New indolic non-peptidic HIV protease inhibitors from (S)-glycidol: synthesis and preliminary biological activity

Article abstract of DOI:10.1016/j.tet.2009.05.089

A series of non-peptidic HIV protease inhibitors were synthesized starting from the same optically active precursor, (S)-glycidol. The substrate was easily converted into different indolic sulfonamides or amines by regioselective reactions. The preliminary inhibitory activity was evaluated.

Full text of DOI:10.1016/j.tet.2009.05.089

Tetrahedron 65 (2009) 5984–5989
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New indolic non-peptidic HIV protease inhibitors from (S)-glycidol:
synthesis and preliminary biological activity
,
a
a
b
Lucia Chiummiento a, Maria Funicello a , Paolo Lupattelli , Francesco Tramutola , Pietro Campaner
*
a
`
Dipartimento di Chimica, Universita della Basilicata, Via N.Sauro 85, 85100 Potenza, Italy
Dipartimento di Scienze Chimiche, Universita di Trieste, Via Giorgieri 1, 34127 Trieste, Italy
b
`
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of non-peptidic HIV protease inhibitors were synthesized starting from the same optically active
precursor, (S)-glycidol. The substrate was easily converted into different indolic sulfonamides or amines
by regioselective reactions. The preliminary inhibitory activity was evaluated.
Ó 2009 Elsevier Ltd. All rights reserved.
Received 3 February 2009
Received in revised form 11 May 2009
Accepted 28 May 2009
Available online 6 June 2009
Dedicated to Professor Carlo Bonini on the
occasion of his 60th birthday
Keywords:
HIV PR inhibitors
Peptidomimetics
Indole
Biological activity
1. Introduction
reported,5 all inhibitors tailored after substrate-based peptide of an
enzyme are referred to as peptidomimetics and show a structure
Acquired immunodeficiency syndrome (AIDS) represents one of
the most important problems in medicine and the interest in de-
veloping inhibitors targeting the different steps in the life cycle of
the virus is still significant. More recent therapies for AIDS1,2 are
essentially directed against important enzymes that regulate the
HIV vital life cycle: reverse transcriptase (RT), aspartyl protease
(PR)3 and more recently integrase (IN).4 In particular, the early
recognition of the central role of HIV protease in viral maturation
has made it an important target for the treatment of HIV/AIDS.
Since Saquinavir, the first protease inhibitor (PI), appeared on
the market, a number of PIs have been introduced in the regimens
of highly active antiretroviral therapy (HAART). Nevertheless, the
first generation anti-protease therapeutics have shown several
drawbacks. These include: (1) severe side effects and drug toxicity;
(2) higher therapeutic doses and low bioavailability due to ‘pep-
tide-like’ character; (3) costly synthesis, which leads to high
treatment cost; (4) rapid emergence of drug resistance. To alleviate
these problems, great interest has been directed to design and
synthesize new inhibitors with a non-peptidic skeleton. As recently
where aminoacidic residues are numbered starting from the scissile
0
0
0
peptidic bond, such as Pn.P2–P1–P1 –P2 .Pn . All the commercially
available drugs against HIV protease are peptidomimetics, but
Tipranavir,6 a recently approved drug, possesses a non-peptidomi-
metic structure.
The need to overcome viral resistance and to have a simpler
synthetic sequence prompted us to investigate new non-peptidic
compounds with simplified structure, bearing the central iso-
propanolamine unit found in the commercially available peptidic
drugs and in some non-peptidic ones such as Darunavir.7 In our
continuing investigation on new inhibitors bearing heteroarylic
groups,8 the attention was focused on some reported structures9 in
which the presence of the indole ring, mimicking the aminoindane
group of Indinavir, seems to be important for the activity.
In a first synthetic approach various aspects can be considered:
(1) in drugs such as Saquinavir and Nelfinavir a perhydroisoquino-
line (PHIQ) residue appears important10 as P2 ligand for a good
0
0
interaction at the S2 subsite of the protease; (2) in most of the
commercially available anti-HIV PR drugs and in other compounds
with good inhibitory activity an isopropanolamine core is present
with R configuration of the hydroxyl function; (3) some non-pep-
tidic inhibitors, such as Darunavir, possess the arylsulfonamide
moiety.
* Corresponding author. Tel.: þ39 0971 202252; fax: þ39 0971 202223.
E-mail address: maria.funicello@unibas.it (M. Funicello).
0040-4020/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2009.05.089

L. Chiummiento et al. / Tetrahedron 65 (2009) 5984–5989
5985
On the basis of the previous considerations, we designed a series
of new indolic compounds as potential HIV PIs starting from the
commercially available (S)-glycidol, fundamental in generation of
the core. At one end of this central structure we hypothesized to
link a suitable indolic derivative by changing functionality and/or
its position on the heterocyclic ring, while at the other end we
considered the introduction of PHIQ or an arylsulfonamide moiety
(Scheme 1).
In particular, when we opened the epoxide ring with the chiral
amine PHIQ the final products 6, 7 and 8 were obtained in excellent
chemical yields and as single diastereomers.
For the preparation of indolic inhibitors with an arylsulfon-
amidic moiety, we then decided to use only epoxide 4 in reaction
with isobutylamine to afford compound 9 in excellent yield.13,14
This compound was then transformed into 10, 11 and 12 in high
yields by reacting with suitable arylsulfonyl chlorides.
It is noteworthy that we could differentiate the final target
molecule by introducing arylsulfonic residues with different elec-
tronic properties in order to evaluate their possible effect on in-
hibitory activity.
OH
R
N
O
X
HN
HO
R'
1
These six compounds were tested for anti-HIV PR activity and
their biological results are reported in Table 1. From these pre-
liminary biological results, it appeared clear that the 5-indolic
derivatives possess better activity than the 4- and 6-indolic ones
(Table 1, entries 1–3). Moreover, comparison of the results to
compounds 10–12 (Table 1, entries 4–6) showed that electron
donating properties of the arylsulfonamide ring could have some
effects on inhibition.
Nu
X = NH, O
(ArO-, ArNH2, RR'NH, etc..)
Scheme 1.
By this synthetic approach the influence of both the type and
the position of the substituent on the indole ring can be tested
using the commercial 4-, 5- and 6-hydroxyindole and 4- and 5-
aminoindole, largely used in many drugs synthesis,11 as starting
material.
