A Click-Chemistry Linked 2′3′-cGAMP Analogue

Article abstract of DOI:10.1002/chem.201805409

2′3′-cGAMP is an uncanonical cyclic dinucleotide where one A and one G base are connected via a 3′-5′ and a unique 2′-5′ linkage. The molecule is produced by the cyclase cGAS in response to cytosolic DNA binding. cGAMP activates STING and hence one of the most powerful pathways of innate immunity. cGAMP analogues with uncharged linkages that feature better cellular penetrability are currently highly desired. Here, the synthesis of a cGAMP analogue with one amide and one triazole linkage is reported. The molecule is best prepared via a first Cu^I-catalyzed click reaction, which establishes the triazole, while the cyclization is achieved by macrolactamization.

Full text of DOI:10.1002/chem.201805409

A Journal of
Accepted Article
Title: A Click-Chemistry Linked 2’3’-cGAMP Analog
Authors: Clemens Reto Dialer, Samuele Stazzoni, David J. Drexler,
Felix Moritz Müller, Simon Veth, Alexander Pichler, Hidenori
Okamura, Gregor Witte, Karl-Peter Hopfner, and Thomas
Carell
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of the final Version of Record (VoR). This work is currently citable by
using the Digital Object Identifier (DOI) given below. The VoR will be
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to this Accepted Article as a result of editing. Readers should obtain
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to ensure accuracy of information. The authors are responsible for the
content of this Accepted Article.
To be cited as: Chem. Eur. J. 10.1002/chem.201805409
Link to VoR: http://dx.doi.org/10.1002/chem.201805409
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A Click-Chemistry Linked 2’3’-cGAMP Analog
Clemens Reto Dialer,[a] Samuele Stazzoni,[a] David Jan Drexler,[b] Felix Moritz Müller,[a] Simon Veth,[a]
Alexander Pichler,[a] Hidenori Okamura,[a] Gregor Witte,[b] Karl-Peter Hopfner,[b] and Thomas Carell*[a]
Abstract: 2’3’-cGAMP is an uncanonical cyclic dinucleotide where
one A and one G base are connected via a 3’-5’ and a unique 2’-5’
linkage. The molecule is produced by the cyclase cGAS in response
to cytosolic DNA binding. cGAMP activates STING and hence one of
the most powerful pathways of innate immunity. cGAMP analogs with
uncharged linkages that feature better cellular penetrability are
currently highly desired. Here, we report the synthesis of a cGAMP
analog with one amide and one triazole linkage. The molecule is best
prepared via a first Cu(I) catalysed click reaction which establishes
the triazole, while the cyclization is achieved by macrolactamization.
Alternative targeting of STING with small molecules is also
known.[24-26] Particularly, compounds which lack the negatively
charged phosphodiester linkages are discussed as new immune-
regulatory pharmaceuticals.[27] While such derivatives are known
for symmetric 3’-5’ dinucleotides[28-32], to the best of our
knowledge, uncharged cGAMP 2 analogs do not exist.
In this article, we describe the modular synthesis of a neutral
cGAMP analog 4 that features one triazole and one amide linkage.
The triazole was generated by a Cu(I) catalysed alkyne-azide
click reaction (CuAAC) that was found to be particularly efficient
on nucleotides and oligonucleotides.[33-35]
Introduction
Cyclic dinucleotides (CDNs) are important cellular messenger
molecules in a variety of organisms.[1] The compounds play a
crucial role in a wide range of biological processes, such as signal
transduction, control of biofilm formation or quorum sensing.[2]
Bacteria produce molecules in which two purine bases are linked
via two 3’-5’ phosphate linkages to give symmetrical cyclophane
structures.[3] One main example for such a molecule is the
c-di-GMP compound 1 shown in Fig. 1.[4,5] Biochemically the
compound is generated from the corresponding nucleotide-5’-
triphosphates. Recently, an unsymmetrical cyclic dipurine
molecule (cGAMP, 2) was discovered in mammalian cells.[6,7] In
this molecule, the two purines are connected via one 3’-5’ and
another 2’-5’ linkage.[8] The dinucleotide 2 is produced by the
cyclase cGAS (cyclic GMP-AMP synthase). cGAS is a cytosolic
DNA sensor and part of the innate immune system.[9,10]
2’3’-cGAMP (2) binds to the transmembrane receptor
STING (stimulator of interferon genes) with nanomolar affinity
(kd = 4.59 nM),[11] which activates the type 1 interferon (IFN)
pathway.[12-14] Subsequent degradation of cGAMP 2 occurs by the
specific cleavage of the 2’-5’ phosphodiester bond by ENPP1
highlighting the importance of this unusual connection.[15,16]
There is currently tremendous interest to develop synthetic routes
towards analogs of cGAMP 2 as potential agonists or antagonists
for cGAS and STING.[17-19] The bisphosphorothioate cGAMP
derivative 3,[20,21] for example, is already in clinical trials.[22,23]
Figure 1: Depiction of the symmetrical microbial c-di-GMP 1, the
unsymmetrical STING activator cGAMP 2, as well as the
bisphosphorothioate analog 3, together with the molecule 4
targeted here.
Results and Discussion
We decided to start our synthetic study by synthesizing the
cGAMP analog 4, in which the 5’-G-3’-A linkage is replaced by a
triazole unit and the 2’-G-5’-A linkage is substituted by an amide
bond.
[a]
[b]
C. R. Dialer, S. Stazzoni, F. M. Müller, S. Veth, A. Pichler, Dr. H.
Okamura, Prof. Dr. T. Carell
Department of Chemistry, Ludwig-Maximilians-Universität München,
Butenandtstraße 5-13, 81377 Munich, Germany
D. J. Drexler, Dr. G. Witte, Prof. Dr. K.-P. Hopfner
Gene Center and Department of Biochemistry, Ludwig-Maximilians-
Universität München, Feodor-Lynen-Straße 25, 81377 Munich,
Germany
*Corresponding author: thomas.carell@lmu.de
Homepage: www.carellgroup.de
Supporting information of this article can be found under:
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Molecular modeling (Fig. 2) showed that the analog 4 is able to
adopt a conformation that is similar to the natural ligand bound to
STING.[11,36]
Figure 3: Synthetic strategy towards compound 4. dpc =
diphenylcarbamoyl, iBu = isobutyryl.
After stereoselective hydroboration (BH3·DMS, dr: 9:1) of 8 and
Swern oxidation, we obtained the carbonyl compound 9, which
we subjected to a Corey-Fuchs alkinylation (CBr4, BuLi). TBS
deprotection and conversion of the primary hydroxyl group into
the azide gave the key intermediate 10. X-ray analysis of the
structure of 10 proved the right configuration of the compound
(recrystallisation from isohexanes/ethyl acetate).
Figure 2: 3D representation showing the potential conformational
similarity between compound 4 (left) and natural 2’3’-cGAMP
(right, conformation of 2 bound to STING, PDB: 4LOH).
In both cases, the macrocycle is thought to force the bases into a
shifted parallel orientation with the imidazole part of the
nucleobases pointing towards each other. This requires anti-
conformations of both glycosidic bonds. The preferred
conformation of compound 4 will be governed by aromatic the
triazole unit. For the conformation of the amide we assume a syn-
conformation due to the small ring size.
Analysis of potential synthetic accesses of 4 shows that it can be
generated by Cu(I) catalyzed azide alkyne reaction plus a
preceding or following lactamization. We developed the synthesis
based on the A-half 5 and the corresponding G-half 6 as depicted
in Fig. 3. For the synthesis of the A-half 5, we started with the
commercially available 1,2-acetonide protected xylofuranoside 7
(two steps from D-xylose), which we converted in three steps into
the 5-TBS-1,2-acetonide protected 3-methylene xylofuranoside 8.
Scheme 1: Synthesis of the A-half 5 in 14 steps. a) TBSCl, Py,
RT, 2h, 97%; b) (COCl)2, DMSO, NEt3, DCM, -60°C, 3h; c)
CH3PPh3Br, BuLi, THF, RT, 6h, 81% (over two steps); d)
BH3*DMS, THF, RT, 12h then 30% H2O2, 2N NaOH, RT, 2h, 76%;
e) (COCl)2, DMSO, NEt3, DCM, -60°C, 3h, 93%; f) CBr4, PPh3,
DCM, 0°C, 1h then RT, 12h, 85%; g) BuLi, THF, -78°C, 1.5h, 83%;
h) TBAF, THF, RT, 4h, 95%; i) TsCl, Py, RT, 18h, 87%; j) NaN3,
DMF, 80°C, 3h, 94%; k) HOAc/Ac2O, H2SO4 (cat.), RT, 5h, 78%;
l) 6-N-Benzoyladenine, BSA, TMSOTf, DCE, 80°C, 4h, 61%; m)
PMe3, H2O, THF, 40 °C, then RT, 12h, 66%; n) Boc2O, NEt3, DCM,
RT, 16h, 64%. Overall yield starting from 7: 6%
Subsequent cleavage of the isopropylidene group and acetyl
protection of the hydroxyl groups provided compound 11, which
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was the sugar building block for the following glycosylation step.
The Vorbrüggen reaction to 12 was found to be most efficient
The resulting alcohol 17 was tosylated and reacted with NaN3 to
give azide 19. The absolute configuration of the compounds was
again proven with a crystal structure of 18 (SI).
under BSA/TMSOTf conditions with
a benzoyl protected
A-heterocycle (α/β: 1:12). Finally, we converted the azide via a
Staudinger reduction (PMe3 worked better than PPh3) into the
corresponding amine, which was Boc-protected afterwards to
give the A-half 5.
We subsequently opened the lactone ring to compound 20 via
hydroxide-mediated benzyl protection and converted it into its
1-O-acetyl derivative 21. The glycosylation reaction to the
G-half 6 was performed by a so far unreported Vorbrüggen
pattern in high β-selectivity (α/β: 1:14) and good yields (79%).
The assembly of nucleoside building blocks A (5) and G (6) was
initiated by a CuAAC reaction. This reaction went smoothly and
provided the dinucleotide 22 in fair yield of 80% (Scheme 3). We
noticed that click-approaches with the Boc-deprotected amine
compound A gave rise of several side products as monitored by
thin-layer chromatography (TLC).
Scheme 3. The assembly towards cyclic dinucleotide 4 in 6 steps.
a) CuSO4, Na-Ascorbate, THF/tBuOH/H2O, RT, 24h, 80%; b)
TFA/DCM (1:1), 0°C, 1h, 81%; c) H2, Pd/C, EtOH, 36h; d) HATU,
DIPEA, DMF (1mM), RT, 24h, 52% (over two steps); e) BCl3,
DCM, -40°C, 3d; f) NH3, H2O/MeOH, 50°C, 20h, 48% (over two
steps). Overall yield starting from 5 and 6: 16%.
Scheme 2. Synthesis of the G-half 6 in 13 steps. a) AcCl, BnOH,
60°C, 5h, 80%; b) Me2C(OMe)2, Me2CO, p-TsOH (cat.), 60°C, 2h,
84%; c) (COCl)2, DMSO, NEt3, DCM, -60°C, 3h; d) Ph3PCHCO2Et,
DCM, RT, 12h, 86% (over two steps); e) H2, Raney-Ni, EtOH, RT,
20h, 90%; f) H2, Pd/C, EtOH/THF, 36h, 88%; g) 80% HOAc, RT,
24h; h) H2SO4 (cat.), MeOH, 4°C, 3d, 72% (over two steps); i)
TsCl, Py, RT, 18h, 76%; i) NaN3, DMF, 80°C, 3h, 75%; k) BnBr,
KOH, THF, reflux, 5h, 91%; l) HOAc/Ac2O, H2SO4 (cat.), RT, 3h,
TFA treatment of dinucleotide 22 resulted in the cleavage of both
the Boc and the diphenylcarbamoyl (dpc) group. Besides, this was
the last step of the consecutive synthesis where purification could
be easily conducted by flash column chromatography
(DCM/MeOH, 10:1) due to the increasing polarity of the following
compounds. A palladium catalyzed hydrogenation reaction
deprotected the benzyl ester by leaving the secondary 3’’’’-O-
benzyl ether intact. Final macrolactamization with HATU
furnished the cyclized dinucleotide 24.
Deprotection of the 3’’’’-O-benzyl ether under BCl3/DCM
conditions (-40°C) proved to be the best option even though
solubility in organic solvents decreased with ongoing removal of
protective groups. Final ammonolysis revealed our target
molecule 4 in 2% overall yield starting from the G-pathway (19
steps) and 1% starting from the A-pathway (20 steps),
respectively. Compounds 24, 25 and 4 were purified by RP-HPLC
and subjected to further NMR-studies.
85%;
m)
6-O-(Diphenylcarbamoyl)-2-N-isobutyrylguanine
(Gdpc/iBu), BSA, TMSOTf, DCE, 80°C, 2h, 72%. Overall yield
starting from 13: 10%.
The desired G-half (Scheme 2) was synthesized starting from
D-arabinose (13). 1-O-Benzyl and 3,4-acetonide protection
yielded alcohol 14.
Subsequent Swern oxidation and Wittig homologation provided
the intermediate 15 (E/Z: 4:1). Employing the acetonide protective
group as a stereoselective directing group, compound 16 was
almost exclusively obtained in R-configuration via a Raney-Ni-
assisted hydrogenation (dr: 20:1).
Under these reduction conditions the 1-O-benzyl group remained
unaffected – keeping the sugar in its pyranoside configuration.
Removal of the protective groups and treatment with catalytic
amounts of acid furnished at
4
°C selectively the
ribofuranoside 17. This was followed by an in situ lactonization.
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Figure 4: NOESY spectrum of the final compound 4 in DMSO-d6
Potential binding of compound 4 to STING was tested in vitro by
nanoDSF assays and analysis of thermal unfolding of the STING
constructs hSTING_L139 (human STING AA139-379) and
mSTING_L138 (mouse STING AA138-378). We used the
physiological ligand 2′3′-cGAMP and a ligand with lower affinity,
3′3′-cGAMP, as positive controls. As expected after the
conformational analysis of compound 4, binding to hSTING or
mSTING could not be detected. This result was confirmed with
ITC experiments (see supporting information). Based on the more
open structure of the here prepared compound 4, we believe that
interaction studies with cGAS or ENPP1 may be more promising.
Investigations in this direction are on the way.
Conformational Analysis and Conclusion
We performed detailed NOESY experiments in order to determine
the conformation preferences of target compound 4 in respect to
potential STING binding. The spectrum is shown in Fig. 4. The
most informative NOE contacts together with a depiction of the
modelling results of 4 in solution is shown in Fig. 5. The NMR data
confirm the overall structure with two β-configured glycosidic
bonds both in anti-conformation. Most interesting, however, is the
large shielding of proton H-2’’’’, which shifts from
(compound 23) = 4.12 ppm to (compound 4) = -0.47 ppm. This
dramatic shift indicates that the proton is positioned just on top of
the aromatic triazole ring.
In summary, we report the first synthesis of a 2’3’-cGAMP analog
which features uncharged bridges that should provide membrane
crossing properties. The synthetic strategy involved first linking of
the two nucleotides by a Cu(I)-catalyzed click reaction followed by
a macrolactamization to close the cycle. The synthesis of medium
size ring structures is always difficult. We believe that the here
described strategy will open the access to a variety of derivatives
of 4. This allows systematic scanning of the conformational space
of the two nucleobases relative to each other regarding the
binding to the involved proteins STING, cGAS and ENPP1.
Experimental Section
Unless otherwise specified, all reactions were magnetically stirred under
an N2 atmosphere. Reaction vessels were dried under high vacuum at
550 °C prior to use. Dry solvents and reagents were purchased from
commercial suppliers, such as Sigma-Aldrich, Acros Organics, Carbosynth,
TCI Europe, ABCR, VWR, stored under septum over molecular sieves and
used as received. The reaction progress and fractions during column
chromatography were monitored by TLC on silica gel 60-F254 plates
purchased from Merck and visualized by irradiation with UV-light (254 nm
or 366 nm) and p-anisaldehyde staining solution (p-anisaldehyde (3.7 mL),
EtOH (135 mL), conc. H2SO4 (5 mL), conc. AcOH (1.5 mL)). Purification
was performed using flash column chromatography with silica gel (Merck,
particle size 0.063 – 0.200 mm). The eluents used were determined by
TLC. Purification of the crude dinucleotides 24, 25 and 4 was operated by
Figure 5: Selected NOE contacts of compound 4 (in DMSO-d6)
and modelling of the preferred conformation based on the NOE
data.
