Synthesis, biological activity and modelling studies of two novel anti HIV PR inhibitors with a thiophene containing hydroxyethylamino core

Article abstract of DOI:10.1016/j.tet.2005.04.048

An efficient method has been developed for the synthesis of a versatile intermediate bearing azido, hydroxyl and ester functions, a useful precursor for peptidomimetic compounds. The two main features for this synthesis were the use of the Sharpless asymm

Full text of DOI:10.1016/j.tet.2005.04.048

Tetrahedron 61 (2005) 6580–6589
Synthesis, biological activity and modelling studies of two novel
anti HIV PR inhibitors with a thiophene containing
hydroxyethylamino core
a
a
a
Carlo Bonini,a, Lucia Chiummiento, Margherita De Bonis, Maria Funicello,
*
Paolo Lupattelli,a Gerardina Suanno,a Federico Bertib and Pietro Campanerb
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
Received 13 January 2005; revised 15 April 2005; accepted 21 April 2005
Abstract—An efficient method has been developed for the synthesis of a versatile intermediate bearing azido, hydroxyl and ester functions, a
useful precursor for peptidomimetic compounds. The two main features for this synthesis were the use of the Sharpless asymmetric
dihydroxylation on thiophene acrylate and the subsequent regioselective ring opening by sodium azide of the cyclic sulfite. Highly
chemoselective reduction of the azido alcohol led to a key compound which was utilized for the synthesis of two analogues of commercial
anti HIV PR such as nelfinavir and saquinavir. The biological activity and molecular modelling study on these two new potential drugs have
been evaluated.
q 2005 Elsevier Ltd. All rights reserved.
1. Introduction
All the commercially available anti HIV PR drugs possess a
peptidomimetic structure based on the tetrahedral transition
state mimetic concept, in which a non-hydrolysable
hydroxyethylene or dihydroxyethylene or hydroxyethyl-
amine moiety is used as central core of the molecule
(Fig. 1).1,2
More recent therapies for AIDS1,2 are essentially directed
against the most important enzymes which regulate HIV:
reverse transcriptase (RT) and aspartyl protease (HIV PR).3
In particular, significant results have been achieved in the
development of drugs as HIV PR inhibitors, based upon
early recognition of the central role of this enzyme in viral
maturation.
HIV PR is an aspartyl protease acting as a C2 symmetric
homodimer with a single active site in the free form and
with 99 residues for each monomer. The C2 axis correlates
the catalytic aspartate (Asp-25 and Asp-250) in the active
site of the enzyme. Using the standard nomenclature, the S1
0
and S1 subsites are structurally equivalent.
Although peptide-derived compounds could be potent HIV
PR inhibitors, they are typically not suitable drug candi-
dates.4 Their use is limited by some undesirable factors such
as poor solubility, low metabolic stability towards degra-
dative enzymes and poor bioavailability after oral
ingestion.5
Figure 1.
In fact, the central core in this class of anti HIV PR
inhibitors is an isostere replacement at the scissile peptidic
bond involved in the proteolytic reaction of the precursor
polypeptide and therefore, it has been found to be essential
in all the drugs with anti HIV PR activity. The development
of such a new class of inhibitors has been also well
Keywords: HIV PR inhibitors; Thiophene; Hydroxyethylamino core;
Biological activity.
*
Corresponding author. Tel.: C39 0971 202254; fax: C39 0971 202223;
e-mail: bonini@unibas.it
0040–4020/$ - see front matter q 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2005.04.048

C. Bonini et al. / Tetrahedron 61 (2005) 6580–6589
6581
supported by the improvement in the field of peptido-
mimetic synthesis.6
hydroxyl3,4 and the same relative anti configuration at the
two vicinal stereocentres.
Currently six drugs are commercially available as HIV PR
inhibitors: these compounds are indeed furnishing good
results in AIDS therapy (together in combination with
other drugs)3b but viral resistance is often increasing due to
mutations in the protease.1,7
We therefore, found interesting a modification in the core of
both the drug candidates, with the introduction of a simple
thiophene moiety in this class of HIV PR inhibitors. With
the intention of testing the influence of a thiophene ring on
the activity of potential drugs we have therefore, defined the
first goal of our synthetic approach as the building of the
novel amino alcohol core in nelfinavir and saquinavir
structures.
The development of drug resistance and the consequent lack
of a final decisive therapy for the HIV infection, makes the
elaboration and the synthesis of innovative anti HIV PR
drugs urgent.
To achieve this objective the two designed targets
(compounds 3 and 4 in Scheme 1) could be derived, in a
retrosynthetic analysis, from the same chiral thienyl
derivative 5 which possesses the correct stereochemistry
and the appropriate functionalizations for the subsequent
elaboration to the final products.
To this aim we have been exploring the possibility of
introducing a thienyl ring in the ‘core’ of some anti HIV PR
drugs. It is known that the thienyl ring mimics the phenyl
group of phenylalanine in peptidomimetics6 in many
drugs.8–10 It was also observed that the biological half-life
of antidepressant drugs such as viloxazine became longer
when a benzene ring is replaced by thiophene. Furthermore,
a recent study on C2-symmetric diol-based HIV PR
inhibitors,11 has clearly shown that the introduction of a
thienyl ring instead of a phenyl group, greatly enhances
antiviral activity of the compound.
We have focused our attention on the structures of two of
the commercially available drugs namely nelfinavir (1)12
and saquinavir (2)13 (Fig. 2) and in this paper, we report a
short and highly stereocontrolled synthesis of modified
nelfinavir and saquinavir compounds starting from a single
thiophene containing chiral precursor, as well as the
preliminary results in the evaluation of biological activities
backed by a molecular modelling study.
Scheme 1. Retrosynthetic analysis of compounds 3 and 4.
This key chiral synthon 5 could be derived from the easily
available 3-thiophen-2-yl-acrylic acid ethyl ester 6.14 With
the crucial goal of introducing the correct chirality in the core
structure we hypothesized a new strategy never before utilised
for the synthesis of this class of HIV PR inhibitors.1,12,13
To this end, a suitable acrylic derivative, such as A, can be
easily transformed into the chiral diol B via Sharpless AD.15
The introduction of the nitrogen function through inversion
of configuration and high regioselectivity represents, in the
chosen strategy, the key step in order to obtain the highly
functionalized chiral compound C, ready for the transform-
ation into different potential analogues of known HIV PR
inhibitors (Scheme 2).
