 (1 C, CH₂–

CO), 74.7 (1 C, 5b-C), 76.7 (1 C, 4a-C), 79.0 (1 C, 3a-C), 81.8 (1 C,

2b-C), 101.2 (1 C, 1b-C), 101.5 (1 C, 1a-C) ppm. MALDI-MS (pos.

mode, matrix DHB, THF): m/z = 1979 [M + Na]⁺, 1995 [M + K]⁺.

1

1:1). [α]²_D⁰= –17.8 (c = 0.5, CHCl₃). H NMR (600 MHz, CDCl₃):

δ = 0.81 (m, 3 H, Ile-CH₃), 0.84 (m, 3 H, Ile-CH₃), 0.87 (m, 6 H,

2ϫ Leu-CH₃), 1.05 (m, 1 H, Ile-CHH), 1.38 (m, 10 H, tert-butyl,

Ile-CHH), 1.43 (s, 9 H, tert-butyl), 1.48 (s, 9 H, tert-butyl), 1.51

(m, 1 H, Leu-β-H), 1.65 [m, 1 H, Leu-CH(CH₃)₂], 1.66 (m, 1 H,

Leu-βЈ-H), 1.70 (m, 1 H, Ile-β-H), 2.43, 2.66 (2 m, 2 H, Cys-β-H),

3.09 (dd, ³J_1,2= ³J_2,3= 8.1 Hz, 1 H, 2b-H), 3.60–3.67 (m, 3 H, 3b-

H, 4b-H, 6a-H), 3.70–3.75 (m, 2 H, 5b-H, 6aЈ-H), 3.82 (m, 2 H,

2a-H, 5a-H), 3.86 (m, 2 H, 4a-H, Fmoc-CHH), 4.04 (m, 1 H, 3a-

H), 4.07–4.09 (m, 3 H, Fmoc, CH₂C=O), 4.17 (m, 1 H, Fmoc),

4.18–4.23 (3 H, Ile-α-H, CH₂C=O), 4.27 (m, 1 H, CHHC=O),

4.29–4.35 (m, 3 H, Cys-α-H, CH₂C=O), 4.42 (m, 1 H, Leu-α-H),

4.56 (d, ³J_1,2= 7.7 Hz, 1 H, 1b-H), 4.69 (d, ³J_1,2= 8.4 Hz, 1 H, 1a-

Carbamoylmethyl 2-O-(Carboxymethyl)-β-

yl( -leucyl- -isoleucyl- -hemicystine)-(1Ǟ4)-3-O-(carboxymethyl)-

2-[( -hemicystinyl)amido]-2-deoxy-β- -allopyranoside (33): A solu-

D-glucopyranosyluron-

L

D

tion of 50 (40 mg, 0.020 mmol) in CH₂Cl₂/MeOH (7:1, 20 mL) was

added dropwise to a solution of iodine (26 mg, 0.102 mmol) in

CH₂Cl₂/MeOH (7:1, 20 mL) at 0 °C. After stirring at 0 °C for

30 min, a 0.01  sodium thiosulfate solution was added until decol-

oration occurred, and the reaction mixture was extracted three

times with CHCl₃. The combined organic phases were dried with

MgSO₄, filtered and concentrated under reduced pressure. The cy-

clized intermediate was then dissolved in a mixture of TFA

(9.5 mL), Et₃SiH (0.3 mL) and H₂O (0.2 mL) and the mixture

stirred at 0 °C for 3.5 h. After removal of the TFA in vacuo, the

residue was dried in vacuo and purified with flash chromatography

(RP-18 silica gel, CH₃CN/H₂O, 1:2.5). RP-18 HPLC yielded 33

(15 mg, 0.016 mmol, 78%) as colorless lyophilisate. HPLC (prep.

RP-18: 0–5 min isocratic 7% CH₃CN + 0.1% TFA, 5–60 min linear

3

H), 5.91 (d, J = 7.6 Hz, 1 H, Cys-NH), 6.43 (d, J = 7.9 Hz, 1 H,

3

Leu-NH), 6.85 (d, J = 8.4 Hz, 1 H, Ile-NH), 7.11–7.35 (m, 38 H,

phenyl, Fmoc), 7.52 (d, ³J = 8.5 Hz, 1 H, 2a-NH), 8.00 (s, 1 H,

C=ONHTrt) ppm. ¹³C NMR (150.8 MHz, CDCl₃, selected data):

