Combination of click chemistry and sulfonamides to develop three-armed triazole compounds

Article abstract of DOI:10.1016/j.tet.2014.06.125

Fragment-based drug discovery is a valuable tool in hit identification, as well as the combination of different small fragments showing a minimal binding activity against biological receptors or enzymes to give merged hits. A high number of fragments on the same scaffold improve the probability to find a candidate showing single- or multi-target affinities. A rapid and versatile approach for synthesizing libraries of densely fragment-functionalized scaffolds is reported. Many fragments were assembled in few steps around a triazole ring starting from amino alcohols and other readily available building blocks. A binding assay against integrin α_vβ₃ was used as a test-bed in order to demonstrate the potential of such an approach in hit discovery strategies.

Full text of DOI:10.1016/j.tet.2014.06.125

Tetrahedron xxx (2014) 1e11
Contents lists available at ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Combination of click chemistry and sulfonamides to develop
three-armed triazole compounds
,
b
a,
Pierangelo Fabbrizzi a , Francesca Bianchini , Gloria Menchi c, Silvia Raspanti b,
*
Antonio Guarna a c, Andrea Trabocchi a
,
,c,
*
a Department of Chemistry “Ugo Schiff”, University of Florence, Via della Lastruccia 13, 50019 Sesto Fiorentino, Florence, Italy
b Department of Biomedical, Experimental and Clinical Sciences “Mario Serio”, University of Florence, Viale Morgagni 50, 50134 Florence, Italy
c Interdepartmental Center for Preclinical Development of Molecular Imaging (CISPIM), University of Florence, Viale Morgagni 85, 50134 Florence, Italy
a r t i c l e i n f o
a b s t r a c t
Article history:
Fragment-based drug discovery is a valuable tool in hit identification, as well as the combination of
different small fragments showing a minimal binding activity against biological receptors or enzymes to
give merged hits. A high number of fragments on the same scaffold improve the probability to find
a candidate showing single- or multi-target affinities. A rapid and versatile approach for synthesizing
libraries of densely fragment-functionalized scaffolds is reported. Many fragments were assembled in
few steps around a triazole ring starting from amino alcohols and other readily available building blocks.
A binding assay against integrin avb3 was used as a test-bed in order to demonstrate the potential of such
an approach in hit discovery strategies.
Received 7 March 2014
Received in revised form 18 June 2014
Accepted 30 June 2014
Available online xxx
Keywords:
Click chemistry
Azides
Ó 2014 Elsevier Ltd. All rights reserved.
Alkynes
Medicinal chemistry
Cycloadditions
1. Introduction
In the present work we propose a versatile ‘click-based’ ap-
proach in developing libraries of densely functionalized scaffolds
The interactions between biological targets (either receptors
or enzymes) and pharmacophoric subunits are the fundamental
concept of fragment-based approach to drug discovery issues.
Many repeating subunits responsible of such useful interactions
can be identified in bioactive molecules.1e14 Thus, the
combination of different ‘binder’ fragments within a single
molecule dramatically increase the probability of finding new
promising ‘hit’ candidates, both for single- and multi-target
purposes.15,16
Copper (I) catalyzed Azide-Alkyne Cycloaddition (CuAAC),
the main tool of the so-called ‘click-chemistry’, is a versatile
reaction allowing for the synthesis of functionalized triazole
ring in high yield and mild reaction conditions, and nowadays
is one of the most widespread used reaction in chemical syn-
thesis.17e24 Furthermore, the triazole ring has been proposed as
a stable isostere of the peptide bond,25e27 thus, click chemistry
is considered a very promising tool in the synthesis of bioactive
ligands.28,29
containing three fragments.30
2. Results and discussion
As CuAAC is usually intended to couple an azide and an alkyne, it
can be easily exploited in a basic approach in order to achieve
a difunctional scaffold. In view of further expanding the combina-
tion of multiple fragments in a single molecule, our aim was to
develop highly functionalized structures still applying the click
chemistry (Fig. 1), as versatility and mild reaction conditions are
fundamental requirements in a useful library development tech-
nique. Thus, we envisaged the synthesis of building blocks con-
taining three more pharmacophoric moieties in
approach, and specifically we focused our attention on single-
fragment-bearing alkyne and multi-fragment-bearing azides.
a modular
* Corresponding authors. Fax: þ39 055 4574913; e-mail addresses: pfabbrizzi@
gmail.com (P. Fabbrizzi), andrea.trabocchi@unifi.it (A. Trabocchi).
Fig. 1. Schematic of a high-density fragments bearing triazole.
http://dx.doi.org/10.1016/j.tet.2014.06.125
0040-4020/Ó 2014 Elsevier Ltd. All rights reserved.
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2
P. Fabbrizzi et al. / Tetrahedron xxx (2014) 1e11
In choosing our model bioactive fragments, we opted for one
performing the reactions at 120 ꢀC for 60 min. A final acidic
deprotection step yielded the water-soluble form of compounds
18e26, without need of further purification, except for solvent
evaporation. The overall total yields for the five-step process were
about 10e40% starting from the amino alcohols, appearing to be
definitely fit for purpose.
basic group and one acidic group, which can be found in many
peptidic and non-peptidic drug-candidates.31 Such moieties were
used in their protected forms, in order to perform reaction and
purification steps with the benefits of organic solvent-soluble
substances. Additional common fragments found in enzymes and
receptors ligands are
p-stacking capable aromatic rings and
heteroatom-based bonds, one the most widespread being the sul-
fonamidic bond. Sulfonamides are widely used in medicinal
chemistry for targeting many different receptors, thus, using this
versatile scaffold increase the probability to find a multi-target hit,
which is a noteworthy additional advantage in designing drug
candidates.32e34
The first phase of our approach was the synthesis of a sub-
library differing only in the size of the backbone-chain
(Scheme 1). Intermediates 34e39 were synthesized starting from
amino alcohols and esters derivatives. Amino alcohols appeared to
be the ideal starting materials, taking advantage of the orthogo-
nality of oxygen and nitrogen atoms with respect to the reactivity
with a
-bromo acetates.35 Similar profile was exploited for the Mi-
chael addition to acrylate ester, which was considered in order to
achieve the isolation of the homologous products not achievable
from bromo-propionic ester.36
Scheme 2. CuAAC and deprotection steps two achieve the three-fragment-bearing
scaffolds.
In order to further explore the broad spectrum of functionalities,
which can be included in such molecules, another subset of
structures was synthesized, varying the basic and the aromatic
moieties. Two different groups were chosen, representing many
useful characteristics in bioactive candidates, including the fluo-
rogenic dansyl ring and a 2-amino-pyridine as a less basic guani-
dine isostere (31 and 32, see Scheme 3).
Scheme 1. Modular approach to azides containing two fragments.
The alkylated amino alcohols were subsequently reacted with
the chosen aromatic sulfonyl chloride (yields ranging from 40% to
quantitative) and converted into azides 1e8 via a Mitsunobu re-
action. Then, the azides bearing two fragments were reacted via
CuAAC with two different Boc-protected alkynes (27, 28) bearing
the basic moiety (Scheme 2). Full-organic reaction conditions
(THF/CuP(OEt)3I) were preferred for the CuAAC process to mini-
mize the formation of copper-complexes, based upon our previous
experience.37 The cycloaddition step was performed under mi-
crowave irradiation in order to shorten the reaction times and
improve yields, all being in the medium-to-excellent range after
Scheme 3. Scope of the modular approach employing a different alkyne and the
dansyl group as additional fragments.
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P. Fabbrizzi et al. / Tetrahedron xxx (2014) 1e11
3
The application of such chemical moieties was accomplished
through the same synthetic pathway, with ‘click’ step yields rang-
ing from 54% to >95%.
To demonstrate the value of the reported approach in the field of
bioactive molecules, the library was screened using integrin avb3 as
a test-bed in a competition assay versus [125I]-echistatin. The avb3 is
a well-known membrane receptor being subject in many research
studies in the fields of cancer research and part of our group
expertise.37e43 Specifically, in vitro solid-phase assays were per-
account the binding interactions as observed in the crystal struc-
ture of the bound ligandeprotein complex. Docking calculations of
compound 21 resulted in a cluster of conformations characterized
by key interactions with Asp218, Asp150 and the MIDAS site of the
integrin (Fig. 2). Specifically, the COOH group was found in the
MIDAS site and interacting with Ser121 and Mn2þ, and the guani-
dinium group undergoing a salt-bridge interaction with Asp218 and
Asp150. The main conformation of 21 showed two
p-stacking in-
teractions experienced by the aromatic ring facing Tyr-122, and the
triazole ring interacting with Tyr178. This binding profile was in
agreement with the ligand properties of Cilengitide with the ex-
formed at 10 and 1 mM concentrations of the test compounds for
their ability to compete with 125I-echistatin for binding to pure avb3
integrin isolated from human placenta, and % binding activity is
reported in Table 1. Preliminar structure-activity-relationship
considerations were formulated starting from the binding assay
results. A major influence of the backbone-chain length on the
binding activity was easily assessed, with 19 (n¼2, m¼1, l¼3) and
ception of the characteristic
p-stacking interactions between the
triazole ring of 21 and Tyr178. Also, an additional catione
p
in-
teraction46 experienced by Arg214 and the phenyl ring of 21 was
found. Such evidences on the binding mode confirmed l¼3 and
m¼1 as a must-have requisite for the compounds in order to ach-
ieve a good binding affinity towards the integrin.
21 (n¼3, m¼1, l¼3) being the most active ligands at one and 10
mM
conc. Also, the analysis of all the reported compounds suggested
l¼3 and m¼1 as a must-have requisite for the compounds in order
to achieve a good binding affinity towards the integrin. In addition,
the comparison between 19 and the inactive pair 26 and 25 readily
showed the primary importance of the aromatic moiety in tuning
the binding properties. Finally, the amino-pyridine derivatives 32
and 31 showed no activity at 1 mM (37% and 17% at 10 mM, re-
spectively), revealing the significant influence of different basic
groups in tuning the binding affinity.
Table 1
Competition studies of triazole ligands versus [125I]-echistatin against avb3 integrin,
reporting % binding activity at 10 and 1 mM conc. of the test compounds
Cmpd
n
m
R
l
% Binding at 1
m
M
% Binding at 10 mM
Fig. 2. RGD ligand docked into the binding region of avb3 integrin, highlighting protein
residues (magenta) that form key interactions: Asp218/Asp150 versus guanidine or
isostere, Mn2þ versus COOH group, Tyr122 and Arg214 versus phenyl moiety, Tyr178
and triazole. Coordinates of reference compound Cilengitide, c[RGDf(Me)V] (data from
PDB: 1L5G) are shown in red for comparison. Non-polar hydrogen atoms are omitted
for clarity.
18
19
20
21
22
23
24
25
26
2
2
2
3
3
4
4
2
2
1
1
2
1
2
1
2
1
1
CH3
CH3
CH3
CH3
CH3
CH3
CH3
F
2
3
3
3
3
3
3
3
3
6.5
39
11.5
31
25
29
18.8
0
0
15.9
66.5
48.8
88
47
47
44
0
30.3
As regarding to druggability features of the compounds, a pre-
liminar evaluation of the physico-chemical properties of all the
compounds was carried out, as shown in Table 2. Specifically, se-
lected physico-chemical parameters (MW, logP, 2D and 3D surface
areas, no. of rotatable bonds, and H-bond acceptors/donors) were
evaluated with calculator plugins within the framework of Mar-
vinSketch v6.2.2 (Chemaxon Ltd., http://www.chemaxon.com). All
compounds but 31 and 32 followed the Lipinski rule of MW <500.
Torsional, H-bond capability and 2D surface area values were found
to be similar for all library members. These parameters suggested
the need of further structural improvement, for example, by
inserting structural constraints, so as to reduce the number of ro-
tatable bonds, H-bond acceptors and donors count and polar sur-
face area, according to Veber rules, provided the maintenance of
the bioactivity profile.47
Ph
In order to assess the binding mode of these triazole-containing
compounds to avb3 integrin, a docking simulation was carried out.