According to this low chemical shift, it is assumed that H-2’’’’
points directly to the triazole ring within the cyclized structures of
compounds 24, 25 and 4. Unraveling of the conformation just
based on the NOE data shows that compound 4 likely adopts a
more open conformation in solution (DMSO-d6) compared to
cGAMP 2, with the two heterocycles being not parallel to each
other.
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Waters 2695 reversed phase high performance liquid chromatography
(RP-HPLC) using Nucleosil columns (250/4 mm, C18ec, particle size 3 μm
for analysis or 250/10 mm, C18ec, 5 μm for purification) from Machery-
Nagel with a bufferfree H2O/MeCN eluent system. Water was purified by a
Milli-Q Plus system from Merck Millipore. NMR-spectra were measured on
a Bruker Ascend 400 or Bruker ARX 600 at room temperature operating
at 400 MHz or 600 MHz for 1H-nuclei and at 101 MHz or 151 MHz for 13C-
nuclei. The chemical shift (δ) in the NMR-spectra is reported in parts per
million (ppm) and referenced by the residual solvent signal. Measurements
were performed in CDCl3 and DMSO-d6. The spectra were referenced to
the residual protons and carbons of the solvent (CHCl3: δ(1H) =7.26 ppm,
δ(13C) = 77.16 ppm; DMSO-d6: δ(1H) =2.50 ppm, δ(13C) = 39.52 ppm).
Proton-spectra also show the integral intensity, the multiplicity,
abbreviated with s (singlet), d (doublet), t (triplet), q (quartet), m (multiplet)
and the coupling constant (J in Hz). Assignments of the signals were
performed using 2D-NMR techniques such as homonuclear correlation
spectroscopy (COSY), nuclear Overhauser effect spectroscopy (NOESY),
heteronuclear single quantum coherence (HSQC) and heteronuclear
multiple bond coherence (HMBC). All spectra were analysed with the
software MestReNOVA 10.0 from Mestrelab Research S. L. Atom labelling
and nomenclature are not in correspondence with IUPAC. High resolution
mass spectra (HRMS) were measured on a Thermo Finnigan MAT 95 (EI)
and a Thermo Finnigan LTQ FTICR (ESI). IR-measurements were
performed on a Perkin Elmer Spectrum BX FT-IR spectrometer with a
diamond-ATR (Attenuated Total Reflection) setup. Uncorrected melting
points were determined with an automated Stanford Research Systems
EZ-Melt apparatus (digital image processing technology). Samples were
loaded in open capillary tubes. X-ray crystallography of single crystals was
performed on an Oxford XCalibur diffractometer and further analysis by
the software Ortep-3.[37] The structure of the synthesized analog 4 in Fig. 2
was obtained using the geometry optimization tool of the open source
software Avogadro and visualized by PyMol.
Nano differential scanning fluorimetry (nanoDSF): Thermal melting
experiments of STING constructs were performed using a Tycho NT.6
instrument (NanoTemper Technologies). In brief, the samples were heated
up in a glass capillary and while heating, the internal fluorescence at 330
nm and 350 nm was recorded. Data analysis, data smoothing and
calculation of derivatives was done using the internal evaluation features
of the NT.6 instrument. All measurements were repeated to confirm
robustness of the assay. Isothermal titration calorimetry: ITC
experiments were performed using a Malvern PEAQ-ITC system with
20 µM protein in ITC-buffer (20 mM HEPES pH = 7.5, 150 mM NaCl) in
the cell. The positive controls of cGAMP ligands (Biolog) were titrated in a
concentration of 200 µM into the cell by 19 injections of 2 µL, spaced 150 s
apart, at 25°C. Compound 4 was used in a concentration of 291 µM for
titration. The results were analyzed using the MicroCal PEAQ-ITC analysis
software provided with the instrument. All titrations were repeated to
confirm robustness of the assay. Cloning, Expression and Purification:
Human STING AA139-379 and mouse STING AA138-378 constructs were
cloned according to previous studies.[38] The plasmids were used to
transform E. coli Rosetta (DE3) protein expression strain cells (Novagen).
The cells were grown in 1 L of Turbo Broth™ media (Molecular
(39.0 g, 128 mmol, 1.00 eq.) in dry DCM (125 mL) was added to the
reaction mixture over 5 min and stirred for 2 h. Triethylamine (53.6 mL,
38.9 g, 384 mmol, 3.00 eq.) was added and the suspension was stirred for
a further hour at -60 °C. The reaction mixture was warmed to RT,
quenched with saturated aqueous NaHCO3 (200 mL) and extracted with
DCM (3x200 mL). The combined organic layers were dried over
anhydrous MgSO4, filtered and concentrated in vacuo to obtain 5-O-(tert-
butyldimethylsilyl)-3-oxo-1,2-O-isopropylidene-α-D-xylofuranose[41] as
a
waxy yellow solid. The compound was used in the next step without further
purification.
Methyl triphenyl phosphonium bromide (86.2 g, 241 mmol, 2.00 eq.) was
suspended in THF (360 mL) and cooled to -78 °C under N2. n-Butyl lithium
(96.5 mL, 241 mmol, 2.5M in hexanes, 2.00 eq.) was carefully added
dropwise and the resulting red suspension (LiBr precipitates) was stirred
for 1 h at 0°C. Subsequent addition of a solution of the crude ketone
(36.5 g, 121 mmol, 1.00 eq.) in THF (60 mL) over 10 min gave a slurry
which was stirred at RT for 6h. The reaction mixture was quenched with
saturated aqueous NH4Cl (50 mL) and then extracted with ethyl acetate
(3 x 200 mL). The combined organic layers were dried over anhydrous
MgSO4, filtered and concentrated in vacuo. The crude product was purified
by flash-column chromatography (silica gel, isohexanes/EtOAc, 9:1→4:1)
to yield compound 8 as a colorless syrup (31.1 g, 104 mmol, 81% over 2
steps). Rf = 0.68 (silica, isohexanes/EtOAc = 4:1). IR (ATR): ṽ = 2930, 1463,
1372, 1252, 1071, 1018, 775 cm-1. 1H NMR, COSY (400 MHz, CDCl3): δ
3
4
= 5.85 (d, J = 4.1 Hz, 1H, H-1), 5.42 (dd, J = 2.2 Hz, 4J = 1.1 Hz, 1H,
C=CHa), 5.26 (m, 1H, C=CHb), 4.88 (dd, 3J = 4.1 Hz, 4J = 1.4 Hz, 1H, H-
2), 4.75 (ddd, 3J = 4.2 Hz, 3J = 3.8 Hz, 4J = 2.2 Hz, 1H, H-4), 3.75 (dd, 2J
= 10.6 Hz, 3J = 4.2 Hz, 1H, Ha-5), 3.67 (dd, 2J = 10.6 Hz, 3J = 3.8 Hz, 1H,
Hb-5), 1.49 (s, 3H, C(CH3)2), 1.38 (s, 3H, C(CH3)2), 0.87 (s, 9H, SiC(CH3)3),
0.043 (s, 3H, Si(CH3)2), 0.040 (s, 3H, Si(CH3)2) ppm. 13C NMR, HSQC,
HMBC (101 MHz, CDCl3): δ = 147.7 (C-3),112.6 (C(CH3)2), 111.7 (C=CH2),
105.1 (C-1), 82.1 (C-2), 81.0 (C-4), 65.9 (C-5), 27.7 (C(CH3)2), 27.5
(C(CH3)2), 26.0 (SiC(CH3)3), 18.4 (SiC(CH3)3), -5.24 (Si(CH3)2), -5.31
(Si(CH3)2) ppm. ESI-HRMS calcd. for [C15H28O4Si + NH4]+ 318.2095,
found: 318.2098.
5-O-(tert-Butyldimethylsilyl)-3-deoxy-3-(hydroxymethyl)-1,2-O-
isopropylidene-α-D-ribofuranose (9a): The title compound was
prepared according to a modified procedure of Betkekar et al.[39] To a
solution of vinyl compound 8 (29.5 g, 98.2 mmol, 1.00 eq.) in dry THF
(300 mL) was added borane dimethyl sulfide complex (73.6 mL, 147 mmol,
2M in THF, 1.50 eq.) at 0 °C. After the solution was stirred for 12 h at RT,
aqueous 2 N NaOH (225 mL) was carefully added under strong gas
evolution at 0 °C to give a turbid suspension. The reaction mixture was
treated slowly with 30% aqueous hydrogen peroxide (98.0 mL) at the same
temperature to avoid heat development. The suspension was stirred for
further 2 h at RT, quenched with saturated aqueous Na2S2O3 solution
(200 mL) at 0 °C and finally extracted with EtOAc (3 x 300 mL). The
combined organic layers were washed with brine, dried over anhydrous
MgSO4, filtered and concentrated under reduced pressure. The crude
product was purified by flash-column chromatography (silica gel,
isohexanes/EtOAc, 4:1→2:1→1:1) to afford alcohol 9a (23.8 g, 74.7 mmol,
76%) as a colorless oil. Rf = 0.76 (isohexanes/EtOAc = 1:1). IR (ATR): ṽ =
3456, 2931, 1463, 1381, 1253, 1105, 1019, 778 cm-1. 1H NMR, COSY
(400 MHz, CDCl3): δ = 5.79 (d, 3J = 3.7 Hz, 1H, H-1), 4.75 (dd, 3J = 4.9 Hz,
3J = 3.7 Hz, 1H, H-2), 4.08 (ddd, 3J = 9.9 Hz, 3J = 6.6 Hz, 3J = 3.5 Hz, 1H,
H-4), 3.88 (dd, 2J = 10.6 Hz, 3J = 3.5 Hz, 1H, Ha-5), 3.86 (d, 3J = 6.2 Hz,
Dimensions)
supplemented
with
Kanamycin
(50ꢀmg/L)
and
Chloramphenicol (34ꢀmg/L) at 37°C to an OD600=1.3 and expression was
induced by adding IPTG to a final concentration of 0.2 mM. Purification of
the STING constructs has been performed as described previously.[38]
3
5-O-(tert-Butyldimethylsilyl)-3,3-deoxymethylene-1,2-O-
2H, CH2OH), 3.66 (dd, 2J = 10.6 Hz, J = 6.6 Hz, 1H, Hb-5), 3.06 (s, 1H,
isopropylidene-α-D-xylofuranose (8): The title compound was prepared
according to a modified procedure of Betkekar et al.[39] To a solution of
oxalyl chloride (12.3 mL, 17.9 g, 141 mmol, 1.10 eq.) in dry DCM (450 mL)
was slowly added DMSO (20.0 mL, 22.0 g, 282 mmol, 2.20 eq.) under N2
at -78 °C. The temperature was maintained below -60 °C and evolving gas
was purged. After the mixture was stirred for 1 h at -60 °C, a solution of 5-
O-(tert-butyldimethylsilyl)-1,2-O-isopropylidene-α-D-xylofuranose[40]
OH), 2.19-2.09 (m, 1H, H-3), 1.52 (s, 3H, C(CH3)2), 1.32 (s, 3H, C(CH3)2),
0.90 (s, 9H, SiC(CH3)3), 0.091 (s, 3H, Si(CH3)2), 0.087 (s, 3H, Si(CH3)2)
ppm. 13C NMR, HSQC, HMBC (101 MHz, CDCl3): δ = 112.2 (C(CH3)2),
105.0 (C-1), 82.6 (C-2), 80.7 (C-4), 64.0 (C-5), 59.6 (CH2OH), 50.0 (C-3),
26.9 (C(CH3)2), 26.4 (C(CH3)2), 26.0 (SiC(CH3)3), 18.4 (SiC(CH3)3), -5.32
(Si(CH3)2) ppm. ESI-HRMS calcd. for [C15H30O5Si + NH4]+: 336.2201,
found: 336.2204.
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10.1002/chem.201805409
Chemistry - A European Journal
FULL PAPER
NH4Cl (100 mL) and extracted with EtOAc (3 x 200 mL). The combined
organic layers were dried over anhydrous MgSO4, filtered and
concentrated in vacuo. Purification of the residue was conducted by flash-
column chromatography (silica gel, isohexane/EtOAc, 9:1→4:1) to afford
ethynyl compound 10b (13.7 g 43.8 mmol, 83%) as a yellowish oil. Rf =
0.60 (isohexanes/EtOAc, 4:1). IR (ATR): ṽ = 3279, 2930, 1472, 1373,
5-O-(tert-Butyldimethylsilyl)-3-deoxy-3-formyl-1,2-O-isopropylidene-
α-D-ribofuranose (9): The title compound was prepared according to a
modified procedure of Parr et al.[41] To a solution of oxalyl chloride
(6.91 mL, 10.1 g, 79.6 mmol, 1.10 eq.) in dry DCM (380 mL) was slowly
added DMSO (11.3 mL, 12.4 g, 159 mmol, 2.20 eq.) under N2 at -78 °C.
The temperature was maintained below -60 °C and evolving gas was
purged. After the mixture was stirred for 1 h at -60 °C, a solution of alcohol
9a (23.0 g, 72.2 mmol, 1.00 eq.) in dry DCM (70 mL) was added to the
reaction mixture over 5 min and stirred for 2 h. Triethylamine (30.2 mL,
21.9 g, 217 mmol, 3.00 eq.) was added and the suspension was stirred for
a further hour at -60 °C. The reaction mixture was warmed to RT,
quenched with saturated aqueous NaHCO3 (150 mL) and extracted with
DCM (3x150 mL). The combined organic layers were dried over
anhydrous MgSO4, filtered and volatile components removed under
reduced pressure. Purification of the crude product was performed by
flash-column chromatography (silica gel, isohexanes/EtOAc, 9:1→4:1) to
give aldehyde 9 (21.2 g 67.0 mmol, 93%) as a colorless oil. It was also
possible to use the crude product in the next step without further
purification. Rf = 0.61 (isohexanes/EtOAc = 4:1). IR (ATR): ṽ = 2931, 1726,
1472, 1382, 1253, 1100, 1019, 778 cm-1. 1H NMR, COSY (400 MHz,
CDCl3): δ = 9.78 (d, 4J = 1.2 Hz, 1H, CHO), 5.87 (d, 3J = 3.7 Hz, 1H, H-1),
5.03 (dd, J = 5.2 Hz, J = 3.7 Hz, 1H, H-2), 4.55 (ddd, J = 9.5 Hz, J =
3.6 Hz, 3J = 3.2 Hz, 1H, H-4), 3.86 (dd, 2J = 11.3 Hz, 3J = 3.6 Hz, 1H, Ha-
5), 3.78 (dd, 2J = 11.3 Hz, 3J = 3.2 Hz, 1H, Hb-5), 3.02 (dd, 3J = 9.5 Hz, 3J
= 5.2 Hz, 4J = 1.2 Hz, 1H, H-3), 1.48 (s, 3H, C(CH3)2), 1.33 (s, 3H, C(CH3)2),
0.86 (s, 9H, SiC(CH3)3), 0.04 (s, 3H, Si(CH3)2), 0.03 (s, 3H, Si(CH3)2) ppm.
13C NMR, HSQC, HMBC (101 MHz, CDCl3): δ = 198.0 (CHO), 113.1
(C(CH3)2), 105.7 (C-1), 80.8 (C-2), 78.2 (C-4), 62.8 (C-5), 56.6 (C-3), 26.8
(C(CH3)2), 26.6 (C(CH3)2), 26.0 (SiC(CH3)3), 18.5 (SiC(CH3)3), 5.31
(Si(CH3)2), -5.31 (Si(CH3)2) ppm. EI-HRMS calcd. for [C15H28O5Si - CH3]+:
301.1466, found: 301.1475.