Table 1
IC50 values for compounds 6–8 and 10–12
2. Results and discussion
Entry
Compound IC50
(m
M)
Entry
Compound IC50 (mM)
H
N
O
Glycidol offers two potential sites of electrophilic reactivity so
we can realize a first displacement and then ring opening, or we can
invert the reaction sequence. In a preliminary consideration both
routes were possible even if the direct displacement of the acti-
vated hydroxyl function by 4- or 5-aminoindole seemed more
difficult due to the decreased nucleophilicity of the heteroaromatic
amino group. After preliminary attempts by using different nucle-
ophiles, it was clear that two different approaches should be fol-
lowed depending on the type of functionality present on the
heterocyclic ring: in fact, the activation of glycidol by a nosyl group
was fundamental with hydroxyindoles since it allowed a regiose-
lective displacement without epoxide opening (Scheme 2), as
reported by Sharpless and co-workers.12
NO2
OH
OH
O
H
O
N
SO2
1
4
HN
N
H
NHtBu
10 (10.3 +/- 0.6)
6 (23.7 +/- 7)
H
OCH3
OCH3
OH
OH
H
O
N
O
O
N
2
3
5
6
SO2
N
H
NHtBu
N
H
11 (1.04 +/- 0.4)
7 (14 +/- 0.2)
H
N
O
NH2
OH
O
OH
H
O
N
SO2
O2N
NsCl, Et3N,
K2CO3, Het-OH
DMF, rt
O
O
N
H
NHtBu
HO
O
NH
12 (12.3 +/- 0.2)
CH2Cl2, -10 °C
80%
S
1
8 (138 +/- 33)
O2
2
H
With the aim toexplore the effectof differentfunctionality on the
indole ring we considered using 4- or 5-aminoindole. In this case
a different synthetic approach must be followed: in fact, PHIQ was
firstly introduced byepoxide ring opening of compound 1. Then, diol
13 was selectively activated on the primary hydroxyl function in
moderate yields and subsequently transformed into the new ep-
oxide 15. Finally 4- or 5-aminoindole was introduced under mild
reaction conditions with acceptable chemical yields (Scheme 3).
OH
O
H
N
O
PHIQ,
O
i-PrOH, rt
N
H
N
O
NHtBu
H
3: 4-hydroxyindole 63%
4: 5-hydroxyindole 72%
5: 6-hydroxyindole 61%
6: 4-hydroxyindole 91%
7: 5-hydroxyindole 87%
8: 6-hydroxyindole 93%
i-BuNH2,
OH
OH
i-PrOH, rt
O
NH
O
N
99%
ArSO2Cl, Et3N,
CH2Cl2, rt
SO2Ar
N
H
9
N
H
H
N
O
H
N
O
10 Ar: 4-NO2-C6H4, 80%
11 Ar: 3,4-diMeO-C6H3, 94%
12 Ar: 4-NH2-C6H4, 74%
H2, Pd/C 10%,
EtOAc
OH
OH
TsCl, Py,
CH2Cl2, rt
45%
PHIQ,
H
H
1
HO
TsO
i-PrOH, rt
96%
13
14
Scheme 2. Preparation of compounds 6–8 and 10–12.
NHtBu
NHtBu
H
N
O
OH
H
N
H
Hence, (S)-glycidol was treated with m-nosyl chloride to obtain
compound 2 in 80% yield: the subsequent displacement of the nosyl
group by suitable hydroxyindoles was performed affording the
corresponding 3, 4 and 5 in good yields and in mild reaction con-
ditions. At last, epoxide ring opening was easily realized to give the
desired final products.
H
HN
Het-NH2, i-PrOH,
O
K2CO3,
H
N
O
NHtBu
reflux
CH3OH, rt
15
16: 4-aminoindole 35%
17: 5-aminoindole 30%
NHtBu
Scheme 3. Preparation of compounds 16 and 17.

5986
L. Chiummiento et al. / Tetrahedron 65 (2009) 5984–5989
Biological assays were performed on these two new compounds
synthesis of compounds 6, 7 and 8 a suitable activation of the
primary hydroxyl function by nosyl chloride was necessary to ob-
tain the final products in only three steps and with 46%, 50% and
45% overall yields, respectively. Indeed, the preparation of com-
pounds 16 and 17, according to the second strategy, needed an
additional step of reaction due to the necessity to control the
regioselectivity in the epoxide ring opening (overall yields were 15
and 13%, respectively).
(Table 2, entries 1 and 2), but the results were unsatisfactory and
prompted us to consider other compounds. Hence, the amino group
on the 5-position of indole wastransformed into a carbamate function
and readily introduced into our inhibitor structure as the P2 ligand.
Table 2
IC50 for compounds 16, 17, 22 and 24
Entry
1
Compound IC50
(
m
M)
Entry
3
Compound IC50 (mM)
Moreover better biological activity resulted when an arylsulfo-
0
namide moiety was introduced in the P2 position.
H
H
Work is in progress in attempting to enlarge this series of non-
peptidic HIV PIs in order to improve the biological activity and to
evaluate the structure–activity relationships through a theoretical
approach.
OH
H
OH
H
N
H
H
N
O
N
O
N
HN
O
N
H
NHtBu
O
NHtBu
22(606)15
16 (157 +/- 12)
4. Experimental
4.1. General
H
NO2
OH
H
OH
O
H
N
H
N
N
O
N
2
4
SO2
O
24 (2.4 +/- 0.7)
N
H
N
H
NHtBu
Column chromatography was carried out on Merck silica gel
(0.063–0.200 mm particle size) by progressive elution with op-
portune solvent mixtures. 1H and 13C NMR spectra were normally
carried out in CDCl3 solutions on a VARIAN INOVA 500. Mass
spectra were obtained with a Hewlett–Packard 5971 mass-selective
detector on a Hewlett–Packard 5890 gas chromatograph [(OV-1
capillary column between 70 and 250 ꢀC (20 ꢀC minꢁ1)]. The optical
purity was evaluated by using a polarimeter JASCO Mod Dip-370.
Dichloromethane was dried by distillation over anhydrous CaCl2 in
inert atmosphere. Dry dimethylformamide was commercially
available. Compound 2 was prepared according to the literature12
starting from the commercially available (S)-(ꢁ)-glycidol.
17 (176 +/- 2)
Two parent compounds (22 and 24) bearing PHIQ or arylsulfo-
0
namide as the P2 unit were synthesized in good to excellent yields,
according to literature conditions reported for similar structures16
(Scheme 4).