The crystal structure of the complex formed by c[RGDf(Me)V], also
known as Cilengitide (Fig. 2, red structure), and the extracellular
fragment of avb3 integrin (PDB code: 1L5G) provides a general
mode of interaction between the integrin and its ligands.44 Asp
carboxylate and Arg guanidinium moieties of RGD-based ligands
are key structural elements for receptor recognition. Specifically,
the carboxylate group interacts with the metal ion-dependent ad-
hesion site (MIDAS) of the Mn2þ ion and Ser121/Ser123, whereas
the Arg guanidinium group is responsible for salt-bridge in-
teractions with Asp218 and Asp150 side chains. Additional
3. Conclusions
ligandereceptor contacts occur between Tyr122 in hydrophobic p-
stacking and Asn215. The docking program Autodock 4.045 was
used to evaluate the binding energies of selected conformations of
ligand 21. Docked conformations were analyzed by taking into
A modular strategy involving only reactions performed in mild
conditions starting from readily available building blocks was de-
veloped, in order to synthesize highly functionalized molecules
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4
P. Fabbrizzi et al. / Tetrahedron xxx (2014) 1e11
Table 2
4.2. General procedure Adalkylation of amino alcohols with
bromides
Physico-chemical properties prediction of triazole ligands
Cmpd MW
log P
PSA (2D) VdWSA Rot.
H-bond
H-bond
(3D)
bonds acceptors donors
The selected amino alcohol (3 equiv) was dissolved in dry THF
18
19
20
21
22
23
24
25
26
31
32
409.46 ꢂ0.02 167.29
575.43
606.56
636.39
638.04
668.22
667.93
698.54
581.53
681.79
781.70
664.19
9
10
11
11
12
12
13
10
11
12
11
9
9
9
9
9
9
9
9
9
9
8
4
4
4
4
4
4
4
4
4
2
2
(0.25 M), and methyl or ethyl bromoacetate (1 equiv) was slowly
added at 0 ꢀC. The reaction was stirred for 4 h at 0 ꢀC, then the
solvent was evaporated in vacuo. For further characterization of 35
and 36 see Ref. 35.
423.49
437.52
437.52
451.54
451.54
465.57
0.27 167.29
0.50 167.29
0.33 167.29
0.56 167.29
0.84 167.29
1.08 167.29
4.2.1. Ethyl N-(2-hydroxyethyl)glycinate (34). Synthesized accord-
ing to literature procedure.35,48
427.45 ꢂ0.1
167.29
485.56
537.63
537.63
1.40 167.29
2.63 133.55
2.63 130.31
4.2.2. Methyl
N-(3-hydroxypropyl)glycinate
(35). Synthesized
according to general procedure A, starting from propanolamine and
methyl bromoacetate. Purified via FCC starting elution with
DCMeMeOH 10:1, then increasing polarity to 5:1. 60% yield, col-
orless oil. 1H NMR (200 MHz, CDCl3)
d
3.80 (t, J¼5.7 Hz, 2H), 3.74 (s,
suitable as ‘hit’ candidates for high-throughput screening and
multiple receptor targeting in a drug discovery. Those structures
were designed around an 1,2,3 triazole ring as a non-classical iso-
stere of the peptide bond. Amino alcohols revealed to be versatile
starting materials, due to the wide possibilities of derivatization
offered by the concurrent presence of easily functionalizable oxy-
gen and nitrogen atoms. The selected approach did not require any
protecting-group strategy, except for the differentiation between
organic-soluble intermediates and water-soluble final products.
The possibility to readily vary each segment of the model scaffold
except for the triazole ring was demonstrated, and the library was
screened against integrin avb3 in order to prove the suitability of
the compounds in the field of bioactive ligands. A preliminary SAR
analysis showed a major influence of the combination of the type
and position of the three selected functional groups on binding
properties. Further high-throughput assays against different re-
ceptors and enzymes will elucidate the full potential of such
a synthetic approach.
3H), 3.45 (s, 2H), 2.86 (t, J¼6 Hz, 2H), 1.78e169 (m, 2H). Anal. Calcd
For C6H13NO3: C, 48.97; H, 8.90; N, 9.52. Found C, 48.86; H, 8.78; N,
9.43.
4.2.3. Ethyl N-(4-hydroxybutyl)glycinate (36). Synthesized accord-
ing to general procedure A, starting from butanolamine and ethyl
bromoacetate. Purified via FCC, eluting with DCMeMeOH 6:1. 89%
yield, colorless oil. (TLC DCMeMeOH 5:1, Rf¼0.43). 1H NMR
(300 MHz, CDCl3)
d
4.20 (q, J¼7.2 Hz, 2H), 3.63e3.58 (m, 2H), 3.41
(s, 2H), 3,21 (br s, 1H), 2.71e2.66 (m, 2H), 1.68e1.61 (m, 4H), 1.28 (t,
J¼6.9 Hz, 3H). Anal. Calcd For C8H17NO3: C, 54.84; H, 9.78; N, 7.99.
Found C, 54.69; H, 9.62; N, 7.88.
4.3. General procedure BdMichael addition to amino
alcohols
The amino alcohol (1 equiv) was cooled to 0 ꢀC and methyl ac-
rylate (1 equiv) was slowly added. The reaction was stirred 1 h at
4. Experimental section
4.1. General remarks
0
ꢀC, then the mixture was allowed to warm up to room temper-
ature and stirred for additional 24 h, to afford the corresponding
pure product.
1H NMR and 13C NMR spectra were recorded on a Varian Mer-
cury 400 (1H: 400 MHz, 13C:100 MHz), a Varian Gemini 200 (1H:
200 MHz, 13C: 50 MHz), or a Varian Gemini 300 (1H: 300 MHz, 13C:
4.3.1. Ethyl
N-(2-hydroxyethyl)-
b
-alaninate
(37). Synthesized
according to general procedure B, starting from ethanolamine and
ethyl acrylate. Quantitative yield, white oil. 1H NMR (300 MHz,
75 MHz). The chemical shifts (
d
) and coupling constants (J) are
CDCl3)
d
4.44 (q, J¼7.2 Hz, 2H), 3.62 (t, J¼5.4 Hz, 2H), 2.90 (t,
expressed in parts per million (ppm) and hertz (Hz), respectively.
Flash column chromatography (FCC) purifications were performed
manually using glass columns using Merck silica gel
(0.040e0.063 mm), or using Biotage Isolera system and SNAP silica
cartridges. TLC analyses were performed on Merck silica gel 60 F254
J¼6.3 Hz, 2H), 2.78 (t, J¼5.4 Hz, 2H), 2.50 (t, J¼6.3 Hz, 2H), 1.26 (t,
J¼7.2 Hz, 3H). Anal. Calcd For C7H15NO3: C, 52.16; H, 9.38; N, 8.69.
Found C, 51.95; H, 9.32; N, 8.63.
4.3.2. Methyl N-(3-hydroxypropyl)-b-alaninate (38). Synthesized
€
plates. Melting points were recorded on a BUCHI B-540 instrument
according to general procedure B, starting from propanolamine and
and were uncorrected. IR spectra were recorded with a FTIR-1600
PerkineElmer spectrophotometer. Elemental analyses were recor-
ded on a Perkin Elmer 240 C,H,N analyzer. ESI mass spectra were
recorded on a Thermo LCQ-Fleet. Tetrahydrofuran (THF) was dis-
tilled over Na/benzophenone, and dichloromethane (DCM) was
distilled over CaH2. Microwave heated reactions were performed in
a Biotage Initiator instrument. All commercially available reagents
and solvents were used as received, unless otherwise specified.
Semipreparative HPLC was performed on a BeckmaneCoulter
methyl acrylate. Quantitative yield, white oil. 1H NMR (200 MHz,
CDCl3)
d
3.79 (t, J¼5.4 Hz, 2H), 3.68 (s, 3H), 2.95e2.82 (m, 4H), 2.51
(t, J¼6.2 Hz, 2H), 1.77e1.61 (m, 2H). Anal. Calcd For C7H15NO3: C,
52.16; H, 9.38; N, 8.69. Found C, 51.92; H, 9.34; N, 8.66.
4.3.3. Methyl N-(4-hydroxybutyl)-b-alaninate (39). Synthesized
general procedure B, starting from buthanolamine and methyl ac-
rylate. Quantitative yield, white oil. 1H NMR (300 MHz, CDCl3)
d
3.56 (s, 3H), 3.48e3.40 (m, 2H), 2.76 (t, J¼6.3 Hz, 2H), 2.67 (t,
System GOLD (column Grace Alltima C18 10
analytical HPLC purity tests were performed on a Dionex UltiMate
3000 (columns: Phenomenex Synergi Fusion-RP 80A,
150ꢁ4.6 mm and Phenomenex Synergi-RP 80A,
mm, 250ꢁ10 mm),
J¼7.2 Hz, 2H), 2.44e2.40 (m, 2H), 1.51e1.49 (m, 2H), 1.48e1.44 (m,
2H). Anal. Calcd for C8H17NO3: C, 54.84; H, 9.78; N, 7.99. Found C,
54.65; H, 9.69; N, 7.87.
4
mm
4
mm
150ꢁ4.6 mm), using 0.1% TFA-buffered elution gradient
CH3CNeH2O from 5:95 to 95:5. All tested compounds were >95%
pure. Other abbreviations: DPPA: Diphenyl phosphoryl azide, DIAD:
Diisopropyl azodicarboxylate.
4.4. General procedure Cdsynthesis of sulfonamides
The alkylated amino alcohol (1 equiv) was dissolved in THF
(0.75 M), and Et3N was added (1 equiv). Then, the solution was
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P. Fabbrizzi et al. / Tetrahedron xxx (2014) 1e11
5
cooled to 0 ꢀC and the corresponding sulfonyl chloride (1 equiv)
was added portionwise. The mixture was stirred for 4 h at 0 ꢀC, then
water was added (0.4 mL for each mmol of sulfonyl chloride) and
2 N HCl was added until pHz2 was reached. The solution was
extracted twice with ethyl acetate (5 mL for each mmol of sulfonyl
chloride), the organic phase was dried over Na2SO4 and the solvent
evaporated under vacuum. Flash column chromatography afforded
the desired products.
(m),1333 (m),1075 (w),1021 (s) cmꢂ1. Anal. Calcd For C18H24N2O5S:
C, 56.82; H, 6.36; N, 7.36. Found C, 56.67; H, 6.30; N, 7.32.
4.4.5. Ethyl N-(2-hydroxyethyl)-N-[(4-methylphenyl)sulfonyl]-b-ala-
ninate (44). Obtained from general procedure C, starting from 37
and tosyl chloride. Purified via FCC using Biotage Isolera system
(TLC to gradient, TLC eluant P. E.eEtOAc 1:1, Rf¼0.39). 67% Yield,
colorless oil. 1H NMR (300 MHz, CDCl3)
d
7.70 (d, J¼8.4 Hz, 2H), 7.32
(d, J¼8.4 Hz, 2H), 4.14 (q, J¼7.2 Hz, 2H), 3.81e3.74 (m, 2H), 3.43 (t,
J¼7.2 Hz, 2H), 3.23 (t, J¼5.1 Hz, 2H), 2.71 (t, J¼7.2 Hz, 2H), 2.43 (s,
4.4.1. Ethyl N-(2-hydroxyethyl)-N-[(4-methylphenyl)sulfonyl]glyci-
nate (40). Obtained from general procedure C, starting from 34 and
tosyl chloride. Purified via FCC using Biotage Isolera system (P.