1252, 1215, 1110, 1017, 815, 777 cm−1 1H NMR, COSY (400 MHz,
.
CDCl3): δ = 5.81 (d, 3J = 3.6 Hz, 1H, H-1), 4.72 (dd, 3J = 4.1 Hz, 3J = 3.6
Hz, 1H, H-2), 4.11 (ddd, 3J = 10.1 Hz, 3J = 2.9 Hz, 3J = 2.0 Hz, 1H, H-4),
3.97 (dd, 2J = 12.0, 3J = 2.0 Hz, 1H, Ha-5), 3.79 (dd, 2J = 12.0, 3J = 2.9 Hz,
1H, Hb-5), 2.97 (ddd, 3J = 10.1 Hz, 3J = 4.1 Hz, 4J = 2.6 Hz, 1H, H-3), 2.21
4
(d, J = 2.6 Hz, 1H, CCH), 1.57 (s, 3H, C(CH3)2), 1.36 (s, 3H, C(CH3)2),
0.89 (s, 9H, SiC(CH3)3), 0.08 (s, 3H, Si(CH3)2), 0.07 (s, 3H, Si(CH3)2) ppm.
13C NMR, HSQC, HMBC (101 MHz, CDCl3): δ = 112.3 (C(CH3)2), 105.1
(C-1), 81.9 (C-4), 81.2 (C-2), 78.2 (CCH), 72.3 (CCH) , 61.1 (C-5), 36.3
(C-3), 26.8 (C(CH3)2), 26.5 (C(CH3)2), 26.1 (SiC(CH3)3), 18.6 (SiC(CH3)3),
-5.1 (Si(CH3)2), -5.2 (Si(CH3)2) ppm. ESI-HRMS calcd. for [C16H28O4Si +
NH4]+: 330.2095, found: 330.2098.
3-Deoxy-3-ethynyl-1,2-O-isopropylidene-α-D-ribofuranose (10c): The
title compound was prepared according to a modified procedure of
Betkekar et al.[39] To a yellow solution of ethynyl compound 10b (13.3 g,
42.6 mmol, 1.00 eq.) in THF (200 mL) was added TBAF (55.3 mL,
55.3 mmol, 1M in THF, 1.30 eq.) at RT. The resulting dark solution was
stirred for 4 h at this temperature. The reaction mixture was quenched with
silica and the solvent was concentrated under reduced pressure. The
crude product was purified by flash-column chromatography (silica gel,
isohexanes/EtOAc, 2:1→1:1→2:3) to yield alcohol 10c (8.03 g, 40.5 mmol,
95%) as colorless crystals. M.p. = 43 – 45 °C. Rf = 0.38 (isohexanes/EtOAc
= 1:1). IR (ATR): ṽ = 3456, 3279, 2936, 1375, 1249, 1215, 1105, 1007,
871 cm−1. 1H NMR, COSY (400 MHz, CDCl3): δ = 5.83 (d, 3J = 3.5 Hz, 1H,
H-1), 4.75 (dd, 3J = 4.1 Hz, 3J = 3.5 Hz, 1H, H-2), 4.17 (ddd, 3J = 10.3 Hz,
3J = 3.2 Hz, 3J = 3.0 Hz, 1H, H-4), (4.00 (dd, 2J = 12.2 Hz, 3J = 3.2 Hz, 1H,
3
3
3
3
2
3
3
5-O-(tert-Butyldimethylsilyl)-3-deoxy-3-(2,2-dibromovinyl)-1,2-O-
isopropylidene-α-D-ribofuranose (10a): The title compound was
prepared according to a modified procedure of Betkekar et al.[39] A solution
of tetrabromomethane (43.0 g, 130 mmol, 2.00 eq.) in DCM (350 mL) was
mixed with triphenylphosphine (68.0, 259 mmol, 4.00 eq.) under N2 at 0 °C
and stirred for 1 h at this temperature. The resulting orange solution was
treated with a solution of aldehyde 9 (20.5 g, 64.8 mmol, 1.00 eq.) in DCM
(120 mL). The dark suspension was stirred at RT for 12 h. After removal
of volatile materials, the crude product was purified by flash-column
chromatography (silica gel, isohexanes/EtOAc, 9:1→4:1) to provide
dibromo compound 10a (26.1 g, 55.3 mmol, 85%) as a slightly yellow oil.
Rf = 0.83 (isohexanes/EtOAc = 4:1). IR (ATR): ṽ = 2929, 1471, 1382, 1252,
1097, 1020, 777 cm-1. 1H NMR, COSY (400 MHz, CDCl3): δ = 6.49 (d, 3J
= 9.4 Hz, 1H, Br2CCH), 5.82 (d, J = 3.5 Hz, 1H, H-1), 4.68 (dd, J = 4.5
Hz, 3J = 3.5 Hz, 1H, H-2), 4.03 (ddd, 3J = 9.9 Hz, 3J = 3.6 Hz, 3J = 3.4 Hz,
1H, H-4), 3.81 (dd, 2J = 11.5 Hz, 3J = 3.4 Hz, 1H, Ha-5), 3.66 (dd, 2J = 11.5
Hz, 3J = 3.6 Hz, 1H, Hb-5), 3.00 (td, 3J = 9.9 Hz, 3J = 4.5 Hz, 1H, H-3), 1.53
(s, 3H, C(CH3)2), 1.32 (s, 3H, C(CH3)2), 0.89 (s, 9H, SiC(CH3)3), 0.06 (s,
6H, Si(CH3)2) ppm. 13C NMR, HSQC, HMBC (101 MHz, CDCl3): δ = 133.2
(Br2CCH), 112.2 (C(CH3)2), 105.2 (C-1), 91.8 (Br2CCH), 81.7 (C-2), 80.9
(C-4), 62.3 (C-5), 49.2 (C-3), 26.9 (C(CH3)2), 26.4 (C(CH3)2), 26.1
(SiC(CH3)3), 18.5 (SiC(CH3)3), 5.16 (Si(CH3)2), -5.22 (Si(CH3)2) ppm. EI-
HRMS calcd. for [C16H28 Br2O4Si - CH3]+: 454.9884, found: 454.9882
Ha-5), 3.71 (ddd, J = 12.2 Hz, J = 8.8 Hz, J = 3.0 Hz, 1H, Hb-5), 2.96
(ddd, 3J = 10.3 Hz, 3J = 4.1 Hz, 4J = 2.5 Hz, 1H, H-3), 2.23 (d, 4J = 2.5 Hz,
1H, CCH), 1.84 (dd, 3J = 8.8 Hz, 4.2 Hz, 1H, OH), 1.58 (s, 3H, C(CH3)2),
1.37 (s, 3H, C(CH3)2) ppm. 13C NMR, HSQC, HMBC (101 MHz, CDCl3): δ
= 112.7 (C(CH3)2), 105.1 (C-1), 81.4 (C-4), 81.3 (C-2), 77.6 (CCH), 72.6
(CCH), 60.6 (C-5), 36.3 (C-3), 26.7 (C(CH3)2), 26.5 (C(CH3)2) ppm. EI-
HRMS calcd. for [C10H14O4 - CH3]+: 183.0652, found: 183.0652.
3-Deoxy-3-ethynyl-1,2-O-isopropylidene-5-O-tosyl-α-D-ribofuranose
(10d): p-Toluenesulfonyl chloride (15.5 g, 56.8 mmol, 1.50 eq.) was
dissolved in dry pyridine (50.0 mL) and added to a solution of alcohol 10c
(7.5 g 37.8 mmol, 1.00 eq.) in pyridine (150 mL) at 0 °C. The reaction
mixture was stirred for 18 h at RT and finally quenched with MeOH (10 mL).
After removal of volatile components, purification of the residue by flash-
column chromatography (silica gel isohexanes/EtOAc, 2:1→1:1) gave
tosyl compound 10d (11.7 g, 33.2 mmol, 87%) as colorless crystals.
Crystallization from isohexanes/EtOAc (vapor diffusion) provided suitable
single crystals for X-ray characterization. M.p. = 113 – 114 °C. Rf = 0.73
(isohexanes/EtOAc = 1:1). IR (ATR): ṽ = 3296, 2990, 1598, 1450, 1360,
3
3
1
1176, 1097, 958, 813 665 cm−1. H NMR, COSY (400 MHz, CDCl3): δ =
7.80 (d, 3J = 8.2 Hz, 2H, aryl-CH-CSO3), 7.34 (d, 3J = 8.2 Hz, 2H, aryl-CH-
3
CCH3), 5.72 (d, 3J = 3.5 Hz, 1H, H-1), 4.68 (dd, 3J = 4.0 Hz, J = 3.5 Hz,
1H, H-2),, 4.40 – 4.31 (m, 1H, Ha-5), 4.22 – 4.14 (m, 2H, H-4, Hb-5), 2.84
(ddd, 3J = 9.7 Hz, 3J = 4.0 Hz, 4J = 2.4 Hz, 1H, H-3), 2.44 (s, 3H, aryl-CH3),
2.21 (d, 4J = 2.4 Hz, 1H, CCH), 1.52 (s, 3H, C(CH3)2), 1.34 (s, 3H, C(CH3)2)
ppm. 13C NMR, HSQC, HMBC (101 MHz, CDCl3): δ = 145.1 (aryl-C-SO3),
132.7 (aryl-C-CH3), 130.0 (aryl-CH-CCH3), 128.2 (aryl-CH-CSO3), 112.8
(C(CH3)2), 105.0 (C-1), 80.7 (C-2), 78.5 (C-4), 76.5 (CCH), 73.3 (CCH),
67.4 (C-5), 37.0 (C-3), 26.7 (C(CH3)2), 26.4 (C(CH3)2), 21.8 (aryl-CH3) ppm.
ESI-HRMS calcd. for [C17H20O6S + H]-: 353.1054, found: 353.1057. ESI-
HRMS calcd. for [C17H20O6S + NH4]+: 370.1319, found: 370.1317.
5-O-(tert-Butyldimethylsilyl)-3-deoxy-3-ethynyl-1,2-O-
isopropylidene-α-D-ribofuranose (10b): The title compound was
prepared according to a modified procedure of Betkekar et al.[39] Dibromo
compound 10a (25.0 g, 52.9 mmol, 1.00 eq.) was dissolved in dry THF
(270 mL) and cooled to -78 °C under N2. n-Butyl lithium (48.7 mL,
122 mmol, 2.5M in hexanes, 2.30 eq.) was added dropwise over a period
of 10 min until a red solution was formed. After stirring for 1.5 h at this
temperature, the reaction mixture was quenched with saturated aqueous
This article is protected by copyright. All rights reserved.

10.1002/chem.201805409
Chemistry - A European Journal
FULL PAPER
5-Azido-3,5-dideoxy-3-ethynyl-1,2-O-isopropylidene-α-D-
performed on a 4 g scale of the diacetate starting material 11 (yield: 50%).
M.p. = 110 °C (decomp.). Rf = 0.20 (isohexanes/EtOAc = 1:2). Rf = 0.24
(DCM/MeOH, 100:5). IR (ATR): ṽ = 3296, 3060, 2932, 2103, 1747, 1698,
1581, 1218, 1072, 797 cm−1. 1H NMR, COSY (400 MHz, CDCl3): δ = 9.27
(s, 1H, NH), 8.74 (s, 1H, H-2), 8.16 (s, 1H, H-8), 7.98 (d, 3J = 7.7 Hz, 2H,
ribofuranose (10): A mixture of tosyl compound 10d (10.5 g, 29.8 mmol,
1.00 eq.) and sodium azide (8.86 g, 95.3 mmol, 3.20 eq.) was suspended
in DMF (300 mL) and stirred under N2 at 80 °C for 3 h. The yellow
suspension was diluted with brine (200 mL) and extracted with
EtOAc (4 x 300 mL). The combined organic layers were dried over
anhydrous MgSO4, filtered and concentrated in vacuo. Purification of the
crude product by flash-column chromatography (silica gel,
isohexanes/EtOAc, 9:1→4:1) yielded azide compound 10 (6.25 g,
28.0 mmol, 94%) as a colorless oil. Rf = 0.67 (isohexanes/EtOAc = 2:1).
3
aryl-o-CH), 7.58 – 7.53 (m, 1H, aryl-p-CH), 7.46 (t, J = 7.7 Hz, 2H, aryl-
m-CH), 6.09 (s, 1H, H-1’), 5.87 (d, 3J = 5.7 Hz, 1H, H-2’), 4.38 (ddd, 3J =
10.0 Hz, 3J = 4.8 Hz, 4J = 2.7 Hz, 1H, H-4’), 4.07 (ddd, 3J = 10.0 Hz, 3J =
5.7 Hz, 4J = 2.4 Hz, 1H, H-3’), 3.79 (dd, 2J = 13.6 Hz, 3J = 2.7 Hz, 1H, Ha-
5’), 3.57 (dd, 2J = 13.6 Hz, 3J = 4.8 Hz, 1H, Hb-5’), 2.26 (d, 4J = 2.4 Hz, 1H,
CCH), 2.18 (s, 3H, CH3) ppm. 13C NMR, HSQC, HMBC (101 MHz, CDCl3):
δ = 169.6 (OCOCH3), 164.8 (N-CO-aryl), 152.8 (C-2), 151.2 (C-4), 149.9
(C-6), 142.0 (C-8), 133.6 (aryl-C-CO-N), 132.9 (aryl-p-CH), 128.9 (aryl-m-
CH), 128.0 (aryl-o-CH), 123.7 (C-5), 89.7 (C-1’), 83.0 (C-4’), 76.9 (C-2’),
75.7 (CCH), 74.4 (CCH), 51.2 (C-5’), 36.2 (C-3’), 20.7 (OCOCH3) ppm.
ESI-HRMS calcd. for [C21H18N8O4 + H]+: 447.1524, found: 447.1529.
IR (ATR): ṽ = 3280, 2989, 2100, 1375, 1216, 1166, 1105, 1012, 871 cm−1
.
1H NMR, COSY (400 MHz, CDCl3): δ = 5.86 (d, 3J = 3.6 Hz, 1H, H-1), 4.75
(dd, 3J = 4.1 Hz, 3J = 3.6 Hz, 1H, H-2), 4.24 (ddd, 3J = 10.2 Hz, 3J = 3.9 Hz,
3J = 2.7 Hz, 1H, H-4), 3.76 (dd, 2J = 13.6 Hz, 3J = 2.7 Hz, 1H, Ha-5), 3.37
(dd, 2J = 13.6 Hz, 3J = 3.9 Hz, 1H, Hb-5), 2.88 (ddd, 3J = 10.2 Hz, 3J = 4.1
Hz, 4J = 2.5 Hz, 1H, H-3), 2.25 (d, 4J = 2.5 Hz, 1H, CCH), 1.57 (s, 3H,
C(CH3)2), 1.37 (s, 3H, C(CH3)2) ppm. 13C NMR, HSQC, HMBC (101 MHz,
CDCl3): δ = 112.8 (C(CH3)2), 105.1 (C-1), 81.0 (C-2), 79.7 (C-4), 76.9
(CCH), 73.2 (CCH), 50.7 (C-5), 37.9 (C-3), 26.7 (C(CH3)2), 26.4 (C(CH3)2)
ppm. EI-HRMS calcd. for [C10H13N3O3 - CH3]+: 208.0717, found: 208.0717.