H
N
O
NH2
ClOCO
Et3N,
O
+
N
H
CH2Cl2, rt
NO2
N
H
NO2
20
19
18
1, Et3N,
CH2Cl2, rt
90%
4.2. Nosyl displacement with hydroxyindoles: general
procedure
H
N
O
O
H
N
O
OH
PHIQ,
i-PrOH, rt
75%
H
N
H
O
O
O
K2CO3 (0.1433, 1.04 mmol) was added to a stirred solution of
hydroxyindole (0.0461 g, 0.35 mmol) in dry DMF (4 mL) at room
temperature under argon atmosphere; after 1 h a DMF solution
(3 mL) of compound 2 (0.0815 g, 0.31 mmol) was added and the
mixture was stirred overnight. After above 14 h (TLC control, CHCl3/
CH3OH 99:1) the reaction mixture was quenched by adding am-
monium chloride (saturated aqueous solution), then was extracted
with diethyl ether and the organic layer washed with brine. After
drying over Na2SO4, the organic layer was concentrated under
vacuo and the crude was purified by column chromatography on
silica gel (eluent: CH2Cl2/EtOAc 99:1).
N
H
21
N
H
i-BuNH,
i-PrOH, rt
92%
NHtBu
22
OH
H
NO2
N
O
NH
OH
H
N
4-NO2-C6H4-SO2Cl,
Et3N, CH2Cl2, rt
90%
O
N
O
SO2
N
H
23
O
N
H
24
Scheme 4. Preparation of compounds 22 and 24.
The values of HIV PR inhibitory activity are shown in Table 2
(entries 3 and 4): the obtained results appear difficult to explain. In
fact while compound 22 shows the worst value of the biological
activity, structure 24 shows an improvement of IC50 with respect to
the structures 16 and 17 (see Table 2): in our opinion, the simple
substitution of the PHIQ with sulfonamidic group is unable to
rationalize such data.
The results of inhibitory assays led us to make further consid-
erations: in particular it is clear that the 5-hydroxyindolic inhibitors
are the most active with both the PHIQ residue and with the aryl-
4.2.1. (ꢁ)-4-[((R)-Oxiran-2-yl)methoxy]-1H-indole (3)
Compound 3 was isolated as a brown thick oil (0.0375 g, 63%).
20
[a
]
ꢁ6.4 (c 1.6, CHCl3); Rf 0.5 (CH2Cl2/EtOAc 99:1); dH (500 MHz,
D
CDCl3) 8.25 (1H, s), 7.14–7.05 (3H, m), 6.71 (1H, t, J 2.5), 6.55 (1H, d, J
8.0), 4.38 (1H, dd, J 3.0 and 11.0), 4.17 (1H, dd, J 6.0 and 11.0), 3.49–
3.47 (1H, m), 2.96 (1H, t, J 5.0), 2.85 (1H, dd, J 2.5 and 5.0); dC
(125 MHz, CDCl3) 152.1, 133.2, 127.1, 120.5, 117.6, 112.3, 103.4, 102.7,
70.5, 50.9, 44.3. MS (EI) m/z: 189 (Mþ) (100),132 (63),104 (50). Anal.
Calcd for C11H11NO2: C, 69.83; H, 5.86%. Found: C, 69.85; H, 5.84.
0
sulfonamidic one as the P2 unit.
On the other hand, the aminoindolic structures show generally
low activity, but transformation of the amino group into a carba-
mate function gave an improvement in IC50 value when a sulfona-
midic framework is present, but a lowering with a PHIQ residue.
4.2.2. (ꢁ)-5-[((R)-Oxiran-2-yl)methoxy]-1H-indole (4)
Compound 4 was isolated as a brown thick oil (0.0450 g, 72%).
20
[a
]
ꢁ2.2 (c 1.2, CHCl3); Rf 0.5 (CH2Cl2/EtOAc 99:1); dH (500 MHz,
D
CDCl3) 8.16 (1H, s), 7.31–7.14 (3H, m), 6.92 (1H, dd, J 1.5 and 8.5), 6.5
(1H, s), 4.27 (1H, dd, J 3.5 and 12.0), 4.04 (1H, dd, J 6 and 11.0), 3.43–
3.42 (1H, m), 2.942 (1H, t, J 4.5), 2.81 (1H, dd, J 3.0 and 5.5); dC
(125 MHz, CDCl3) 153.0, 131.2, 128.1, 125.0, 112.8, 111.7, 103.8, 102.3,
69.6, 50.4, 44.9; MS (EI) m/z: 189 (Mþ) (100),132 (80),104 (54). Anal.
Calcd for C11H11NO2: C, 69.83; H, 5.86%. Found: C, 69.82; H, 5.87.
3. Conclusion
Ten new indolic non-peptidic inhibitors were prepared starting
from (S)-glycidol in two different synthetic approaches. For the

L. Chiummiento et al. / Tetrahedron 65 (2009) 5984–5989
5987
4.2.3. (ꢁ)-6-[((R)-Oxiran-2-yl)methoxy]-1H-indole (5)
temperature for 26 h. Then solvent was removed under reduced
pressure and the crude was purified by column chromatography on
Compound 5 was isolated as a yellow solid (0.0357 g, 61%). Mp
20
105 ꢀC; [
a
]
D
ꢁ3.4 (c 1, CHCl3); Rf 0.4 (CH2Cl2/EtOAc 99:1); dH
silica gel (CHCl3/CH3OH 9:1) affording compound 9 as a colourless
20
(500 MHz, CDCl3) 8.15 (1H, s), 7.54 (1H, d, J 11.0), 7.09 (1H, s), 6.85
(2H, d, J 6.5), 6.50 (1H, d, J 1.0), 4.24 (1H, dd, J 1.5 and 13.5), 3.96 (1H,
dd, J 7.0 and 14.0), 3.39 (1H, m), 2.93–2.78 (2H, m); dC (125 MHz,
CDCl3) 155.1, 136.3, 123.4, 122.5, 121.2, 110.2, 102.2, 95.9, 69.3, 50.3,
44.7; MS (EI) m/z: 189 (Mþ) (100), 132 (80), 104 (54). Anal. Calcd for
C11H11NO2: C, 69.83; H, 5.86%. Found: C, 69.82; H, 5.87.
thick oil (0.358 g, 99%). [
a
]
þ6.5 (c 1.3, CH3OH); Rf 0.5 (CHCl3/
D
CH3OH 9:1); dH (500 MHz, CDCl3) 8.66 (1H, s), 7.25–7.10 (3H, m),
6.85 (1H, dd, J 1.5 and 8.5), 6.45 (1H, s), 4.22–4.18 (1H, m), 4.02–3.97
(2H, m), 3.72–3.68 (2H, m), 2.87 (2H, d, J 7.0), 1.85–1.80 (1H, m),
0.94 (3H, d, J 3.5), 0.93 (3H, d, J 3.5); dC (125 MHz, CDCl3) 152.9,
131.2, 128.2, 125.1, 112.4, 111.7, 103.6, 101.9, 71.3, 67.9, 57.4, 51.8, 27.7,
20.4. IR (cmꢁ1) 3402, 3313, 3050, 2958, 1455, 1159. Anal. Calcd for
C15H22N2O2: C, 68.67; H, 8.45%. Found: C, 68.69; H, 8.43.