E.eEtOAc gradient). 56% yield, colorless oil. 1H NMR (300 MHz,
3H), 1.26 (t, J¼7.2 Hz, 3H). 13C NMR (100 MHz, CDCl3)
d 171.8, 143.7,
135.5, 129.8, 127.2, 61.4, 60.9, 52.1, 45.9, 34.5, 21.4, 14.1. MS (ESI): m/
z (%) 338 (100) [MþNaþ], 316 (12) [MþHþ]. IR (CHCl3) 3626 (s),
2943 (m), 2835 (m), 2253 (m), 1750 (s), 1604 (m), 1341 (s), 1217 (m),
1159 (s), 1088 (m), 1072 (m), 1022 (s), 814 (m) cmꢂ1. Anal. Calcd For
CDCl3)
d
7.40 (d, J¼8.4 Hz, 2H), 7.32 (d, J¼8.4 Hz, 2H), 4.18 (q,
J¼7.2 Hz, 2H), 4.04 (s, 2H), 3.71 (t, J¼4.8 Hz, 2H), 3.34 (t, J¼4.8 Hz),
2.43 (s, 3H), 1.26 (t, J¼7.2 Hz, 3H). 13C NMR (50 MHz, CDCl3)
d
170.3,
C14H21NO5S: C, 53.32; H, 6.71; N, 4.44. Found C, 53.26; H, 6.67; N,
4.41.
143.8, 136, 130.2, 127, 61.7, 60.5, 51.9, 49.9, 21.4, 13.9. MS (ESI): m/z
(%) 324 [MþNaþ] (100), 318.8 (85), 302 (50) [MþHþ]. IR (CHCl3)
3622 (s), 2943 (m), 2835 (m), 2253 (m), 1750 (s), 1604 (m), 1341 (s),
1217 (m), 1158 (s), 1088 (m), 1072 (m), 1024 (s), 814 (m) cmꢂ1. Anal.
Calcd For C13H19NO5S: C, 51.81; H, 6.35; N, 4.65. Found C, 51.65; H,
6.29; N, 4.62.
4.4.6. Methyl N-(3-hydroxypropyl)-N-[(4-methylphenyl)sulfonyl]gly-
cinate (45). Obtained from general procedure C with addition of
20% methanol to solubilize the product, starting from 35 and tosyl
chloride. Purified via FCC using Biotage Isolera system (TLC to
gradient, TLC eluant P. E.eEtOAc 1:1, Rf¼0.32). >95% Yield, colorless
4.4.2. Ethyl
N-[(4-fluorophenyl)sulfonyl]-N-(2-hydroxyethyl)glyci-
oil. 1H NMR (200 MHz, CDCl3)
d
7.69 (d, J¼8.2 Hz, 2H), 7.28 (d,
nate (41). Obtained from general procedure C, starting from 34 and
4-fluorobenzenesulfonyl chloride. Purified via FCC using Biotage
Isolera system (TLC to gradient, TLC eluant P. E.eEtOAc 1:1,
Rf¼0.36). 43% Yield, white solid. Mp 83.5e84.5 ꢀC. 1H NMR
J¼8.2 Hz), 4.00 (s, 2H), 3.75e3.70 (m, 2H), 3.60 (s, 3H), 3.33 (t,
J¼6.6 Hz, 2H), 2.0 (s, 3H), 1.75e1.68 (m, 2H). 13C NMR (50 MHz,
CDCl3)
d 169.5, 143.7, 136.1, 129.6, 127.3, 58.8, 52.1, 48.4, 45.4, 30.4,
21.5. MS (ESI): m/z (%) 302.1 (100) [MþHþ]. IR (CHCl3) 3623 (s),
2940 (m), 2835 (m), 2253 (m), 1751 (s), 1604 (m), 1341 (s), 1217 (m),
1158 (s), 1090 (m), 1072 (m), 1024 (s), 814 (m) cmꢂ1. Anal. Calcd For
(300 MHz, CDCl3)
d 7.90e7.84 (m, 2H), 7.22e7.15 (m, 2H), 4.17 (q,
J¼7.2 Hz, 2H), 4.08 (s, 2H), 3.72e3.68 (m, 2H), 3.36 (t, J¼4.8 Hz, 2H),
2.99e2.97 (m, 1H), 1.25 (t, J¼7.2 Hz, 3H). 13C NMR (50 MHz, CDCl3)
C13H19NO5S: C, 51.81; H, 6.35; N, 4.65. Found C, 51.64; H, 6.28; N,
d
170.3, 167.8, 135.4, 130.3, 116.5, 116, 61.9, 60.6, 52.2, 49.8, 14. MS
4.62.
(ESI): m/z (%) 328 (100) [MþNaþ], 322.9 (48), 305.9 (18) [MþHþ]. IR
(CHCl3) 3622 (s), 2954 (m), 2911 (m), 2835 (w), 2253 (s), 1744 (m),
1604 (m),1497 (w),1373 (w),1346 (w),1160 (w),1154 (w),1094 (m),
1018 (s), 809 (m) cmꢂ1. Anal. Calcd For C12H16FNO5S: C, 47.21; H,
5.28; N, 4.59. Found C, 47.09; H, 5.22; N, 4.63.
4.4.7. Methyl N-(3-hydroxypropyl)-N-[(4-methylphenyl)sulfonyl]-b-
alaninate (46). Obtained from general procedure C starting from 38
and tosyl chloride. Purified via FCC using Biotage Isolera system
(TLC to gradient, TLC eluant P. E.eEtOAc 1:1, Rf¼0.66). 69% Yield,
colorless oil. 1H NMR (200 MHz, CDCl3)
d
7.69 (d, J¼8.4 Hz, 2H), 7.31
4.4.3. Ethyl N-(biphenyl-4-ylsulfonyl)-N-(2-hydroxyethyl)glycinate
(42). Obtained from general procedure C, starting from 34 and
[1,10-biphenyl]-4-sulfonyl chloride. Purified via FCC using Biotage
Isolera system (TLC to gradient, TLC eluant P. E.eEtOAc 1:1,
Rf¼0.55). 41% Yield, white solid. Mp 92.6e94.5 ꢀC. 1H NMR
(d, J¼8.4 Hz, 2H), 3.78e3.69 (m, 2H), 3.66 (s, 3H), 3.41 (t, J¼7.2 Hz,
2H), 3.24 (t, J¼6.6 Hz, 2H), 2.63 (t, J¼7.2 Hz, 2H), 2.42 (s, 3H),
1.78e1.72 (m, 2H). MS (ESI): m/z (%) 301.6 (96) [MþHþ]. IR (CHCl3)
3623 (s), 2943 (m), 2835 (m), 2253 (m), 1755 (s), 1601 (m), 1344 (s),
1217 (m), 1158 (s), 1088 (m), 1072 (m), 1024 (s), 814 (m) cmꢂ1
.
(300 MHz, CDCl3)
d
7.93e7.89 (m, 2H), 7.74e7.70 (m, 2H), 7.63e7.58
Anal. Calcd for C14H21NO5S: C, 53.32; H, 6.71; N, 4.44. Found C,
53.27; H, 6.68; N, 4.40.
(m, 2H), 7.52e7.45 (m, 3H), 4.23e4.11 (m, 2H), 4.09 (s 2H),
3.76e3.71 (m, 2H), 3.41 (t, J¼4.8 Hz, 2H), 3.11 (t, J¼6.3 Hz, 2H), 1.26
(t, J¼7.2 Hz, 3H). 13C NMR (100 MHz, CDCl3)
d
170.4, 145.7, 137.6,
4.4.8. Methyl N-(4-hydroxybutyl)-N-[(4-methylphenyl)sulfonyl]-b-
132.3,129, 128.5, 127.9, 127.7, 127.3, 61.9, 60.7, 52.4, 50.1, 14. MS
(ESI): m/z (%)386.1 (100) (MþNaþ), 363.1 (14) [Mþ]. IR (CHCl3) 3622
(s), 2943 (m), 2921 (m), 2835, 2253, 1740 (w), 1599 (w), 1500 (w),
1330 (m),1158 (w), 1072 (m), 1018 (s), 809 (m) cmꢂ1. Anal. Calcd For
alaninate (48). Obtained from general procedure C starting from 39
and tosyl chloride. Purified via FCC using Biotage Isolera system
(TLC to gradient, TLC eluant P. E.eEtOAc 1:1, Rf¼0.60). 56% Yield,
colorless oil. 1H NMR (300 MHz, CDCl3)
d
7.69 (d, J¼8.4 Hz, 2H), 7.30
C
18H21NO5S: C, 59.49; H, 5.82; N, 3.85. Found C, 59.25; H, 5.73; N,
(d, J¼8.4 Hz, 2H), 3.67e3.62 (m, 4H), 3.40 (t, J¼7.2 Hz, 2H), 3.15 (t,
J¼7.2 Hz, 2H), 2.65 (t, J¼7.2 Hz, 2H), 2.42 (s, 3H), 1.65e1.56 (m, 4H).
MS (ESI): m/z (%) 330.3 (100) [MþHþ]. IR (CHCl3) 3621 (s), 2943 (m),
2838 (m), 2253 (m), 1751 (s), 1605 (m), 1341 (s), 1218 (m), 1158 (s),
1089 (m), 1072 (m), 1024 (s), 814 (m) cmꢂ1. Anal. Calcd for
3.79.
4.4.4. Ethyl N-{[5-(dimethylamino)-1-naphthyl]sulfonyl}-N-(2-
hydroxyethyl)glycinate (43). Obtained from general procedure C,
starting from 34 and dansyl chloride. Purified via FCC using Biotage
Isolera system (TLC to gradient, TLC eluant P. E.eEtOAc 1:1,
C15H23NO5S: C, 54.69; H, 7.04; N, 4.25. Found C, 54.45; H, 6.98; N,
4.23.
Rf¼0.43). 49% Yield, yellow oil. 1H NMR (300 MHz, CDCl3)
d 8.55 (d,
J¼8.4 Hz, 1H), 8.32 (d, J¼8.4 Hz, 1H), 8.23 (dd, J¼7.5, 1.2 Hz, 1H),
7.59e7.48 (m, 2H), 7.18 (d, J¼7.8 Hz, 1H), 4.16 (s, 2H), 4.11 (q,
J¼7.2 Hz, 2H), 3.70e3.65 (m, 2H), 3.46 (t, J¼5.1 Hz, 2H), 2.89 (s, 6H),
4.4.9. Ethyl N-(4-hydroxybutyl)-N-[(4-methylphenyl)sulfonyl]glyci-
nate (47). Obtained from general procedure C starting from 36 and
tosyl chloride. Purified via FCC using Biotage Isolera system (TLC to
gradient, TLC eluant P. E.eEtOAc 1:1, Rf¼0.34). 90% Yield, colorless
1.19 (t, J¼7.2 Hz, 3H). 13C NMR (50 MHz, CDCl3)
d 170.1, 151.7, 134.7,
130.5, 130, 129.3, 128.2, 123.1, 119.2, 115.2, 61.5, 60.5, 51.6, 49.5, 45.3,
13.8. MS (ESI): m/z (%) 381.1 (100) [MþHþ], 335.1 (34). IR (CHCl3)
3638 (s), 3477 (bm), 2948 (m), 2851 (m), 2247 (m), 1602 (m), 1467
oil. 1H NMR (300 MHz, CDCl3)
d 4.13e4.01 (m, 4H), 3.68e3.62 (m,
2H), 3.27 (d, J¼6.9 Hz, 2H), 2.42 (s, 3H), 1.63e1.59 (m, 4H). MS (ESI):
m/z (%) 352.3 (47) [MþNaþ], 329.3 (100) [MþHþ]. IR (CHCl3) 3627
Please cite this article in press as: Fabbrizzi, P.; et al., Tetrahedron (2014), http://dx.doi.org/10.1016/j.tet.2014.06.125

6
P. Fabbrizzi et al. / Tetrahedron xxx (2014) 1e11
(s), 2943 (m), 2835 (m), 2255 (m), 1751 (s), 1607 (m), 1341 (s), 1218
(m), 1158 (s), 1088 (m), 1072 (m), 1026 (s), 814 (m) cmꢂ1. Anal. Calcd
for C15H23NO5S: C, 54.69; H, 7.04; N, 4.25. Found C, 54.45; H, 6.99;
N, 4.25.
Calcd For C18H23N5O4S: C, 53.32; H, 5.72; N,17.27. Found C, 53.44; H,
5.78; N, 17.14.
