Synthetic route to a benzooxazole derivative with heparanase inhibitory activity

Article abstract of DOI:10.1080/00397910902994186

The synthesis of a benzooxazol-5-yl acetic acid derivative (9) with strong heparanase and angiogenesis inhibitory activity, and thus possible commercial interest, is described in detail. Copyright Taylor & Francis Group, LLC.

Full text of DOI:10.1080/00397910902994186

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Synthetic Route to a Benzooxazole
Derivative with Heparanase Inhibitory
Activity
Chiara Da Pieve a , Pravin Patel a & Sotiris Missailidis a
a Open University , Milton Keynes, United Kingdom
Published online: 03 Feb 2010.
To cite this article: Chiara Da Pieve , Pravin Patel & Sotiris Missailidis (2010) Synthetic Route to
a Benzooxazole Derivative with Heparanase Inhibitory Activity, Synthetic Communications: An
International Journal for Rapid Communication of Synthetic Organic Chemistry, 40:4, 518-522, DOI:
10.1080/00397910902994186
To link to this article: http://dx.doi.org/10.1080/00397910902994186
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Synthetic Communications1, 40: 518–522, 2010
Copyright # Taylor & Francis Group, LLC
ISSN: 0039-7911 print=1532-2432 online
DOI: 10.1080/00397910902994186
SYNTHETIC ROUTE TO A BENZOOXAZOLE
DERIVATIVE WITH HEPARANASE INHIBITORY
ACTIVITY
Chiara Da Pieve, Pravin Patel, and Sotiris Missailidis
Open University, Milton Keynes, United Kingdom
The synthesis of a benzooxazol-5-yl acetic acid derivative (9) with strong heparanase and
angiogenesis inhibitory activity, and thus possible commercial interest, is described in detail.
Keywords: Angiogenesis; benzooxazole; heparanase; inhibitor; metastasis
INTRODUCTION
Heparanase is an enzyme implicated in several physiological and pathological
processes including tumor angiogenesis and metastasis.[1–3] Few compounds with
heparanase inhibitory activity have been reported in the literature,[4–6] and most of
these are sugar derivatives, which have modest activity. Compound 9, however, is
a small molecule (Fig. 1) that shows potent and selective heparanase, as well as
angiogenic, inhibitory activities.[7]
Despite its biological activity, no detailed synthesis or data for 9 are available.
Herewith, we report an efficient protocol for the synthesis of 9.
The seven steps of our synthesis are outlined in Scheme 1. 4-Hydroxy-3-
nitrophenylacetic acid was first protected as its methyl ester in 90% yield in two steps
according to a published procedure, and the nitro group of 1 was then reduced to
obtain the amino derivative 2.[8] Compound 2 was then coupled to 3-fluoro-4-nitro
benzoyl chloride (3). Refluxing the amide 4 with p-toluenesulfonic acid (p-TsOH)
in toluene[9] afforded the ring-closed product 5 in 60% yield. Reduction of the nitro
group in 5 was accomplished using standard catalytic hydrogenation conditions.
When a scale larger than 300 mg of the starting material 5 was used, the yield of
the product 6 was in fact improved from <50% to quantitative using H2Pd=C instead
the Zn=acetic acid procedure suggested in literature.[7] Compound 6 was coupled to
4-bromocinnamoyl chloride (7) in CH2Cl2. The product 8 was carefully purified, after
washing with tetrahydrofuran (THF) water, by chromatography on silica gel starting
with CH2Cl2 as eluent and then gradually increasing the polarity to 50:1 CH2Cl2–
methanol. Because of the limited solubility of 8 in most organic solvents, hydrolysis
Received March 27, 2009.
Address correspondence to Sotiris Missailidis, The Open University, Department of Chemistry and
Analytical Sciences, Walton Hall, MK7 6AA, Milton Keynes, UK. E-mail: s.missailidis@open.ac.uk
518

HEPARANASE INHIBITOR
519
Figure 1. (2-{4-[(E)-3-(4-Bromophenyl)acryloylamino]-3-fluoro-phenyl}benzooxazol-5-yl) acetic acid.
Scheme 1. Synthesis of (2-{4-[(E)-3-(4-bromophenyl)acryloylamino]-3-fluoro-phenyl}benzooxazol-5-yl)
acetic acid (9).
of the methyl ester was performed using LiOH in N,N-dimethylformamide. The final
product (9) was obtained as the free benzooxazolyl acetic acid and purified by cation
exchange column chromatography using a mixture of methanol and formic acid as
eluent. Other similar compounds, having different functional groups, can be obtained
by the same synthetic route.
EXPERIMENTAL
All chemicals were purchased from Sigma Aldrich (Gillingham, Dorset, UK)
and used without further purification. Thionyl chloride was distilled before use.