4.3. Ring opening of the oxyranylmethoxy-1H-indole with
PHIQ: general procedure
4.5. Synthesis of arylsulfonamides starting from compound
9: general procedure
PHIQ (0.0433 g, 0.18 mmol) was added to a stirred solution of
suitable epoxide (0.0314 g, 0.15 mmol) in i-PrOH (2 mL) at room
temperature. After above 20 h the solvent was removed under re-
duced pressure and the crude purified by column chromatography
on silica gel (CHCl3/CH3OH 95:5).
Dry triethylamine (0.14 mL, 1.01 mmol) and the opportune
arylsulfonyl chloride (0.93 mmol) were added to a solution of
stirred compound 9 (0.206 g, 0.78 mmol) in dry CH2Cl2 (40 mL) at
room temperature under argon atmosphere. After about 24 h (TLC
control in CHCl3/CH3OH 99:1) the reaction mixture was quenched
by adding a 5% solution of H2SO4 and extracted by CH2Cl2. The or-
ganic layer was washed with a NaHCO3 (saturated aqueous solu-
tion) and brine, then it was dried over Na2SO4 and concentrated
under reduced pressure. The crude was purified by column chro-
matography on silica gel (CHCl3/CH3OH 99:1).
4.3.1. (ꢁ)-(3S,4aS,8aS)-2-[(R)-3-(1H-Indol-4-yloxy)-2-hydroxy-
propyl]-N-tert-butyl-decahydroisoquinoline-3-carboxamide (6)
Compound 6 was isolated as a pink solid (0.0613 g, 91%); mp
20
85 ꢀC; [
a
]
ꢁ61.2 (c 1.5, CHCl3); Rf 0.4 (CHCl3/CH3OH 95:5); dH
D
(500 MHz, CDCl3) 8.51 (1H, s), 7.14–7.06 (3H, m), 6.65 (1H, d, J 2.5),
6.53 (1H, d, J 7.5), 6.25 (1H, s, NH), 4.42 (1H, d, J 9.0), 4.07 (1H, t, J
8.0), 3.12 (1H, s, OH), 2.99 (1H, d, J 7.5), 2.78 (1H, d, J 8.0), 2.66 (1H, d,
J 8.0), 2.44 (1H, d, J 7.5), 2.31 (1H, d, J 8.0), 1.92–1.39 (9H, m), 1.34
(9H, s), 1.28–1.16 (4H, m); dC (125 MHz, CDCl3) 171.5, 152.1, 137.3,
122.8, 122.5, 118.6,105.1, 100.7, 99.5, 70.4, 68.2, 59.5, 58.4, 50.6, 35.7,
33.1, 30.8, 30.6, 28.6, 26.1, 25.7, 20.4. IR (cmꢁ1) 3311, 2925, 1652,
1365. Anal. Calcd for C25H37N3O3: C, 70.22; H, 8.72%. Found: C,
70.20; H, 8.73.
4.5.1. (þ)-(R)-N-[3-(1H-Indol-5-yloxy)-2-hydroxypropyl]-
N-isobutyl-4-nitrobenzenesulfonamide (10)
Compound 10 was isolated as a yellow solid (0.279 g, 80%). Mp
20
135 ꢀC; [
a]
þ18.0 (c 0.8, CHCl3); Rf 0.7 (CHCl3/CH3OH 99:1); dH
D
(500 MHz, CDCl3) 8.29 (1H, d, J 8.5), 8.16 (1H, s), 7.99 (2H, d, J 8.5),
7.27 (1H, d, J 7.0), 7.20 (1H, d, J 2.5), 7.07 (1H, s), 6.80 (1H, dd, J 1.5
and 8.5), 6.47 (1H, s), 4.22–4.18 (1H, m), 4.00–3.99 (2H, m), 3.42–
3.38 (2H, m), 3.09–3.05 (2H, m), 2.86 (2H, d, J 4.5), 2.01–1.98 (1H,
m), 0.92 (3H, d, J 2.5), 0.90 (3H, d, J 2.5); dC (125 MHz, CDCl3) 152.6,
149.9, 145.1, 131.3, 128.5, 128.3, 124.0, 112.3, 111.8, 103.8, 102.4, 70.2,
68.8, 57.4, 51.8, 26.7, 19.9. IR (cmꢁ1) 3417, 3102, 2961, 1529, 1349,
1159. Anal. Calcd for C21H25N3O6S: C, 56.36; H, 5.63%. Found: C,
56.38; H, 5.61.
4.3.2. (ꢁ)-(3S,4aS,8aS)-2-[(R)-3-(1H-Indol-5-yloxy)-2-hydroxy-
propyl]-N-tert-butyl-decahydroisoquinoline-3-carboxamide (7)
Compound 7 was isolated as a yellow thick oil (0.0632 g, 87%);
20
[a]
ꢁ70.3 (c 2, CHCl3); Rf 0.4 (CHCl3/CH3OH 95:5); dH (500 MHz,
D
CDCl3) 8.45 (1H, s), 7.29(1H, t, J 9.0), 7.21 (1H, t, J 2.5), 7.11 (1H, d, J 2.5),
6.87 (1H, dd, J 2.5 and 9.0), 6.47 (1H, t, J 2.5), 6.29 (1H, s, NH), 4.19 (1H,
s, OH), 4.11 (1H, dd, J4.0and9.5), 3.95(1H, dd, J 7.0and9.0), 3.01–2.22
(5H, m), 1.91–139 (10H, m), 1.35 (9H, s), 1.22–1.18 (3H, m); dC
(125 MHz, CDCl3) 173.9, 152.9, 131.3, 128.2, 125.1, 112.6, 111.8, 103.7,
102.7, 71.2, 70.3, 68.3, 59.4, 58.6, 50.7, 35.7, 35.7, 33.1, 30.8, 30.6, 29.6,
28.6, 26.1, 25.7, 20.5. IR (cmꢁ1) 3311, 2925,1652,1365. Anal. Calcd for
C25H37N3O3: C, 70.22; H, 8.72%. Found: C, 70.19; H, 8.76.