Figure 2.
2. Results and discussion
The chemical structures of nelfinavir (1) and saquinavir (2)
(Fig. 2) show the presence of a perhydroisoquinoline
0
0
(PHIQ) residue as P2 –Pn unit which seems of particular
importance for a good interaction with the corresponding
subsites of the enzyme.4
Scheme 2.
On the other hand, the P2–Pn termini of the two structures
are very different: in nelfinavir a 2-methyl-3-hydroxy-
benzoic acid residue is present at this position, while in
saquinavir a dipeptide unit (asparagine–quinaldic acid)
extends the structure toward the S3 subsite of the enzyme.4
This general synthetic strategy was first applied to the
synthesis of thiophene containing nelfinavir and saquinavir
analogues.16
Also the crucial core is slightly different for the two drugs:
in nelfinavir there is a phenyl moiety, while in saquinavir a
phenylalanine residue. However, they posses the same (R)
absolute configuration at the carbon bearing the central
3. Synthesis of the thiophene azido alcohol 5
The known (E) olefin 614 (Scheme 3), prepared from

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commercially available 2-thiophencarboxaldehyde, was
subjected to the Sharpless dihydroxylation following a
recently modified procedure.17 Further, improvements of
overall yields of the purified diol 7 have been achieved by
adding 20% more AD mix-b and ligand (66% yield and
eeO98).
Scheme 4. (a) 3-Acetoxy-2-methyl benzoic acid, DCC, DMAP, CH2Cl2, rt,
3 h; (b) (i) Pd/C, H2, MeOH, rt, 7 h; (ii) MeOH, rt, 72 h; (c) BH3$SMe2,
NaBH4, THF, rt, 2 h.
Scheme 3. (a) AD mix-b, [DHQD]2PHAL, MeSO2NH2, H2O/t-BuOH,
0–15 8C, 15 h; (b) SOCl2, Pyr, CH2Cl2, 0 8C, 15 h; (c) NaN3, DMF/CH3CN,
15 8C, 15 h.
Having built the diol moiety the subsequent step was the
introduction of the nitrogen with inversion of configuration,
that could be, in principle, better achieved from the
corresponding cyclic sulfate,18 obtained via oxidation of
the corresponding cyclic sulfite. However, the possibility to
oxidize the thienyl ring during the conversion of the sulfite
into the sulfate prompted us to directly perform the ring
opening reaction with sodium azide, using sulfite 8 as crude
material. Pleasingly, the reaction on compound 8 furnished
the isolated and purified azido alcohol 5 in 70% yield
starting from diol 7 (two steps) (Scheme 3). The reaction
was totally regio- and stereoselective as demonstrated in
comparison with known compounds.19
Scheme 5. (a) EtOAc, Pd/C, rt, 14 h. (b) (i) NaBH4, MeOH, rt, 24 h or
(ii) LiAlH4, THF, rt, 21 h or (iii) DIBAL, toluene, rt, 6 h.
good yield. However, different reaction conditions (NaBH4,
LiAlH4, DIBAL) for reduction of the carboxylic ester only
afforded degradation products or starting amino alcohol 12.
Azido alcohol 5 is a very interesting chiral synthon due to
the presence of three different functions on vicinal carbons:
an amino precursor as an azido group, a secondary hydroxyl
function and a carboxylic group.
Best results were finally obtained by planning a reduction of
the carboxylic function on the azido alcohol 5. The use of
BH3$SMe2, a known reagent for reduction of a-hydroxy
esters,23 afforded quite nicely the expected azido diol 14 in
90% yield with complete chemoselectivity (Scheme 6). This
reaction is noteworthy since it has never been applied to
such particularly functionalized compounds.
4. Synthesis of nelfinavir analogue 3
With this key compound 5 in hand we next turned our
attention to the synthesis of the nelfinavir analogue 3, with
the introduction on the hydroxyl group of a benzoic acid
derivative to obtain compound (rac)-9 (Scheme 4). The
proposed sequence was firstly achieved on racemic
compound 5. As expected, the esterification step20 pro-
ceeded smoothly in high yield (96% yield): the subsequent
reduction of azido group to the corresponding amino alcohol
with the rearrangement shift of carboxylic substituent on the
amino group21 was then achieved in good yield (90%).
To complete the functional group differentiation on
compound 14 it was necessary to regioselectively activate
the primary hydroxyl group for the subsequent introduction
of the PHIQ unit. Both activations with tosyl or mesi-
tylenesulfonyl (MesSO2, 2,4,6-trimethylbenzenesulfonyl)
chloride were tested, but much better result (80% yield)
were obtained with the more hindered mesitylene derivative
15b.
The first amino side chain was easily introduced by treating
the azido alcohols 1524 with commercially available PHIQ
thus, obtaining the product 16 in good chemical yield
(67%). The azido alcohol was quantitatively transformed
into amino alcohol 17 by hydrogenation with 5% Pd/C at rt.
However, quite disappointingly, in the last step of the
sequence to the target diol (rac)-11 all attempts to
chemoselectively reduce the ester function (by
BH3$SMe2/NaBH4 and similar reagents)22 gave nearly no
reaction. Therefore, we had to change our synthetic strategy
in order to reduce the ester function.
This compound 17 was purified either by column chroma-
tography or by crystallization and represents an important
intermediate for the preparation of possible new HIV
protease inhibitors containing a thiophene ring. In fact,
In a second approach (Scheme 5) the selective hydrogen-
ation of the azido alcohol 5 provided the amino alcohol 12 in

C. Bonini et al. / Tetrahedron 61 (2005) 6580–6589
6583
used in the preparation of saquinavir 2.1,13 The first strategy
provides initial coupling between the commercially avail-
able N-Cbz asparagine 19 and fragment 17, N-Cbz through
hydrogenation with Pd/C, and final coupling with quinaldic
acid.
Using this procedure we obtained, as expected, compound
20 (Scheme 8), but all subsequent attempts to deprotect
the amino group in 21 under various conditions failed.
Scheme 6. (a) BH3 SMe2, NaBH4, THF, MeOH, rt, 3 h; (b) (i) RZTs, TsCl,
Pyr, 50 8C, 4 h; (ii) RZSO2Mes, MesSO2Cl, Pyr, CH2Cl2, rt, 22 h;
(c) K2CO3, PHIQ, i-PrOH, 50 8C, 21 h; (d) H2, Pd/C, MeOH, rt, 3 h. MesZ
2,4,6-trimethylbenzene.
starting from compound 17, nelfinavir and saquinavir
analogues (see below) were finally prepared.
