Novel synthetic route to the C-nucleoside, 2-deoxy benzamide riboside

Article abstract of DOI:10.1016/j.bmcl.2012.06.069

2-Deoxy-C-nucleosides are a subcategory of C-nucleosides that has not been explored extensively, largely because the synthesis is less facile. Flexible synthetic procedures giving access to 2-deoxy-C-nucleosides are therefore of interest. To exemplify the versatility and highlight the limitations of a synthetic route recently developed to that effect, the first synthesis of 2-deoxy benzamide riboside is reported. Biological properties of this novel C-nucleoside are also discussed.

Full text of DOI:10.1016/j.bmcl.2012.06.069

Bioorganic & Medicinal Chemistry Letters 22 (2012) 5204–5207
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Novel synthetic route to the C-nucleoside, 2-deoxy benzamide riboside
Rebecca R. Midtkandal a, Philip Redpath a, Samuel A. J. Trammell b, Simon J. F. Macdonald c,
Charles Brenner b, Marie E. Migaud a,
⇑
a Queen’s University Belfast, John King Laboratory, School of Pharmacy, Lisburn Road, Belfast, BT9 7BL, Northern Ireland, UK
b University of Iowa, Department of Biochemistry, Carver College of Medicine, 51 Newton Rd, 4-403 BSB, Iowa City, IA 52242, USA
c GlaxoSmithKline, Stevenage, SG1 2NY, UK
a r t i c l e i n f o
a b s t r a c t
Article history:
2-Deoxy-C-nucleosides are a subcategory of C-nucleosides that has not been explored extensively, largely
because the synthesis is less facile. Flexible synthetic procedures giving access to 2-deoxy-C-nucleosides
are therefore of interest. To exemplify the versatility and highlight the limitations of a synthetic route
recently developed to that effect, the first synthesis of 2-deoxy benzamide riboside is reported. Biological
properties of this novel C-nucleoside are also discussed.
Received 16 May 2012
Revised 20 June 2012
Accepted 22 June 2012
Available online 28 June 2012
Ó 2012 Elsevier Ltd. All rights reserved.
Keywords:
C-Nucleoside
Benzamide riboside
Intramolecular cyclisation
Inosine monophosphate dehydrogenase
Nicotinamide adenine dinucleotide
C-Nucleosides are an important class of hydrolytically and enzy-
matically stable compounds and have been found to be important
probes in the pursuit of novel antiviral and antiproliferative drugs.1
There are several known ribose-based C-nucleosides that are potent
antiproliferatives. Famous examples are formycin, tiazofurin and
benzamide riboside (Fig. 1).2 Carboxamide derivatives of C-nucleo-
sides, such as, benzamide riboside and tiazofurin, are based on
nicotinamide riboside (NR), one of three NAD precursor vitamins.3
As prodrugs, these compounds must gain entry to target cells, be
phosphorylated by NR kinase 1 or NR kinase 23a and then be ade-
nylylated by NMN adenylyltransferase (NMNAT)4 to a derivative
of NAD.5 Benzamide riboside (BR) has been shown, as its benzamide
adenine dinucleotide (BAD) metabolite, to exhibit significant cyto-
toxicity against a variety of human tumour cell lines.6 The mecha-
nism of cytotoxicity of BAD and TAD depends on inhibition of
inosine monophosphate dehydrogenase (IMPDH),5 thus resulting
in the depletion of the intracellular GTP pool.7
the possibility for synthetic desymmetrisation, if either or both
analogues show interesting antiproliferative properties.8
The syntheses of C-nucleosides are, in general, quite difficult,
low yielding and do not allow access to large series of derivatives
for SAR studies, in particular with regards to the aromatic moiety.9
Strategies have remained linear, resulting in a limited number of
options with regards to the possible structural and functional com-
binations that can potentially enhance pharmacodynamic/pharma-
cokinetic properties of the novel compounds.