6-Benzoylamino-9-(2-O-acetyl-5-amino-3,5-dideoxy-3-ethynyl-β-D-
ribofuranosyl)-9H-purine (5a): Trimethylphosphine (4.48 mL, 4.48 mmol,
1.0M in THF, 2.00 eq.) was added to a stirred solution of nucleoside
12 (1.00 g, 2.24 mmol, 1.00 eq.) in THF (25 mL). After 5 min the reaction
mixture turned turbid under N2 evolution and was heated to 40 °C for 1.5
h. The reaction mixture was treated with water (0.44 mL, 24.6 mmol, 11.0
eq.) and stirred for 10 h at RT. Volatile materials were removed under
reduced pressure and the residue was purified by flash-column
chromatography (silica gel, DCM/MeOH, 100:2→100:5) to give amino
compound 5a (0.62 g, 1.48 mmol, 66%) as a colorless foam. The reaction
could also be performed on a 4 g scale of the azide starting material 12
(yield: 56%). Rf = 0.34 (DCM/MeOH = 5:1). IR (ATR): ṽ = 3366, 3275,
1,2-di-O-Acetyl-5-azido-3,5-dideoxy-3-ethynyl-D-ribofuranose (11): A
stirred solution of azide compound 10 (6.00 g, 26.9 mmol, 1.00 eq.) in
acetic acid (100 mL) and acetic anhydride (50 mL) was treated with
concentrated sulfuric acid (96%, 1.30 mL) at 0 °C. The reaction mixture
turned dark and was stirred for 5 h at RT. After careful quenching with
saturated aqueous NaHCO3 solution (200 mL) and solid NaHCO3 until
CO2 evolution stopped, the reaction was extracted with DCM (4 x 200 mL).
The organic phase was washed with brine (200 mL), dried over anhydrous
MgSO4, filtered and concentrated in vacuo. The crude product was purified
by flash-column chromatography (silica gel, isohexanes/EtOAc, 9:1) to
afford diacetate compound 11 (5.61 g, 21.0 mmol, 78%) as colorless
crystals. α/β = 1:6. β anomer could be isolated for analysis. Crystallization
from isohexanes/EtOAc (vapor diffusion) provided suitable single crystals
of the β anomer for X-ray characterization. M.p. = 81 – 82 °C. Rf (α
anomer) = 0.55 (isohexanes/EtOAc = 4:1). Rf (β anomer) = 0.46
(isohexanes/EtOAc, 4:1). IR (ATR, β anomer): ṽ = 3281, 2934, 2099, 1743,
1438, 1371, 1205, 1097, 1024, 959, cm−1. Major β anomer: 1H NMR,
COSY (400 MHz, CDCl3): δ = 6.12 (s, 1H, H-1), 5.37 (d, 3J = 4.5 Hz, 1H,
H-2), 4.38 (ddd, 3J = 10.0 Hz, 3J = 3.2 Hz, 3J = 3.0 Hz, 1H, H-4), 3.75 (dd,
2J = 13.7 Hz, 3J = 3.0 Hz, 1H, Ha-5), 3.39 (ddd, 3J = 10.0 Hz, 3J = 4.5 Hz,
4J = 2.5 Hz, 1H, H-3), 3.24 (dd, 2J = 13.7 Hz, 3J = 3.2 Hz, 1H, Hb-5), 2.16
2918, 1747, 1640, 1422, 1296, 1138, 943, 860 cm−1
.
1H NMR, COSY
3
(400 MHz, CDCl3): δ = 8.75 (s, 1H, H-2), 8.27 (s, 1H, H-8), 8.00 (d, J =
7.7 Hz, 2H, aryl-o-CH), 7.61 – 7.55 (m, 1H, aryl-p-CH), 7.49 (t, 3J = 7.7 Hz,
2H, aryl-m-CH), 6.08 (s, 1H, H-1’), 5.81 (d, 3J = 5.9 Hz, 1H, H-2’), 4.27
3
3
4
3
(ddd, J = 9.9 Hz, J = 4.8 Hz, J = 3.0 Hz, 1H, H-4’), 4.02 (ddd, J = 9.9
Hz, 3J = 5.9 Hz, 4J = 2.4 Hz, 1H, H-3’), 3.24 (dd, 2J = 14.0 Hz, 3J = 3.0 Hz,
1H, Ha-5’), 3.01 (dd, 2J = 14.0 Hz, 3J = 4.8 Hz, 1H, Hb-5’), 2.23 (d, 4J = 2.4
Hz, 1H, CCH), 2.19 (s, 3H, CH3) ppm. 13C NMR, HSQC, HMBC (101 MHz,
CDCl3): δ = 169.8 (OCOCH3), 164.9 (N-CO-aryl), 152.8 (C-2), 151.3 (C-4),
149.9 (C-6), 142.2 (C-8), 133.6 (aryl-C-CO-N), 132.9 (aryl-p-CH), 128.9
(aryl-m-CH), 128.0 (aryl-o-CH), 123.7 (C-5), 89.5 (C-1’), 85.4 (C-4’), 77.4
(C-2’), 76.7 (CCH), 73.9 (CCH), 42.6 (C-5’), 35.8 (C-3’), 20.9 (OCOCH3)
ppm. ESI-HRMS calcd. for [C21H20N6O4 + H]+: 421.1619, found: 421.1623.
4
(d, J = 2.5 Hz, 1H, CCH), 2.14 (s, 3H, C-2-OCOCH3), 2.08 (s, 3H, C-1-
OCOCH3) ppm. 13C NMR, HSQC, HMBC (101 MHz, CDCl3): δ = 169.5 (C-
2-OCOCH3), 169.2 (C-1-OCOCH3), 98.7 (C-1), 83.7 (C-4), 76.8 (C-2), 76.0
(CCH), 73.6 (CCH), 51.0 (C-5), 34.5 (C-3), 21.12 (C-1-OCOCH3), 20.73
(C-2-OCOCH3) ppm. ESI-HRMS calcd. for [C11H13N3O5 + H]+: 268.0928,
found: 268.0930. ESI-HRMS calcd. for [C11H13N3O5 + NH4]+: 285.1193,
found: 285.1196.
6-Benzoylamino-9-(2-O-acetyl-5-(tert-butoxycarbonyl)amino-3,5-
dideoxy-3-ethynyl-β-D-ribofuranosyl)-9H-purine (5):
A mixture of
amine compound 5a (2.00 g, 4.76 mmol, 1.00 eq.), triethylamine (1.99 mL,
1.44 g, 14.3 mmol, 3.00 eq.) and di-tert-butyldicarbonate (1.53 mL, 1.56 g,
7.14 mmol, 1.50 eq.) in dry DCM (40 mL) was stirred at RT for 16 h. MeOH
(3 mL) was added and volatile materials were removed in vacuo. The
residue was purified by flash-column chromatography (silica gel,
DCM/MeOH, 100:2→100:3) to give the title compound 5 as a colorless
foam (1.59 g, 3.05 mmol, 64%). M.p. = 143 °C (decomp.). Rf = 0.20
(DCM/MeOH = 100:5). IR (ATR): ṽ = 3265, 2977, 1749, 1699, 1610, 1516,
1455, 1248, 1227, 1086 cm−1. 1H NMR, COSY (400 MHz, CDCl3): δ = 9.15
6-Benzoylamino-9-(2-O-acetyl-5-azido-3,5-dideoxy-3-ethynyl-β-D-
ribofuranosyl)-9H-purine (12): N,O-Bis(trimethylsilyl)acetamid (BSA)
(3.66 mL, 3.05 g, 15.0 mmol, 4.00 eq.) was added under N2 to a stirred
suspension of diacetate compound 11 (1.00 g, 3.74 mmol, 1.00 eq.) and
3
6-N-benzoyladenine
(1.79 g,
7.48 mmol,
2.00 eq.)
in
(s, 1H, NHBz), 8.79 (s, 1H, H-2), 8.03 (s, 1H, H-8), 8.01 (d, J = 7.7 Hz,
3
dichloroethane (40 mL) and heated to 80 °C for 1 h until a clear solution
was obtained. The reaction mixture was brought to RT and treated with
trimethylsilyl triflate (TMSOTf) (1.36 mL, 1.66 g, 7.48 mmol, 2.00 eq.). The
dark red solution was stirred at 80 °C for 4 h and additional 8 h at RT. The
reaction was quenched with saturated aqueous NaHCO3 (30 mL) and
extracted with DCM (4 x 50 mL). The combined organic layers were dried
over anhydrous MgSO4, filtered and concentrated in vacuo. The residue
2H, aryl-o-CH), 7.63 – 7.56 (m, 1H, aryl-p-CH), 7.49 (t, J = 7.7 Hz, 2H,
aryl-m-CH), 6.26 (dd, 3J = 7.0 Hz, 4J = 3.7 Hz, 1H, NHBoc), 5.99 (d, 3J =
2.4 Hz, 1H, H-1’), 5.66 (dd, 3J = 6.6 Hz, 3J = 2.4 Hz, 1H, H-2’), 4.43 (ddd,
3J = 9.4 Hz, 3J = 3.7 Hz, 3J = 3.2 Hz, 1H, H-4’), 4.03 (ddd, 3J = 9.4 Hz, 3J
= 6.6 Hz, 4J = 2.4 Hz, 1H, H-3’), 3.67 (ddd, 2J = 14.5 Hz, 3J = 7.0 Hz, 3J =
2
3
3.7 Hz, 1H, Ha-5’), 3.57 (ddd, J = 14.5 Hz, J = 7.0 Hz, 3J = 3.2 Hz, 1H,
Hb-5’), 2.28 (d, 4J = 2.4 Hz, 1H, CCH), 2.18 (s, 3H, OCOCH3), 1.46 (s, 9H,
C(CH3)3) ppm. 13C NMR, HSQC, HMBC (101 MHz, CDCl3): δ = 170.0
(OCOCH3), 164.7 (N-CO-aryl), 156.5 (N-CO-O), 152.9 (C-2), 151.1 (C-4),
150.2 (C-6), 142.4 (C-8), 133.5 (aryl-C-CO-N), 133.0 (aryl-p-CH), 129.0
was
purified
by
flash-column
chromatography
(silica
gel,
isohexanes/EtOAc, 2:1→1:1→1:2→EtOAc) to provide nucleoside 12
(1.02 g, 2.29 mmol, 61%) as a colorless foam. The reaction could also be
This article is protected by copyright. All rights reserved.

10.1002/chem.201805409
Chemistry - A European Journal
FULL PAPER
(aryl-m-CH), 128.1 (aryl-o-CH), 124.2 (C-5), 90.5 (C-1’), 83.5 (C-4’), 79.6
(C(CH3)3), 77.4 (C-2’), 76.4 (CCH), 74.4 (CCH), 41.6 (C-5’), 35.8 (C-3’),
28.6 (C(CH3)3), 20.8 (OCOCH3) ppm. ESI-HRMS calcd. for [C26H28N6O6 +
H]+: 521.2143, found: 521.2150.
CH), 128.0 (aryl-o-CH), 124.4 (C=CH), 110.6 (C(CH3)2), 95.8 (C-1), 75.2
(C-4), 69.5 (PhCH2O), 68. 6 (C-3), 63.2 (C-5), 60.8 (OCH2CH3), 26.4
(C(CH3)2), 25.3 (C(CH3)2), 14.3 (OCH2CH3) ppm. ESI-HRMS calcd. for
[C19H24O6 + NH4]+: 366.1911, found: 366.1911.
Benzyl 3,4-O-isopropylidene-β-D-arabinopyranoside (14): The title
compound was prepared according to a modified procedure of Shing et
al.[42] Benzyl β-D-arabinopyranoside (48.0 g, 200 mmol, 1.00 eq.) was
suspended in acetone (500 mL) and 2,2-dimethoxypropane (49.0 mL,
41.6 g; 400 mmol, 2.00 eq.). After addition of p-toluenesulfonic acid
monohydrate (1.14 g, 5.99 mmol, 0.03 eq.), the reaction mixture was
stirred at 60 °C for 2 h to obtain a clear solution. The reaction was
neutralized by treatment with triethylamine (0.84 mL, 0,61 g, 5.99 mmol,
1.00 eq.). Volatile components were removed in vacuo and the residue
Benzyl 2-deoxy-2-C-[(ethoxycarbonyl)methyl]-3,4-O-isopropylidene-
β-D-ribopyranoside (16): The title compound was prepared according to
a modified procedure of Kaiya et al.[43] Vinyl compound 15 (45.0 g,
129 mmol, 1.00 eq.) was dissolved in EtOH (300 mL) and Raney-Ni (ca.
15 mL) was added to the solution at RT. The reaction vessel was
evacuated and flushed with hydrogen three times. Subsequently, the
mixture was stirred under hydrogen atmosphere for 20 h. Upon completion
of the reaction as monitored by TLC, the reaction mixture was filtered
through celite. Volatile materials were removed in vacuo and the residue
was
purified
by
flash-column
chromatography
(silica
gel,
was
purified
by
flash-column
chromatography
(silica
gel,
isohexanes/EtOAc, 2:1→3:2→1:1, gradient elution) to furnish the title
compound 14 (46.8 g, 167 mmol, 84%) as a colorless oil. Rf = 0.54
(isohexanes/EtOAc = 1:1). IR (ATR): ṽ = 3221, 2938, 1499, 1453, 1315,
1252, 1048, 1001, 848, 783, 701 cm−1. 1H NMR, COSY (400 MHz, CDCl3):
δ = 7.42 – 7.28 (m, 5H, aryl-H), 4.94 (d, 3J = 3.6 Hz, 1H, H-1), 4.79 (d, 2J
= 11.7 Hz, 1H, PhCH2O), 4.55 (d, 2J = 11.7 Hz, 1H, PhCH2O), 4.24 (ddd,
3J = 6.2 Hz, 3J = 2.6 Hz, 3J = 1.2 Hz, 1H, H-4), 4.21 (q, 3J = 6.2 Hz, 1H, H-
3), 4.01 (dd, 2J = 13.2 Hz, 3J = 2.6 Hz, 1H, Ha-5), 3.93 (dd, 2J = 13.2 Hz,
3J = 1.2 Hz, 1H, Hb-5), 3.80 (dd, 3J = 6.2 Hz, 3.6 Hz, 1H, H-2), 2.22 (br s,
1H, OH), 1.53 (s, 3H, CH3), 1.36 (s, 3H, CH3) ppm. 13C NMR, HSQC,
HMBC (101 MHz, CDCl3): δ = 137.1 (aryl-C-CH2), 128.7 (aryl-m-CH),
128.22 (aryl-p-CH), 128.15 (aryl-o-CH), 109.4 (C(CH3)2), 97.0 (C-1), 76.1
(C-4), 73.1 (C-3), 70.1 (C-2), 69.9 (PhCH2O), 59.9 (C-5), 28.0 (CH3), 26.1
(CH3) ppm. ESI-HRMS calcd. for [C15H20O5 + NH4]+: 298.1649, found:
298.1651. EI-HRMS calcd. for [C15H20O5 - CH3]+: 265.1071, found:
265.1084.
isohexanes/EtOAc, 4:1) to yield reduced compound 16 (40.8 g, 116 mmol,
90%) as a colorless oil. dr = 13:1. Rf = 0.33 (isohexanes/EtOAc = 4:1). IR
(ATR): ṽ = 2983, 1732, 1455, 1370, 1212, 1071, 1021, 870, 738, 699 cm−1
.
1H NMR, COSY (400 MHz, CDCl3): δ = 7.38 – 7.25 (m, 5H, aryl-H), 4.82
(d, 3J = 11.7 Hz, 1H, OCH2Ph), 4.64 (d, 3J = 8.0 Hz, 1H, H-1), 4.49 (dd, 3J
= 7.4 Hz, 3J = 2.8 Hz, 1H, H-3), 4.48 (d, 3J = 11.7 Hz, 1H, OCH2Ph), 4.24
(ddd, 3J = 7.4 Hz, 3J = 2.6 Hz, 3J = 2.3 Hz, 1H, H-4), 4.10 (q, 3J = 7.2 Hz,
2H, OCH2CH3), 3.84 (dd, 2J = 12.8 Hz, 3J = 2.6 Hz, 1H, Ha-5), 3.63 (dd, 2J
= 12.8 Hz, 3J = 2.3 Hz, 1H, Hb-5), 2.60 – 2.55 (m, 2H, CH2COO), 2.28 (ddd,
3J = 8.0 Hz, 3J = 6.5 Hz, 3J = 2.8 Hz, 1H, H-2), 1.47 (s, 3H, C(CH3)2), 1.31
(s, 3H, C(CH3)2), 1.22 (t, 3J = 7.2 Hz, 3H, OCH2CH3) ppm. 13C NMR, HSQC,
HMBC (101 MHz, CDCl3): δ = 172.2 (C=O), 138.1 (aryl-C-CH2), 128.5
(aryl-m-CH), 128.1 (aryl-p-CH), 127.8 (aryl-o-CH), 109.1 (C(CH3)2), 99.0
(C-1), 73.2 (C-4), 72.6 (C-3), 69.7 (PhCH2O), 62.6 (C-5), 60.7 (OCH2CH3),
37.2 (C-2), 33.6 (CH2COO), 26.7 (C(CH3)2), 25.0 (C(CH3)2), 14.3
(OCH2CH3) ppm. EI-HRMS calcd. for [C19H26O6 - CH3]+:335.1489, found:
335.1481.