For the synthesis of thienyl nelfinavir 3, two additional steps
were required (Scheme 7). The peptide coupling of 17 with
3-acetoxy-2-methyl benzoic acid furnished compound 18.
Then final deacetylation of the phenolic group with Na/
MeOH afforded the target compound 3 in 50% overall yield
(two steps).
Scheme 8. (a) DCC, DMAP, CH2Cl2, rt, 6 h; (b) H2, Pd/C, EtOH, rt, 48 h.
Therefore, an alternative synthetic sequence was followed
starting with the preparation of dipeptide 24, obtained by
coupling of asparagine t-butyl ester 22 and quinaldic acid
23. Then dipeptide 2413c was hydrolyzed to acid 25,25 and
coupling with fragment 17 (Scheme 9) afforded the desired
product 4 in 65% yield.
Scheme 7. (a) 3-Acetoxy-2-methyl benzoic acid, DCC, DMAP, CH2Cl2, rt,
2 h; (b) Na, MeOH, rt, 1 h.
5. Synthesis of saquinavir analogue 4
Key compound 17 has also been utilized for the synthesis of
the thienyl saquinavir 4, where a dipeptide unit containing
asparagine N-bonded to quinaldic acid has to be introduced
to the P2–Pn end.
Two possible approaches could be followed for the
synthesis of saquinavir analogue 4 according to the protocol
Scheme 9. (a) CH2Cl2, TFA, rt, 6 h; (b) DCC, DMF, DMAP, CH2Cl2, rt,
7 h.

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6. Biological activity and molecular modelling studies
The activity of compounds 3 and 4 was evaluated on the
inhibition of the catalytic activity of a recombinant wild-
type HIV PR. IC50 values were obtained by measuring the
initial velocities of hydrolysis of the fluorogenic substrate
Abz-Thr-Ile-Nle-Phe(NO2)-Gln-Arg, and are reported in
Table 1, column 2, together with the IC50 values measured
in the same experiment for the reference inhibitors
nelfinavir 1 and saquinavir 2.
Table 1
Compound
IC50 (mM)
DEcompl,rela (Kcal/mol)
1
2
3
4
0.0019
0.0004
30.0
2.6
0.0
64.9
87.6
5.0
a DEcompl,relZ(EPL-E8L)–(EPsaq-E8saq), where EPL, E8L are the Amber
energies for the optimized geometries of the protein–ligand complexes
and for the ground state conformations of the free ligands, respectively.
EPsaq and E8saq are the corresponding values for the reference ligand
saquinavir.
Both candidates, 3 and 4, show a clear inhibitory activity,
with IC50 values in the micromolar range (30 and 5 mM,
respectively, Table 1), but they are much less efficient than
nelfinavir and saquinavir, which show, under our experi-
mental conditions, IC50 values in the low nanomolar range.
A preliminary molecular modelling study was carried out by
docking compound 3 onto the structure of the nelfinavir
-HIV-PR complex, while the complex of 4 was modelled
over the structure of the saquinavir-HIV-PR complex. After
a first optimization, molecular dynamics runs were carried
out for all the complexes and went to equilibration,
indicating a good fit of the inhibitors into the active site.
Final post-dynamics optimizations gave the relative com-
plexation energies reported in Table 1, column 3, and the
final geometries of the complexes. The energy differences
are largely overestimated at this stage, mainly due to the
lack of accounting for solvation of the free ligands, but
the trend of such relative complexation energies, which
are commonly used in the first ligand screening stages,26
are qualitatively in agreement with the trend of the
measured IC50.
Figure 3. RMS overlay of the optimized complexes of HIV PR with
nelfinavir and 3 (a), and saquinavir with and 4 (b).
developed, with the preparation of key chiral compounds
which can be utilized for the synthesis of different potential
HIV PR inhibitors. The two target compounds 3 and 4 have
been obtained in nine and eight steps starting from acrylate
6 with 10 and 13% in overall yields, respectively.
The biological experimental and the theoretical data
furnished information of the low activity of 3 and 4 with
respect to the unmodified molecules 1 and 2. Further studies
are in progress for improving the activity of these drugs
containing a thiophene unit.
Both the geometries of the 3 and 4 complexes, when
compared with nelfinavir and saquinavir, respectively,
suggest the same explanation for the difference of activity.
3 and nelfinavir are almost perfectly superimposable as to
the core hydroxyethylene unit and the residues at P2, P3, P20
and P30 (Fig. 3 a), and the same occurs for 4 and saquinavir
(Fig 3 b).
8. Experimental
8.1. General
Unless otherwise specified, materials were purchased from
commercial suppliers and used without further purification.
THF, toluene, diethyl ether were distilled from sodium/
benzophenone ketyl immediately before use. Dichloro-
methane was distilled from P2O5. DMF was freshly distilled
However, in both the complexes the thiophene moiety at P1
enters the S1 protease subsite less deeply, due to the lack of
the phenylalanine benzylic carbon.
˚
and stored over 4 A sieves. Moisture-sensitive reactions
were conducted in oven- or flame-dried glassware under an
argon atmosphere. All reactions were magnetically stirred
and monitored by thin-layer chromatography using pre-
coated silica gel (60 F254) plates. Column chromatography
was performed with the indicated solvents using silica gel-
7. Conclusions
In conclusion, a general strategy for the synthesis of
thiophene containing HIV PR inhibitor analogues has been
˚
60 A. Mass spectra were obtained by GC/MS with electron

C. Bonini et al. / Tetrahedron 61 (2005) 6580–6589
6585
impact ionization. NMR spectra were recorded in CDCl3
solution at rt (1H at 300 and 500 MHz and 13C at 75 and
125 MHz, respectively). Chemical shifts (d) were expressed
in ppm and coupling constant (J) in Hz. Optical rotations
were determined operating at the sodium D line at 25 8C.
HPLC analyses were conducted using Chiralcel OJ-H
column with UV detection at 235 nm.