4.5.5. Ethyl N-(2-azidoethyl)-N-[(4-methylphenyl)sulfonyl]-b-alani-
nate (2). Obtained from general procedure D, starting from 44. Puri-
fied via FCC using Biotage Isolera system (TLC to gradient, TLC eluant P.
E.eEtOAc 2:1, Rf¼0.51). 83% Yield, pale yellow oil. 1H NMR (300 MHz,
4.5. General procedure Ddconversion to azides (Mitsunobu
reaction)
CDCl3)
d
7.71 (d, J¼8.4 Hz, 2H), 7.32 (d, J¼8.4 Hz, 2H), 4.13 (q, J¼7.2 Hz,
Sulfonamidic adduct (1 equiv) was dissolved in anhydrous THF
(0.1 M) in a microwave vial, and the solution cooled to ꢂ10 ꢀC, then
PPh3 (1.2 equiv), DIAD (1.2 equiv), and DPPA (1.2 equiv) were suc-
cessively added. The mixture was stirred for 10 min at ꢂ10 ꢀC, then
heated under microwave irradiation at 60 ꢀC for 30 min. Solvent
was evaporated under vacuum and the product purified via FCC.
2H), 3.54e3.41 (m, 4H), 2.43 (s, 3H), 1.27 (t, J¼7.2 Hz, 3H). 13C NMR
(100 MHz, CDCl3) d 171, 143.6, 137.7, 129.7, 129.6, 127, 60.4, 50.2, 48.3,
45.2, 34.2, 21.1,13.9. MS (ESI): m/z (%) 363.1 (100) [MþNaþ], 357.8 (92),
340.7 (42) [MþHþ], 290.8 (62), 287.1 (40). IR (CHCl3) 3628 (s), 2958
(m), 2916 (m), 2830 (m), 2247 (m), 1742 (w), 1601 (w), 1338 (m), 1161
(w), 1097 (m), 1015 (s), 811 (m) cmꢂ1. Anal. Calcd For C14H20N4O4S: C,
49.40; H, 5.92; N, 16.46. Found C, 49.51; H, 5.99; N, 16.37.
4.5.1. Ethyl N-(2-azidoethyl)-N-[(4-methylphenyl)sulfonyl]glycinate
(1). Obtained from general procedure D, starting from 40. Purified
via FCC using Biotage Isolera system (TLC to gradient, TLC eluant P.
E.eEtOAc 3:1, Rf¼0.35). 86% Yield, colorless oil. 1H NMR (300 MHz,
4.5.6. Methyl N-(3-azidopropyl)-N-[(4-methylphenyl)sulfonyl]glyci-
nate (3). Obtained from general procedure D, starting from 45. Puri-
fied via FCC using Biotage Isolera system (TLC to gradient, TLC eluant P.
E.eEtOAc 2:1, Rf¼0.47). 88% Yield, pale yellow oil. 1H NMR (200 MHz,
CDCl3)
d
7.72 (d, J¼8.4 Hz, 2H), 7.31 (d, J¼8.4 Hz, 2H), 4.15 (s, 2H), 4.09
(q, J¼7.2 Hz, 2H), 3.56 (t, J¼6 Hz, 2H), 3.39 (t, J¼6 Hz, 2H), 2.43 (s, 3H),
1.21 (t, J¼7.2 Hz, 3H). MS (ESI): m/z (%)365 [MþKþ] (25), 349
[MþNaþ] (100), 343.9 (25), 326.8 (5) [Mþ]. IR (CHCl3) 3628 (s), 2959
(m), 2916 (m), 2830 (m), 2247 (m),1741 (w),1603 (w),1338 (m),1161
(w),1096 (m),1016 (s), 811 (m) cmꢂ1. Anal. Calcd for C13H18N4O4S: C,
47.84; H, 5.56; N, 17.17. Found C, 47.72; H, 5.49; N, 17.08.
CDCl3)
d
7.65 (d, J¼8 Hz, 2H), 7.24 (d, J¼8 Hz, 2H), 3.96 (s, 2H), 3.57 (s,
3H), 3.32 (t, J¼6.4 Hz, 2H), 3.23 (t, J¼6.4 Hz, 2H), 2.35 (s, 3H),1.82e1.65
(m, 2H). MS(ESI):m/z(%)327.4(100)[MþHþ]. IR(CHCl3) 3627 (s), 2960
(m), 2917 (m), 2831 (m), 2247 (m), 1736 (w), 1602 (w), 1338 (m), 1161
(w), 1094 (m), 1016 (s), 811 (m) cmꢂ1. Anal. Calcd For C13H18N4O4S: C,
47.84; H, 5.56; N, 17.17. Found C, 47.99; H, 5.68; N, 17.04.
4.5.2. Ethyl
N-(2-azidoethyl)-N-(biphenyl-4-ylsulfonyl)glycinate
4.5.7. Methyl
N-(3-azidopropyl)-N-[(4-methylphenyl)sulfonyl]-b-
(8). Obtained from general procedure D, starting from 42. Purified
via FCC using Biotage Isolera system (TLC to gradient, TLC eluant P.
E.eEtOAc 3:1, Rf¼0.56). 87% Yield, waxy solid. 1H NMR (300 MHz,
alaninate (4). Obtained from general procedure D, starting from 46.
Purified via FCC using Biotage Isolera system (TLC to gradient, TLC
eluant P. E.eEtOAc 3:1, Rf¼0.47). 90% Yield, pale yellow oil. 1H NMR
CDCl3)
d
7.92e7.88 (m, 2H), 7.75e7.69 (m, 2H), 7.63e7.58 (m, 2H),
(300 MHz, CDCl3)
d
7.69 (d, J¼8.4 Hz, 2H), 7.32 (d, J¼8.4 Hz, 2H),
7.51e7.42 (m, 3H), 4.19 (s, 2H), 4.10 (q, J¼7.2 Hz, 2H), 3.59 (t, J¼6 Hz,
3.68 (s, 3H), 3.43e3.35 (m, 4H), 3.18 (t, J¼7.2 Hz, 2H), 2.64 (t,
J¼7.2 Hz, 2H), 2.43 (s, 3H),1.85e1.79 (m, 2H). MS (ESI): m/z (%) 341.3
(100) [MþHþ]. IR (CHCl3) 3626 (s), 2959 (m), 2916 (m), 2828 (m),
2245 (m), 1743 (w), 1601 (w), 1336 (m), 1161 (w), 1094 (m), 1016 (s),
811 (m) cmꢂ1. Anal. Calcd For C14H20N4O4S: C, 49.40; H, 5.92; N,
16.46. Found C, 49.32; H, 5.89; N, 16.43.
2H), 3.43 (t, J¼6 Hz, 2H), 1.20 (t, J¼7.2 Hz, 2H). 13C NMR (100 MHz,
CDCl3)
d 168.7, 145.9, 139.2, 137.8, 132.3, 129, 128.9, 128.5, 127.9,
127.6,127.3, 61.5, 50.8, 49.7, 47.8,14. MS (ESI): m/z (%) 388.7 [MþHþ]
(95). IR (CHCl3) 3681 (m), 3628 (m), 2948 (m), 2840 (m), 2247 (s),
2097 (s), 1747 (s), 1596 (m), 1478 (w), 1376 (w), 1349 (m), 1215 (w),
1155 (s), 1096 (m), 1016 (s). Anal. Calcd For C18H20N4O4S: C, 55.66;
H, 5.19; N, 14.42. Found C, 55.51; H, 5.13; N, 14.37.
4.5.8. Methyl N-(4-azidobutyl)-N-[(4-methylphenyl)sulfonyl]-b-ala-
ninate (6). Obtained from general procedure D, starting from 47.
Purified via FCC using Biotage Isolera system (TLC to gradient, TLC
eluant P. E.eEtOAc 3:1, Rf¼0.42). 39% Yield, colorless oil. 1H NMR
4.5.3. Ethyl N-(2-azidoethyl)-N-[(4-fluorophenyl)sulfonyl]glycinate
(7). Obtained from general procedure D, starting from 41. Purified
via FCC using Biotage Isolera system (TLC to gradient, TLC eluant P.
E.eEtOAc 3:1, Rf¼0.51). 83% Yield, colorless oil. 1H NMR (300 MHz,
(200 MHz, CDCl3)
d
7.68 (d, J¼8.4 Hz, 2H), 7.32 (d, J¼8.4 Hz, 2H),
3.63 (s, 3H), 3.40e3.25 (m, 4H), 3.20e3.12 (m, 2H), 2.64 (t, J¼6.6 Hz,
2H), 2.39 (s, 3H), 1.71e1.55 (m, 4H). MS (ESI): m/z (%) 355.2 (100)
[MþHþ]. IR (CHCl3) 3628 (s), 2958 (m), 2917 (m), 2831 (m), 2247
CDCl3)
d 7.90e7.84 (m, 2H), 7.23e7.15 (m, 2H), 4.18 (s, 2H), 4.10 (q,
J¼7.2 Hz, 2H), 3.56 (t, J¼5.7 Hz, 2H), 3.40 (t, J¼5.7 Hz, 2H), 1.22 (t,
J¼7.2 Hz, 3H). 13C NMR (50 MHz, CDCl3)
d
168.6, 162.7, 135.5, 130.2,
(m), 1741 (w), 1601 (w), 1335 (m), 1161 (w), 1098 (m), 1016 (s) cmꢂ1
.
116.4, 116, 61.5, 50.7, 49.5, 47.6, 14. MS (ESI): m/z (%) 353.1 (100)
[MþNaþ]. IR (CHCl3) 3611 (s), 2943 (m), 2835 (m), 2253 (m), 2102
(m), 1744 (m), 1644 (m), 1336 (m), 1163 (w), 1153 (w), 1088 (m), 811
(m). Anal. Calcd for C12H15FN4O4S: C, 43.63; H, 4.58; N, 16.96. Found
C, 43.75; H, 4.62; N, 16.83.
Anal. Calcd For C15H22N4O4S: C, 50.83; H, 6.26; N, 15.81. Found C,
51.08.72; H, 6.32; N, 15.72.
4.5.9. Ethyl N-(4-azidobutyl)-N-[(4-methylphenyl)sulfonyl]glycinate
(5). Obtained from general procedure D, starting from 47. Purified via
FCC using Biotage Isolera system (TLC to gradient, TLC eluant P.
E.eEtOAc 3:1, Rf¼0.61). 79% Yield, colorless oil. 1H NMR (200 MHz,
4.5.4. Ethyl
N-(2-azidoethyl)-N-{[5-(dimethylamino)-1-naphthyl]
sulfonyl}glycinate (30). Obtained from general procedure D, start-
ing from 43. Purified via FCC using Biotage Isolera system (TLC to
gradient, TLC eluant P. E.eEtOAc 4:1, Rf¼0.29). 67% Yield, yellow oil.
CDCl3)
d
7.70 (d, J¼8.4 Hz, 2H), 7.28 (d, J¼8.4 Hz, 2H), 4.09e3.95 (m,
4H), 3.33e3.17 (m, 4H), 2.40(s, 3H),1.67e1.51 (m, 4H), 1.18 (t, J¼7.2 Hz,
3H). MS (ESI): m/z (%) 354.8 (100) [MþHþ]. IR (CHCl3) 3628 (s), 2957
(m), 2914 (m), 2830 (m), 2247 (m),1739 (w),1601 (w),1336 (m),1159
(w), 1096 (m), 1013 (s), 809 (m) cmꢂ1. Anal. Calcd For C15H22N4O4S: C,
50.83; H, 6.26; N, 15.81. Found C, 50.96; H, 6.31; N, 15.64.
1H NMR (300 MHz, CDCl3)
d
8.55 (d, J¼8.7 Hz,1), 8.30e8.24 (m, 2H),
7.59e7.48 (m, 2H), 7.18 (d, J¼7.8 Hz, 1H), 4.25 (s, 2H), 3.99 (q,
J¼7.2 Hz, 2H), 3.53e3.47 (m, 2H), 2.87 (s, 6H), 1.10 (t, J¼7.2 Hz, 3H).