4.5.2. (þ)-(R)-N-[3-(1H-Indol-5-yloxy)-2-hydroxypropyl]-N-
isobutyl-3,4-dimethoxybenzenesulfonamide (11)
Compound 11 was isolated as a white solid (0.34 g, 94%). Mp
20
118 ꢀC; [
a
]
þ4.1 (c 1.6, CHCl3); Rf 0.6 (CHCl3/CH3OH 99:1); dH
D
(500 MHz, CDCl3) 8.14 (1H, s), 7.45 (1H, dd, J 1.5 and 8.0), 7.29–7.26
(2H, m), 7.19 (1H, d, J 2.5), 7.09 (1H, s), 6.93 (1H, d, J 8.5), 6.83 (1H,
dd, J 2.0 and 8.5), 6.47 (1H, t, J 1.0), 4.26–4.22 (1H, m), 4.06–3.99
(2H, m), 3.93 (3H, s), 3.91 (3H, s), 3.33–3.21 (2H, m), 3.04–2.92 (2H,
m), 1.98–1.96 (1H, m), 0.94 (3H, d, J 8.5), 0.90 (3H, d, J 8.5); dC
(125 MHz, CDCl3) 152.8, 152.6, 149.1, 131.3, 130.5, 128.3, 125.1, 121.3,
112.4, 111.7, 110.6, 109.9, 103.7, 102.4, 70.3, 69.2, 58.3, 56.2, 56.0,
52.7, 27.0, 20.0. IR (cmꢁ1) 3390, 2954, 1509, 1262, 1137. Anal. Calcd
for C22H30N2O6S: C, 59.72; H, 6.54%. Found: C, 59.70; H, 6.51.
4.3.3. (ꢁ)-(3S,4aS,8aS)-2-[(R)-3-(1H-Indol-6-yloxy)-2-hydroxy-
propyl]-N-tert-butyl-decahydroisoquinoline-3-carboxamide (8)
Compound 8 was isolated as a brown thick oil (0.0633 g, 93%);
20
[
a]
ꢁ66.0 (c 1.3, CH3OH); Rf 0.6 (CHCl3/CH3OH 95:5); dH
D
(500 MHz, CDCl3) 8.53 (1H, s), 7.51 (1H, d, J 8.5), 7.21 (1H, t, J 2.5),
6.90 (1H, d, J 2.0), 6.79 (1H, dd, J 2.5 and 9.0), 6.48 (1H, t, J 2.0), 6.24
(1H, s, NH), 4.16 (1H, t, J 5.0), 4.08 (1H, dd, J 4.5 and 9.0), 3.95 (1H,
dd, J 6.5 and 9.5), 2.95 (1H, dd, J 2.0 and 11.5), 2.76–2.64 (2H, m), 2.4
(1H, dd, J 5.0 and 13.5), 2.28 (1H, dd, J 3.0 and 12.0), 1.92–1.40 (11H,
m), 1.37 (9H, s), 1.34–1.21 (3H, m); dC (125 MHz, CDCl3) 174.0, 155.0,
136.4, 123.4, 122.5, 121.1, 110.1, 102.1, 95.9, 70.9, 70.2, 68.3, 59.5,
58.5, 50.7, 35.7, 33.1, 30.8, 30.6, 28.6, 26.1, 25.7, 20.5. IR (cmꢁ1) 3311,
2925, 1652, 1365. Anal. Calcd for C25H37N3O3: C, 70.22; H, 8.72%.
Found: C, 70.20; H, 8.71.
4.6. (D)-(R)-4-Amino-N-[3-(1H-indol-5-yloxy)-2-hydroxy-
propyl]-N-isobutyl-benzenesulfonamide (12)
Compound 15 (0.1041 g, 0.23 mmol) was added to a suspension
of 10% Pd/C (14 mg) in ethyl acetate (20 mL) under hydrogen at-
mosphere. After 28 h the reaction mixture was filtered on a plate of
Celite, concentrated under vacuo and purified by column chroma-
4.4. (D)-(R)-1-(1H-Indol-5-yloxy)-3-(isobutylamino)-
propan-2-ol (9)
tography on silica gel (CHCl3/CH3OH 95:5) to afford compound 12
20
as a violet thick oil (0.071 g, 74%). [
a]
þ12.0 (c 1, CH3OH); Rf 0.3
D
(CHCl3/CH3OH 95:5); dH (500 MHz, CD3OD) 7.64 (2H, d, J 9.0), 7.26
(1H, d, J 8.5), 7.18 (1H, d, J 3.0), 7.06 (1H, d, J 2.5), 6.99 (1H, d, J 9.0),
6.79 (1H, dd, J 2.0 and 9.0), 6.36 (1H, d, J 2.5), 4.18–4.14 (1H, m), 4.00
i-ButNH2 (1.12 g, 1.48 mmol) was added to a stirred solution of
compound 4 (0.287 g, 1.38 mmol) in i-PrOH (30 mL) at room

5988
L. Chiummiento et al. / Tetrahedron 65 (2009) 5984–5989
(1H, dd, J 4.0 and 10.0), 3.94 (1H, dd, J 5.0 and 9.5), 3.42 (1H, dd, J 4.5
and 15.0), 3.11 (1H, dd, J 7.0 and 14.5), 2.96 (1H, dd, J 7.5 and 13.5),
2.88 (1H, dd, J 7.5 and 13.5), 2.01–1.97 (1H, m), 0.93 (3H, d, J 3.0),
0.91 (3H, d, J 3.0); dC (125 MHz, CD3OD) 157.1, 154.1, 133.1, 129.8,
126.2, 114.5, 113.2, 112.9, 112.7, 104.4, 102.2, 71.9, 70.3, 59.1, 53.1,
28.0, 20.5. IR (cmꢁ1) 3465, 3312, 3065, 2954, 1509, 1262, 1137. Anal.
Calcd for C21H27N3O4S: C, 60.41; H, 6.52%. Found: C, 60.39; H, 6.53.
(3 mL). The mixture was heated at reflux temperature until disap-
pearance of epoxide (about 10 h, TLC control, CHCl3/CH3OH 9:1).
After cooling the solvent was removed under reduced pressure and
the crude was purified by column chromatography on silica gel
(eluent: CHCl3/CH3OH 9:1).