8.1.4. 3-Acetoxy-2-methyl-benzoic acid 2-azido-1-ethoxy-
carbonyl-2-thiophen-2-yl-ethyl ester [(rac)-9]. To a
mixture of azido alcohol (rac)-5 (200 mg, 0.83 mmol),
3-acetoxy-2-methylbenzoic acid (308 mg, 1.66 mmol) and
DMAP (32 mg, 0.29 mmol) in dry CH2Cl2 (4 mL) a solution
of DCC (320 mg, 1.66 mmol) in dry CH2Cl2 (1.2 mL) was
slowly added. The reaction, monitored by TLC, was
quenched after 3 h by adding EtOH (2.4 mL) and the solid
was filtered on Buckner funnel, washed with petroleum
ether and the filtrated was treated with 10% HCl. The
organic layer was washed with saturated aqueous solution of
NaHCO3, brine and after solvent removal the crude product
was purified by column chromatography (petroleum ether/
EtOAc 8:2) affording ester (rac)-9 (333 mg, 96% yield) as a
yellow oil: RfZ0.4 (petroleum ether/EtOAc 8:2); 1H NMR
(500 MHz, CDCl3) d 7.92 (d, JZ8.3 Hz, 1H), 7.34 (m, 1H),
7.31 (m, 1H), 7.24 (d, JZ8.3 Hz, 1H), 7.20 (m, 1H), 7.00
(m, 1H), 5.66 (d, JZ15 Hz, 1H), 5.42 (d, JZ15 Hz, 1H),
4.17 (q, JZ10 Hz, 2H), 2.42 (s, 3H), 2.33 (s, 3H), 1.18
(t, JZ10 Hz, 3H); 13C NMR (125 MHz, CDCl3) d 169.3,
166.8, 165.9, 150.2, 135.6, 133.3, 130.6, 128.8, 128.6,
127.6, 127.0, 126.8, 126.7, 74.5, 62.3, 60.7, 20.9, 14.2, 13.6.
Anal. Calcd for C19H19N3O6S: C, 54.67; H, 4.59. Found: C,
54.63; H, 4.61.
8.1.1. (C)-(2S,3S)-3-(2-Thienyl)-2,3-dihydroxy ethyl
propanoate (7). Prepared according to the standard
procedure for the AD,15 using 1.68 g of AD-mix b and
adding 9.6 mg of (DHQD)2PHAL for 1 mmol of olefin 6.
The reaction residue was purified by column chromato-
graphy on silica gel (CHCl3/MeOH 98:2) to give the pure
diol 7 (142 mg, 66%) as a yellow oil: RfZ0.2 (CHCl3/
MeOH 98:2); [a]2D0 K2.7 (c 4, MeOH); eeZ99.87
(Chiralcel OJ-H, Hexane/i-PrOH 95:5, 0.5 mL/min, lZ
235 nm, tRZ59.76, tRZ65.16); 1H NMR (500 MHz,
CDCl3) d 7.32 (d, JZ4.8 Hz, 1H), 7.11 (d, JZ3.4 Hz,
1H), 7 (m, 1H), 5.27 (d, JZ1.8 Hz, 1H), 4.44 (d, JZ2.4 Hz,
1H), 4.31 (q, JZ7 Hz, 2H), 1.32 (t, JZ7 Hz, 3H), 2.85 (br s,
1H), 3.3 (br s, 1H); 13C NMR (125 MHz, CDCl3) d 172.0,
143.8, 126.8, 125.8, 125.4, 74.6, 71.1, 62.5, 14.3. EI-MS
m/z: MC, 216, (100), 113. Anal. Calcd for C9H12O4S: C,
49.99; H, 5.59. Found: C, 50.01; H, 6.02.
8.1.5. 3-(3-Acetoxy-2-methyl-benzoylamino)-2-hydroxy-
3-thiophen-2-yl-propionic acid ethyl ester [(rac)-10]. To
a solution of 10% Pd/C (67 mg) in methanol (5 mL)
previously activated in hydrogen atmosphere for 30 min, a
solution of substrate (rac)-9 (94 mg, 0.224 mmol) in MeOH
(15 mL) was added. The reaction was monitored by TLC
(petroleum ether/EtOAc 7:3) until substrate disappearance
(7 h). After hydrogen removal, the mixture was filtered and
the solution was stirred for 72 h. The solvent was evaporated
affording the pure amide (rac)-10 (285 mg, 91%) as a
yellow oil that was used without any purification: RfZ0.3
8.1.2. 2-Oxo-5-thiophen-2-yl-2l4-[1,3,2]dioxathiolane-4-
carboxylic acid ethyl ester (8). To a cold (0 8C) stirred
solution of diol 7 (21.4 mg, 0.1 mmol) in dry CH2Cl2
(0.28 mL) under nitrogen atmosphere, pyridine (22.7 mg,
0.27 mmol) and SOCl2 (0.01 mL, 0.13 mmol) were added.
After 15 h few millilitres of Hexane/EtOAc 2:1 were added,
the mixture was filtered and the residue was washed with
EtOAc. After solvent removal the brown crude product was
used in the next step without purification for the preparation
1
of compound 5. H NMR (300 MHz, CDCl3) d 7.45–6.95
1
(petroleum ether/EtOAc 7:3); H NMR (500 MHz, CDCl3)
(m, 6H), 6.43 (d, JZ7.6 Hz, 1H), 5.85 (d, JZ7.6 Hz, 1H),
5.35 (d, JZ7.6 Hz, 1H), 4.91 (d, JZ7.6 Hz, 1H), 4.40–4.25
(m, 4H), 1.45–1.30 (m, 6H); 13C NMR (75 MHz, CDCl3) d
166.0, 165.0, 138.0–125.0, 82.5, 80.9, 79.7, 82.6, 82.3, 14.1,
13.9. Anal. Calcd for C9H10O5S2: C, 41.21; H, 3.84. Found:
C, 41.19; H, 3.86.
d 9.24 (br s, 1H), 8.19 (m, 1H), 7.24 (m, 4H), 6.86 (m, 1H),
6.20 (d, JZ3.5 Hz, 1H), 5.30 (d, JZ3.5 Hz, 1H), 4.22 (q,
JZ7 Hz, 2H), 2.36 (s, 6H), 1.13 (t, JZ7 Hz, 3H); 13C NMR
(125 MHz, CDCl3) d 173.9, 172.0, 158.4, 136.2, 130.8,
129.0, 127.4, 125.7, 123.3, 122.5, 121.7, 112.9, 80.5, 61.5,
59.4, 57.9, 14.0, 13.6. Anal. Calcd for C19H21NO6S: C,
58.30; H, 5.41. Found: C, 58.32; H, 5.45.