Benzyl
2-deoxy-2-C-[(ethoxycarbonyl)methylene]-3,4-O-
isopropylidene-β-D-arabinofuranoside (15): The title compound was
prepared according to a modified procedure of Kaiya et al.[43] Oxalyl
chloride (15.3 mL, 22.4 g, 176 mmol, 1.15 eq.) was dissolved in dry DCM
(600 mL). After cooling to −78 °C, dry DMSO (25.1 mL, 27.6 g, 352 mmol,
2.30 eq.) was added dropwise and the mixture was stirred at −60 °C for
1 h until no further gas development was observed. Subsequently, a
solution of acetonide compound 14 (43.0 g, 153 mmol, 1.00 eq.) in dry
DCM (150 mL) was added slowly over 10 min and the mixture was stirred
at -60 °C for 2 h. The reaction mixture was treated with triethylamine
(64.1 mL, 46.6 g, 460 mmol, 3.00 eq.), stirred at -60 °C for 1 h, quenched
upon addition of saturated aqueous NaHCO3 (300 mL) and extracted with
DCM (3 x 300 mL). The combined organic layers were washed with brine,
dried over anhydrous MgSO4, filtered and concentrated under reduced
pressure. Ketone 15a was obtained as a waxy syrup which was used in
the next step without further purification. EI-HRMS calcd. for
[C15H18O5 - CH3]+: 263.0914, found: 263.0914.
2-Deoxy-3,4-O-isopropylidene-2-C-[(ethoxycarbonyl)methyl]-D-
ribopyranose (17a): To a stirred solution of ester compound 16 (38.0 g,
108 mmol, 1.00 eq.) in EtOH (250 mL) and THF (100 mL) was added Pd/C
(10 wt.%, 1.70 g) under N2 at RT. The reaction vessel was evacuated and
flushed with hydrogen three times. The mixture was stirred under hydrogen
atmosphere for 24 h and then filtered through celite. The solution was
concentrated to dryness under reduced pressure and the residue was
purified by flash-column chromatography (silica gel, isohexanes/EtOAc,
4:1→1:1) to furnish anomeric alcohol 17a as a colorless oil (24.8 g,
95.3 mmol, 88%). α/β = 1:9 (inseparable mixture by fcc). Rf = 0.36
(isohexanes/EtOAc = 1:1). IR (ATR, α/β-mixture): ṽ = 2984, 1731, 1458,
1371, 1213, 1109, 1056, 1020, 868 cm-1. Major β anomer: 1H NMR, COSY
3
(400 MHz, CDCl3): δ = 4.88 (dd, 3J = 8.3 Hz, J = 4.8 Hz, 1H, H-1), 4.44
(dd, 3J = 7.1 Hz, 3J = 2.9 Hz, 1H, H-3), 4.23 (ddd, 3J = 7.1 Hz, 3J = 3.7 Hz,
3J = 3.4 Hz, 1H, H-4), 4.16 (q, 3J = 7.1 Hz, 2H, OCH2CH3), 3.88 (dd, 2J =
12.6 Hz, 3J = 3.4 Hz, 1H, Ha-5), 3.60 (dd, 2J = 12.6, 3J = 3.7 Hz, 1H, Hb-5),
3.26 (d, 3J = 4.8 Hz, 1H, OH), 2.66 (dd, 2J = 16.9 Hz, 3J = 5.9 Hz, 1H,
CH2COO), 2.59 (dd, 3J = 16.9 Hz, 2J = 8.3 Hz, 1H, CH2COO), 2.20 (ddd,
3J = 8.3 Hz, 3J = 5.9 Hz, 3J = 2.9 Hz, 1H, H-2), 1.45 (s, 3H, (CH3)2), 1.30
(s, 3H, C(CH3)2), 1.26 (t, J = 7.1 Hz, 3H, OCH2CH3) ppm. 13C NMR, HSQC,
HMBC (101 MHz, CDCl3): δ = 172.8 (C=O), 109.2 (C(CH3)2), 94.4 (C-1),
73.4 (C-3), 72.7 (C-4), 63.3 (C-5), 61.0 (OCH2CH3), 38.5 (C-2), 33.5
(CH2COO), 27.0 (C(CH3)2), 25.27 (C(CH3)2), 14.3 (OCH2CH3) ppm. EI-
HRMS calcd. for [C12H20O6 - CH3]+: 245.1020, found: 245.1023.
A
mixture of crude ketone 15a (42.0 g, 151 mmol, 1.00 eq.) and
(carbethoxymethylene)triphenylphosphorane[44]
(68.3 g, 196 mmol,
1.30 eq.) in DCM (400 mL) was stirred at RT for 12 h. Volatile components
were evaporated and the residue was purified by flash-column
chromatography (silica gel, isohexanes/EtOAc, 4:1) to afford the title
compound 15 as a colorless oil (46.0 g, 132 mmol, 86% over 2 steps). E/Z
= 4:1 (inseparable mixture by fcc). Rf = 0.84 (isohexanes/EtOAc = 2:1). IR
(ATR, E/Z-mixture): ṽ = 2983, 1718, 1372, 1214, 1150, 1020, 853, 736,
699 cm−1. Major E-Isomer: 1H NMR, COSY (400 MHz, CDCl3): δ = 7.41 –
7.32 (m, 5H, aryl-H), 6.41 (d, 4J = 1.8 Hz, 1H, C=CH), 6.06 (d, 3J = 7.5 Hz,
1H, H-3), 5.44 (d, 4J = 1.8 Hz, 1H, H-1), 4.86 (d, 2J = 11.9 Hz, 1H, OCH2Ph),
4.62 (d, 2J = 11.9 Hz, 1H, OCH2Ph), 4.34 (dd, 3J = 7.5 Hz, 3J = 1.7 Hz, 1H,
Methyl
2-C-carboxymethyl-2-deoxy-2,3-lactone-D-ribofuranoside
(17): The title compound was prepared according to a modified procedure
of Li et al.[45] Anomeric alcohol 17a (20.7 g, 79.5 mmol, 1.00 eq.) was
dissolved in AcOH/H2O (v/v 4:1, 400 mL) and stirred at RT for 24 h. Tthe
mixture was heated to 40 °C for additional 2 h. Volatile materials were
evaporated and the crude product 17b was co-evaporated with
toluene (3 x 300 mL) and used in the next step without further purification.
3
H-4), 4.20 (q, 3J = 7.1 Hz, 2H, OCH2CH3), 3.70 (d, J = 1.7 Hz, 2H, H-5),
1.53 (s, 3H, C(CH3)2), 1.40 (s, 3H, C(CH3)2), 1.30 (t, 3J = 7.1 Hz, 3H,
OCH2CH3) ppm. 13C NMR, HSQC, HMBC (101 MHz, CDCl3): δ = 165.5
(C=O), 147.9 (C-2), 137.6 (aryl-C-CH2), 128.6 (aryl-m-CH), 128.1 (aryl-p-
This article is protected by copyright. All rights reserved.

10.1002/chem.201805409
Chemistry - A European Journal
FULL PAPER
Rf
=
0.05 (isohexanes/EtOAc
=
1:1). EI-HRMS calcd. for
isohexanes/EtOAc, 4:1→2:1→1:1) to afford azide compound 19 (6.08 g,
28.5 mmol, 75%) as a colorless oil. α/β = 2:3 (inseparable mixture by fcc).
Rf = 0.46 (isohexanes/EtOAc = 1:1). IR (ATR, α/β-mixture): ṽ = 2936, 2100,
1776, 1444, 1282, 1160, 1110, 1031, 920 cm-1. β anomer: 1H NMR, COSY
[C12H20O6 - 2 x H2O]+: 184.0730, found: 184.0726.
Concentrated sulfuric acid (0.56 mL) was added to a stirred solution of triol
compound 17b in dry methanol (450 mL) at 0 °C. The reaction mixture was
stirred at 4 °C for 72 h and then neutralized by addition of solid sodium
bicarbonate. The resulting suspension was filtered through celite and the
filtrate was concentrated to dryness under reduced pressure. Purification
by flash-column chromatography (silica gel, isohexanes/EtOAc,
2:1→1:1→1:3) yielded lactone 17 (10.8 g, 57.4 mmol, 72% over 2 steps)
as colorless crystals. α/β = 2:3. β anomer could be isolated for analysis.
M.p. = 45 – 47 °C. Rf (α anomer) = 0.44 (DCM/MeOH = 100:5). Rf (β
anomer) = 0.30 (DCM/MeOH = 100:5). IR (ATR, β anomer): ṽ = 3442,
2940, 1775, 1172, 1102, 1031, 1003, 932 cm−1. Major β anomer: 1H NMR,
COSY (400 MHz, CDCl3): δ = 5.16 (dd, 3J = 7.0 Hz, 3J = 0.9 Hz, 1H, H-3),
4.90 (d, 3J = 1.4 Hz, 1H, H-1), 4.49 – 4.44 (m, 1H, H-4), 3.73 (dd, 2J = 12.8
Hz, 3J = 2.9 Hz, 1H, Ha-5), 3.68 (dd, 2J = 12.8, 3J = 3.6 Hz, 1H, Hb-5), 3.41
(s, 3H, OCH3), 3.13 – 3.05 (m, 1H, H-2), 2.87 (dd, 2J = 18.6 Hz, 3J = 11.0
Hz,1H, CH2COO), 2.55 (dd, 2J = 18.6 Hz, 3J = 3.7 Hz, 1H, CH2COO) ppm.
13C NMR, HSQC, HMBC (101 MHz, CDCl3): δ = 175.6 (C=O), 111.8 (C-1),
86.9 (C-4), 84.8 (C-3), 63.7 (C-5), 55.8 (OCH3), 46.6 (C-2), 32.4 (CH2COO)
ppm. EI-HRMS calcd. for [C8H12O5 - CH2OH]+: 157.0495, found: 157.0494.
3
3
(400 MHz, CDCl3): δ = 4.95 (dd, J = 7.3 Hz, J = 1.3 Hz, 1H, H-3), 4.92
(d, 3J = 1.3 Hz, 1H, H-1), 4.41 – 4.36 (m, 1H, H-4), 3.53 (dd, 2J = 12.7 Hz,
3J = 7.2 Hz, 1H, Ha-5), 3.39 (s, 3H, OCH3), 3.38 (m, 1H, Hb-5), 3.18 – 3.11
(m, 1H, H-2), 2.86 (dd, 2J = 18.6 Hz, 3J = 10.9 Hz, 1H, CH2COO), 2.56 (dd,
2J = 18.6 Hz, 3J = 4.7 Hz, 1H, CH2COO) ppm. 13C NMR, HSQC, HMBC
(101 MHz, CDCl3): δ = 175.3 (C=O), 111.9 (C-1), 84.7 (C-3), 84.0 (C-4),
1
55.7 (OCH3), 53.1 (C-5), 45.8 (C-2), 31.8 (CH2COO) ppm. α anomer: H
3
NMR, COSY (400 MHz, CDCl3): δ = 5.11 (d, J = 5.2 Hz, 1H, H-1), 4.81
(dd, 3J = 7.8 Hz, 3J = 2.9 Hz, 1H, H-3), 4.37 – 4.33 (m, 1H, H-4), 3.68 (dd,
3
2
2J = 13.2 Hz, J = 3.4 Hz, 1H, Ha-5), 3.43 (dd, J = 13.2 Hz, 3J = 3.8 Hz,
1H, Hb-5), 3.38 (s, 3H, OCH3), 3.12 – 3.05 (m, 1H, H-2), 2.73 (dd, 2J = 17.7
3
2
3
Hz, J = 1.6 Hz, 1H, CH2COO), 2.54 (dd, J = 17.7 Hz, J = 9.1 Hz, 1H,
CH2COO) ppm. 13C NMR, HSQC, HMBC (101 MHz, CDCl3): δ = 176.3
(C=O), 104.4 (C-1), 83.8 (C-3), 81.6 (C-4), 55.3 (OCH3), 52.1 (C-5), 44.4
(C-2), 29.0 (CH2COO) ppm. ESI-HRMS calcd. for [C8H11N3O4 + NH4]+:
231.1088, found: 231.1088.
Methyl
5-azido-3-O-benzyl-2,5-dideoxy-2-C-
Methyl
2-C-carboxymethyl-2,5-dideoxy-2,3-lactone-5-tosyl-D-
[(benzyloxycarbonyl)methylene]-D-ribofuranoside (20): The title
compound was prepared according to a modified procedure of Webber et
al.[46] Azide compound 19 (5.05 g, 23.7 mmol, 1.00 eq.) was mixed with
KOH (10.6g, 190 mmol, 8.00 eq.) in THF (250 mL). The stirred suspension
was treated with benzyl bromide (28.1 mL, 40.5 g, 237 mmol, 10.0 eq.)
and refluxed for 5 h. After cooling to 0 °C, the reaction was diluted with
water (250 mL) and extracted with EtOAc (3 x 300 mL). The combined
organic layers were washed with brine (500 mL), dried over anhydrous
MgSO4, filtered and concentrated in vacuo. The crude product was purified
by flash-column chromatography (silica gel, isohexanes/EtOAc,
9:1→4:1→2:1) to furnish benzylated compound 20 as a colorless oil
(8.89 g, 21.6 mmol, 91%). α/β = 2:3. β anomer could be isolated for
analysis. Rf (α anomer) = 0.57 (isohexanes/EtOAc = 4:1). Rf (β anomer) =
0.48 (isohexanes/EtOAc = 4:1). IR (ATR, β anomer): ṽ = 2931, 2100, 1733,
1455, 1282, 1168, 1057, 910, 738, 698 cm−1. β anomer: 1H NMR, COSY
(400 MHz, CDCl3): δ = 7.40 – 7.26 (m, 10H, aryl H), 5.11 (d, 2J = 12.3 Hz,
1H, COOCH2Ph), 5.06 (d, 2J = 12.3 Hz, 1H, COOCH2Ph), 4.82 (d, 3J = 2.3
Hz, 1H, H-1), 4.43 (s, 2H, OCH2Ph), 4.16 – 4.11 (m, 1H, H-3), 4.14 – 4.08
(m, 1H, H-4), 3.37 (s, 3H, OCH3), 3.32 (dd, 2J = 12.7 Hz, 3J = 6.2 Hz, 1H,
Ha-5), 3.25 (dd, 2J = 12.7 Hz, 3J = 4.3 Hz, 1H, Hb-5), 2.86 – 2.79 (m, 1H,
H-2), 2.74 (dd, 2J = 16.5 Hz, 3J = 7.6 Hz, 1H, CH2COO), 2.43 (dd, 2J = 16.5
Hz, 3J = 7.4 Hz, 1H, CH2COO) ppm. 13C NMR, HSQC, HMBC (101 MHz,
CDCl3): δ = 172.2 (C=O), 137.5 (OCH2Ph-C-1), 135.9 (COOCH2Ph-C-1),
128.7, 128.6, 128.41, 128.38, 128.08, 127.8 (aryl 10C), 109.4 (C-1), 81.4
(C-4), 80.3 (C-3), 72.6 (OCH2Ph), 66.6 (COOCH2Ph), 55.8 (OCH3), 54.3
(C-5), 44.2 (C-2), 30.5 (CH2COO) ppm. ESI-HRMS calcd. for [C22H25N3O5
+ NH4]+: 429.2132, found: 429.2138.
ribofuranoside (18): To a stirred solution of lactone compound 17 (10.0 g,
53.1 mmol, 1.00 eq.) in dry pyridine (300 mL) was added a solution of p-
toluenesulfonyl chloride (13.2 g, 69.1 mmol, 1.30 eq.) in pyridine (40 mL)
at 0 °C. After stirring at RT for 18 h, the reaction was quenched by
treatment with MeOH (20 mL). Solvents were removed in vacuo and the
residue was purified by flash-column chromatography (silica gel,
hexane/EtOAc, 3:2→1:1→1:3) to give tosyl compound 18 as colorless
crystals (13.9 g, 40.6 mmol, 76%). α/β = 2:3 (inseparable mixture by fcc).