Our goal is to synthesise analogues of BR to probe each step
in the process of NR to NAD biotransformation: stability of the
glycosidic linkage to hydrolysis/phosphorolysis3c import, phos-
phorylation by NR kinases,3a,d adenylylation,4 and inhibition of sub-
sequent target enzymes. Definition of NR analogue specificity may
allow probes to be developed for cancer or for dissection of the met-
abolic effects of NR.3f,h Analogues with variation on the aromatic
moiety, such as tiazofurin and its selenium equivalent, selenofurin
have yielded precursors to potent IMPDH inhibitors. Similarly, the
2’ position of tiazofurin was shown to be tolerant to manipulation,
with various compounds retaining cytotoxicity, for example, the
20-amido-tiazofurin analogue.10 Similarly the epoxide functionality
across the 20–30 positions displayed the most pronounced increase
in cytotoxic activity against certain lymphoma cell lines, being al-
most 30 times more potent than the parent compound tiazofurin.11
While BR and tiazofurin share similar biological profiles and poten-
cies, similar analogues have not been reported for BR. We therefore
proposed that BR-analogues incorporating different carbohydrate
substitution could also offer novel biological opportunities. As a
Additionally, the discovery that select nucleosides with L-con-
figuration have biological activity has lead to a breakthrough in
antiviral chemotherapy. The antiviral mechanism has shown that
such L-nucleosides were phosphorylated by cellular kinases, and
selectively interacted with viral polymerases, while remaining
relatively innocuous to normal cellular processes.8 These results
highlight the need for new synthetic strategies generating both
enantiomers of nucleoside analogues equally well, while retaining
⇑
Corresponding author.
E-mail address: m.migaud@qub.ac.uk (M.E. Migaud).
0960-894X/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2012.06.069

R. R. Midtkandal et al. / Bioorg. Med. Chem. Lett. 22 (2012) 5204–5207
5205
H
N
N
O
O
NH2
H O
HO
NH2
N
N
H O
HO
OH
Benzamide Riboside
O
OH
Formycin
S
NH2
O
H O
N
O
H O
OH
Tiazofurin
Figure 1. Examples of potent antiproliferatives. Formycin benzamide riboside and tiazofurin.
result, 2- and 4-deoxypyranoside analogues of BR have been gener-
ated.12 We also proposed to test whether biological properties
might be retained in the 2-deoxyriboside series of C-nucleosides.
To access this type of novel 2-deoxy C-nucleoside furanosides,
we have devised a synthetic methodology involving key exo-alkene
intermediates.13 Herein, we report the synthesis of the meta-vinyl
substituted phenyl exo-alkene intermediate which provided the
necessary chemical flexibility to access the meta-carboxamide sub-
stituent essential to BR analogues. The meta-vinyl group was cho-
sen as a direct result of the electron withdrawing nature of the
preferred amide or nitrile aromatic functionality, which had pro-
ven incompatible to the TMSNTf2 mediated cyclization conditions
necessary to access the exo-alkene furanoside intermediates.13
The procedure began with the synthesis of acetal 3 from the
reaction between easily accessible intermediates allylsilane 1 and
chloroacetal 2 in 59% yield.13 Subsequently, using optimized condi-
tions, acetal 3 was intramolecularly cyclised to provide a 1.3:1 (cis:-
trans) mixture of furanoside 4 using 2 equiv of TMSNTf2 in 54%
combined yield.14 With this intermediate mixture in hand, the dou-
ble oxidation of the unsaturated functionalities was performed
using optimized conditions for ozonolysis,13 to achieve the cis and
trans isomers of keto-aldehyde product 5 in 87% combined yields.
This diastereomeric mixture of 5 was easily separated on silica with
the desired cis-isomer isolated in 50% yield. Unfortunately, at-
tempts failed to convert the trans-isomer to the cis-isomer by epi-
merisation. Density Functional Theory (DFT) calculations showed
that the trans-isomer was 16 kcal/mol lower in energy relative to
the cis-isomer,15 and this was therefore the preferred configuration
for the compound, with too high an energy difference to easily
achieve efficient epimerisation from trans- to cis-configuration
without decomposition.