8.1.3. (C)-(2S,3R)-3-Azido-2-hydroxy-3-thiophen-2-yl-
propionic acid ethyl ester (5). The crude sulfite 8
(26 mg, 0.1 mmol) in CH3CN (1.5 mL)/DMF (1.5 mL)
was treated with NaN3 (0.32 g, 0.5 mmol) and stirred at
15 8C for 16 h. Then a solution of H2SO4 20% (0.3 mL) was
added and the solid residue was extracted with EtOAc. The
organic layer was washed with 10% H2SO4, water, saturated
aqueous solution of NaHCO3 and brine. After drying over
Na2SO4 and solvent removal the crude product was purified
by column chromatography (hexane/EtOAc 8:2) affording
the pure azido alcohol 5 as a brown oil (17 mg, 70%): Rf1Z
0.4 (hexane/EtOAc 8:2); [a]2D0 C36.0 (c 2, MeOH); H
NMR (500 MHz, CDCl3) d 7.35 (d, JZ5.5 Hz, 1H), 7.13 (d,
JZ3.5 Hz, 1H), 7.04 (m, 1H), 5.1 (d, JZ3 Hz, 1H), 4.6 (d,
JZ3 Hz, 1H), 4.207 (q, JZ8 Hz, 2H), 3.22 (s, 1H, OH),
1.21 (t, JZ8 Hz, 3H); 13C NMR (125 MHz, CDCl3) d 170.9,
135.1, 127.6, 127.0, 127.0, 73.6, 73.4, 62.4, 14.0. EI-MS
m/z: MCKN2, 213, (100), 110. Anal. Calcd for
C9H11N3O3S: C, 44.80; H, 4.60. Found: C, 44.82; H, 4.61.
8.1.6. (K)-(2S,3R)-3-Amino-2-hydroxy-3-thiophen-2-yl-
propionic acid ethyl ester (12). To a solution of ethanol
(1.5 mL) of 10% Pd/C (15 mg), previously activated in
hydrogen atmosphere for 30 min a solution of substrate 5
(55 mg, 0.23 mmol) in EtOH (3.5 mL) was added. The
reaction was monitored by TLC (petroleum ether/EtOAc
8:2) until substrate disappearance (21 h). After hydrogen
removal, the mixture was filtered and the solvent was
evaporated in vacuo. The crude product was purified by
column chromatography (CHCl3/MeOH 9:1) affording the
amino alcohol 12 (40 mg, 80%) as a yellow solid: RfZ0.69
(CHCl3/MeOH 9:1); [a]2D0 K6.8 (c 1, MeOH); mp 50–
1
53 8C; H NMR (300 MHz, CDCl3) d 7.22 (d, JZ5.5 Hz,
1H), 6.95 (m, 2H), 4.60 (d, JZ3 Hz, 1H), 4.48 (d, JZ3 Hz,
1H), 4.15 (q, JZ7 Hz, 2H), 1.2 (t, JZ7 Hz, 3H); 13C NMR
(125 MHz, CDCl3) d 172.4, 144.1, 126.7, 125.0, 124.9,
74.7, 62.0, 54.7, 14.3. EI-MS m/z: MCKHC, 214, 112,

6586
C. Bonini et al. / Tetrahedron 61 (2005) 6580–6589
(100), 113. Anal. Calcd for C9H13N3O3S: C, 50.21; H, 6.09.
Found: C, 50.24; H, 6.10.
137.2, 132.1, 130.3, 130.9, 128.2, 128.1, 127.4, 127.2, 72.2,
69.2, 62.6, 62.3, 23.1, 22.8, 21.2. Anal. Calcd for
C16H19N3O4S2: C, 50.38; H, 5.02. Found: C, 50.41; H, 5.05.
8.1.7. (C)-(2R,3R)-3-Azido-3-thiophen-2-yl-propane-
1,2-diol (14). To a cold (0 8C) stirred solution of azido
alcohol 5 (400 mg, 1.6 mmol) in dry THF (8 mL), a solution
of BH3$SMe2 (399 mg, 3.2 mmol) in dry THF (0.32 mL)
was slowly added under argon atmosphere and then the
temperature was allowed to warm to rt. After 45 min,
NaBH4 (3.2 mg, 0.08 mmol) was added and the mixture was
stirred for 3 h. The reaction was slowly quenched adding
MeOH (1 mL) at 0 8C and stirred for 45 min. After solvent
removal the crude material was purified by column
chromatography (CHCl3/MeOH 95:5) affording the azido
diol 14 (297 mg, 90%) as a deep oil: RfZ0.3 (CHCl3/MeOH
8.1.10. (K)-(3S,4aS,8aS,20R,30R)-[(30-Azido-20-hydroxy-
30-(thiophen-2-yl)-propyl]-decahydro-isoquinoline-3-
carboxylic acid tert-butylamide (16). The same procedure
was followed for 15a and 15b. To a solution of 15a
(200 mg, 0.57 mmol) or 15b (217 mg, 0.57 mmol) in
i-PrOH (10.8 mL) PHIQ (231 mg, 0.97 mmol) and K2CO3
(157 mg, 1.14 mmol) were added. The mixture was stirred
at 50 8C for 21 h, then i-PrOH was evaporated in vacuo and
the residue dissolved in EtOAc (20 mL). The organic layer
was washed with saturated solution of NH4Cl and brine then
was dried over Na2SO4 and concentrated in vacuo. The
resulting residue was purified by chromatography (petro-
leum ether/EtOAc 7:3) to give amine 16 as a pale yellow oil
(160 mg, 67%): RfZ0.37 (petroleum ether/EtOAc 7:3);
[a]2D0 K10.2 (c 0.5, EtOAc); 1H NMR (500 MHz, CDCl3) d
7.37–6.98 (m, 3H), 6.07–6.04 (br s, 1H), 4.76–4.74 (m, 1H),
3.97–3.93 (m, 1H), 3.42–3.61 (br s, 1H), 2.99–2.95 (m, 1H),
2.72–2.70 (m, 2H), 2.40–2.31 (m, 2H), 1.93–1.27 (m, 12H),
1.35 (s, 9H); 13C NMR (125 MHz, CDCl3) d 173.4, 135.0,
127.4, 126.7, 126.6, 72.9, 70.0, 64.4, 60.1, 58.1, 50.9,
35.8, 33.2, 30.8, 30.5, 29.7, 26.0, 25.9, 20.8. EI-MS m/z:
MCKTh–CHNC3 , 280, 182, (100). Anal. Calcd for
C21H33N5O2S: C, 60.11; H, 7.93. Found: C, 60.13; H, 7.95.