13C NMR (50 MHz, CDCl3)
d 168.7, 151.9, 134.9, 130.7, 130.1, 129.5,
128.3, 123.2, 119.2, 115.3, 61.2, 50.4, 49.3, 47.5, 45.3, 14.2, 13.8. MS
(ESI): m/z (%) 428 (96) [MþNaþ], 406.2 (100) [MþHþ]. IR (CHCl3)
3622 (s), 2943 (m), 2835 (m), 2253 (m), 2102 (m), 1750 (m), 1604
(m), 1459 (m), 1330, 1206 (w), 1158 (m), 1142 (m), 1013 (s). Anal.
4.6. General procedure EdCuAAC
Azide (1.1 equiv) was dissolved in THF (0.15 M solution). Alkyne
(1 equiv) and CuP(OEt)3I (0.11 equiv) were added. The reaction was
Please cite this article in press as: Fabbrizzi, P.; et al., Tetrahedron (2014), http://dx.doi.org/10.1016/j.tet.2014.06.125

P. Fabbrizzi et al. / Tetrahedron xxx (2014) 1e11
7
performed under microwave irradiation (temperature 120 ꢀC, time
1 h). THF was evaporated, the crude dissolved in AcOEt or DCM,
washed twice with concentrated ammonia solution (40 mL for each
mmol of CuP(OEt)3I) and brine. The organic layer was dried over
Na2SO4, evaporated under vacuum and purified via flash column
chromatography.
d
7.85e7.78 (m, 2H), 7.60 (s,1H), 7.22e7.13 (m, 2H), 4.60 (t, J¼6.3 Hz,
2H), 4.06e3.93 (m, 4H), 3.86 (s, 2H), 3.75 (t, J¼6.3 Hz, 2H), 2.76 (t,
J¼7.5 Hz, 2H), 2.01e1.96 (m, 2H), 1.51 (s, 9H), 1.17 (t, J¼7.2 Hz, 3H).
13C NMR (50 MHz, CDCl3)
d 168.3, 167.8, 163.8, 162.7, 160.7, 154.9,
147.6, 134.9, 130.2, 130, 122.3, 116.5, 116, 83.8, 78.7, 61.5, 49.6, 49.4,
48.7, 44.1, 28.3, 28.1, 27.9, 22.9, 13.9. MS (ESI): m/z (%) 677.9 (22)
[MþNaþ], 655.9 (100) [MþHþ]. IR (CHCl3) 3682 (m), 3617 (m), 3044
(s), 2980 (m), 2942 (m), 2836 (w), 2257 (m),1722 (m),1603 (s), 1421
(m), 1367 (m), 1329 (m), 1153 (s), 1021 (s) cmꢂ1. Anal. Calcd for
4.6.1. Ethyl N-(2-{4-[3-({[(tert-butoxycarbonyl)amino][(tert-butox-
ycarbonyl)imino]methyl}amino)propyl]-1H-1,2,3-triazol-1-yl}ethyl)-
N-[(4-methylphenyl)sulfonyl]glycinate (10). Obtained from general
procedure E starting from 1 and 28 via FCC using Biotage Isolera
system (TLC to gradient, TLC eluant P. E.eEtOAc 1:2, Rf¼0.38). 84%
C
28H42FN7O8S: C, 51.29; H, 6.46; N, 14.95. Found C, 51.36; H, 6.48; N,
14.90.
Yield, pale yellow oil. 1H NMR (200 MHz, CDCl3)
d
7.62 (d,
4.6.5. Ethyl N-(biphenyl-4-ylsulfonyl)-N-(2-{4-[3-({(Z)-[(tert-butox-
ycarbonyl)amino][(tert-butoxycarbonyl)imino]methyl}amino)pro-
pyl]-1H-1,2,3-triazol-1-yl}ethyl)glycinate
J¼8.4 Hz, 2H), 7.57 (s, 1H), 7.29 (d, J¼8.4 Hz, 2H), 4.61 (t, J¼6.2 Hz,
2H), 4.03e3.92 (m, 4H), 2.05e1.82 (m, 2H), 1.50 (s, 18H), 1.15 (t,
(17). Obtained
from
J¼7.2 Hz, 3H). 13C NMR (50 MHz, CDCl3)
d
168.4, 163.7, 160.6, 154.9,
general procedure E starting from 8 and 28. Purified via FCC using
Biotage Isolera system (TLC to gradient, TLC eluant P. E.eEtOAc 1:1,
Rf¼0.29). 55% Yield, colorless oil. 1H NMR (300 MHz, CDCl3)
143.9, 135.6, 129.7, 127.3, 83.8, 78.7, 61.4, 49.8, 49.6, 48.9, 44.1, 28.3,
28.1, 28, 23, 21.5, 21, 14. MS (ESI): m/z (%)673.9 [MþNaþ] (15),
651.9 (100) [Mþ]. IR (CHCl3) 3686 (m), 3619 (m), 3046 (s), 2979
(m), 2939 (m), 2826 (w), 2257 (m), 1724 (m), 1601 (s), 1424 (m),
1367 (m), 1329 (m), 1152 (s), 1020 (s) cmꢂ1. Anal. Calcd For
d
7.88e7.83 (m, 2H), 7.73e7.68 (m, 2H), 7.61e7.56 (m, 3H),
7.50e7.40 (m, 3H), 4.63 (t, J¼6.3 Hz, 2H), 4.11 (q, J¼7.2 Hz, 2H), 3.88
(s, 2H), 3.79 (t, J¼6.3 Hz, 2H), 2.75 (t, J¼7.5 Hz, 2H), 1.99e1.94 (m,
2H), 1.51 (s, 18H), 1.14 (t, J¼7.2 Hz, 2H). 13C NMR (100 MHz, CDCl3)
C
29H45N7O8S: C, 53.44; H, 6.96; N, 15.04. Found C, 53.32; H, 6.89;
N, 14.98.
d 168.4, 163.8, 160.7, 155, 147.6, 146, 139, 137.2, 129, 128.6, 127.9,
127.7, 127.3, 122.4, 83.8, 78.8, 61.5, 44.1, 30.9, 28.4, 28, 23, 14. MS
(ESI): m/z (%) 736.1 (100) [MþNaþ], 714.2 (94) [MþHþ]. IR (CHCl3)
3940 (m), 3757 (w), 3692 (m), 3056 (s), 2981 (s), 2679 (m), 2528
(w), 2398 (w), 2301 (m), 2247 (m), 2150 (w), 2053 (w), 1790 (w),
1602 (w), 1548 (w),1446 (m),1424 (s),1370 (m),1263 (vs),1155 (m),
1096 (w), 1010 (w) cmꢂ1. Anal. Calcd For C34H47N7O8S: C, 57.21; H,
6.64; N, 13.74. Found C, 57.43; H, 6.71; N, 13.64.
4.6.2. Ethyl N-(2-{4-[2-({[(tert-butoxycarbonyl)amino][(tert-butox-
ycarbonyl)imino]methyl}amino)ethyl]-1H-1,2,3-triazol-1-yl}ethyl)-
N-[(4-methylphenyl)sulfonyl]glycinate (9). Obtained from general
procedure E starting from 1 and 27. Purified via FCC using Biotage
Isolera system (TLC to gradient, TLC eluant P. E.eEtOAc 1:1,
Rf¼0.25). 86% Yield, pale yellow oil. 1H NMR (300 MHz, CDCl3)
d
7.68 (d, J¼2.1 Hz, 2H), 7.54 (s, 1H), 7.30 (d, J¼2.1 Hz, 2H), 4.53 (t,
J¼6.4 Hz, 2H), 4.25e4.08 (m, 4H), 3.93 (s, 3H), 3.70 (d, J¼6.4 Hz, 2H),
3.09e2.95 (m, 2H), 2.43 (s, 3H), 1.60e1.43 (m,18H), 1.28 (t, J¼8.4 Hz,
4.6.6. Ethyl N-[2-(4-{3-[(tert-butoxycarbonyl)(pyridin-2-yl)amino]
propyl}-1H-1,2,3-triazol-1-yl)ethyl]-N-{[5-(dimethylamino)-1-
naphthyl]sulfonyl}glycinate (34). Obtained from general pro-
cedure E starting from 30 and 29. Purified via FCC using Biotage
Isolera system (TLC to gradient, TLC eluant P. E.eEtOAc 1:2,
3H). 13C NMR (100 MHz, CDCl3)
d 168.5, 160.3, 154.8, 144.1, 135.6,
129.7, 127.4, 127.4, 84, 78.8, 77.3, 77, 76.7, 61.5, 50, 49.7, 49, 44.3,
28.3, 28.3, 27.9, 25.3, 21.5, 13.9. MS (ESI): m/z (%) 660 (46) [MþNaþ],
638.1 (100) [MþHþ]. IR (CHCl3) 3684 (m), 3619 (m), 3045 (s), 2980
(m), 2942 (m), 2838 (w), 2255 (m), 1724 (m), 1599 (s), 1424 (m),
1367 (m), 1329 (m), 1152 (s), 1020 (s) cmꢂ1. Anal. Calcd For
Rf¼0.30). 54% Yield, yellow oil. 1H NMR (300 MHz, CDCl3)
d 8.54
(d, J¼8.4 Hz, 1H), 8.39e8.35 (m, 1H), 8.26e8.19 (m, 2H),
7.66e7.46 (m, 4H), 7.21 (s, 1H), 7.16 (d, J¼7.5 Hz, 1H), 7.02e6.97
(m, 1H), 4.50 (t, J¼6.3 Hz, 2H), 3.99 (t, J¼7.2 Hz, 2H), 3.95e3.80
(m, 6H), 2.85 (s, 6H), 2.68 (t, J¼7.8 Hz, 2H), 2.01e1.94 (m, 2H),
1.49 (s, 18H), 1.03 (t, J¼7.2 Hz, 3H). 13C NMR (50 MHz, CDCl3)
C
28H43N7O8S: C, 52.73; H, 6.80; N, 15.37. Found C, 52.84; H, 6.85; N,
15.35.
4.6.3. Ethyl N-[2-(4-{3-[(tert-butoxycarbonyl)(pyridin-2-yl)amino]
propyl}-1H-1,2,3-triazol-1-yl)ethyl]-N-[(4-methylphenyl)sulfonyl]
glycinate (33). Obtained from general procedure E starting from 1
and 29. Pale yellow oil >95% Yield. FCC using Biotage Isolera
system (TLC to gradient, TLC eluant P. E.eEtOAc 1:2, Rf¼0.26). 1H
d 168.4, 154.5, 154.1, 151.9, 147.5, 136.8, 134.4, 130.8, 130, 129.6,
128.3, 123.1, 121.9, 119.9, 119.5, 119, 115.3, 81, 61.3, 49.6, 49.1,
48.7, 46.2, 45.3, 28.4, 28.2, 23, 13.8. MS (ESI): m/z (%) 688.1 (100)
[MþNaþ], 666 (56) [MþHþ]. IR (CHCl3) 3687 (m), 3617 (m), 3045
(s), 2982 (s), 2942 (m), 2838 (w), 2254 (m), 1724 (m), 1605 (s),
1424 (m), 1368 (m), 1329 (m), 1153 (s), 1021 (s) cmꢂ1. Anal. Calcd
For C33H43N7O6S: C, 59.53; H, 6.51; N, 14.73. Found C, 59.46; H,
6.48; N, 14.68.