Difficulties in accessing the essential amide functionality di-
rectly from aldehyde 5 led us to proceed through the carboxylic
acid derivative 6. An oxidising system of sodium chlorite, hydrogen
peroxide and sodium dihydrogen phosphate was successfully
implemented to obtain 6 in high yields.18 At this point some con-
flicting issues arose: the amide formation was inevitably going to
involve an amine, which had the potential to epimerise the stereo-
centre adjacent to the ketone, as observed in previous studies.13
The most straightforward route from a carboxylic acid to an amide
is via an acid chloride and subsequent reaction with the required
amine. The latter was successfully carried out using aqueous
ammonia. Gratifyingly, this reaction did not cause noticeable epi-
merisation of the vulnerable stereocentre and quantitatively pro-
vided compound 7.17
sulfonic acid, the silyl protecting group was cleaved at 60 °C in
DMF, albeit in 56% yield. The final reduction using sodium triacet-
oxyborohydride proceeded smoothly, giving the final compound,
2-deoxy-benzamide riboside cis-8, in 95% yields, with the relative
stereochemistry confirmed by NOE experiments, (Scheme 1).13,19
To the best of our knowledge, this compound has not yet been
synthesised, and thus never been tested for targeted biological and
enzymatic activity. It was anticipated that the removal of the 2’-
hydroxyl group on the ribose would affect the ability of the com-
pound to hydrogen bond with polar residues of the binding pockets
in NRK,3d,g NMNAT20 and IMPDH.21 Therefore to retain activity, it is
vital that the resulting NAD analogue structure not only fit into the
IMPDH pocket to achieve inhibition, but that its metabolic precur-
sors be recognised and metabolised effectively by the enzymes that
convert it into an NAD analogue.7,22
The target molecule cis-8 was therefore tested for activity
against a number of cancerous cell lines. The cell lines were:
RKO—Human colon carcinoma, HT29 - Human colorectal adeno-
carcinoma, HCT116—Human colorectal carcinoma, H460 - Human
large lung cell carcinoma, MCF7—Human breast adenocarcinoma
(pleural effusion metastatic site), T47D Human breast ductal carci-
noma, PC3—Human prostate adenocarcinoma (bone metastatic
site) and DU145—Human prostate carcinoma (brain metastatic
site). Some of these cell lines have been shown to be sensitive to
tiazofurin and benzamide riboside at concentrations that range
from 50 lM (benzamide riboside in MCF7) to 500 lM (cell line
PC-3).23 Despite incubation at concentrations up to 1 mM, no cell
line displayed a significant level of inhibition upon incubation with
cis-8 compared to the blank sample of pure DMSO.
In work preceding the discovery of the NRK-dependent NAD
biosynthetic pathway3 and the evidence that tiazofurin was an
excellent substrate for NRK,3d tiazofurin and benzamide riboside
were proposed to enter cells via nucleoside transporters. While a
nicotinamide riboside specific transporter has been identified in
yeast, the human homolog still remains undiscovered. To test the
hypothesis that cis-8 fails as a cytotoxic agent because of a deficit
in the biotransformation to NAD rather than or in addition to lim-
ited transport, we incubated the compound at 0.5 mM with 1 mM
ATP and sufficient NR kinase 2 for complete conversion of 0.5 mM
NR to NMN in 30 min. Reactions monitored for 2 h by HPLC and
LC–MS indicated that the compound is totally resistant to conver-
sion to the corresponding monophosphate. Earlier a 2-deoxy deriv-
ative of tiazofurin24 was synthesized and shown to be lacking in
cytotoxic activity. Our data strongly suggest that 2-deoxy substitu-
tions are not tolerated by NR kinases,3a,d thereby severely compro-
mising their pharmacological activities.