1
95:5); [a]2D0 C4.9 (c 1.5, MeOH); H NMR (500 MHz,
CDCl3) d 7.4 (d, JZ5 Hz, 1H), 7.14 (d, JZ3.5 Hz, 1H),
7.08–7.06 (m, 1H), 4.91 (d, JZ6.5 Hz, 1H), 3.91 (dd, JZ
6.5, 3.5 Hz, 1H), 3.77–3.69 (m, 2H); 2.40 (s, 1H, OH); 2.06
(s, 1H, OH); 13C NMR (125 MHz, CDCl3) d 138.4, 127.8,
127.7, 127.1, 74.3, 63.1, 62.87. EI-MS m/z: MCKHC, 199,
110, (100). Anal. Calcd for C7H9N3O2S: C, 42.20; H, 4.55.
Found: C, 42.19; H, 4.57.
8.1.8. (C)-(2R,3R)-Toluene-4-sulfonic acid 3-azido-2-
hydroxy-3-thiophen-2-yl-propyl ester (15a). To a solution
of azido diol 14 (150 mg, 0.75 mmol) and pyridine (1.5 mL)
TsCl (173 mg, 0.91 mmol) in portions was added at 0 8C
under argon atmosphere, and the solution was stirred at
50 8C for 4 h. After pyridine elimination in vacuo, the crude
mixture was dissolved in EtOAc and washed with cold 2 M
HCl, water, saturated aqueous solution of NaHCO3 and
brine. The organic layer was dried over Na2SO4, concen-
trated and the mixture was purified by column chromato-
graphy (petroleum ether/EtOAc 8:2) affording product 15a
as a colourless oil (132 mg, 50%): RfZ0.3 (petroleum ether/
EtOAc 8:2); [a]D20 C93.8 (c 0.5, CHCl3); 1H NMR
(500 MHz, CDCl3) d 7.81–7.01 (m, 7H), 4.87 (d, JZ
6.5 Hz, 1H), 4.11–4.08 (m, 2H), 4.08–4.06 (m, 1H), 2.46 (s,
3H), 2.6 (s, 1H, OH); 13C NMR (125 MHz, CDCl3) d 145.6,
137.2, 132.5, 130.3, 130.2, 128.4, 128.2, 127.4, 127.2, 72.2,
70.1, 62.4, 62.2, 22.0. Anal. Calcd for C14H15N3O4S2: C,
47.58; H, 4.28. Found: C, 47.60; H, 4.26.
8.1.11. (K)-(3S,4aS,8aS,20R,30R)-[(30-Amino-20-hydroxy-
30-(thiophen-2-yl)-propyl]-decahydro-isoquinoline-3-
carboxylic acid tert-butylamide (17). Prepared as
described for 12, starting from 16 using methanol instead
of ethanol. After column chromatography (CHCl3/MeOH
9:1) compound 17 (91%) was obtained as a deep brown oil:
RfZ0.3 (CHCl3/MeOH 9:1); [a]2D0 K76.9 (c 0.55, MeOH);
1H NMR (500 MHz, CDCl3) d 7.25–7.24 (m, 1H), 6.98–
6.97 (m, 2H), 6.34–6.32 (br s, 1H, NH), 4.32–4.29 (m, 1H),
3.93–3.89 (m, 1H), 3.07–3.00 (m, 1H), 2.70–2.20 (m, 4H),
2.10–0.90 (m, 15H), 1.36 (s, 9H); 13C NMR (125 MHz,
CDCl3) d 173.4, 145.4, 126.5, 124.8, 124.8, 73.1, 69.8, 59.4,
59.1, 56.1, 50.7, 35.6, 33.2, 30.5, 30.5, 29.7, 25.9, 25.9,
20.8. Anal. Calcd for C21H35N3O2S: C, 64.08; H, 8.96.
Found: C, 64.10; H, 8.94.
8.1.9. (C)-(2R,3R)-2,4,6-Trimethyl-benzenesulfonic acid
3-azido-2-hydroxy-3-thiophen-2-yl-propyl ester (15b).
To a cold (0 8C) stirred solution of azido diol 14 (280 mg,
1.43 mmol) in CH2Cl2 (3.7 mL), pyridine (236 mg,
2.86 mmol) and mesitylenesulfonyl chloride (344 mg,
1.57 mmol) were added. The reaction mixture was stirred
at 0 8C for 4 h and then was warmed at 20 8C for 20 h. The
solvent was eliminated in vacuo, the crude mixture was
dissolved in EtOAc and washed with cold 2 M HCl (5 mL),
water, saturated aqueous solution of NaHCO3 and brine.
The organic layer was dried over Na2SO4, concentrated and
the crude product was purified by column chromatography
(petroleum ether/EtOAc 8:2) affording the pure compound
15b (430 mg, 80%) as a deep yellow oil: RfZ0.5 (petroleum
8.1.12. (K)-(3S,4aS,8aS,10R,20R)-[Acetic acid 3-[30-(3-
tert-butylcarbamoyl-decahydro-isoquinolin-2-yl)-20-
hydroxy-10-thiophen-2-yl-propylcarbamoyl]-2-methyl-
phenyl ester (18). Prepared as described for (rac)-9, using
alcohol 17, 1.01 equiv of acid, 1.3 equiv of DCC. The crude
product was purified by column chromatography (petroleum
ether/EtOAc 7:3) affording amide 18 (62% yield) as a
viscous white oil: RfZ0.4; [a]2D0 K27.2 (c 0.5, CHCl3); 1H
NMR (500 MHz, CDCl3) d 7.35–7.33 (m, 1H), 7.26–7.20
(m, 2H), 7.16–7.13 (m, 1H, NH), 7.12–7.11 (m, 1H), 7.08–
7.06 (m, 1H), 6.99–6.97 (m, 1H), 6.00 (br s, 1H, NH), 5.49–
5.46 (m, 1H), 4.22–4.19 (m, 1H), 2.75–2.60 (m, 3H), 2.30
(s, 3H), 2.2 (s, 3H), 1.9–1.00 (m, 15H), 1.33 (s, 9H); 13C
NMR (125 MHz, CDCl3) d 173.6, 168.9, 168.3, 149.6,
140.7, 139.2, 128.6, 126.7, 126.5, 126.4, 125.2, 124.6,
123.5, 71.1, 69.7, 59.3, 59.1, 53.0, 49.9, 35.5, 33.2, 30.6,
30.3, 28.5, 25.7, 25.5, 24.8, 20.8, 12.8. Anal. Calcd for
C31H43N3O5S: C, 65.35; H, 7.61. Found: C, 65.37; H, 7.59.