4.10.1. (ꢁ)-(3S,4aS,8aS)-2-[(S)-3-(1H-Indol-4-ylamino)-
2-hydroxypropyl]-N-tert-butyl-decahydroisoquinoline-3-
carboxamide (16)
4.7. (L)-(3S,4aS,8aS)-N-tert-Butyl-2-[(R)-2,3-dihydroxy-
propyl]decahydroisoquinoline-3-carboxamide (13)
Compound 16 was isolated as a violet thick oil (0.0334 g, 35%
20
from compound 14). [
a
]
D
ꢁ96.4 (c 1.4, CHCl3); Rf 0.3 (CHCl3/CH3OH
PHIQ (1.1442 g, 4.8 mmol) was added to a stirred solution of (S)-
(ꢁ)-glicydol 1 (0.2922 g, 4 mmol) in i-PrOH (45 mL), at room tem-
perature for about 34 h until complete disappearance of epoxide
(TLC control, EtOAc). Evaporation of the solvent and crystallization
9:1); dH (500 MHz, CDCl3) 8.68 (1H, s), 7.10 (1H, t, J 2.5), 7.01 (1H, t, J
7.5), 6.87 (1H, d, J 8.0), 6.46–6.36 (3H, m), 3.84–2.57 (7H, m), 2.15
(1H, dd, J 3.0 and 11.5), 1.99 (1H, d, J 12.0), 1.88–1.41 (9H, m), 1.36
(9H, s), 1.33–1.24 (4H, m); dC (125 MHz, CDCl3) 173.7, 139.1, 136.8,
122.8, 122.5, 117.3, 103.9, 102.3, 98.4, 69.7, 65.7, 64.6, 58.1, 57.8, 50.8,
35.7, 33.1, 30.8, 30.7, 28.5, 26.2, 25.4, 20.1. IR (cmꢁ1) 3286, 3100,
2944, 1659, 1368, 1262, 1140. Anal. Calcd for C25H38N4O2: C, 70.39;
H, 8.98%. Found: C, 70.41; H, 8.95.
(EtOAc/hexane) afforded compound 13 (1.2 g, 96%) as a white solid.
20
Mp 166–168 ꢀC; [
a]
ꢁ142.0 (c 1, CHCl3); Rf 0.4 (EtOAc); dH
D
(500 MHz, CDCl3) 6.04 (1H, s), 3.83 (2H, d, J 9.0), 3.63 (1H, dd, J 4.0
and 11.5), 2.82 (1H, d, J 11.0), 2.58 (1H, d, J 10.0), 2.52 (1H, dd, J 9.5 and
12.5), 2.24 (2H, dt, J 4.5 and 12.5), 1.97–1.59 (4H, m), 1.53–1.32 (6H,
m), 1.30 (9H, s), 1.28–1.20 (2H, m); dC (125 MHz, CDCl3) 174.0, 70.2,
68.7, 64.7, 59.1, 58.2, 51.2, 35.8, 33.3, 30.8, 30.8, 28.6, 26.1, 25.5, 20.3.
IR (cmꢁ1) 3400, 2954, 1680, 1429, 1262, 1137. Anal. Calcd for
C17H32N2O3: C, 65.35; H, 10.32%. Found: C, 65.32; H, 10.34.
4.10.2. (ꢁ)-(3S,4aS,8aS)-2-[(S)-3-(1H-Indol-5-ylamino)-
2-hydroxypropyl]-N-tert-butyl-decahydroisoquinoline-3-
carboxamide (17)
Compound 17 was isolated as a violet solid (0.029 g, 30% from
20
compound 14). Mp 128 ꢀC; [
a
]
ꢁ87.3 (c 1.1, CHCl3); Rf 0.3 (CHCl3/
D
4.8. (L)-(R)-3-[(3S,4aS,8aS)-3-(tert-Butylcarbamoyl)-
octahydroisoquinolin-2(1H)-yl]-2-hydroxypropyl 4-
methylbenzenesulfonate (14)
CH3OH 9:1); dH (500 MHz, CDCl3) 8.24 (1H, s), 7.18 (1H, d, J 8.5), 7.11
(1H, d, J 3.0), 6.94 (1H, s), 6.66–6.64 (1H, m), 6.35 (1H, s), 6.04 (1H,
s), 3.81–3.62 (3H, m), 3.81–3.37 (7H, m), 2.77 (1H, d, J 11.0), 2.55–
2.47 (2H, m), 2.21–2.15 (2H, m), 1.92–1.37 (5H, m), 1.35 (9H, s), 1.34–
1.20 (2H, m); dC (125 MHz, CDCl3) 174.1, 139.2, 130.7, 128.7, 124.7,
112.8, 111.5, 105.5, 101.3, 70.2, 65.6, 64.7, 59.1, 58.2, 51.1, 35.7, 33.2,
30.8, 30.7, 28.6, 26.1, 25.5, 20.2. IR (cmꢁ1) 3281, 3090, 2950, 1642,
1388, 1256, 1123. Anal. Calcd for C25H38N4O2: C, 70.39; H, 8.98%.
Found: C, 70.37; H, 8.96.
Dry pyridine (0.284 mL, 3.49 mmol) and tosyl chloride (0.666 g,
3.49 mmol) were added to a stirred solution of compound 13
(0.992 g, 3.17 mmol) in dry CH2Cl2 (43 mL) at room temperature.
The mixture was stirred in argon atmosphere for 20 h, then it was
washed with diluted hydrochloric acid (0.1 M, 20 mL), with a sat-
urated aqueous solution of NaHCO3 and finally with brine. The
organic layer was dried and evaporated under reduced pressure to
leave a crude purified by crystallization (Et2O/hexane). Compound
4.11. 4-Nitrophenyl 1H-indol-5-ylcarbamate (20)
14 was obtained as a pink solid (0.666 g, 45%). Mp 110–111 ꢀC;
Dry triethylamine (0.54 mL, 3.9 mmol) and compound 18
(0.7860 g, 3.9 mmol) were added to a stirred solution of 19
(0.396 g, 3 mmol) in dry CH2Cl2 (30 mL) at room temperature
and in argon atmosphere. After 2 h a TLC control (CH2Cl2/EtOAc
99:1) showed the disappearance of 19 and the reaction was
quenched by adding water (15 mL). The organic layer was
washed with brine, dried over Na2SO4 and concentrated under
vacuo affording compound 20 (0.862 g, 97%) as a yellow solid.
This compound was used as crude for the following reaction. Rf
0.6 (CH2Cl2/EtOAc 99:1).
20
[
a]
ꢁ34.0 (c 1.14, CHCl3); Rf 0.5 (petroleum ether/EtOAc 1:1); dH
D
(500 MHz, CDCl3) 7.78 (2H, d, J 8.5), 7.34 (2H, d, J 8.0), 5.97 (1H, s),
4.57–4.43 (2H, m), 4.09 (1H, dd, J 8.0 and 12.5), 3.94 (1H, t, J 9.5),
3.86 (1H, dd, J 9.0 and 12.0), 2.43 (3H, s), 2.78–2.53 (3H, m), 2.29–
2.15 (3H, m), 1.84–1.35 (6H, m), 1.30 (9H, s), 1.28–1.15 (4H, m); dC
(125 MHz, CDCl3) 167.5, 144.9, 132.4, 129.8, 127.9, 71.9, 69.9, 67.1,
57.6, 51.9, 50.8, 35.6, 32.9, 31.5, 30.5, 28.5, 26.0, 25.3, 21.5, 21.2.