Crystallization from isohexanes/EtOAc (vapor diffusion) provided suitable
β single crystals for X-ray characterization. M.p. = 78 – 80 °C. Rf = 0.36
(isohexanes/EtOAc = 1:1). IR (ATR, α/β-mixture): ṽ = 2938, 1781, 1598,
1358, 1173, 1111, 979, 815, 665 cm−1. β anomer: 1H NMR, COSY
(400 MHz, CDCl3): δ = 7.82 – 7.75 (m, 2H, aryl H-2-2’), 7.40 – 7.32 (m, 2H,
aryl H-3,-3’), 4.98 (dd, 3J = 7.0 Hz, 3J = 0.9 Hz, 1H, H-3), 4.86 (d, 3J = 1.1
Hz, 1H, H-1), 4.41 – 4.34 (m, 1H, H-4), 4.10 (dd, 2J = 10.3 Hz, 3J = 7.6 Hz,
1H, Ha-5), 4.06 (dd, 2J = 10.3 Hz, 3J = 6.4 Hz, 1H, Hb-5), 3.24 (s, 3H, OCH3),
3.13 – 3.05 (m, 1H, H-2), 2.82 (dd, 3J = 18.6 Hz, 3J = 11.0 Hz, 1H,
CH2COO), 2.56 – 2.47 (m, 1H, CH2COO), 2.45 (s, 3H, aryl CH3) ppm. 13
C
NMR, HSQC, HMBC (101 MHz, CDCl3): δ = 175.3 (C=O), 145.5 (aryl C-
4), 132.5 (aryl C-1), 130.13 (aryl C-3,-3’), 128.02 (aryl C-2,-2’), 111.6 (C-
1), 83.9 (C-3), 82.2 (C-4), 68.3 (C-5), 55.5 (OCH3), 45.5 (C-2), 31.6
(CH2COO), 21.8 (aryl CH3) ppm. α anomer: 1H NMR, COSY (400 MHz,
CDCl3): δ = 7.82 – 7.75 (m, 2H, aryl H-2-2’), 7.40 – 7.32 (m, 2H, aryl H-3-
3’), 4.99 (d, 3J = 3.8 Hz, 1H, H-1), 4.82 (dd, 3J = 7.6 Hz, 3J = 2.7 Hz, 1H,
H-3), 4.32 – 4.28 (m, 1H, H-4), 4.23 (dd, 2J = 11.0 Hz, 3J = 3.1 Hz, 1H, Ha-
5), 4.20 (dd, 2J = 11.0 Hz, 3J = 3.4 Hz, 1H, Hb-5), 3.31 (s, 3H, OCH3), 3.05
3
3
– 2.98 (m, 1H, H-2), 2.67 (dd, J = 17.7 Hz, J = 1.5 Hz, 1H, CH2COO),
2.56 – 2.47 (m, 1H, CH2COO), 2.45 (s, 3H, aryl CH3) ppm. 13C NMR,
HSQC, HMBC (101 MHz, CDCl3): δ = 176.2 (C=O), 145.4 (aryl C-4), 132.4
(aryl C-1), 130.11 (aryl C-3,-3’), 128.03 (aryl C-2,-2’), 104.5 (C-1), 83.1 (C-
3), 80.2 (C-4), 69.0 (C-5), 55.4 (OCH3), 44.1 (C-2), 29.0 (CH2COO), 21.8
(aryl CH3) ppm. ESI-HRMS calcd. for [C12H20O6 + NH4]+: 360.1111, found:
360.1109.
Acetyl
5-azido-3-O-benzyl-2,5-dideoxy-2-C-
[(benzyloxycarbonyl)methylene]-D-ribofuranoside (21): To a solution
of benzylated compound 20 (8.02 g, 19.5 mmol, 1.00 eq.) in AcOH
(70 mL) and Ac2O (70 mL), was added concentrated H2SO4 (0.20 mL) at
0 °C. The solution was warmed to RT and stirred for 3 h. After careful
quenching with saturated NaHCO3 solution (150 mL) and solid NaHCO3
until CO2 evolution stopped, the reaction was extracted with DCM
(4 x 200 mL), washed with brine (400 mL), dried over anhydrous MgSO4
and filtered. Volatile materials were removed in vacuo and the residue was
co-evaporated with toluene (2 x 100 mL). The crude product was purified
by flash-column chromatography (silica gel, isohexanes/EtOAc,
9:1→4:1→2:1) to obtain acetylated compound 21 as a colorless oil (7.27 g,
16.5 mmol, 85%). α/β = 3:2 (inseparable mixture by fcc). Rf = 0.25
(isohexanes/EtOAc = 4:1). IR (ATR): ṽ = 2101, 1733, 1455, 1366, 1230,
1170, 1007, 899, 738, 698 cm−1. β anomer: 1H NMR, COSY (400 MHz,
Methyl
5-azido-2-C-carboxymethyl-2,5-dideoxy-2,3-lactone-D-
ribofuranoside (19): A mixture of tosyl compound 18 (13.0 g, 38.0 mmol,
1.00 eq.) and sodium azide (14.1 g, 152 mmol, 4.00 eq.) was suspended
in DMF (300 mL) and stirred under N2 at 80 °C for 3 h. The yellow
suspension was diluted with brine (200 mL) and extracted with EtOAc
(4x300 mL). The combined organic layers were dried over anhydrous
MgSO4, filtered and concentrated under reduced pressure. The residue
was
purified
by
flash-column
chromatography
(silica
gel,
This article is protected by copyright. All rights reserved.

10.1002/chem.201805409
Chemistry - A European Journal
FULL PAPER
CDCl3): δ = 7.26 (s, 10H, aryl H), 6.09 (d, J = 2.4 Hz, 1H, H-1), 5.09 (d, 2J
= 12.2 Hz, 1H, COOCH2Ph), 5.04 (d, 2J = 12.2 Hz, 1H, COOCH2Ph), 4.45
(s, 2H, OCH2Ph), 4.28 – 4.24 (m, 1H, H-3), 4.16 (ddd, 3J = 9.6 Hz, 3J = 5.0
ribofuranosyl}-6’’’-O-(diphenylcarbamoyl)-2’’’-N-isobutyrylguanin-
5’’’’-yl}-1,2,3-triazole (22): The title compound was prepared according to
a modified procedure of Singh et al.[49] A-half 5 (1.30 g, 2.50 mmol,
1.00 eq.) and G-half 6 (2.39 g, 3.00 mmol, 1.20 eq.) were dissolved in
THF/tert-BuOH/H2O (2:2:1, 80 mL) under N2 at RT. Subsequently, a
solution of sodium ascorbate (0.41 g, 2.00 mmol, 0.80 eq.) in water (3 mL)
and a solution of copper(II) sulfate (0.16 g, 1.00 mmol, 0.40 eq.) in
water (2 mL) was added. The mixture was stirred at RT for 12 h. Volatile
components were evaporated and the residue was purified by flash-
column chromatography (silica gel, DCM/MeOH, 100:2→100:5→10:1) to
provide dinucleotide 22 (2.62 g, 2.00 mmol, 80%) as a colorless foam.
M.p. = 183 °C (decomp.). Rf = 0.46 (DCM/MeOH = 10:1). IR (ATR): ṽ =
3268, 2976, 2106, 1738, 1707, 1584, 1452, 1216 1167, 732 cm−1. 1H NMR,
COSY (400 MHz, CDCl3): δ = 9.14 (s, 1H, NHBz), 8.86 (s, 1H, H-2‘), 8.25
(s, 1H, NHiBu), 8.09 (s, 1H, H-8‘), 8.02 (d, 3J = 7.6 Hz, 1H, Bz-o-CH), 7.85
(s, 1H, H-8‘‘‘), 7.82 (s, 1H, H-5), 7.62 – 7.56 (m, 1H, Bz-p-CH), 7.50 (t, 3J
3
3
Hz, J = 4.6 Hz, 1H, H-4), 3.43 (dd, 2J = 13.2 Hz, J = 4.6 Hz, 1H, Ha-5),
2
3
3.23 (dd, J = 13.2 Hz, J = 5.0 Hz, 1H, Hb-5), 3.00 – 2.92 (m, 1H, H-2),
2.78 (dd, 2J = 16.9 Hz, 3J = 8.4 Hz, 1H, CH2COO), 2.61 (dd, 2J = 16.9 Hz,
3J = 7.1 Hz, 1H, CH2COO), 2.07 (s, 3H, CH3COO) ppm. 13C NMR, HSQC,
HMBC (101 MHz, CDCl3): δ = 171.8 (COOBn), 170.1 (COOCH3), 137.3
(OCH2Ph-C-1), 135.7 (COOCH2Ph-C-1), 128.71, 128.64, 128.51, 128.47,
128.20, 127.84 (aryl 10C), 101.3 (C-1), 82.2 (C-4), 78.9 (C-3), 72.8
(OCH2Ph), 66.8 (COOCH2Ph), 52.5 (C-5), 43.7 (C-2), 30.3 (CH2COO),
1
21.3 (CH3COO) ppm. α anomer: H NMR, COSY (400 MHz, CDCl3): δ =
3
7.41 – 7.22 (m, 10H, aryl H), 6.36 (d, J = 4.8 Hz, 1H, H-1), 5.10 (s, 2H,
COOCH2Ph), 4.46 (d, 2J = 12.0 Hz, 1H, OCH2Ph), 4.42 (d, 2J = 12.0 Hz,
1H, OCH2Ph), 4.32 – 4.27 (m, 1H, H-4), 4.01 (dd, 3J = 7.0, 3J = 2.3 Hz, 1H,
H-3), 3.35 (dd, 2J = 12.9 Hz, 3J = 5.1 Hz, 1H, Ha-5), 3.15 (dd, 2J = 12.9 Hz,
3J = 4.2 Hz, 1H, Hb-5), 2.89 – 2.82 (m, 1H, H-2), 2.77 (dd, 2J = 16.7 Hz, 3J
= 6.6 Hz, 1H, CH2COO), 2.61 (dd, 2J = 16.7 Hz, 3J = 5.8 Hz, 1H, CH2COO),
2.05 (s, 3H, CH3COO) ppm. 13C NMR, HSQC, HMBC (101 MHz, CDCl3):
δ = 172.0 (COOBn), 170.5 (CH3COO), 137.8 (OCH2Ph-C-1), 135.8
(COOCH2Ph-C-1), 128.74, 128.59, 128.52, 128.51, 128.05, 127.80 (aryl
10C), 98.2 (C-1), 84.4 (C-4), 79.2 (C-3), 72.6 (OCH2Ph), 66.7
(COOCH2Ph), 52.7 (C-5), 43.1 (C-2), 28.5 (CH2COO) 21.3 (CH3COO) ppm.
ESI-HRMS calcd. for [C23H25N3O6 + NH4]+: 457.2081, found: 457.2082.
3
= 7.6 Hz, 2H, Bz-m-CH), 7.48 – 7.12 (m, 20H, aryl-H), 6.81 (dd, J = 7.3
Hz, 4J = 3.2 Hz, 1H, NHBoc), 6.19 (d, 3J = 3.9 Hz, 1H, H-1‘‘), 5.95 (d, 3J =
8.2 Hz, 1H, H-1‘‘‘‘), 5.71 (dd, 3J = 7.3 Hz, 3J = 3.9 Hz, 1H, H-2‘‘), 5.15 (dd,
2J = 14.0 Hz, 3J = 6.5 Hz, 1H, Ha-5’’’’), 4.93 (dd, 2J = 14.0 Hz, 3J = 8.0 Hz,
1H, Hb-5’’’’), 4.88 (d, 2J = 12.3 Hz, 1H, COOCH2Ph), 4.87 – 4.83 (m, 1H,
H-4’’’’), 4.83 – 4.78 (m, 1H, H-4‘‘), 4.79 (d, 2J = 12.3 Hz, 1H, COOCH2Ph),
4.61 (d, 2J = 11.6 Hz, 1H, OCH2Ph), 4.40 (d, 3J = 5.3 Hz, 1H, H-3’’’’), 4.37
– 4.32 (m, 1H, H-3’’), 4.32 (d, 2J = 11.6 Hz, 1H, OCH2Ph), 4.15 – 4.04 (m,
1H, H-2’’’’), 3.61 – 3.51 (m, 1H, Ha-5’’), 3.53 – 3.44 (m, 1H, Hb-5’’), 2.80
(dd, 2J = 16.6 Hz, 3J = 8.3 Hz, 1H, CH2COO), 2.59 (hept, 3J = 6.9 Hz, 1H,
CH(CH3)2), 2.45 (dd, 2J = 16.7 Hz, 3J = 7.1 Hz, 1H, CH2COO), 1.70 (s, 3H,
OCOCH3), 1.45 (s, 9H, C(CH3)3), 1.25, (d, 3J = 6.9 Hz, 3H, CH(CH3)2), 1.21,
(d, 3J = 6.9 Hz, 3H, CH(CH3)2) ppm. 13C NMR, HSQC, HMBC (101 MHz,
CDCl3): δ = 174.3 (iBu-CONH), 171.0 (COOBn), 169.8 (OCOCH3), 164.7
(N-CO-Ph), 156.7 (N-CO-OC(CH3)3), 156.4 (C-2‘‘‘), 154.1 (C-4‘‘‘), 152.7
(C-2‘), 151.6 (OCONPh2), 151.1 (C-4‘), 150.4 (C-6‘), 150.1 (C-6‘‘‘), 144.4
(C-8‘‘‘), 142.7 (C-8‘), 141.7 (OCON-Ph2-C1), 140.7 (C-4), 137.1 (OCH2Ph-
C-1), 135.4 (COOCH2Ph-C-1), 133.5 (Bz-C-CO-N), 133.0 (Bz-p-CH),
129.3, 129.0, 128.61, 128.55, 128.42, 128.25, 128.17, 128.03, 127.98 (aryl
25C), 125.2 (C-5), 124.2 (C-5‘), 122.8 (C-5‘‘‘), 90.9 (C-1‘‘‘‘), 90.2 (C-1‘‘),
83.3 (C-4‘‘), 82.1 (C-4‘‘‘‘), 79.6 (C-3‘‘‘‘), 79.4 (C(CH3)3), 77.1 (C-2‘‘), 71.8
(OCH2Ph), 66.7 (COOCH2Ph), 51.2 (C-5‘‘‘‘), 42.4 (C-5‘‘), 40.5 (C-2‘‘‘‘),
39.9 (C-3‘‘), 36.9 (CH(CH3)2), 30.3 (CH2COO), 28.6 (C(CH3)3), 20.3
(OCOCH3), 19.5 (CH(CH3)2), 19.40 (CH(CH3)2) ppm. ESI-HRMS calcd. for
[C69H69N15O13 + H]+: 1316.5272, found: 1316.5330. ESI-HRMS calcd. for
[C69H69N15O13 + Na]+: 1338.5091, found: 1338.5151.
9-{5-Azido-3-O-benzyl-2,5-dideoxy-2-C-
[(benzyloxycarbonyl)methylene]-β-D-ribofuranosyl}-6-O-
(diphenylcarbamoyl)-2-N-isobutyrylguanine
(6):
N,O-
Bis(trimethylsilyl)acetamide (BSA) (2.23 mL, 1.85 g, 9.10 mmol, 4.00 eq.)
was added under N2 to a stirred suspension of compound 21 (1.00 g,
2.28 mmol,
1.00 eq.)
and
6-O-(diphenylcarbamoyl)-2-N-
isobutyrylguanine[47,48] (1.90 g, 4.55 mmol, 2.00 eq.) in dichloroethane
(30 mL) and heated to 80 °C for 30 min until a clear solution was obtained.