1
ether/EtOAc 8:2); [a]2D0 C81.3 (c 0.5, EtOAc); H NMR
(500 MHz, CDCl3) d 7.36 (d, JZ4.5 Hz, 1H), 7.10 (d, JZ
3.5 Hz, 1H), 7.04–7.03 (m, 1H), 6.99 (s, 2H), 4.89 (d, JZ
6.5 Hz, 1H), 4.06–4.04 (m, 3H), 2.75 (br s, 1H), 2.63 (s,
6H), 2.33 (s, 3H); 13C NMR (125 MHz, CDCl3) d 144.0,

C. Bonini et al. / Tetrahedron 61 (2005) 6580–6589
6587
8.1.13. (K)-(3S,4aS,8aS,20R,30R)-2-[20-Hydroxy-30-(3-
hydroxy-2-methyl-benzoylamino)-30-(thiophen-2-yl)-
propyl]-decahydro-isoquinoline-3-carboxylic acid tert-
butylamide (3). To a solution of amino alcohol 18
(30 mg, 0.053 mmol) in MeOH (1 mL) a catalytic amount
of sodium was added and after 45 min the reaction was
stopped by solvent removal. The crude mixture dissolved in
EtOAc (10 mL) was washed with water and the aqueous
layers extracted twice with EtOAc (10 mL). The combined
organic layers were dried over Na2SO4, filtered and
concentrated in vacuo. The mixture was purified by column
chromatography (EtOAc/petroleum ether 7:3) affording
amide 3 (22 mg 81%): RfZ0.46; [a]2D0 K27.0 (c 0.6,
EtOAc); 1H NMR (500 MHz, CDCl3) d 7.26–7.24 (m, 1H),
7.13–7.12 (m, 1H), 7.05–7.02 (m, 1H, NH), 6.99–6.98 (m,
2H), 6.90–6.85 (m, 2H), 6.05 (br s, 1H, NH), 5.47–5.43 (m,
1H), 4.22–4.19 (m, 1H), 2.68–1.20 (m, 19H), 2.14 (s, 3H),
1.34 (s, 9H); 13C NMR (125 MHz, CDCl3) d 179.0, 169.8,
155.1, 140.2, 137.9, 127.3, 126.8, 125.8, 122.6, 119.0,
116.9, 71.2, 70.1, 60.0, 59.4, 53.1, 51.5, 35.9, 33.4, 30.9,
30.7, 29.9, 28.8, 26.2, 25.9, 21.3, 21.1, 12.6. Anal. Calcd for
C29H41N3O4S: C, 66.00; H, 7.83. Found: C, 66.03; H, 7.82.
on silica gel (CHCl3/MeOH 95:5) affording compound 4
(43 mg, 65%) as a paled yellow oil: RfZ0.35 (CHCl3/
MeOH 95:5); [a]D20 C21.4 (c 0.65, CHCl3); 1H NMR
(500 MHz, CD3OD) d 9.26 (d, JZ7 Hz, 1H), 8.26 (d, JZ
8 Hz, 1H), 8.19 (d, JZ8.5 Hz, 1H), 8.15 (d, JZ9 Hz, 1H),
7.85 (d, JZ8 Hz, 1H), 7.79–7.74 (m, 2H), 7.64–7.61 (m,
1H), 7.16 (d, JZ5 Hz, 1H), 7.08 (d, JZ3 Hz, 1H), 6.90 (t,
JZ4 Hz, 1H), 6.41 (s, 1H), 6.19 (s, 1H), 5.82 (s, 1H), 5.31–
5.30 (m, 1H), 5.06–5.04 (m, 1H), 4.26–4.24 (m, 1H), 3.04–
2.90 (m, 3H), 2.68–2.59 (m, 2H), 2.32–2.22 (m, 2H), 1.85–
1.66 (m, 6H), 1.62–1.58 (m, 1H), 1.48–1.37 (m, 6H), 1.33
(s, 9H); 13C NMR (125 MHz, CD3OD) d 174.0, 173.2,
169.9, 165.0, 149.1, 146.8, 139.6, 137.7, 130.3, 130.3,
129.6, 128.4, 127.8, 127.1, 126.8, 125.9, 118.9, 71.4, 69.6,
59.9, 53.9, 53.8, 50.9, 50.7, 50.6, 38.5, 35.6, 33.4, 30.8,
30.7, 30.0, 28.9, 28.9, 26.2, 21.2. Anal. Calcd for
C35H46N6O5S: C, 63.42; H, 6.99. Found: C, 63.40; H, 6.95.
8.1.17. Enzyme assays. Recombinant HIV-1 protease was
purchased from Bioczech (Prague, Czech Republic); the
fluorogenic substrate Abz-Thr-Ile-Nle-Phe(NO2)-Gln-Arg
was purchased from Bachem (Bubendorf, Switzerland). The
extent of inhibition of the activity of the aspartic protease
was evaluated as previously reported for other series of
mono- and dihydroxy pseudopeptide HIV-1-PR inhibi-
tors.27 The assays were performed at 25 8C in 400 mM
NaCl, 1 mM EDTA, 1 mM DTT, 100 mM MES-NaOH
buffer at pH 5.5, and 30 mM fluorogenic substrate. Titration
of HIV PR was carried out with two reference inhibitors
structurally related to our series, namely nelfinavir and
saquinavir, using the equation IC50Z[E]/2CKi(1C[S]/
Km)2, where Ki are the literature values of the inhibition
constants for nelfinavir and saquinavir,28 and Km was
25 mM, as determined under our conditions. The active
enzyme concentration in the assay, [E], was estimated to be
3.6G2.3 nM.