Deprotection of the TBDPS moiety could not be carried out using
standard protocols, either by using a fluoride source or other basic
conditions without causing the epimerisation.16 Indeed, attempts
to use TBAF gave a mixture of products in low yields, with complete
scrambling of the stereocentre. However, when using p-toluene
The present work provides the direction which the chemistry of
C-nicotinamide riboside modified on the sugar moiety must take
and that is attempting to by-pass the NRK step in accessing
in situ NAD analogues, putative inhibitors of IMPDH and other

5206
R. R. Midtkandal et al. / Bioorg. Med. Chem. Lett. 22 (2012) 5204–5207
Cl
O H
O
O
TBDPSO
TMS
(i)
TBDPSO
TMS
(iv)
2
OMe
OMe
(ii)
(iii)
O
3
59%
1
89%
( )
TBDPSO
( )
TBDPSO
( )
TBDPSO
O
O
O
(vii)
(v)
(vi)
O
O
O
O
H O
4
54% 1.3:1 cis: trans
6
90%
5
87% (50% cis isolated yield)
Diastereomers separable
( )
TBDPSO
( )
O
O
(viii)
(ix)
(x) 56%
(xi)
H O
O
O
95%
H O
O
H
N
H N
2
2
7
cis-8
95%
Scheme 1. Complete synthetic route to the novel C-nucleoside 2-deoxy benzamide riboside, cis-8. (i) TMSCH2Li, THF, ꢁ78 °C; (ii) TBDPSCl, Et3N, DMAP. DCM, rt; (iii) Et3N,
DCM, 0 °C ꢁrt; (iv) TMSNTf2, DCM, ꢁ78 °C; (v) O3; (vi) PPh3, DCM; (vii) NaClO2, H2O2, NaH2PO4, MeCN, H2O; (viii) SOCl2, PhMe, (ix) NH4OH, (x) pTSA, DMF, 60 °C, (xi)
NaBH(OAc)3, AcOH, MeCN, 0 °C.
M.; Watanabe, K. A. J. Med. Chem. 1997, 40, 1287; (l) Štefko, M.; Pohl, R.; Hocek,
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2. (a) Declercq, E.; Balzarini, J.; Madej, D.; Hansske, F.; Robins, M. J. Med. Chem.
NAD dependent enzymes. Pre-forming the monophosphate of cis-8
and synthesising the NAD analogue incorporating cis-8 will give an
indication as to whether the dinucleotide compound has the ability
to inhibit IMPDH, and would substantiate the proposal made by
Robins.24 This work is on-going in our laboratory, and will be com-
bined to isolated enzyme kinetic experiments so to obtain a clear
understanding of the activation pathway requirements.
The present results demonstrate the first successful synthesis of
2-deoxy benzamide riboside, cis-8. The vinyl group on the key
intermediate 4 was successfully converted to the amide group in
the meta-position of the aromatic moiety, proving that the cyclisa-
tion reaction developed in our previous work can provide interme-
diates for a wide range of biologically relevant C-nucleosides. This
interesting advancement in the methodology of useful C-nucleo-
sides will increase the existing pool of synthetic procedures avail-
able for the pursuit of novel C-nucleosides.
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Acknowledgments
8. Zemlicka, J. Pharmacol. Ther. 2000, 85, 251.
9. (a) Wu, Q. P.; Simons, C. Synthesis 2004, 1533; (b) Štambasky´, J.; Hocek, M.;
This work was supported by the John King’s funds (School of
Pharmacy, QUB) and an FP6 EST Marie Curie Scholarship (MEST-
CT-2005-020351). Dr C. Higgins from the Centre for Cancer Research
and Cell Biology at Queen’s University Belfast is thanked for con-
ducting the antiproliferative assays.
ˇ
´
Kocovsky, P. Chem. Rev. 2009, 109, 6729.
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10. Popsavin, M.; Torovic, L.; Svircev, M.; Kojic, V.; Bogdanovic, G.; Popsavin, V.
Bioorg. Med. Chem. Lett. 2006, 16, 2773.
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Popsavin, V. Tetrahedron 2009, 65, 7637.
12. Redpath, P.; Macdonald, S. J. F.; Migaud, M. E. Org. Lett. 2008, 10, 3323.
13. Midtkandal, R. R.; Macdonald, S. J. F.; Migaud, M. E. Chem. Commun. 2010, 46,
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References and notes
14. Acetal 3 (0.50 g, 0.9 mmol, 1 equiv) was dissolved in dry DCM (ꢀ1.5 mL/mmol)
and the reaction mixture cooled to ꢁ78 °C. TMSNTf2 (2 equiv) was added and
the reaction mixture stirred for 5 min. It was then filtered through a pad of
silica gel using 25% EtOAc in hexane or pet. ether as eluent. The filtrate was
concentrated in vacuo and the residue purified by column chromatography (0–
5% EtOAc in hexane on silica gel) and yielded 4 (0.22 g, 54%) as a colourless oil.