The reaction mixture was brought to RT and treated with trimethylsilyl
triflate (TMSOTf) (1.07 mL, 1.32 g, 5.92 mmol, 2.60 eq.). The dark red
solution was stirred at 80 °C for 2 h. The reaction was quenched with
saturated aqueous NaHCO3 (30 mL) at RT and extracted with DCM
(4 x 50 mL). The combined organic layers were dried over anhydrous
MgSO4, filtered and concentrated in vacuo. The residue was purified by
flash-column
chromatography
(silica
gel,
isohexanes/EtOAc,
4:1→2:1→1:1) to give nucleoside 6 (1.31 g, 1.65 mmol, 72%) as a
colorless foam. The reaction could also be performed on a 5 g scale of the
starting material 21 (yield: 59%). Rf = 0.68 (isohexanes/EtOAc = 1:1). IR
(ATR): ṽ = 3321, 2933, 2102, 1731, 1584, 1492, 1268, 1166, 1047, 694
4-[6’-Benzoylamino-9’-(2’’-O-acetyl-5’’-amino-3’’,5’’-dideoxy-β-D-
ribofuranosyl)-9’H-purin-3’’-yl]-1-{9’’’-[3’’’’-O-benzyl-2’’’’,5’’’’-
dideoxy-2’’’’-C-carboxymethyl-β-D-ribofuranosyl]-2’’’-N-
1
cm−1. H NMR, COSY (400 MHz, CDCl3): δ = 7.99 (s, 1H, H-8), 7.92 (s,
1H, NH), 7.47 – 7.18 (m, 20H, aryl-H), 6.00 (d, 3J = 8.2 Hz, 1H, H-1’), 4.94
(d, 2J = 12.3 Hz, 1H, COOCH2Ph), 4.88 (d, 2J = 12.3 Hz, 1H, COOCH2Ph),
4.58 (d, 2J = 11.6 Hz, 1H, OCH2Ph), 4.443 (d, 2J = 11.6 Hz, 1H, OCH2Ph),
4.441 (d, 3J = 5.8 Hz, 3J = 2.1 Hz, 1H, H-3’), 4.27 – 4.21 (m, 1H, H-4’), 3.80
isobutyrylguanin-5’’’’-yl}-2’’’’,5’’-lactame-1,2,3-triazole (24): To
a
stirred solution of dinucleotide 22 (2.12 g, 1.61 mmol, 1.00 eq.) in dry DCM
(40 mL) was added TFA (20 mL) at 0 °C under N2. The mixture was stirred
for 1 h at this temperature and then concentrated in vacuo. The brown
residue was purified by flash-column chromatography (silica gel,
DCM/MeOH, 100:2→100:5 →5:1) to give amino compound 23 as a
colorless solid (1.33 g, 1.30 mmol, 81%). M.p. = 128 °C (decomp.). Rf =
0.39 (DCM/MeOH = 5:1). ESI-HRMS calcd. for [C51H52N14O10 + H]+:
1021.4064, found: 1021.4038. ESI-HRMS calcd. for [C51H52N14O10 – H]-:
1019.3918, found: 1019.3918.
To a solution of amino compound 23 (1.08 g, 1.06 mmol, 1.00 eq.) in EtOH
(50 mL) was added Pd/C (10 wt.%, 0.30 g) under nitrogen stream at RT.
The reaction vessel was evacuated and flushed with hydrogen three times.
The mixture was stirred under hydrogen atmosphere for 36 h and then
filtered through celite. The solution was concentrated to dryness under
reduced pressure. The residue was used in the next step without further
purification. ESI-HRMS calcd. for [C44H46N14O10 + H]+: 931.3594, found:
931.3594. ESI-HRMS calcd. for [C44H46N14O10 – H]-: 929.3448, found:
929.3450.
3
(dd, 2J = 13.0 Hz, J = 6.3 Hz, 1H, Ha-5), 3.74 – 3.63 (m, 1H, H-2’), 3.70
(dd, 2J = 13.0 Hz, 3J = 5.3 Hz, 1H, Hb-5), 2.85 (dd, 2J = 16.7 Hz, 3J = 8.3
Hz, 1H, CH2COO), 2.80 – 2.70 (m, 1H, CH(CH3)2), 2.48 (dd, 2J = 16.7 Hz,
3J = 6.5 Hz, 1H, CH2COO), 1.26 (s, 3H, CH(CH3)2), 1.24 (s, 3H, CH(CH3)2)
ppm. 13C NMR, HSQC, HMBC (101 MHz, CDCl3): δ = 174.7 (CONH),
171.3 (COOBn), 156.3 (C-2), 154.6 (C-4), 151.9 (OCONPh2), 150.4 (C-6),
143.4 (C-8), 141.8 (OCON-Ph-C1), 137.4 (OCH2Ph-C-1), 135.5
(COOCH2Ph-C-1), 129.3, 128.70, 128.67, 128.46, 128.41, 128.28, 128.17
(aryl 20C), 122.0 (C-5), 89.5 (C-1’), 82.8 (C-4’), 80.1 (C-3’), 72.4
(OCH2Ph), 66.8 (COOCH2Ph), 52.6 (C-5’), 42.7 (C-2’), 36.5 (CH(CH3)2),
30.2 (CH2COO), 19.5 (CH(CH3)2), 19.4 (CH(CH3)2) ppm. ESI-HRMS calcd.
for [C43H41N9O7 + H]+: 796.3202, found: 796.3214.
4-{6’-Benzoylamino-9’-[2’’-O-acetyl-5’’-(tert-butoxycarbonyl)amino-
3’’,5’’-dideoxy-β-D-ribofuranosyl]-9’H-purin-3’’-yl}-1-{9’’’-{3’’’’-O-
benzyl-2’’’’,5’’’’-dideoxy-2’’’’-C-[(benzyloxycarbonyl)methylene]-β-D-
This article is protected by copyright. All rights reserved.

10.1002/chem.201805409
Chemistry - A European Journal
FULL PAPER
Finally, the title compound was prepared according to a modified
procedure of Horne et al.[50] and Kinzie et al.[51] To a yellow solution of the
hydrogenated compound 23 and HATU (0.60 g, 1.58mmol, 1.50 eq.) in dry
DMF (1000 mL) was added DIPEA (0.72 mL, 0.54 g, 4.21 mmol, 4.00 eq.)
at RT. The solution turned orange and was stirred at RT for 24 h. After
addition of MeOH (5 mL), volatile materials were removed under reduced
pressure and the crude product was purified by flash-column
chromatography (silica gel, DCM/MeOH, 100:2→100:5→5:1) to yield
cyclized compound 24 as a colorless solid (506 mg, 0.55 mmol, 52% over
2 steps). An analytical sample was provided by RP-HPLC. M.p. = 185 °C
(decomp.). Rf = 0.57 (DCM/MeOH = 5:1). Rt = 16.1 min (RP-HPLC, 15%
to 80% MeCN gradient elution). IR (ATR): ṽ = 3220, 1682, 1608, 1454,
(m, 1H, Hb-5’’), 2.73 (hept, 3J = 6.8 Hz, 1H, CH(CH3)2), 2.09 (s, 3H,
OCOCH3), 2.02 (t, 3J = 12.0 Hz, 1H, CH2CONH), 1.67 – 1.60 (m, 1H,
CH2CONH), 1.11, (d, 3J = 6.8 Hz, 3H, CH(CH3)2), 1.10, (d, 3J = 6.8 Hz, 3H,
CH(CH3)2), -0.39 – -0.49 (m, 1H, H-2‘‘‘‘) ppm. 13C NMR, HSQC, HMBC
(151 MHz, DMSO-d6): δ = 180.1 (iBu-CONH), 171.0 (CH2CONHCH2),
169.2 (OCOCH3), 165.7 (N-CO-Ph), 154.8 (C-2‘‘‘), 151.8 (C-2‘), 151.5 (C-
4‘), 150.7 (C-6‘), 149.1 (C-6‘‘‘), 148.30 (C-4‘‘‘), 144.0 (C-8‘), 142.6 (C-4),
138.2 (C-8‘‘‘), 133.3 (Bz-C-CO-N), 132.5 (Bz-p-CH), 128.52 (Bz-o-CH),
128.49 (Bz-m-CH), 126.6 (C-5), 126.2 (C-5‘), 119.7 (C-5‘‘‘), 89.3 (C-1‘‘),
83.2 (C-1‘‘‘‘),83.0 (C-4‘‘‘‘), 81.0 (C-4‘‘), 77.4 (C-2‘‘), 70.6 (C-3‘‘‘‘), 52.5 (C-
5‘‘‘‘), 47.1 (C-2‘‘‘‘), 44.5 (C-3‘‘), 42.5 (C-5‘‘), 34.8 (CH(CH3)2), 29.2
(CH2CONH), 20.6 (OCOCH3), 18.85 (CH(CH3)2), 18.83 (CH(CH3)2) ppm.
ESI-HRMS calcd. for [C37H38N14O9 + H]+: 823.3019, found: 823.3015. ESI-
HRMS calcd. for [C37H38N14O9 - H]-: 821.2873, found: 821.2873.
1403, 1222, 1049, 797, 708 cm−1 1H NMR, COSY, NOESY (600 MHz,
.
DMSO-d6): δ = 12.06 (s, 1H, NH), 11.59 (s, 1H, NH), 11.27 (s, 1H, NH),
8.83 (s, 1H, H-2‘), 8.68 (s, 1H, H-8‘), 8.54 (s, 1H, H-5), 8.35 (s, 1H, H-8‘‘‘),
8.08 – 8.03 (m, 2H, Bz-o-CH), 8.06 – 8.03 (s, 1H, CH2CONHCH2), 7.68 –
7.63 (m, 1H, Bz-p-CH), 7.59 – 7.53 (m, 2H, Bz-m-CH), 7.52 – 7.49 (m, 2H,
Bn-o-CH), 7.44 – 7.40 (m, 2H, Bn-m-CH), 7.38 – 7.34 (m, 1H, Bn-p-CH),
6.46 (d, 3J = 1.1 Hz, 1H, H-1‘‘), 5.92 (dd, 3J = 5.9 Hz, 3J = 1.1 Hz, 1H, H-
2‘‘), 5.66 (d, 3J = 10.2 Hz, 1H, H-1‘‘‘‘), 4.85 (dd, 2J = 15.0 Hz, 3J = 3.3 Hz,
1H, Ha-5’’’’), 4.80 – 4.77 (m, 1H, Hb-5’’’’), 4.80 (d, 2J = 10.9 Hz, 1H,
OCH2Ph), 4.76 (dd, 3J = 10.6 Hz, 3J = 6.0 Hz, 1H, H-3’’), 4.68 (d, 2J = 10.9
Hz, 1H, OCH2Ph), 4.68 – 4.66 (m, 1H, H-4’’’’), 4.57 (td, 3J = 10.5 Hz, 3J =
4.1 Hz, 1H, H-4‘‘), 4.15 (d, 3J = 3.4 Hz, 1H, H3‘‘‘‘), 3.87 – 3.81 (m, 1H, Ha-
4-[6’-Amino-9’-(5’’-amino-3’’,5’’-dideoxy-β-D-ribofuranosyl)-9’H-
purin-3’’-yl]-1-{9’’’-[2’’’’,5’’’’-dideoxy-2’’’’-C-carboxymethyl-β-D-
ribofuranosyl]-guanin-5’’’’-yl}-2’’’’,5’’-lactame-1,2,3-triazole (4): The
crude compound 25 was dissolved in MeOH (15 mL) and aqueous
ammonia (25%, 15 mL) in a sealed vessel at RT. The mixture was stirred
at 50 °C for 20 h. Volatile components were removed under reduced
pressure. The residue was purified by preparative RP-HPLC to provide the
final compound 4 as a colorless solid (109 mg, 0.18 mmol, 48% over 2
steps). M.p. = 270 °C (decomp.). Rt = 7.8 min (RP-HPLC, 15% to 80%
MeCN gradient elution). IR (ATR): ṽ = 3338, 1639, 1599, 1477, 1419,
3
5’’), 2.91 – 2.83 (m, 1H, Hb-5’’), 2.67 (hept, J = 6.9 Hz, 1H, CH(CH3)2),
2.11 (s, 3H, OCOCH3), 2.04 (t, 3J = 12.0 Hz, 1H, CH2CONH), 1.76 – 1.70
(m, 1H, CH2CONH), 1.07, (d, 3J = 6.9 Hz, 3H, CH(CH3)2), 1.06, (d, 3J = 6.9
Hz, 3H, CH(CH3)2), -0.19 – -0.26 (m, 1H, H-2‘‘‘‘) ppm. 13C NMR, HSQC,
1209, 1089, 1047, 1005, 730 cm−1. H NMR, COSY, NOESY (600 MHz,
1
DMSO-d6): δ = 10.60 (s, 1H, Guanine-NH), 8.34 (s, 1H, H-8‘), 8.19 (s, 1H,
H-2‘), 8.12 (s, 1H, H-5), 8.07 (s, 1H, H-8‘‘‘), 7.73 – 7.69 (m, 1H,
CH2CONHCH2), 7.32 (s, 2H, A-NH2), 6.50 (s, 2H, G-NH2), 6.07 (d, 3J = 1.1
Hz, 1H, H-1‘‘), 5.97 (d, 3J = 5.1 Hz, 1H, H-2‘‘), 5.64 (d, J = 10.3 Hz, 1H, H-
1‘‘‘‘), 5.58 (d, J = 3.6 Hz, 1H, OH-3‘‘‘‘), 4.77 (dd, 2J = 14.9 Hz, 3J = 3.1 Hz,
HMBC (151 MHz, DMSO-d6):
δ
=
180.1 (iBu-CONH), 170.8
(CH2CONHCH2), 169.1 (OCOCH3), 165.7 (N-CO-Ph), 154.8 (C-2‘‘‘), 151.8
(C-2‘), 151.5 (C-4‘), 150.7 (C-6‘), 149.1 (C-6‘‘‘), 148.30 (C-4‘‘‘), 144.0 (C-
8‘), 142.8 (C-4), 138.2 (OCH2Ph-C-1), 137.9 (C-8‘‘‘), 133.3 (Bz-C-CO-N),
132.5 (Bz-p-CH), 128.53 (Bz-o-CH), 128.49 (Bz-m-CH), 128.47 (Bn-m-
CH), 128.3 (Bn-o-CH), 127.9 (Bn-p-CH), 126.8 (C-5), 126.2 (C-5‘), 119.7
(C-5‘‘‘), 89.3 (C-1‘‘), 83.3 (C-1‘‘‘‘), 81.2 (H-4‘‘), 79.9 (H-3‘‘‘‘), 79.8 (H-4‘‘‘‘),
77.5 (C-2‘‘), 72.0 (OCH2Ph), 53.0 (C-5‘‘‘‘), 46.9 (C-2‘‘‘‘), 44.5 (C-3‘‘), 42.4
(C-5‘‘), 34.7 (CH(CH3)2), 29.7 (CH2CONH), 20.7 (OCOCH3), 18.81
(CH(CH3)2), 18.79 (CH(CH3)2) ppm. ESI-HRMS calcd. for [C44H44N14O9 +
H]+: 913.3489, found: 913.3495. ESI-HRMS calcd. for [C44H44N14O9 - H]-:
911.3343, found: 911.3348.