Anal. Calcd for C24H38N2O5S: C, 61.77; H, 8.21%. Found: C, 61.73;
H, 8.24.
4.9. (3S,4aS,8aS)-N-tert-Butyl-2-[(R)-oxiran-2-yl methyl]deca-
4.12. (D)-(R)-Oxiran-2-ylmethyl 1H-indol-5-ylcarbamate (21)
hydroisoquinoline-3-carboxamide (15)
Dry triethylamine (0.7 mL, 5 mmol) and S-glycidol 1 (0.279 g,
3.8 mmol) were added to a stirred solution of compound 20
(0.8621 g, 2.9 mmol) in dry CH2Cl2 (30 mL) at room temperature
and under argon atmosphere. After the disappearance of glycidol
1 (about 10 h by TLC control, CH2Cl2/EtOAc 8:2) the mixture was
diluted with CH2Cl2. The organic layer was washed with brine,
dried over Na2SO4 and concentrated under vacuo. The crude was
purified by column chromatography (CH2Cl2/EtOAc 8:2) afford-
Potassium carbonate was added to a stirred solution of com-
pound 14 (0.208 g, 0.45 mmol) in methanol (26 mL) at room tem-
perature. After disappearance of the starting material, the reaction
was quenched by adding ammonium chloride (saturated aqueous
solution), then extracted with diethyl ether and washed with brine.
The combined extracts were dried over Na2SO4 and evaporated
under reduced pressure to give the crude 15 (0.107 g): this com-
pound was used in subsequent reaction without any purification. Rf
0.6 (petroleum ether/EtOAc 3:2); dH (500 MHz, CDCl3) 6.61 (1H, s),
3.01–2.57 (8H, m), 1.75–1.46 (10H, m), 1.30 (9H, s).
ing compound 21 as a yellow solid (0.606 g, 90%). Mp 109 ꢀC;
20
[
a
]
þ7.0 (c 1, CHCl3); Rf 0.3 (CH2Cl2/EtOAc 8:2); dH (500 MHz,
D
CDCl3) 8.18 (1H s), 7.69 (1H, s), 7.33–7.13 (3H, m), 6.70 (1H, s),
6.51 (1H, d, J 2.5), 4.56 (1H, dd, J 3.0 and 12.5), 4.01 (1H, dd, J 5.5
and 11.0), 3.30–2.71 (3H, m); dC (125 MHz, CDCl3) 153.7, 133.0,
130.0, 128.1, 125.3, 115.7, 111.3, 111.1, 102.7, 65.5, 49.8, 44.7. Anal.
Calcd for C12H12N2O3: C, 62.06; H, 5.21%. Found: C, 62.09;
H, 5.24.
4.10. Ring opening of epoxide 15: general procedure
Aminoindole (0.0285 g, 0.22 mmol) was added to a stirred so-
lution of the crude epoxide 15 (0.0534 g, 0.18 mmol) in i-PrOH

L. Chiummiento et al. / Tetrahedron 65 (2009) 5984–5989
5989
4.13. (L)-(R)-3-[(3S,4aS,8aS)-3-(tert-Butylcarbamoyl)-
octahydroisoquinolin-2(1H)-yl]-2-hydroxypropyl -
1H-indol-5-ylcarbamate (22)
7.8ꢂ0.3 mMꢁ1) substrate’s hydrolysis by a commercially available
HIV-PR, at a lexc and at a lem of 325 nm and 420 nm, respectively;
114
obtained diluting 10
of substrate in DMSO with 1.99 mL of pH 5.5 MES buffer) were put
in a thermostated cuvette (25 ꢀC) with 75
L of MES buffer (con-
m
L of the fluorogenic substrate (with a concentration of 53
mM,
m
L of a stock solution containing 10 mg mLꢁ1
PHIQ (0.040 g, 0.19 mmol) was added to a stirred solution of
compound 21 (0.030 g, 0.13 mmol) in i-PrOH (5 mL) at room tem-
perature. After about 28 h the mixture was concentrated in vacuo
and the crude was purified by column chromatography (CHCl3/
m
taining 100 mM 2-[N-morpholino]ethansulfonic acid (MES)/NaOH,
pH 5.5; 400 mM NaCl; 1 mM ethylendiaminotetracetic acid (EDTA);
CH3OH 9:1) affording compound 22 as a brown powder (0.046 g,
1 mM dithiotreitol (DTT)), obtaining a final concentration of 30
and starting measuring the fluorescence. After 1.5 min the HIV-PR
was added (11 L of a solution obtained diluting 1:100, with a MES/
mM
20
75%). Mp 139 ꢀC; [
a
]
D
þ42.0 (c 1.2, CH3OH); Rf 0.6 (CHCl3/CH3OH
9:1); dH (500 MHz, CDCl3) 8.31 (1H, s), 7.68 (1H, s), 7.31 (1H, d, J 9.0),
7.21 (1H, t, J 2.5), 7.11 (1H, d, J 8.0), 6.85 (1H, br s), 6.47 (1H, t, J 2.5),
6.50 (1H, s), 6.16 (1H, s), 4.39 (1H, d, J 8.0), 4.08–4.12 (2H, m), 2.92
(1H, d, J 11.5), 2.63 (1H, t, J 5.0), 2.28 (2H, t, J 12.0), 2.22–1.39 (11H,
m), 1.35 (9H, s), 1.22–1.18 (3H, m); dC (125 MHz, CDCl3) 173.7, 154.4,
133.1, 130.0, 129.7, 128.1, 125.2, 115.7, 111.3, 102.6, 70.1, 68.1, 59.4,
58.8, 50.8, 35.7, 33.0, 31.9, 30.7, 30.6, 29.7, 29.3, 28.7, 26.1, 25.7, 20.6.
IR (cmꢁ1) 3410, 3280, 3046, 2939,1730,1642,1308,1245,1128. Anal.
Calcd for C26H38N4O4: C, 66.36; H, 8.14%. Found: C, 66.39; H, 7.15.
m
BSA buffer, a stock solution of 0.4 mg mLꢁ1 of HIV-PR in a 10 mM
sodium phosphate pH¼6.5, 1 mM EDTA/10% glycerol/0.05% mer-
captoethanol/50–100 mM NaCl), obtaining an enzymatic concen-
tration of 10 nm. The increase in the fluorescence was then
measured; after 1 min the inhibitor containing solution was added
(2 mL) and the fluorescence measured for other additional 10 min.