The diastereomers were not separated. Diastereomeric ratio was measured at a
ratio 1.3:1 (cis:trans) based on 1H NMR.
1. (a) Wang, L.; Schultz, P. G. Chem. Commun. 2002, 1; (b) Henry, A. A.; Romesberg,
F. E. Curr. Opin. Chem. Biol. 2003, 7, 727; (c) Kool, E. T.; Morales, J. C.; Guckian, K.
M. Angew. Chem., Int. Ed. 2000, 39, 990; (d) Kool, E. T. Acc. Chem. Res. 2002, 35,
936; (e) Matsuda, S.; Fillo, J. D.; Henry, A. A.; Rai, P.; Wilkens, S. J.; Dwyer, T. J.;
Geierstanger, B. H.; Wemmer, D. E.; Schultz, P. G.; Spraggon, G.; Romesberg, F.
E. J. Am. Chem. Soc. 2007, 129, 10466; (f) Leconte, A. M.; Hwang, G. T.; Matsuda,
S.; Capek, P.; Hari, Y.; Romesberg, F. E. J. Am. Chem. Soc. 2008, 130, 2336; (g)
Leconte, A. M.; Joubert, N.; Hocek, M.; Romesberg, F. E. Chem. Bio. Chem 2008, 9,
2796; (h) Franchetti, P.; Cappellacci, L.; Griffantini, M.; Barzi, A.; Nocentini, G.;
Yang, H. Y.; O’Connor, A.; Jayaram, H. N.; Carrell, C.; Goldstein, B. M. J. Med.
Chem. 1995, 38, 3829; (i) Walker, J. A.; Liu, W.; Wise, D. S.; Drach, J. C.;
Townsend, L. B. J. Med. Chem. 1998, 41, 1236; (j) Aketani, S.; Tanaka, K.;
Yamamoto, K.; Ishihama, A.; Cao, H.; Tengeiji, A.; Hiraoka, S.; Shiro, M.;
Shionoya, M. J. Med. Chem. 2002, 45, 5594; (k) Pankiewicz, K. W.; Lesiak, K.;
Zatorski, A.; Goldstein, B. M.; Carr, S. F.; Sochacki, M.; Majumdar, A.; Seidman,
(S⁄,R⁄)-4 (cis): dH (400 MHz; CDCl3): 1.06 (9H, s, Si-C-CH3), 2.57–2.64 (1H, m,
CH(Ar)-CH2), 2.89 (1H, dd, J = 5.4, 15.1 Hz, CH(Ar)-CH2), 3.83 (1H, dd, J = 4.5,
10.7 Hz, TBDPSO-CH2), 3.90 (1H, dd, J = 4.2, 10.7 Hz, TBDPSO-CH2), 4.56-4.58
(1H, m, O-CH-C@), 4.92 (1H, dd, J = 5.4, 10.7 Hz, O-CH-Ar), 4.99–5.01 (1H, m,
C@CH2), 5.09–5.11 (1H, m, C@CH2), 5.21 (1H, d, J = 11.0 Hz, CH = CH2), 5.71 (1H,
d, J = 17.6 Hz, CH@CH2), 6.68 (1H, dd, J = 11.0, 17.6 Hz, CH = CH2), 7.22–7.47
(10H, m, Ar-H), 7.71–7.75 (4H, m, Ar-H). dC(100 MHz; CDCl3): 19.3 (Si-C-CH3),
26.8 (Si-C-CH3), 42.6 (CH(Ar)-CH2), 66.8 (TBDPSO-CH2), 80.4 (O-CH-Ar), 82.0
(O-CH-C@), 105.8 (C@CH2), 113.9 (CH@CH2), 124.1 (Ar-CH), 125.4 (Ar-CH),

R. R. Midtkandal et al. / Bioorg. Med. Chem. Lett. 22 (2012) 5204–5207
5207
125.7 (Ar-CH), 127.6 (Ar-CH), 127.7 (Ar-CH), 128.5 (Ar-CH), 129.6 (Ar-CH),