520
C. DA PIEVE, P. PATEL, AND S. MISSAILIDIS
4-Hydroxy-3-nitrophenylacetic acid, 3-fluoro-4-nitrobenzoic acid, and 4-bromoci-
nammic acid were purchased from Apollo Scientific Limited (Stockport, UK).
4-Bromocinnamoyl chloride (7) and compounds 1 and 2 were synthesised according
to literature.[8,10] Thin-layer chromatography (TLC) was carried out on 0.25-mm
silica gel on polyethylene terephthalate (PET) foils (Fluka) with a fluorescent indi-
cator visualized using ultraviolet (UV) light or Ce-P-Mo staining solution. Column
chromatography was performed on silica gel (particle size 0.035–0.070 mm). Anion
exchange chromatography was performed on Dowex Marathon A (Sigma Aldrich).
NMR spectra were recorded on Jeol 300-MHz or Jeol 400-MHz spectrometers.
The chemical shifts are relative to residual solvent protons as reference. Electro-
spray ionization (ESI) mass spectra (MS) were recorded on a VG Quattro
spectrometer (Fisons Instruments). Elemental analyses were performed by Medac
Ltd. (Egham, UK).
Methyl [3-(3-Fluoro-4-nitro benzoylamino)-4-hydroxy-phenyl]
Acetate (4)
3-Fluoro-4-nitrobenzoic acid (2.19 g, 9.93 mmol) was suspended in dry CH2Cl2
(20 ml). SOCl2 (2 ml) was added, followed by few drops of dimethylformamide
(DMF). The mixture was refluxed overnight under N2. The solvent was removed
under vacuum, and the product (3) was used without further purification. Com-
pound 2 (1.8 g, 9.93 mmol) and an equimolar quantity of 3 were dissolved in dry
CH2Cl2 (45 ml) containing triethylamine (3.5 ml). The solution was stirred under
N2 overnight at room temperature and then washed with HCl 1 M (3 ꢀ 25 ml).
The organic phase was dried (MgSO4) and evaporated under vacuum. The product
was recrystallized from CH2Cl2=methanol (100:1) as yellow needles. Yield 70%. Rf
1
0.23 (100:1 CH2Cl2=methanol). H NMR (300 MHz, DMSO) d 3.57 (s, 2H, CH2),
3.59 (s, 3H, CH3), 6.86 (d, 1H, aromatic, J ¼ 8.25), 6.96 (dd, 1H, aromatic,
J ¼ 8.25, 2.01), 7.46 (d, 1H, aromatic, J ¼ 2.01), 7.94–8.31 (m, 3H, aromatic), 9.80
(bs, NH þ OH). 13C (75 MHz, DMSO) d 39.38, 51.63, 115.86, 117.67 (d,
J ¼ 22.30), 124.36 (d, J ¼ 4.37), 124.60, 124.70, 126.20, 126.55 (d, J ¼ 2.41), 127.54,
138.40 (d, J ¼ 7.39), 141.44 (d, J ¼ 7.47), 149.26, 154.29 (d, J ¼ 261), 162.47,
171.89. MS (ESI): m=z (%) ¼ 347 (100) [M]þ. Anal. calcd. for C16H13FN2O6: C,
55.18; H, 3.76; N, 8.04. Found: C, 55.03; H, 3.77; N, 7.88.
Methyl [2-(3-Fluoro-4-nitrophenyl)benzooxazol-5-yl] Acetate (5)
Compound 4 (2.3 g, 6.60 mmol) was added to a solution of p-toluenesulfonic acid
monohydrate (0.95 g, 5 mmol) in dry toluene (200 ml). The mixture was refluxed over-
night using a Dean–Stark apparatus. The solvent was removed under vacuum, and the
product was purified by chromatography on silica gel. Elution was started with 100%
hexane, and the polarity of the eluent was increased gradually to 3:1 hexane=ethyl acet-
ate. Yield: 60%. Rf ¼ 0.64 (3:1 hexane=ethyl acetate). 1H NMR (300 MHz, CDCl3) d
3.65 (s, 3H, CH3), 3.71 (s, 2H, CH2), 7.31 (dd, 1H, aromatic, J ¼ 8.43, 1.65), 7.51
(d, 1H, aromatic, J ¼ 8.43), 7.66 (d, 1H, aromatic, J ¼ 1.65), 8.08–8.17 (m, 3H,
aromatic). 13C (75 MHz, CDCl3) d 40.97, 52.21, 110.90, 117.31 (d, J ¼ 23.61),
121.46, 123.27 (d, J ¼ 4.37), 126.95 (d, J ¼ 2.49), 128.06, 131.53, 133.86 (d, J ¼ 8.67),