For each inhibitor, a stock solution in DMSO was prepared by
weight, then diluted it with DMSO or MES buffer. The amount of the
inhibition was evaluated comparing the initial rates, extrapolated
from the linear parts of the curves obtained plotting fluorescence
versus time, of the catalyzed reaction in the presence of different
inhibitor’s concentration. IC50 values were obtained simply plotting
the different slopes versus inhibitor’s concentrations (expressed
using a logarithmical scale) and interpolating the value corre-
sponding to the 50% of inhibition.
4.14. (D)-(R)-2-Hydroxy-3-(isobutylamino)propyl 1H-
indol-5-ylcarbamate (23)
i-BuNH2 (0.3 mL, 2.9 mmol) was added to a stirred solution of
compound 21 (0.24 g, 0.96 mmol) in i-PrOH (24 mL) at room tem-
perature. After 34 h the solvent was evaporated in vacuo affording
20
product 23 (0.270 g, 92%) as a yellow thick oil. [
a
]
þ4.9 (c 1.4,
D
CH3OH); Rf 0.6 (CHCl3/CH3OH 7:3); dH (500 MHz, CDCl3) 8.45 (1H,
s), 7.66 (1H, s), 7.27–7.04 (3H, m), 6.46 (1H, s), 4.28–4.25 (1H, m),
4.13–3.94 (2H, m), 2.75–2.41 (4H, m), 1.75–1.70 (1H, m), 0.91 (3H, d,
J 3.0), 0.89 (3H, d, J 3.0); dC (125 MHz, CDCl3) 154.4, 133.0, 130.1,
128.0, 125.3, 125.1, 115.5, 111.3, 102.6, 67.9, 67.2, 57.6, 51.3, 28.3, 20.5.
IR (cmꢁ1) 3316, 2958, 1704, 1557, 1481, 1237, 1063. Anal. Calcd for
C16H23N3O3: C, 62.93; H, 7.59%. Found: C, 62.89; H, 7.55.
Acknowledgements
The authors gratefully acknowledge MIUR (Ministry of In-
struction, University and Research-Rome) (Project PRIN 2005) and
University of Basilicata for financial support.
References and notes
4.15. (D)-(R)-2-Hydroxy-3-(N-isobutyl-4-nitrophenylsul-
fonamido)propyl 1H-indol-5-ylcarbamate (24)
1. Ghosh, A. K.; Bilcer, G.; Schiltz, G. Synthesis 2001, 15, 2203–2229.
2. Brik, A.; Wong, C. H. Org. Biomol. Chem. 2003, 1, 5–14.
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5. Tsantrizos, Y. S. Acc. Chem. Res. 2008, 41, 1252–1263.
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rahedron 2005, 61, 6580–6589.
Dry triethylamine (0.04 mL, 0.3 mmol) and 4-nitrobenzene-1-
sulfonyl chloride (0.059 g, 0.3 mmol) were added to a stirred so-
lution of compound 23 (0.075 g, 0.23 mmol) in dry CH2Cl2 at room
temperature under argon atmosphere. After 26 h the reaction was
quenched by adding 5% H2SO4 solution. Hence the mixture was
extracted with CH2Cl2 and the organic layer washed with a satu-
rated aqueous solution of NaHCO3 and brine. After drying over
Na2SO4 the solvent was evaporated in vacuo. The crude was puri-
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Bioorg. Med. Chem. 2002, 10, 2511–2526.
fied by column chromatography on silica gel (CHCl3/CH3OH 9:1)
10. (a) Babine, R. E.; Bender, S. L. Chem. Rev. 1997, 97, 1359–1472; (b) Ripka, A. S.;
Satyshur, K. A.; Bohacek, R. S.; Rich, D. H. Org. Lett. 2001, 3, 2309–2312.
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R.; Smith, D.; Schechter, L. E.; Andree, T. H. J. Med. Chem. 2004, 47, 3823–3842;
(b) Czeskis, B. A.; Clodfelter, D. K.; Wheeler, W. J. J. Labelled Compd. Radiopharm.
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12. (a) Klunder, J. M.; Onami, T.; Sharpless, K. B. J. Org. Chem. 1989, 54, 1295–1304;
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2002, 54, 29–36.
20
affording compound 24 as a yellow thick oil (0.10 g, 90%). [
a
]
þ8.8
D
(c 0.8, CHCl3); Rf 0.6 (CHCl3/CH3OH 99:1); dH (500 MHz, CDCl3) 8.38
(1H, s), 8.24 (2H, br s), 7.95 (2H, br s), 7.63 (1H, s), 7.28–7.00 (4H, m),
6.46 (1H, s), 4.27 (1H, dd, J 5.0 and 11.5), 4.18–4.09 (2H, m), 3.27–
3.01 (4H, m), 1.95–1.91 (1H, m), 0.92 (3H, d, J 3.0), 0.89 (3H, d, J 3.0);
dC (125 MHz, CDCl3) 154.2, 149.9, 144.8, 133.1, 129.8, 128.6, 128.4,
128.1, 125.5, 124.3,115.6, 111.5, 102.4, 68.8, 66.8, 57.4, 51.6, 26.8, 19.9.
IR (cmꢁ1) 3410, 3071, 2957, 1709, 1535, 1346, 1161. Anal. Calcd for
C22H26N4O7S: C, 53.87; H, 5.34%. Found: C, 53.82; H, 5.36.
14. The same sequence reaction was performed also starting from compound 5, but
the IC50 value resulted very unsatisfactory confirming the hypothesis of a bad
influence of the substituent in 6-position of indole ring.
15. The result for such compound is the only possible value because this was just
obtained for the mother solution.
4.16. Protease inhibition assay
16. (a) Wuts, P. G. M.; Greene, T. W. Greene’s Protective Groups in Organic Synthesis;
4th ed.; Wiley: New Jersey, NJ, 2007; 279–298, 416–418, 684–686, 706–771 and
references therein; (b) Miller, J. F.; Brieger, M.; Furfine, E. S.; Hazen, R. J.; Kaldor,
I.; Reynolds, D.; Sherrill, R. G.; Spaltenstein, A. Bioorg. Med. Chem. Lett. 2005, 15,
3496–3500.
Biological assays were performed measuring the increase in the
fluorescence due to the Abz-NF*-6 (Km¼37ꢂ8 M; Vmax¼690
m
ꢂ90 nmol minꢁ1 (mg protease)ꢁ1
;
Kcat¼0.29ꢂ0.03 Sꢁ1
;
Kcat/Km¼