129.6 (Ar-CH), 133.4 (Ar-C), 133.6 (Ar-C), 135.7 (Ar-CH), 135.8 (Ar-CH), 136.8
(CH@CH2), 137.7 (Ar-C), 142.0 (Ar-C), 149.2 (C@CH2). (S⁄,S⁄)-4 (trans): dH
(400 MHz; CDCl3): 1.07 (9H, s, Si-C-CH3), 2.57–2.64 (1H, m, CH(Ar)-CH2), 3.05
(1H, m, CH(Ar)-CH2), 3.78 (1H, dd, J = 5.0, 10.9 Hz, TBDPSO-CH2), 3.82 (1H, dd,
J = 4.0, 10.9 Hz, TBDPSO-CH2), 4.73 (1H, br s, O-CH-C@), 4.98 (1H, dd, J = 2.1,
4.1 Hz, C=CH2), 5..08–5.11 (1H, m, C@CH2), 5.13 (1H, t, J = 7.2 Hz, O-CH-Ar),
5.25 (1H, d, J = 11.0 Hz, CH@CH2), 5.75 (1H, d, J = 17.6 Hz, CH@CH2), 6.73 (1H,
dd, J = 11.0, 17.6 Hz, CH@CH2), 7.22–7.47 (10H, m, Ar-H), 7.71–7.75 (4H, m, Ar-
H). dC (100 MHz; CDCl3): 19.3 (Si-C-CH3), 26.9 (Si-C-CH3), 41.6 (CH(Ar)-CH2),
66.9 (TBDPSO-CH2), 79.6 (O-CH-Ar), 81.8 (O-CH-C@), 106.1 (C@CH2), 113.9
(CH@CH2), 123.8 (Ar-CH), 125.3 (Ar-CH), 125.7 (Ar-H), 127.6 (Ar-CH), 127.7
(Ar-CH), 128.6 (Ar-CH), 129.6 (Ar-CH), 129.6 (Ar-CH), 133.4 (Ar-C), 133.6 (Ar-
C), 135.7 (Ar-CH), 135.8 (Ar-CH), 136.9 (CH@CH2), 137.7 (Ar-C), 142.9 (Ar-C),
CH), 129.8 (Ar-CH), 129.8 (Ar-CH), 129.9 (Ar-CH), 132.6 (Ar-C), 133.3 (Ar-C),
133.8 (Ar-C), 134.8 (Ar-CH), 135.4 (Ar-CH), 135.6 (Ar-CH), 141.3 (Ar-C), 168.7
(CONH2), 213.2 (CO). m
max(film)/cm-1: 3433 (br s), 2929 (s), 1761 (m), 1662 (s),
1428 (m), 1261 (m), 1112 (s), 800 (s), 702 (s). m/z (ES): 496.1939 (M+Na+),
calculated for C28H31NO4Si + Na+: 496.1920 (3.8 ppm).
18. Wuts, P. G. M.; Greene, T. W. Greene’s Protective Groups in Organic Synthesis, 4th
ed.; John Wiley & Sons inc.: Hoboken, 2006.
19. Amide 7 (0.24 g, 0.51 mmol, 1 equiv) was dissolved in DMF (25 mL) and p-
toluenesulfonic acid (0.13 g, 0.8 mmol, 1.5 equiv) was added. The reaction
mixture was stirred at 60 °C for 4 days. The reaction was then quenched with
saturated aqueous NaHCO3 (25 mL) and the aqueous layer extracted with
EtOAc (3 ꢂ 50 mL). The combined organic layers were dried with MgSO4,
filtered and concentrated in vacuo. Purification by column chromatography (0–
15% MeOH in DCM on silica gel) provided the desilylated intermediate.
Subsequently, NaBH(OAc)3 (95 mg, 0.5 mmol, 5 equiv) was dissolved in MeCN
148.6 (C@CH2). m
max(film)/cmꢁ1: 3059 (m), 2930 (s), 1428 (m), 1245 (s), 1113
(s), 907 (m), 702 (s). m/z (ES): 397.1629 (M-tBu), calcd for
397.1624 (1.3 ppm).