3
1H, Ha-5’’’’), 4.65 – 4.62 (m, 1H, OH-2’’), 4.61 (dd, 2J = 14.9 Hz, J = 1.5
Hz, 1H, Hb-5’’’’), 4.43 (td, 3J = 10.5 Hz, 3J = 4.1 Hz, 1H, H-4‘‘), 4.23 – 4.21
(m, 1H, H-4’’’’), 4.23 (dd, 3J = 10.6 Hz, 3J = 5.6 Hz, 1H, H-3’’), 4.10 – 4.07
(m, 1H, H-3‘‘‘‘), 3.77 – 3.71 (m, 1H, Ha-5’’), 2.84 – 2.77 (m, 1H, Hb-5’’), 1.97
(m, 1H, CH2CONH), 1.56 (dd, 2J = 12.0 Hz, 3J = 2.2 Hz,1H, CH2CONH), -
0.44 – -0.41 (m, 1H, H-2‘‘‘‘) ppm. 13C NMR, HSQC, HMBC (151 MHz,
DMSO-d6): δ = 170.9 (CH2CONHCH2), 156.8 (C-2‘‘‘), 156.2 (C-6‘), 153.8
(C-6‘‘‘), 152.6 (C-2‘), 151.7 (C-4‘‘‘), 148.7 (C-4‘), 143.7 (C-4), 139.81 (C-
8‘), 136.0 (C-8‘‘‘), 126.8 (C-5), 119.5 (C-5‘), 116.1 (C-5‘‘‘), 91.9 (C-1‘‘), 83.7
(C-4‘‘‘‘), 82.7 (C-1‘‘‘‘), 80.8 (C-4‘‘), 75.9 (C-2‘‘), 70.6 (C-3‘‘‘‘), 52.2 (C-5‘‘‘‘),
46.7 (C-2‘‘‘‘), 45.9 (C-3‘‘), 42.7 (C-5‘‘), 29.3 (CH2CONH) ppm. ESI-HRMS
calcd. for [C24H26N14O6 + H]+: 607.2233, found: 607.2231. ESI-HRMS
calcd. for [C24H26N14O6 - H]-: 605.2087, found: 605.2090.
4-[6’-Benzoylamino-9’-(2’’-O-acetyl-5’’-amino-3’’,5’’-dideoxy-β-D-
ribofuranosyl)-9’H-purin-3’’-yl]-1-{9’’’-[2’’’’,5’’’’-dideoxy-2’’’’-C-
carboxymethyl-β-D-ribofuranosyl]-2’’’-N-isobutyrylguanin-5’’’’-yl}-
2’’’’,5’’-lactame-1,2,3-triazole (25): To a solution of dinucleotide 24
(340 mg, 0.37 mmol, 1.00 eq.) in dry DCM (300 mL) was added BCl3
(5.96 mL, 5.96 mmol, 1M in DCM, 16.0 eq.) at -40°C. The mixture was
stirred for 3 days at this temperature, quenched by addition of MeOH
(5 mL) and extracted with saturated sodium bicarbonate (20 mL) and DCM
(4 x 50 mL). The combined organic layers were dried over anhydrous
MgSO4, filtered and concentrated in vacuo. The compound was used in
the next step without further purification. An analytical sample was
prepared by RP-HPLC to yield a colorless solid. M.p. = 258 °C (decomp.).
Rt = 12.5 min (RP-HPLC, 15% to 80% MeCN gradient elution). IR (ATR):
ORCID
Clemens Dialer: 0000-0002-5659-2776
Gregor Witte: 0000-0003-0313-0962
Karl-Peter Hopfner: 0000-0002-4528-8357
Thomas Carell: 0000-0001-7898-2831
Conflict of interest
1
ṽ = 3234, 1756, 1677, 1613, 1460, 1403, 1220, 1047, 796, 707 cm−1. H
The authors declare no conflict of interest.
NMR, COSY, NOESY (600 MHz, DMSO-d6): δ = 12.07 (s, 1H, NH), 11.69
(s, 1H, NH), 11.26 (s, 1H, NH), 8.82 (s, 1H, H-2‘), 8.66 (s, 1H, H-8‘), 8.43
(s, 1H, H-5), 8.33 (s, 1H, H-8‘‘‘), 8.08 – 8.03 (m, 1H, Bz-o-CH), 7.93 – 7.87
(m, 1H, CH2CONHCH2), 7.68 – 7.63 (m, 1H, Bz-p-CH), 7.59 – 7.53 (m, 2H,
Bz-m-CH), 6.42 (d, 3J = 1.1 Hz, 1H, H-1‘‘), 5.88 (dd, 3J = 5.9 Hz, 3J = 1.1
Hz, 1H, H-2‘‘), 5.76 (d, J = 10.2 Hz, 1H, H-1‘‘‘‘), 5.68 (d, J = 3.4 Hz, 1H,
Acknowledgement
We thank Milda Nainytė, Dr. Tynchtyk Amatov and Dr. Markus
Müller for scientific discussions, Dr. David Stephenson for NMR
spectroscopy, Dr. Peter Mayer for X-ray analysis and Dr. Werner
Spahl for high-resolution mass spectrometry. We thank the
Deutsche Forschungsgemeinschaft (DFG) for financial support
via the programs SFB1032 (TP-A5), SFB749 (TP-A4) and SPP-
2
3
3
OH-3‘‘‘‘), 4.78 (dd, J = 15.0 Hz, J = 3.3 Hz, 1H, Ha-5’’’’), 4.71 (dd, J =
3
2
3
10.6 Hz, J = 6.0 Hz, 1H, H-3’’), 4.68 (dd, J = 15.0 Hz, J = 1.8 Hz, 1H,
Hb-5’’’’), 4.50 (td, 3J = 10.4 Hz, 3J = 3.9 Hz, 1H, H-4‘‘), 4.32 – 4.29 (m, 1H,
H-4’’’’), 4.17 – 4.13 (m, 1H, H3‘‘‘‘), 3.82 – 3.74 (m, 1H, Ha-5’’), 2.89 – 2.79
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10.1002/chem.201805409
Chemistry - A European Journal
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[24]
[25]
[26]
Haag, S. M.; Gulen, M. F.; Reymond, L.; Gibelin, A.;
Abrami, L.; Decout, A.; Heymann, M.; van der Goot, F. G.;
Turcatti, G.; Behrendt, R.; Ablasser, A. Nature 2018, 559,
269-273.
Khiar, S.; Lucas-Hourani, M.; Nisole, S.; Smith, N.;
Helynck, O.; Bourgine, M.; Ruffié, C.; Herbeuval, J.-P.;
Munier-Lehmann, H.; Tangy, F.; Vidalain, P.-O. Sci. Rep.
2017, 7, 2561.
1784. Additional funding from the Excellence Cluster EXC117
CiPSM is acknowledged. This project has received additional
funding from the European Research Council (ERC) under the
European Union's Horizon 2020 research and innovation
programme (grant agreement n° EPiR 741912).
Keywords: cGAMP analog, cGAS, STING, ENPP1, cyclophane,
Adli, A. D. F.; Jahanban‐Esfahlan, R.; Seidi, K.;
CuAAC, macrolactamization
Samandari‐Rad, S.; Zarghami, N. Chem. Biol. Drug Des.
2018, 1-12.
[27]
[28]
[29]
Farkona, S.; Diamandis, E. P.; Blasutig, I. M. BMC
Medicine 2016, 14, 73.
Fujino, T.; Okada, K.; Isobe, H. Tetrahedron Lett. 2014,
55, 2659-2661.
Pal, C.; Chakraborty, T. K. A. J. Org. Chem. 2017, 6,
1421-1427.
Gaffney, B. L.; Jones, R. A. Org. Lett. 2014, 16, 158-161.
Kline, T.; Jackson, S. R.; Deng, W.; Verlinde, C. L. M. J.;
Miller, S. I. Nucleosides Nucleotides Nucleic Acids 2008,
27, 1282-1300.
Fernicola, S.; Torquati, I.; Paiardini, A.; Giardina, G.;
Rampioni, G.; Messina, M.; Leoni, L.; Del Bello, F.;
Petrelli, R.; Rinaldo, S.; Cappellacci, L.; Cutruzzolà, F. J.
Med. Chem. 2015, 58, 8269-8284.
Gierlich, J.; Burley, G. A.; Gramlich, P. M. E.; Hammond,
D. M.; Carell, T. Org. Lett. 2006, 8, 3639-3642.
El-Sagheer, A. H.; Brown, T. Chem. Soc. Rev. 2010, 39,
1388-1405.
El-Sagheer, A. H.; Sanzone, A. P.; Gao, R.; Tavassoli, A.;
Brown, T. PNAS 2011, 108, 11338-11343.
Gao, P.; Ascano, M.; Zillinger, T.; Wang, W.; Dai, P.;
Serganov, Artem A.; Gaffney, Barbara L.; Shuman, S.;
Jones, Roger A.; Deng, L.; Hartmann, G.; Barchet, W.;
Tuschl, T.; Patel, Dinshaw J. Cell 2013, 154, 748-762.
Farrugia, L. J. Appl. Crystallogr. 2012, 45, 849-854.
Cavlar, T.; Deimling, T.; Ablasser, A.; Hopfner, K. P.;
Hornung, V. The EMBO Journal 2013, 32, 1440-1450.
Betkekar, V. V.; Panda, S.; Kaliappan, K. P. Org. Lett.
2012, 14, 198-201.
Yoshimura, Y.; Sano, T.; Matsuda, A.; Ueda, T. Chem.
Pharm. Bull. 1988, 36, 162-167.
Parr, I. B.; Horenstein, B. A. J. Org. Chem. 1997, 62,
7489-7494.
Shing, T. K. M.; Leung, Y. C.; Yeung, K. W. Tetrahedron
2003, 59, 2159-2168.
Kaiya, Y.; Hasegawa, J.-i.; Momose, T.; Sato, T.; Chida,
N. Chem. Asian J. 2011, 6, 209-219.
Mondal, S.; Mohamed, R. K.; Manoharan, M.; Phan, H.;
Alabugin, I. V. Org. Lett. 2013, 15, 5650-5653.
Li, N.-S.; Lu, J.; Piccirilli, J. A. Org. Lett. 2007, 9, 3009-
3012.
Webber, M. J.; Warren, S. A.; Grainger, D. M.; Weston,
M.; Clark, S.; Woodhead, S. J.; Powell, L.; Stokes, S.;
Alanine, A.; Stonehouse, J. P.; Frampton, C. S.; White, A.
J. P.; Spivey, A. C. Org. Biomol. Chem. 2013, 11, 2514-
2533.
Jenny, T. F.; Schneider, K. C.; Benner, S. A. Nucleosides
and Nucleotides 1992, 11, 1257-1261.
Robins, M. J.; Zou, R.; Guo, Z.; Wnuk, S. F. J. Org. Chem.
1996, 61, 9207-9212.
[1]
[2]
Danilchanka, O.; Mekalanos, John J. Cell 2013, 154, 962-
970.
Römling, U.; Gomelsky, M.; Galperin, M. Y. Mol. Microbiol.
2005, 57, 629-639.
Gomelsky, M. Mol. Microbiol. 2011, 79, 562-565.
Hengge, R. Nat. Rev. Microbiol. 2009, 7, 263.
Shu, C.; Yi, G.; Watts, T.; Kao, C. C.; Li, P. Nat. Struct.
Mol. Biol. 2012, 19, 722.
Ablasser, A.; Goldeck, M.; Cavlar, T.; Deimling, T.; Witte,
G.; Röhl, I.; Hopfner, K.-P.; Ludwig, J.; Hornung, V. Nature
2013, 498, 380.
Wu, J.; Sun, L.; Chen, X.; Du, F.; Shi, H.; Chen, C.; Chen,
Z. J. Science 2013, 339, 826-830.
Gao, P.; Ascano, M.; Wu, Y.; Barchet, W.; Gaffney,
Barbara L.; Zillinger, T.; Serganov, Artem A.; Liu, Y.;
Jones, Roger A.; Hartmann, G.; Tuschl, T.; Patel,
Dinshaw J. Cell 2013, 153, 1094-1107.
Sun, L.; Wu, J.; Du, F.; Chen, X.; Chen, Z. J. Science
2013, 339, 786-791.
Chen, Q.; Sun, L.; Chen, Z. J. Nat. Immun. 2016, 17,
1142.
[3]
[4]
[5]
[30]
[31]
[6]
[32]
[7]
[8]
[33]
[34]
[35]
[36]
[9]
[10]
[11]
[12]
[13]
[14]
[15]
[16]
Zhang, X.; Shi, H.; Wu, J.; Zhang, X.; Sun, L.; Chen, C.;
Chen, Zhijian J. Mol. Cell 2013, 51, 226-235.
Kato, K.; Omura, H.; Ishitani, R.; Nureki, O. Annu. Rev.
Biochem. 2017, 86, 541-566.
Shu, C.; Li, X.; Li, P. Cytokine Growth Factor Rev. 2014,
25, 641-648.
[37]
[38]
[39]
[40]
[41]
[42]
[43]
[44]
[45]
[46]
Cai, X.; Chiu, Y.-H.; Chen, Zhijian J. Mol. Cell 2014, 54,
289-296.
Li, L.; Yin, Q.; Kuss, P.; Maliga, Z.; Millán, J. L.; Wu, H.;
Mitchison, T. J. Nat. Chem. Biol. 2014, 10, 1043.
Kato, K.; Nishimasu, H.; Oikawa, D.; Hirano, S.; Hirano,
H.; Kasuya, G.; Ishitani, R.; Tokunaga, F.; Nureki, O.
Nature Communications 2018, 9, 4424.
Opoku-Temeng, C.; Zhou, J.; Zheng, Y.; Su, J.; Sintim, H.
O. Chem. Commun. 2016, 52, 9327-9342.
Schwede, F.; Genieser, H.-G.; Rentsch, A. Handb. Exp.
Pharmacol. 2015, 251-263.
[17]
[18]
[19]
[20]
Clivio, P.; Coantic-Castex, S.; Guillaume, D. Chem. Rev.
2013, 113, 7354-7401.
Knouse, K. W.; deGruyter, J. N.; Schmidt, M. A.; Zheng,
B.; Vantourout, J. C.; Kingston, C.; Mercer, S. E.;
Mcdonald, I. M.; Olson, R. E.; Zhu, Y.; Hang, C.; Zhu, J.;
Yuan, C.; Wang, Q.; Park, P.; Eastgate, M. D.; Baran, P.
S. Science 2018, 361, 1234-1238.
Lioux, T.; Mauny, M.-A.; Lamoureux, A.; Bascoul, N.;
Hays, M.; Vernejoul, F.; Baudru, A.-S.; Boularan, C.;
Lopes-Vicente, J.; Qushair, G.; Tiraby, G. J. Med. Chem.
2016, 59, 10253-10267.
Fu, J.; Kanne, D. B.; Leong, M.; Glickman, L. H.;
McWhirter, S. M.; Lemmens, E.; Mechette, K.; Leong, J.
J.; Lauer, P.; Liu, W.; Sivick, K. E.; Zeng, Q.; Soares, K.
C.; Zheng, L.; Portnoy, D. A.; Woodward, J. J.; Pardoll, D.
M.; Dubensky, T. W.; Kim, Y. Science Transl. Med. 2015,
7, 283ra252-283ra252.
[21]
[22]
[47]
[48]
[49]
[50]
K. Singh, S.; K. Sharma, V.; Bohra, K.; E. Olsen, C.; K.
Prasad, A. Curr. Org. Synth. 2014, 11, 757-766.
Horne, W. S.; Olsen, C. A.; Beierle, J. M.; Montero, A.;
Ghadiri, M. R. Angew. Chem. Int. Ed. 2009, 48, 4718-
4724.
[51]
Kinzie, C. R.; Steele, A. D.; Pasciolla, S. M.; Wuest, W. M.
Tetrahedron Lett. 2014, 55, 4966-4968.
[23]
Corrales, L.; McWhirter, S. M.; Dubensky, T. W., Jr.;
Gajewski, T. F. J. Clin. Invest. 2016, 126, 2404-2411.
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10.1002/chem.201805409
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FULL PAPER
A Click-Chemistry Linked
2’3’-cGAMP Analog
Clemens R. Dialer, Samuele Stazzoni,
David J. Drexler, Felix M. Müller, Simon
Veth, Alexander Pichler, Hidenori
Okamura, Gregor Witte, Karl-Peter
Hopfner and Thomas Carell*
2’3’-cGAMP is an uncanonical cyclic dinucleotide which consists of one A and one G
nucleobase connected via a 3’-5’ and a unique 2’-5’ phosphodiester linkage. Here,
we report a convergent synthesis of a cGAMP analog with one amide and one triazole
replacing the natural phosphodiester backbone. The molecule is best prepared by a
first CuAAC reaction followed by macrolactamization.
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