NMR (200 MHz, CDCl3)
d
8.36 (d, J¼5.6 Hz, 1H), 7.71e7.58 (m, 4H),
7.41 (s, 1H), 7.27 (d, J¼8.2 Hz, 2H), 7.05e6.84 (m, 1H), 4.58 (t,
J¼6.2 Hz, 2H), 4.11e3.90 (m, 4H), 3.78 (s, 2H), 3.67 (t, J¼6.2 Hz,
2H), 2.73 (t, J¼7.8 Hz, 2H), 2.39 (s, 3H), 2.10e1.92 (m, 2H), 1.50 (s,
9H), 1.12 (t, J¼7.1 Hz, 3H). 13C NMR (50 MHz, CDCl3)
d
168.5, 154.6,
4.6.7. Ethyl N-(2-{4-[3-({[(tert-butoxycarbonyl)amino][(tert-butox-
ycarbonyl)imino]methyl}amino)propyl]-1H-1,2,3-triazol-1-yl}ethyl)-
N-[(4-methylphenyl)sulfonyl]-b-alaninate (11). Obtained from gen-
154.1, 147.6, 144, 136.9, 135.7, 129.7, 127.4, 120.1, 119.6, 81, 67.1, 61.4,
50, 49.7, 49, 46.3, 28.3, 23.1, 21.5, 13.9. MS (ESI): m/z (%) 609 (100)
[MþNaþ], 587 (28) [MþHþ]. IR (CHCl3) 3685 (m), 3619 (m), 3044
(s), 2980 (m), 2942 (m), 2838 (w), 2257 (m), 1724 (m), 1600 (s),
1422 (m), 1367 (m), 1329 (m), 1149 (s), 1020 (s) cmꢂ1. Anal. Calcd
for C28H38N6O6S: C, 57.32; H, 6.53; N, 14.32. Found C, 57.38; H,
6.56; N, 14.30.
eral procedure E starting from 2 and 28. Purified via FCC using
Biotage Isolera system (TLC to gradient, TLC eluant P. E.eEtOAc 1:1,
Rf¼0.26). 83% Yield, colorless oil. 1H NMR (200 MHz, CDCl3)
d 7.66
(d, J¼8.4 Hz, 2H), 7.31 (d, J¼8.4 Hz, 2H), 4.57 (t, J¼6.6 Hz, 2H),
4.13e3.92 (m, 4H), 3.56 (t, J¼6.6 Hz, 2H), 3.29 (t, J¼7.2 Hz, 2H),
2.74 (t, J¼7.8 Hz, 2H), 2.42e2.32 (m, 5H), 2.01e1.92 (m, 2H), 1.49 (s,
4.6.4. Ethyl N-(2-{4-[3-({[(tert-butoxycarbonyl)amino][(tert-butox-
ycarbonyl)imino]methyl}amino)propyl]-1H-1,2,3-triazol-1-yl}ethyl)-
N-[(4-fluorophenyl)sulfonyl]glycinate (16). Obtained from general
procedure E starting from 7 and 28. Purified via FCC using Biotage
Isolera system (TLC to gradient, TLC eluant P. E.eEtOAc 1:1,
Rf¼0.22). 70% Yield, colorless oil. 1H NMR (300 MHz, CDCl3)
18H), 1.27e1.17 (m, 3H). 13C NMR (100 MHz, CDCl3)
d 171, 163.8,
160.7, 155, 144, 135.2, 129.9, 127.3, 83.8, 78.7, 77.4, 77.1, 76.7, 60.8,
49.6, 49.5, 45.9, 44.1, 33.9, 28.3, 28.2, 28, 23, 21.5, 14.1. MS (ESI): m/
z (%) 688 (24) [MþNaþ], 665.9 (100) [MþHþ]. IR (CHCl3) 3684 (m),
3618 (m), 3047 (s), 2980 (m), 2935 (m), 2838 (w), 2257 (m), 1723
(m), 1598 (s), 1424 (m), 1367 (m), 1329 (m), 1153 (s), 1021 (s) cmꢂ1
.
Please cite this article in press as: Fabbrizzi, P.; et al., Tetrahedron (2014), http://dx.doi.org/10.1016/j.tet.2014.06.125

8
P. Fabbrizzi et al. / Tetrahedron xxx (2014) 1e11
Anal. Calcd For C30H47N7O8S: C, 54.12; H, 7.12; N, 14.73. Found C,
54.21; H, 7.20; N, 14.52.
136.4, 129.5, 127.3, 83.8, 78.6, 61.3, 49.3, 48.2, 47.6, 44.1, 28.3, 27.9,
27.1, 24.9, 23, 21.4, 13.9. MS (ESI): m/z (%) 680.2 (100) [MþHþ]. IR
(CHCl3) 3689 (m), 3621 (m), 3046 (s), 2980 (m), 2942 (m), 2838 (w),
2256 (m), 1726 (m), 1600 (s), 1424 (m), 1366 (m), 1325 (m), 1153 (s),
1025 (s) cmꢂ1. Anal. Calcd for C31H49N7O8S: C, 54.77; H, 7.26; N,
14.42. Found C, 54.72; H, 7.24; N, 14.39.
4.6.8. Methyl N-(3-{4-[3-({[(tert-butoxycarbonyl)amino][(tert-bu-
toxycarbonyl)imino]methyl}amino)propyl]-1H-1,2,3-triazol-1-yl}pro-
pyl)-N-[(4-methylphenyl)sulfonyl]glycinate (12). Obtained from
general procedure E starting from 3 and 28. Purified via FCC using
Biotage Isolera system (TLC to gradient, TLC eluant P. E.eEtOAc 1:2,
Rf¼0.38). 26% Yield, colorless oil. 1H NMR (300 MHz, CDCl3)
4.7. General procedure Fddeprotection
d
7.70e7.62 (m, 3H), 7.58 (d, J¼8.4 Hz, 2H), 4.46 (t, J¼6.9 Hz, 2H),
The protected adduct was suspended in 3 N HCl (10 mL/mmol)
and stirred at 30 ꢀC overnight. The solution was evaporated to
dryness in vacuo to afford the pure product in quantitative yield.
4.00e3.64 (m, 4H), 3.63 (s, 2H), 3.20 (t, J¼6.6 Hz, 2H), 2.77 (t,
J¼7.2 Hz, 2H), 2.41 (s, 3H), 2.20 (t, J¼6.6 Hz, 2H), 2.00e1.55 (m, 2H),
1.49 (s, 18H). 13C NMR (50 MHz, CDCl3)
d 169.3, 154.9, 143.8, 135.8,
129.7, 127.3, 127, 83.8, 52.2, 49.1, 47.1, 46.4, 44.1, 29, 28.3, 27.9, 27.8,
23, 21.5. MS (ESI): m/z (%) 651.9 (100) [Mþ]. IR (CHCl3) 3688 (m),
3615 (m), 3044 (s), 2979 (m), 2939 (m), 2831 (w), 2254 (m), 1721
4.7.1. N-{2-[4-(3-{[Amino(imino)methyl]amino}propyl)-1H-1,2,3-
triazol-1-yl]ethyl}-N-[(4-methylphenyl)sulfonyl]glycine monohydro-
chloride (19). Obtained from general procedure F starting from 10.
(m), 1598 (s), 1422 (m), 1361 (m), 1322 (m), 1156 (s), 1022 (s) cmꢂ1
.
Pale yellow solid. 1H NMR (300 MHz, D2O)
d 7.67 (s, 1H), 7.23 (d,
Anal. Calcd For C29H45N7O8S: C, 53.44; H, 6.96; N, 15.04. Found C,
53.48; H, 6.97; N, 15.02.
J¼8.4 Hz, 2H), 7.21 (d, J¼8.4 Hz, 2H), 4.43 (t, J¼3.4 Hz, 2H), 4.00 (s,
2H), 3.67 (d, J¼3.4 Hz, 2H), 3.04 (d, J¼7.2 Hz, 2H), 2.57 (d, J¼7.2 Hz,
2H), 2.27 (s, 3H), 1.82e1.71 (m, 2H). 13C NMR (100 MHz, D2O)
4.6.9. Methyl N-(3-{4-[3-({[(tert-butoxycarbonyl)amino][(tert-bu-
toxycarbonyl)imino]methyl}amino)propyl]-1H-1,2,3-triazol-1-yl}pro-
pyl)-N-[(4-methylphenyl)sulfonyl]-b-alaninate (13). Obtained from
d 172.4,156.7, 145.3, 134.2,129.9,126.8, 48.7, 40, 30.2, 27.1, 21.1, 20.6.
MS (ESI): m/z (%) 424.1 (100) [Mþ]; 410 (28). IR (KBr) 3396 (br s),
3191 (s), 2964 (m), 2921 (m), 2846 (w), 1744 (m), 1733 (m), 1663 (s),
1647 (s), 1626 (m), 1465 (m), 1448 (m), 1379 (m), 1330 (s), 1293 (m),
1260 (m), 1158 (s), 1088 (m), 1045 (m), 932 (w), 814 (m), 803 (m),
663 (m), 572 (m), 551 (m) cmꢂ1. Anal. Calcd For C17H26ClN7O4S: C,
44.39; H, 5.70; N, 21.32. Found C, 44.30; H, 5.74; N, 21.29.
general procedure E starting from 4 and 28. Purified via FCC using
Biotage Isolera system (TLC to gradient, TLC eluant P. E.eEtOAc 1:2,
Rf¼0.58). 47% Yield, colorless oil. 1H NMR (200 MHz, CDCl3)
d
7.68e7.55 (m, 2H), 7.28 (d, J¼8.2 Hz, 2H), 4.37 (t, J¼6.6 Hz, 2H),
3.94 (t, J¼6.6 Hz, 2H), 3.64 (s, 3H), 3.35 (d, J¼7.2 Hz, 2H), 3.10 (t,
J¼6.6 Hz, 2H), 2.75 (t, J¼7.2 Hz, 2H), 2.59 (t, J¼7.8 Hz, 2H), 2.40 (s,
3H), 2.27e2.12 (m, 2H), 2.03e1.88 (m, 2H), 1.48 (s, 18H). 13C NMR
4.7.2. N-{2-[4-(2-{[Amino(imino)methyl]amino}ethyl)-1H-1,2,3-
triazol-1-yl]ethyl}-N-[(4-methylphenyl)sulfonyl]glycine monohydro-
chloride (18). Obtained from general procedure F starting from 9.
(50 MHz, CDCl3)
d 171.6, 163.8, 160.7, 155, 147.5, 144, 135.4, 129.8,
127.1, 126.9, 83.8, 78.7, 51.8, 47.2, 46.8, 44.9, 44.1, 34.2, 29.5, 28.3,
28.0, 27.9, 22.9, 21.4. MS (ESI): m/z (%) 665.4 (100) [Mþ]. IR (CHCl3)
3686 (m), 3619 (m), 3044 (s), 2982 (m), 2941 (m), 2837 (w), 2255
(m), 1721 (m), 1601 (s), 1424 (m), 1366 (m), 1329 (m), 1151 (s), 1019
(s) cmꢂ1. Anal. Calcd For C30H47N7O8S: C, 54.12; H, 7.12; N, 14.73.
Found C, 54.09; H, 7.11; N, 14.74.
Pale yellow solid. 1H NMR (300 MHz, D2O)
d 7.70 (s, 1H), 7.44 (d,
J¼6.9 Hz, 2H), 7.24 (d, J¼6.9 Hz, 2H), 4.44 (t, J¼5.1 Hz, 2H), 3.98 (s,
3H), 3.75e3.62 (m, 2H), 3.33 (d, J¼6.3 Hz, 2H), 2.79 (d, J¼6.3 Hz,
2H), 2.28 (s, 3H). 13C NMR (100 MHz, D2O)
d 172.4, 156.7, 145.3,
134.2, 129.9, 129.9, 126.8, 125.0, 48.7, 48.5, 48.3, 40.4, 24.1, 20.6. MS
(ESI): m/z (%) 410.2 (100) [Mþ], 303.7 (26). IR (KBr) 3396 (br s), 3191
(s), 2964 (m), 2922 (m), 2846 (w), 1744 (m), 1733 (m), 1663 (s), 1647
(s), 1626 (m), 1465 (m), 1448 (m), 1379 (m), 1330 (s), 1293 (m), 1260
(m), 1158 (s), 1081 (m), 1039 (m), 932 (w), 814 (m), 803 (m), 663
(m), 572 (m), 551 (m) cmꢂ1. Anal. Calcd for C16H24ClN7O4S: C, 43.10;
H, 5.42; N, 21.99. Found C, 43.05; H, 5.48; N, 21.95.