15. Calculation carried out using the Gaussian 03, Revision D.01.
C
26H25O2Si:
(4 mL). After cooling the resulting solution to 0 °C, AcOH (60 lL, 1 mmol,
10 equiv) was added. The purified intermediate (21 mg, 0.09 mmol, 1 equiv)
was dissolved in MeCN (3 mL) and added to the cooled solution to be stirred at
0 °C for 1 h. MeOH (3 ꢂ 8 mL) was subsequently added, with volatiles removed
under reduced pressure between each portion added. Purification by column
chromatography (0–15% MeOH in DCM on silica gel) yielded title compound
cis-8 (20 mg, 95%) as a colourless oil.
16. Keto-aldehyde compound
5 (0.36 g, 0.75 mmol, 1 equiv) was dissolved in
MeCN (15 mL) and the resulting solution cooled to 0 °C. A solution of NaH2PO4
(43 mg, 0.4 mmol, 0.5 equiv) in water (1 mL) was added, followed by H2O2
(0.14 mL, 30% in water, 1.3 mmol, 1.8 equiv) and a solution of NaClO2 (0.12 mg,
1.5 mmol, 2 equiv) in water (1 mL). The reaction mixture was stirred at rt for
2 h, before quenching with Na2SO3 (0.20 g, 1.5 mmol, 2 equiv) and extraction
with EtOAc (3 ꢂ 20 mL). The combined organic layers were dried with MgSO4,
filtered and concentrated in vacuo to yield compound 6 (0.32 g, 90%) as a
colorless oil.
dH (400 MHz; CDCl3): 1.07 (9H, s, Si-CH3), 2.63 (1H, dd, J = 8.7, 18.0 Hz, CH(Ar)-
CH2), 3.04 (dd, 1H, J = 6.9, 18.0, Hz, CH(Ar)-CH2), 3.98–4.16 (2H, m, TBDPSO-
CH2), 4.27 (1H, s, O-CH-CO), 5.82 (1H, dd, J = 6.9, 8.7 Hz, O-CH-Ar), 7.39–7.47
(7H, m, Ar-H), 7.53 (1H, t, J = 7.7 Hz, Ar-H), 7.66–7.75 (4H, m, Ar-H), 8.11 (1H, d,
J = 7.7 Hz, Ar-H), 8.18 (1H, s, Ar-H), 10.54 (1H, br s, COOH). dC (100 MHz;
CDCl3): 19.2 (Si-C-CH3), 26.7 (Si-C-CH3), 45.2 (CH(Ar)-CH2), 65.4 (TBDPSO-CH2),
78.5 (O-CH-Ar), 81.6 (O-CH-CO), 127.6 (Ar-CH), 127.9 (Ar-CH), 128.1 (Ar-C),
129.0 (Ar-CH), 129.7 (Ar-CH), 129.9 (Ar-CH), 129.9 (Ar-CH), 131.3 (Ar-CH),
132.3 (Ar-C), 132.7 (Ar-C), 135.5 (Ar-CH), 135.7 (Ar-CH), 142.2 (Ar-C), 171.0
(COOH), 214.5 (CO). m/z (ES): 497.1772 (M+Na+), calculated for C28H30O5Si +
Na+: 497.1760 (2.4 ppm).
dH (500 MHz; MeOH-d4): 1.97 (1H, ddd, J = 6.0, 10.5, 13.1 Hz, CH(Ar)-CH2), 2.26
(1H, ddd, J = 1.6, 5.4, 13.1 Hz, CH(Ph)-CH2), 3.70 (1H, dd, J = 4.9, 11.4 Hz, HO-
CH2), 3.72 (1H, dd, J = 4.9, 11.4 Hz, HO-CH2), 3.99 (1H, dt, J = 2.5, 4.9 Hz, O-CH-
CHOH), 4.35–4.37 (1H, m, HO-CH), 5.20 (1H, dd, J = 5.4, 10.5 Hz, O-CH-Ar), 7.45
(1H, t, J = 7.7 Hz, Ar-H), 7.60 (1H, d, J = 7.7 Hz, Ar-H), 7.80 (1H, td, J = 1.7, 7.7 Hz,
Ar-H), 7.94 (1H, td, J = 1.7, 7.7 Hz, Ar-H). dC (125 MHz; MeOH-d4): 45.2
(CH(Ph)-CH2), 64.1 (HO-CH2), 74.5 (HO-CH), 81.3 (O-CH-CHOH), 89.5 (O-CH-
Ph), 126.4 (Ar-CH), 128.0 (Ar-CH), 129.7 (Ar-CH), 130.9 (Ar-CH), 135.2 (Ar-C),
144.1 (Ar-C), 172.5 (CO). m/z (ES): 260.0887 (M+Na+), calcd for
C
12H15NO5 + Na+: 260.0899 (ꢁ4.6 ppm). HPLC MS-MS analyses indicated a
product purity >92%, with the main contaminant possessing the same exact
mass as cis-8 at MW+H+ (m/z 238.1063 vs m/z 238.1067) and MW+Na+ (m/z
260.0885 vs m/z 260.0895). The data indicated identical fragment masses,
observed with some difference in ratio. This indicates that either some trans-5
intermediate was carried through in the preparation of cis-8 or that some
epimerization occurred post-purification of
undetected by NMR.