HEPARANASE INHIBITOR
521
138.64 (d, J ¼ 9.88), 142.08, 152.07 (d, J ¼ 278), 157.43, 159.84, 171.75. MS (ESI): m=z
(%) ¼ 331 (100) [M þ H]þ, 371 (37) [M þ H þ K]þ. Anal. calcd. for
C16H11FN2O5 ꢁ 0.2H2O: C, 57.54; H, 3.44; N, 8.39. Found: C, 57.54; H, 3.48; N, 8.40.
Methyl [2-(4-Amino-3-fluorophenyl)benzooxazol-5-yl] Acetate (6)
Compound 5 (1.2 g, 3.63 mmol) was dissolved in ethyl acetate (20 ml), then 10%
Pd=C (30 mg) was added and H2 was bubbled through the stirred mixture for 5 h.
The catalyst was removed by filtration, and the filtrate was evaporated under
vacuum. Yield quantitative. Rf 0.55 (3:2 hexane=ethyl acetate). 1H NMR
(400 MHz, CDCl3) d 3.07 (s, 3H, CH3), 3.13 (s, 2H, CH2), 4.79 (bs, NH2),
6.27–7.24 (m, 6H, aromatic). 13C (75 MHz, DMSO) d 40.48, 51.53, 109.65, 113.73
(d, J ¼ 21.12), 114.16, 115.59 (d, J ¼ 4.90), 119.40, 124.21 (d, J ¼ 1.81), 125.26,
130.07, 139.69 (d, J ¼ 13.05), 142.13, 149.15, 149.98 (d, J ¼ 238.27), 162.94 (d,
J ¼ 3.16), 171.39. MS (ESI): m=z (%) ¼ 301 (100) [M þ H]þ. Anal. calcd. for
C16H13FN2O3 ꢁ H2O: C, 60.38; H, 4.75; N, 8.80. Found: C, 60.25; H, 4.52; N, 9.04.
Methyl (2-{4-[(E)-3-(4-Bromo-phenyl)acryloylamino]-
3-fluorophenyl}benzooxazol-5-yl) Acetate (8)
A solution of 7 (817 mg, 3.33 mmol) in 10 ml of dry CH2Cl2 was added drop-
wise to a solution of 6 (1 g, 3.33 mmol) in dry CH2Cl2 (30 ml) and Et3N (1 ml). The
mixture was stirred at room temperature overnight. The solvent was removed under
vacuum and the residue was washed with THF=water (1:1). Product 8 was further
purified by chromatography on silica gel starting with CH2Cl2 as eluent and then
gradually increasing the polarity to 50:1 CH2Cl2=methanol. Yield 90%. Rf 0.31
1
(100:1 CH2Cl2=methanol). H NMR (400 MHz, DMSO) d 3.62 (s, 3H, CH3), 3.83
(s, 2H, CH2), 7.15 (s, 1H, CH), 7.20 (s, 1H, CH), 7.30–8.53 (m, 10H, aromatic),
10.24 (s, 1H, NHCO). 13C (75 MHz, DMSO) d 40.33, 51.70, 110.49, 114.03 (d,
J ¼ 21.73), 120.45, 122.17 (d, J ¼ 8.67), 122.44, 122.81, 123.22, 123.84 (d, J ¼ 3.77),
127.01, 129.74, 129.99 (d, J ¼ 11.16), 131.36, 131.99, 133.88, 140.05, 141.58,
149.23, 152.39 (d, J ¼ 246.34), 161.44 (d, J ¼ 2.49), 164.00, 171.64. MS (ESI): m=z
(%) ¼ 507 (100) [M]þ. Anal. calcd. for C25H18BrFN2O4: C, 58.95; H, 3.56; N, 5.50.
Found: C, 58.24; H, 3.56; N, 5.39.
(2-{4-[(E)-3-(4-Bromophenyl)acryloylamino]-3-fluoro-
phenyl}benzooxazol-5-yl) Acetic Acid (9)
LiOH monohydrate (0.25 g, 5.9 mmol) was added to a solution of 8 (1 g,
1.96 mmol) in DMF (40 ml) and stirred overnight at room temperature. The solvent
was removed under vacuum, and the residue was purified by ion-exchange chroma-
tography over Dowex Marathon A previously washed with water and then
methanol. The product was eluted with methanol=formic acid 3 M (8:2) and recrys-
tallized from methanol.
1
Yield 28.3%. H NMR (300 MHz, DMSO) d 3.71 (s, 2H, CH2), 7.08 (s, 1H,
CH), 7.15 (s, 1H, CH), 7.20–8.53 (m, 10H, aromatic), 10.24 (s, 1H, NHCO), 12.40
(bs, COOH). 13C (75 MHz, DMSO) d 40.30, 110.38, 114.03 (d, J ¼ 21.73), 120.46,

522
C. DA PIEVE, P. PATEL, AND S. MISSAILIDIS
122.19 (d, J ¼ 7.99), 122.46, 122.80, 123.26, 123.84 (d, J ¼ 2.41), 127.11, 129.76,
129.99 (d, J ¼ 11.16), 132.01, 132.07, 133.89, 140.07, 141.53, 149.13, 152.39 (d,
J ¼ 245.74), 161.36 (d, J ¼ 3.09), 164.02, 172.76. MS (ESI): m=z (%) ¼ 493 (100)
[M]þ. Anal. calcd. for C24H16BrFN2O4 ꢁ 1.8H2O: C, 54.62; H, 3.74; N, 5.31. Found:
C, 54.62; H, 3.74; N, 5.10.
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