δ = 11.4, 15.1, 23.0, 24.6 (4 C, 4ϫ CH₃), 24.6 (1 C, Ile-CH₂), 33.9

(1 C, Cys-β-C), 38.5 (1 C, Ile-β-C), 41.3 (1 C, Leu-β-C), 51.9 (1 C,

Cys-α-C), 52.6 (1 C, Leu-α-C), 54.0 (1 C, 2a-C), 57.5 (1 C, Ile-α-

C), 61.0 (1 C, 6a-C), 72.3 (1 C, 4b-C), 73.1 (1 C, 5a-C), 74.4 (1 C,

5b-C), 74.9 (1 C, 3b-C), 75.3 (1 C, 4a-C), 82.7 (1 C, 3a-C), 83.2 (1

C, 2b-C), 101.9 (1 C, 1a-C), 103.4 (1 C, 1b-C) ppm. MALDI-MS

(pos. mode, matrix DHB, THF): m/z = 1755 [M + Na]⁺, 1771 [M

+ K]⁺. C₉₈H₁₁₇N₅O₂₁S·H₂O (1751.1): calcd. C 67.22, H 6.85, N

4.00; found C 66.68, H 6.81, N 4.00.

gradient 7–50 % CH₃CN + 0.1 % TFA, flow 10 mL/min): t_R

=

1

32.4 min. H NMR (600 MHz, [D₆]DMSO): δ = 0.80–0.87 (m, 12

H, 4ϫ CH₃), 1.07 (m, 1 H, Ile-CHH), 1.46 (m, 2 H, Leu-β-H, Ile-

CHH), 1.54 [m, 1 H, Leu-CH(CH₃)₂], 1.78 (m, 1 H, Ile-β-H), 2.99

(m, 2 H, 2b-H, Cys-β-H), 3.07 (m, 1 H, Cys-β-H), 3.24 (m, 1 H,

Cys-βЈ-H), 3.32 (m, 1 H, Cys-βЈ-H), 3.35–3.41 (m, 2 H, 3b-H, 4b-

H), 3.52 (m, 1 H, 6a-H), 3.65 (m, 1 H, 6aЈ-H), 3.68 (m, 1 H, 5b-

H), 3.80 (m, 2 H, 4a-H, 5a-H), 3.88 (m, 1 H, 2a-H), 3.95–4.04

(m, 4 H, 3a-H, Cys-α-H, CH₂C=O), 4.15–4.27 (m, 4 H, Ile-α-H,

CH₂C=O, CHHC=O), 4.34–4.36 (m, 2 H, Leu-α-H, CHHC=O),

4.54 (d, ³J_1,2= 7.7 Hz, 1 H, 1b-H), 4.57 (d, ³J_1,2= 8.5 Hz, 1 H, 1a-

[N-(Triphenylmethyl)carbamoyl]methyl 2-O-[(tert-Butyloxycarbon-

yl)methyl]-β-

ylmethyl)- -cysteine tert-butyl ester]-(1Ǟ4)-3-O-[(tert-butyloxycar-

bonyl)methyl]-2-{[N-(tert-butyloxycarbonyl)-S-(triphenylmethyl)-

cysteinyl]amido}-2-deoxy-β- -allopyranoside (50): Piperidine

D-glucopyranosyluronyl[L-leucyl-L-isoleucyl-S-(triphen-

L

L-

D

(1.3 mL) was added to a solution of 49 (50 mg, 0.029 mmol) in

DMF (5 mL) and the mixture stirred for 4 h, The reaction mixture

was concentrated in vacuo, coevaporated twice with toluene and

the residue purified by column chromatography (silica gel, CHCl₃/

MeOH, 100:1 Ǟ 75:1 Ǟ 25:1). The resulting NH-unprotected com-

pound (42 mg, 0.027 mmol, 99%) was dissolved with Boc-Cys(Trt)-

3

H), 4.61 (m, 1 H, Cys-α-H), 7.47 (d, J = 7.8 Hz, 1 H, Leu-NH),

3

7.90 (d, J = 8.7 Hz, 1 H, Ile-NH), 8.32 (d, J = 8.2 Hz, 1 H, Cys-

3

NH), 8.35 (d, 1 H, Cys-NH), 8.36 (s, 2 H, C=ONH₂), 8.59 (d, J

= 8.8 Hz, 1 H, 2a-NH) ppm. ¹³C NMR (150.8 MHz, [D₆]DMSO,

selected data): δ = 11.0, 15.2, 21.0, 22.9 (4 C, 4ϫ CH₃), 23.9, 24.0

OH (16 mg, 0.033 mmol) and PyBOP (19 mg, 0.036 mmol) in dry [2 C, Ile-CH₂, Leu-CH(CH₃)₂], 36.3 (1 C, Ile-β-C), 40.9 (1 C, Cys-