4.6.10. Methyl N-(4-{4-[3-({[(tert-butoxycarbonyl)amino][(tert-bu-
toxycarbonyl)imino]methyl}amino)propyl]-1H-1,2,3-triazol-1-yl}bu-
tyl)-N-[(4-methylphenyl)sulfonyl]-b-alaninate (15). Obtained from
general procedure E starting from 6 and 28. Purified via FCC using
Biotage Isolera system (TLC to gradient, TLC eluant P. E.eEtOAc 1:2,
Rf¼0.29). 51% Yield, colorless oil. 1H NMR (200 MHz, CDCl3)
d
7.66
4.7.3. N-[(4-Methylphenyl)sulfonyl]-N-(2-{4-[3-(pyridinium-2-
(d, J¼8 Hz, 2H), 7.56 (s, 1H), 7.30 (d, J¼8 Hz, 2H), 4.34 (t, J¼6.6 Hz,
2H), 3.95 (t, J¼7.2 Hz, 2H), 3.64 (s, 2H), 3.34 (t, J¼7.2 Hz, 2H), 3.12 (t,
J¼7.2 Hz, 2H), 2.75 (t, J¼7.2 Hz, 2H), 2.41 (s, 3H), 2.06e1.89 (m, 4H),
ylamino)propyl]-1H-1,2,3-triazol-1-yl}ethyl)glycine
(32). Obtained from general procedure F starting from 33. Pale
yellow solid. 1H NMR (200 MHz, D2O)
7.81 (s, 1H), 7.72e7.61 (m,
chloride
d
1.55e1.40 (m, 20H). 13C NMR (50 MHz, CDCl3)
d
171.6, 160.7, 155,
1H), 7.55 (d, J¼7 Hz, 1H), 7.39 (d, J¼7.8 Hz, 2H), 7.21 (d, J¼7.8 Hz,
2H), 6.81 (d, J¼9.2 Hz, 1H), 6.66 (t, J¼7 Hz, 1H), 4.52e4.43 (m, 2H),
4.00 (s, 2H), 3.72e3.65 (m, 2H), 3.22 (t, J¼7 Hz, 2H), 2.70 (t, J¼7 Hz,
147.6, 143.5, 136.1, 129.8, 127.1, 121.1, 83.8, 78.7, 51.8, 49.2, 48.4, 44.3,
44.1, 34.3, 28.4, 28, 27.2, 25.4, 23, 21.5. MS (ESI): m/z (%) 702.4 (100)
[MþNaþ], 680.2 (100) [MþHþ]. IR (CHCl3) 3687 (m), 3621 (m), 3047
(s), 2980 (m), 2943 (m), 2839 (w), 2254 (m),1724 (m),1605 (s),1424
(m), 1365 (m), 1323 (m), 1153 (s), 1021 (s) cmꢂ1. Anal. Calcd For
2H), 2.26 (s, 3H), 2.05e1.96 (m, 2H). 13C NMR (50 MHz, D2O)
d 152.4,
145.3, 143.5, 134.5, 134.1, 129.9, 126.8, 113, 112.4, 49.9, 48.9, 40.5,
26.4, 20.8, 20.7. MS (ESI): m/z (%) 459.2 (100) [Mþ]. IR (KBr) 3415 (br
s), 3191 (s), 2964 (m), 2921 (m), 2846 (w), 1744 (m), 1733 (m), 1662
(s), 1647 (s), 1625 (m), 1465 (m), 1448 (m), 1379 (m), 1330 (s), 1293
(m), 1260 (m), 1157 (s), 1088 (m), 1045 (m), 932 (w), 814 (m), 803
(m), 793 (m), 771 (m), 668 (w), 647 (w), 665 (m), 588 (m) 572 (m),
551 (m) cmꢂ1. Anal. Calcd For C21H27ClN6O4S: C, 50.96; H, 5.50; N,
16.98. Found C, 50.89; H, 5.55; N, 16.94.
C
31H49N7O8S: C, 54.77; H, 7.26; N, 14.42. Found C, 54.82; H, 7.23; N,
14.38.
4.6.11. Ethyl N-(4-{4-[3-({[(tert-butoxycarbonyl)amino][(tert-butox-
ycarbonyl)imino]methyl}amino)propyl]-1H-1,2,3-triazol-1-yl}butyl)-
N-[(4-methylphenyl)sulfonyl]glycinate (14). Obtained from general
procedure E starting from 6 and 28. Purified via FCC using Biotage
Isolera system (TLC to gradient, TLC eluant P. E.eEtOAc 1:2,
4.7.4. N-{2-[4-(3-{[Amino(imino)methyl]amino}propyl)-1H-1,2,3-
triazol-1-yl]ethyl}-N-[(4-fluorophenyl)sulfonyl]glycine monohydro-
chloride (25). Obtained from general procedure F starting from 16.
Rf¼0.47). 73% Yield, colorless oil. 1H NMR (300 MHz, CDCl3)
d 7.64
(d, J¼8.1 Hz, 2H), 7.52 (s, 1H), 7.24 (d, J¼8.1 Hz, 2H), 4.29 (t, J¼6.9 Hz,
2H), 4.30e3.88 (m, 6H), 3.20 (t, J¼6.9 Hz, 2H), 2.71 (t, J¼7.2 Hz, 2H),
2.36 (s, 3H), 1.96e1.88 (m, 4H), 1.53e1.44 (m, 20H), 1.13 (t, J¼7.2 Hz,
Pale yellow solid. 1H NMR (300 MHz, D2O)
d 7.85 (s, 1H), 7.66e7.60
(m, 2H), 7.19e7.11 (m, 2H), 4.51 (t, J¼5.4 Hz, 2H), 4.05 (s, 2H),
3H). 13C NMR (50 MHz, CDCl3)
d 168.7, 163.8, 160.7, 155, 144.3, 143.5,
3.73e3.68 (m, 2H), 3.08 (t, J¼6.9 Hz, 2H), 2.66 (t, J¼7.5 Hz, 2H),
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P. Fabbrizzi et al. / Tetrahedron xxx (2014) 1e11
9
1.86e1.75 (m, 2H). 13C NMR (50 MHz, D2O)
d
172.2, 167.9, 162.8,
(38). IR (KBr) 3394 (br s), 3192 (s), 2961 (m), 2921 (m), 2847 (w),
1744 (m), 1733 (m), 1663 (s), 1649 (s), 1626 (m), 1464 (m), 1448 (m),
1379 (m), 1330 (s), 1293 (m), 1260 (m), 1158 (s), 1088 (m), 1045 (m),
932 (w), 814 (m), 803 (m), 663 (m), 572 (m), 551 (m) cmꢂ1. Anal.
Calcd For C18H28ClN7O4S: C, 45.61; H, 5.95; N, 20.69. Found C, 45.30;
H, 5.98; N, 20.65.
156.7, 144.5, 133.5, 130, 129.8, 126.3, 116.9, 116.4, 49.8, 48.7, 40, 26.9,
20.6. MS (ESI): m/z (%) 428.1 (100) [Mþ]. IR (KBr) 3396 (br s), 3192
(s), 2964 (m), 2921 (m), 2846 (w), 1743 (m), 1733 (m), 1663 (s), 1647
(s), 1616 (m), 1465 (m), 1448 (m), 1379 (m), 1330 (s), 1293 (m), 1260
(m), 1158 (s), 1088 (m), 1045 (m), 932 (w), 814 (m), 803 (m), 663
(m), 572 (m), 551 (m) cmꢂ1. Anal. Calcd for C16H23ClN7O4S: C, 41.42;
H, 5.00; N, 21.13. Found C, 41.32; H, 5.11; N, 21.05.
4.7.9. N-{3-[4-(3-{[Amino(imino)methyl]amino}propyl)-1H-1,2,3-
triazol-1-yl]propyl}-N-[(4-methylphenyl)sulfonyl]-
hydrochloride (22). Obtained from general procedure F starting
from 13. Pale yellow solid. 1H NMR (200 MHz, D2O)
7.87 (s, 1H),
b-alanine mono-
4.7.5. N-{2-[4-(3-{[Amino(imino)methyl]amino}propyl)-1H-1,2,3-
triazol-1-yl]ethyl}-N-(biphenyl-4-ylsulfonyl)glycine
chloride (26). Obtained from general procedure F starting from 17.
White solid. 1H NMR (200 MHz, D2O)
7.50e7.29 (m, 8H), 7.26e7.21
monohydro-
d
7.46 (d, J¼8.4 Hz, 2H), 7.25 (d, J¼8.4 Hz, 2H), 4.34 (t, J¼6.6 Hz, 2H),
3.28 (d, J¼6.6 Hz, 2H), 3.14e2.94 (m, 2H), 2.70 (d, J¼7.4 Hz, 2H), 2.42
(d, J¼6.9 Hz, 2H), 2.25 (s, 3H), 2.06e2.00 (m, 2H), 1.86e1.79. 13C
d
(m, 3H), 4.40e4.30 (m, 2H), 4.01 (s, 2H), 3.70e3.55 (m, 2H), 2.77 (t,
J¼6 Hz, 2H), 2.29 (t, J¼6 Hz, 2H), 1.62e1.49 (m, 2H). 13C NMR
NMR (50 MHz, D2O)
d 174.9, 156.2, 144.8, 133.2, 129.6, 126.4, 126.1,
(50 MHz, D2O)
d
171.1, 155.7, 144.6, 143.8, 136.8, 135.5, 128, 127.7,
48.3, 45.1, 43.7, 39.7, 39, 33.1, 27.2, 26.7, 25.7, 21.8, 20.6, 20.3. MS
(ESI): m/z (%) 452.2 (100) (Mþ). IR (KBr) 3396 (br s), 3191 (s), 2964
(m), 2928 (m), 2841 (w), 1744 (m), 1733 (m), 1664 (s), 1647 (s), 1626
(m),1468 (m),1448 (m),1379 (m),1330 (s), 1293 (m),1259 (m),1147
(s), 1088 (m), 1041 (m), 932 (w), 814 (m), 88 (m), 664 (m), 575 (m),
542 (m) cmꢂ1. Anal. Calcd For C19H30ClN7O4S: C, 46.76; H, 6.20; N,
20.09. Found C, 46.66; H, 6.32; N, 20.01.
126.5, 126.2, 125.8, 124, 48.4, 39.4, 26.3, 20.4. MS (ESI): m/z (%)
509.2 (44) [MþNaþ], 486.2 (100) [Mþ]. IR (KBr) 3396 (br s), 3191 (s),
2966 (m), 2923 (m), 2846 (w), 1741 (m), 1733 (m), 1661 (s), 1646 (s),
1623 (m), 1465 (m), 1448 (m), 1379 (m), 1330 (s), 1291 (m), 1260
(m), 1158 (s), 1083 (m), 1041 (m), 932 (w), 814 (m), 803 (m), 663
(m), 571 (m), 551 (m) cmꢂ1. Anal. Calcd For C22H28ClN7O4S: C,
50.62; H, 5.41; N, 18.78. Found C, 50.55; H, 5.52; N, 18.69.
4.7.10. N-{4-[4-(3-{[Amino(imino)methyl]amino}propyl)-1H-1,2,3-
4.7.6. N-{[5-(Dimethylamino)-1-naphthyl]sulfonyl}-N-(2-{4-[3-(pyr-
triazol-1-yl]butyl}-N-[(4-methylphenyl)sulfonyl]-
hydrochloride (24). Obtained from general procedure F starting
from 15. Pale yellow solid. 1H NMR (300 MHz, D2O)
7.83 (s, 1H),
b
-alanine
mono-
idin-2-ylamino)propyl]-1H-1,2,3-triazol-1-yl}ethyl)glycine
hydrochloride (31). Obtained from general procedure F starting
from 34. Yellow solid. 1H NMR (300 MHz, D2O)
8.44e8.34 (m, 2H),
mono-
d
d
7.44 (d, J¼8.2 Hz, 2H), 7.18 (d, J¼8.2 Hz, 2H), 4.24 (t, J¼6.6 Hz, 2H),
3.19 (t, J¼6.9 Hz, 2H), 3.02 (t, J¼6.9 Hz, 2H), 2.94 (t, J¼7.2 Hz, 2H),
2.18 (s, 3H), 1.79e1.74 (m, 2H), 1.65e1.60 (m, 2H), 1.31e1.22 (m, 2H).