5 and of 6, which remained
17. Acid 6 (0.36 g, 0.76 mmol, 1 equiv) was dissolved in dry toluene (30 mL) and
added SOCl2 (0.1 mL, 1.4 mmol, 1.5 equiv). This was heated to reflux for 2 h
before removing volatiles in vacuo and adding cold NH4OH (15 mL, 35% NH3 w/
w) to the residue. The reaction mixture was then stirred for 1 h at rt. The
product was extracted with EtOAc (3 ꢂ 15 mL), and the combined organic
layers were dried with MgSO4, filtered and concentrated in vacuo to yield title
compound 7 (0.34 g, 95%) as a colourless oil.
20. (a) Khan, J. A.; Xiang, S.; Tong, L. Structure 2007, 12, 1005; (b) Natsumeda, Y.;
Ohno, S.; Kawasaki, H.; Konno, Y.; Weber, G.; Suzuki, K. J. Biol. Chem. 1990, 265,
5292; (c) Garavaglia, S.; D’Angelo, I.; Emanuelli, M.; Carnevali, F.; Pierella, F.;
Magni, G.; Rizzi, M. J. Biol. Chem 2002, 277, 8524.
21. Colby, T. D.; Vanderveen, K.; Strickler, M. D.; Goldstein, B. M. Proc. Natl. Acad.
Sci. USA 1999, 96, 3531.
dH(400 MHz; CDCl3): 1.02 (9H, s, Si-CH3), 2.55 (1H, dd, J=11.2, 17.5 Hz, CH(Ph)-
CH2), 2.93 (1H, dd, J=5.6, 17.5 Hz, CH(Ph)-CH2), 4.02 (2H, d, J=1.9 Hz, TBDPSO-
CH2), 4.08-4.10 (1H, m, O-CH-CO), 5.28 (1H, dd, J=5.6, 11.2 Hz, O-CH-Ar), 5.48
(1H, br s, NH), 5.82 (1H, br s, NH), 7.36-7.47 (6H, m, Ar-H), 7.67-7.74 (6H, m,
Ar-H), 7.81 (1H, d, J=7.7 Hz, Ar-H), 7.91 (1H, s, Ar-H). dC(100 MHz; CDCl3): 19.3
(Si-C-CH3), 26.7 (Si-C-CH3), 46.4 (CH(Ar)-CH2), 63.1 (TBDPSO-CH2), 76.7 (O-CH-
Ar), 82.7 (O-CH-CO), 124.7 (Ar-CH), 127.5 (Ar-CH), 127.8 (Ar-CH), 129.0 (Ar-
22. Khan, J. A.; Forouhar, F.; Tao, X.; Tong, L. Expert Opin. Ther. Targets 2007, 11, 695.
23. (a) Floryk, D.; Tollaksen, S. L.; Giometti, C. S.; Huberman, E. Cancer Res. 2004, 64,
9049; (b) Damaraju, V. L.; Visser, F.; Zhang, J.; Mowles, D.; Ng, A. M. L.; Young, J.
D.; Jayaram, H. N.; Cass, C. E. Mol. Pharmacol. 2005, 67(1), 273.
24. Goldstein, B. M.; Takusagawa, F.; Berman, H. M.; Srivastava, P. C.; Robins, R. K. J.
Am. Chem. Soc. 1983, 105, 7416.