DMF (1.5 mL). The pH was adjusted to 8 with DIPEA (ca. 15 µL)

and the reaction mixture stirred at room temperature for 1.5 h. It

was then diluted with ethyl acetate and washed with KHSO₄,

NaHCO₃and NaCl solutions. The organic phase was dried with

β-C), 41.2 (1 C, Leu-β-C), 43.1 (1 C, Cys-β-C), 50.7 (1 C, Leu-α-

C), 51.0 (1 C, Cys-α-C), 51.7 (1 C, 2a-C), 53.3 (1 C, Cys-α-C), 56.5

(1 C, Ile-α-C), 60.0 (1 C, 6a-C), 71.5 (1 C, 4b-C), 73.3 (1 C, 5a-C),

74.2 (1 C, 5b-C), 74.5 (1 C, 3b-C), 76.2 (1 C, 4a-C), 80.4 (1 C, 3a-

6032

www.eurjoc.org

Eur. J. Org. Chem. 2007, 6016–6033

Carbohydrate-Based VEGF Inhibitors

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C), 81.7 (1 C, 2b-C), 98.5 (1 C, 1a-C), 102.9 (1 C, 1b-C) ppm.

MALDI-MS (pos. mode, matrix DHB, THF): m/z = 983

[M + Na]⁺, 999 [M + K]⁺.

Carbamoylmethyl 2-O-(Carboxymethyl)-β-

yl( -leucyl- -isoleucyl- -hemicystine)-(1Ǟ4)-3-O-(carboxymethyl)-

2-deoxy-[( -hemicystinylglycyl)amido]-β- -allopyranoside (34): As

D-glucopyranosyluron-

L

D

in the synthesis of 33, for the preparation of 34 the nonapeptide

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1

26.9 min. H NMR (600 MHz, [D₆]DMSO): δ = 0.80–0.87 (m, 12

H, 2ϫ Ile-CH₃, 2ϫ Leu-CH₃), 1.13 (m, 1 H, Ile-CHH), 1.37 (m,

1 H, Leu-β-H), 1.45 (m, 1 H, Ile-CHH), 1.51 [m, 1 H, Leu-

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2

3

CH(CH₃)₂], 1.81 (m, 1 H, Ile-β-H), 2.86 (dd, J_gem= 12.9, J_vic

=

9.6 Hz, 1 H, Cys-β-H), 2.97 (m, 2 H, 2b-H, Cys-β-H), 3.15 (dd,

3

²J_gem= 13.2, J_vic= 5.3 Hz, 1 H, Cys-β-H), 3.18 (dd, ²J_gem= 14.1,

³J_vic= 5.4 Hz, 1 H, Cys-β-H), 3.35 (dd, ³J_2,3= ³J_3,4= 8.9 Hz, 1 H,

3

2

3b-H), 3.46 (dd, J_3,4= J_3,4= 9.2 Hz, 1 H, 4b-H), 3.51 (dd, J_gem

= 11.7, ³J_vic= 4.3 Hz, 1 H, 6a-H), 3.63 (m, 2 H, 6aЈ-H, 5b-H), 3.75

(m, 1 H, 5a-H), 3.83–3.91 (m, 5 H, 2a-H, 3a-H, 4a-H, Gly-α-H),

2.92 (d, 1 H, CHHC=O), 4.05 (d, 1 H, CHHC=O), 4.06 (m, 1 H,

Cys-α-H), 4.12 (m, 1 H, Ile-α-H), 4.12–4.30 (m, 4 H, 2 ϫ

3

CH₂C=O), 4.45 (d, J_1,2= 7.8 Hz, 1 H, 1b-H), 4.50 (m, 1 H, Cys-

3

α-H), 4.56 (d, J_1,2= 8.4 Hz, 1 H, 1a-H), 4.60 (m, 1 H, Leu-α-H),

3

7.71 (d, J = 8.4 Hz, 1 H, Cys-NH), 8.11 (d, J = 8.0 Hz, 1 H, 2a-

NH), 8.20 (m, 2 H, Gly-NH, Ile-NH), 8.27 (br. s, 1 H, Cys-NH),

3

8.47 (d, J = 7.8 Hz, 1 H, Cys-NH) ppm. ¹³C NMR (150.8 MHz,

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35.6 (1 C, Ile-β-C), 38.7 (1 C, Cys-β-C), 38.8 (1 C, Cys-β-C), 41.8

(1 C, Leu-β-C), 42.4 (1 C, Gly-α-C), 50.0 (1 C, Leu-α-C), 50.6 (1

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Published Online: October 29, 2007

Eur. J. Org. Chem. 2007, 6016–6033

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Carbohydrate-based VEGF inhibitors

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