8.18 (d, J¼7.2 Hz, 1H), 8.06 (d, J¼7.2 Hz, 1H), 8.00 (s, 1H), 7.86e7.67
(m, 3H), 7.59 (d, J¼6.6 Hz, 1H), 6.88 (d, J¼9 Hz, 1H), 6.68 (t, J¼6.6 Hz,
1H), 4.65e4.60 (m, 2H), 4.31 (s, 2H), 3.97e3.92 (m, 2H), 3.50 (s, 6H),
13C NMR (50 MHz, D2O)
d 175.4, 145.1, 134.4, 130, 126.8, 124.7, 51,
3.29 (t, J¼6.9 Hz, 2H), 2.71 (t, J¼6.9 Hz, 2H), 2.01e1.94 (m, 2H). 13
C
47.6, 43.8, 40.2, 33.7, 27, 26.1, 24.4, 20.9, 20.6. MS (ESI): m/z (%)
466.2 (100) (Mþ). IR (KBr) 3398 (br s), 3191 (s), 2965 (m), 2919 (m),
2846 (w), 1738 (m), 1736 (m), 1662 (s), 1642 (s), 1621 (m), 14,657
(m), 1449 (m), 1379 (m), 1331 (s), 1297 (m), 1261 (m), 1158 (s), 1089
(m), 1051 (m), 933 (w), 820 (m), 802 (m), 658 (m), 578 (m), 555 (m)
cmꢂ1. Anal. Calcd For C20H32ClN7O4S: C, 47.85; H, 6.42; N, 19.53.
Found C, 47.74; H, 6.54; N, 19.49.
NMR (50 MHz, D2O) d 171.1,152.1,143.3,138, 134.5,134, 130.4,128.5,
128, 126.9, 126.3, 126, 125.3, 119.7, 112.9, 112.2, 50.1, 48.5, 48.2, 46.9,
40.4, 26.2, 20.3. MS (ESI): m/z (%) 538.2 (100) [Mþ]. IR (KBr) 3428
(br s), 3137 (m), 2954 (m), 2921 (m), 2857 (w), 1733 (m), 1653 (s),
1626 (m), 1454 (m), 1379 (m), 1336 (s), 1206 (w), 1169 (w), 1142 (s),
1099 (m), 1045 (m), 1024 (w), 991 (w), 879 (m), 793 (m), 771 (m),
669 (w), 647 (w), 588 (m) cmꢂ1. Anal. Calcd For C26H32ClN7O4S: C,
54.39; H, 5.62; N, 17.08. Found C, 54.32; H, 5.70; N, 17.01.
4.7.11. N-{4-[4-(3-{[Amino(imino)methyl]amino}propyl)-1H-1,2,3-
triazol-1-yl]butyl}-N-[(4-methylphenyl)sulfonyl]glycine monohydro-
chloride (23). Obtained from general procedure F starting from 15.
4.7.7. Ethyl
1,2,3-triazol-1-yl]ethyl}-N-[(4-methylphenyl)sulfonyl]-
monohydrochloride (20). Obtained from general procedure
starting from 11. Pale yellow solid. 1H NMR (300 MHz, D2O)
7.71 (s,
N-{2-[4-(3-{[amino(imino)methyl]amino}propyl)-1H-
b
-alaninate
Pale yellow solid. 1H NMR (300 MHz, D2O)
d 7.87 (s, 1H), 7.52 (d,
F
J¼8.4 Hz, 2H), 7.23 (d, J¼8.4 Hz, 2H), 4.26 (t, J¼6.9 Hz, 2H), 3.87 (s,
2H), 3.11e3.04 (m, 4H), 2.71 (t, J¼7.8 Hz, 2H), 2.24 (s, 3H), 1.85e1.79
(m, 2H), 1.72e1.65 (m, 2H), 1.35e1.28 (m, 2H). 13C NMR (50 MHz,
d
1H), 7.48 (d, J¼7.8 Hz, 2H), 7.28 (d, J¼7.8 Hz, 2H), 4.53e4.49 (m, 2H),
3.60e3.58 (m, 2H), 3.33 (t, J¼6.9 Hz, 2H), 3.06 (t, J¼6.9 Hz, 2H), 2.65
(t, J¼7.5 Hz, 2H), 2.40e2.34 (m, 2H), 2.29 (s, 3H), 1.84e1.67 (m, 2H).
D2O)
d 172.1, 155.8, 144.3, 133.6, 129.1, 126.1, 124.3, 50.6, 47.7, 47.4,
39.4, 26.1, 25.1, 23.1, 19.9. MS (ESI): m/z (%) 452.2 (100) (Mþ). IR
(KBr) 3397 (br s), 3192 (s), 2966 (m), 2926 (m), 2842 (w), 1743 (m),
1731 (m), 1659 (s), 1646 (s), 1623 (m), 1465 (m), 1444 (m), 1373 (m),
1329 (s), 1290 (m), 1258 (m), 1147 (s), 1083 (m), 1040 (m), 927 (w),
814 (m), 803 (m), 657 (m), 566 (m), 555 (m) cmꢂ1. Anal. Calcd For
13C NMR (100 MHz, D2O)
d 174.9, 156.6, 145.2, 133.8, 130.1, 126.8,
50.3, 48.3, 44.4, 40, 33, 30.2, 26.9, 20.7. MS (ESI): m/z (%) 438.2 (100)
[Mþ]. IR (KBr) 3391 (br s), 3191 (s), 2964 (m), 2921 (m), 2846 (w),
1744 (m), 1733 (m), 1663 (s), 1647 (s), 1628 (m), 1465 (m), 1448 (m),
1379 (m), 1333 (s), 1293 (m), 1260 (m), 1158 (s), 1088 (m), 1045 (m),
933 (w), 814 (m), 803 (m), 668 (m), 572 (m), 551 (m) cmꢂ1. Anal.
Calcd For C18H28ClN7O4S: C, 45.61; H, 5.95; N, 20.69. Found C, 45.51;
H, 6.01; N, 20.49.
C19H30ClN7O4S: C, 46.76; H, 6.20; N, 20.09. Found C, 46.64; H, 6.29;
N, 20.01.
4.8. Synthesis of alkynes
4.7.8. N-{3-[4-(3-{[Amino(imino)methyl]amino}propyl)-1H-1,2,3-
N,N0-Di-Boc-N00-(but-3-ynyl)-guanidine (27), N,N0-Di-Boc-N00-
(pent-3-ynyl)-guanidine (28) were synthesized according to liter-
ature procedure.37
triazol-1-yl]propyl}-N-[(4-methylphenyl)sulfonyl]glycine
mono-
hydrochloride (21). Obtained from general procedure F starting
from 12. Partially unstable compound, a small aliquot was isolated
via semipreparative RP-HPLC and used in screening assay and 1H
4.8.1. Pent-4-ynyl-pyridin-2-yl-carbamic acid tert-butyl ester
(29). N-Boc-2-amino-pyridine49 (0.5 g, 2.57 mmol) was dissolved
in anhydrous DMF under N2 atmosphere and cooled to 0 ꢀC. To the
vigorous stirred solution was slowly added NaH (0.129 g, 60% dis-
persion in mineral oil), paying attention to keep temperature below
spectroscopy. 1H NMR (300 MHz, D2O)
d 7.61 (s, 1H), 7.48 (d,
J¼8.4 Hz, 2H), 7.26 (d, J¼8.4 Hz, 2H), 4.29e2.5 (m, 2H), 3.89 (s, 2H),
3.07e3.01 (m, 4H), 2.67e2.61 (m, 2H), 2.27 (s, 3H), 1.96e1.85 (m,
2H), 1.84e1.77 (m, 2H). MS (ESI): m/z (%) 438.2 (100) [Mþ], 380.2
Please cite this article in press as: Fabbrizzi, P.; et al., Tetrahedron (2014), http://dx.doi.org/10.1016/j.tet.2014.06.125

10
P. Fabbrizzi et al. / Tetrahedron xxx (2014) 1e11
ꢀC. The mixture was stirred under these conditions for 20 min,
L,
added. Three-dimensional energy scoring grids of 0.375 A resolu-
ꢀ
5
ꢀ
ꢀ
ꢀ
then (5-chloro-1-pentynyl)trimethylsilane (0.52 g, 530
m
tion and 40 Aꢁ40 Aꢁ40 A dimensions were computed. The center
of the grid was set to be coincident with mass center of ligands
preliminary fitted on the X-ray structure of c[RGDf(Me)V] in the
avb3 complex (1L5G). A total of 50 runs with a maximum of
2,500,000 energy evaluations were carried out for each ligand,
using the default parameters for LGA. Cluster analysis was per-
formed on docked results using a root-mean-square (rms) toler-
2.96 mmol) was added dropwise, and stirring was continued at
ꢃ5 ꢀC for 30 min. The solution was allowed to reach room tem-
perature and stirred for additional 1 h, then 1.33 mL of water was
added. TBAF (1 M solution in THF, 5.15 mL, 5.15 mmol) was added,
and the solution stirred overnight at rt. Finally, the mixture was
diluted with water (6 mL) and extracted with diethyl ether
(40 mLꢁ2), the organic layer was washed with 0.1 M HCl, saturated
NaHCO3 and brine and dried over Na2SO4. The solvent was evap-
orated under vacuum. Purified via FCC P. E.eEtOAc 10:1 Rf¼0.43.
ꢀ
ance of 1.5 A. Analysis of the binding mode, calculation of the
binding energy, and prediction of the binding activity of docked
conformations were carried out using Autodock plugin within
PyMol software.51
Colorless oil, 30% yield, 0.2 g. 1H NMR (200 MHz, CDCl3)
d 8.39e8.32
(m, 1H), 7.65e7.55 (m, 2H), 7.04e6.94 (m, 1H), 4.03 (t, J¼7.1 Hz, 2H),
2.22 (td, J¼7.4 Hz, 2.6 Hz, 2H), 1.94e1.80 (m, 3H), 1.51 (s, 9H). 13
C
4.11. Physico-chemical predictions
NMR (50 MHz, CDCl3) d 147.6, 136.9, 119.9, 119.5, 83.9, 81.1, 68.3, 46,
28.3, 27.8, 16.2. MS (ESI): m/z (%) 260.9 (Mþ, 58%), 283.0 (MꢂNa,
100%). IR (CHCl3) 3680 (m), 3595 (m), 3052 (s), 2985 (m), 2252 (m),
1607 (m), 1422 (m) cmꢂ1. Anal. Calcd for C15H20N2O2: C, 69.20; H,
7.74; N, 10.76. Found C, 69.31; H, 7.76; N, 10.73.
All physico-chemical parameters included in Table 2 were cal-
culated from chemical structures of compounds 18e26, 31, and 32
with calculator plugins within the framework of MarvinSketch
v6.2.2 (Chemaxon Ltd., http://www.chemaxon.com).
4.9. Solid-phase integrin binding assay
Acknowledgements
[
125I]-Echistatin, labeled by the lactoperoxidase method to
This work has received financial support from the Regione
Toscana regional council through POR CRO FSE 2007-2013, UNI-
FI_FSE2012, and MIUR PRIN2010-2011 cod. 2010NRREPL funding
actions.
a specific activity of 2000 Ci/mmol, was purchased from GE
Healthcare, and integrin avb3 from human placenta was purchased
from Chemicon International Inc., Temecula, CA. Purified avb3
integrin was diluted in coating buffer (20 mM Tris (pH 7.4), 150 mM
NaCl, 2 mM CaCl2, 1 mM MgCl2, 1 mM MnCl2) at concentrations of
500 or 1000 ng/mL. An aliquot of diluted integrin (100 mL/well) was
References and notes
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Please cite this article in press as: Fabbrizzi, P.; et al., Tetrahedron (2014), http://dx.doi.org/10.1016/j.tet.2014.06.125