8.1.14. (K)-(3S,4aS,8aS,20R,30R)-{1-[3-(3-tert-Butyl-
carbamoyl-decahydro-isoquinolin-2-yl)-2-hydroxy-1-
thiophen-2-yl-propylcarbamoyl]-2-carbamoyl-ethyl}-
carbamic acid benzyl ester (20). Prepared as described for
18 starting from 19 affording after purification by column
chromatography (CHCl3/MeOH 98:2) compound 20
(68 mg, 80%) as a yellow oil: RfZ0.2 (CHCl3/MeOH
98:2); [a]2D0 K41.0 (c 1.5, MeOH); H NMR (500 MHz,
1
CDCl3) d 7.56 (m, 1H), 7.36 (br s, 5H), 7.19 (m, 1H), 7.00
(m, 1H), 6.91 (m, 1H), 6.27 (m, 2H), 6.09 (s, 1H), 5.65 (s,
1H), 5.23 (m, 1H), 5.08 (s, 2H), 4.61 (m, 1H), 4.16 (m, 1H),
2.81 (m, 2H), 2.55 (m, 3H), 2.33 (m, 2H), 1.55 (m, 12H),
1.34 (s, 9H); 13C NMR (75 MHz, CDCl3) d 174.0, 173.5,
170.0, 156.4, 139.6, 136.3, 128.7, 128.6, 128.2, 128.1,
127.5, 126.8, 126.6, 125.8, 71.3, 69.6, 67.2, 58.9, 53.4, 51.8,
49.2, 38.2, 35.5, 34.0, 33.2, 30.6, 30.5, 28.7, 26.4, 25.9,
25.6, 25.0, 21.4. Anal. Calcd for C33H47N5O3S: C, 61.75; H,
7.38; Found C, 61.78; H, 7.40.
8.1.18. Molecular modelling. Calculations were carried out
on two Silicon Graphics Octane 1 R12000 workstations and
on a Pentium4 2.53GHz/Red Hat Linux machine. The
Cornell version of the Amber force field29 as implemented
in Sybyl6.8 (Tripos Inc.) was used in all energy minimiz-
ations and dynamics runs. New Amber parameters for the
heterocyclic systems of the ligands were developed
according to Geremia and Calligaris30 or following the
original Amber protocol. The required ab initio calculations
on the ligands were carried out with Gaussian03.31 All the
energy minimizations were carried out until a convergence
8.1.15. (2S)-2-[(Quinoline-2-carbonyl)-amino]-suc-
cinamic acid (25). To a stirred cold (0 8C) solution of 24
(100 mg, 0.29 mmol) in CH2Cl2 (1 mL) TFA (0.4 mL)
dropwise was added. The solution was warmed to 25 8C and
after 6 h the solvent was evaporated in vacuo. The product
25 was obtained after crystallization by MeOH/EtOAc
(66 mg, 80%). The compound shows the same spectro-
scopic data as reported in the literature.24
˚
criterion of 0.001 Kcal/mol/A was achieved for all the
energy gradients. The conjugate gradient minimisation
algorithm was always used after running 20 initial steps of
Simplex linear minimization. All the calculations were
carried out in a continuum dielectric of relative permittivity
3Z4rij.32
8.1.16. (C)-N1-(3S,4aS,8aS,10R,20R)-[3-(3-tert-Butyl-
carbamoyl-decahydro-isoquinolin-2-yl)-2-hydroxy-1-
thiophen-2-yl-propyl]-(2S)-2-[(quinoline-2-carbonyl)-
amino]-succinamide (4). To a stirred solution of 17 (40 mg,
0.1 mmol) and DMAP (4.4 mg, 0.035 mmol) in CH2Cl2
(1.5 mL), a solution of 25 (34 mg, 0.12 mmol) in DMF
(0.1 mL) and a solution of DCC (27 mg, 0.13 mmol) in
CH2Cl2 (0.2 mL) were added. The solution was stirred at rt
for 7 h, diluted with CH2Cl2 and washed with 10% HCl,
water, saturated aquoeus solution of NaHCO3 and brine.
The organic layer was dried over Na2SO4, concentrated and
the crude mixture was purified by column chromatography
The structures of the ligands were built with standard bond
lengths and angles, and optimized first at the RHF-6.31G*
level, in order to obtain a charge distribution comparable to
that used in the AMBER parameterization. The ligands were
then submitted to an extensive conformational search driven
by a genetic algorithm operating on all the routable bonds.33
The AMBER energies of the absolute conformational

6588
C. Bonini et al. / Tetrahedron 61 (2005) 6580–6589
9. Zhang, A.; Zhou, G.; Rong, S.-B.; Johnson, K. M.; Zang, M.;
Kozikowski, A. P. Bioorg. Med. Chem. Lett. 2002, 12, 993.
10. Uckun, F. M.; Pendergrass, S.; Maher, D.; Zhu, D.; Tuel-
Ahlgren, L.; Mao, C.; Venkatachalam, T. K. Bioorg. Med.
Chem. Lett. 1999, 9, 3411.
minima thus, obtained were taken as reference energies of
the free ligands.
The crystallographic coordinates of two reference com-
plexes of HIV PR with saquinavir and nelfinavir were
obtained from the Protein Data Bank, Brookhaven National
Laboratory (Saquinavir: Pdb id 1C6Z;34 nelfinavir: Pdb id
1OHR).35 All the hydrogen atoms were added, assuming an
environment pH of 5.5, and a single proton to be shared
between the two catalytic aspartic residues.36 All the
crystallization water molecules were removed, with the
exception of the essential water always present inside
the HIV PR binding site complexed with non ureidic
inhibitors, and hydrogen bonded to Ile50 and 500 residues.
The reference complexes were then allowed to relax by the
multistage minimization procedure described by Levit and
Lifson.37
11. In this paper the authors described the effects on the
substitution of the oxyalkyl and oxyaryl P1/P10 groups with
thioalkyl and thioaryl groups. Muhlman, A.; Classon, B.;
Haliberg, A.; Samuelsson, B. J. Med. Chem. 2001, 44, 3402.
12. For reported syntheses of Nelfinavir see: (a) Kaldor, S. W.;
Kalish, V. J.; Davies, J. F., II; Shetty, B. V.; Fritz, J. E.; Appelt,
K.; Burgess, J. A.; Campanale, K. M.; Chirgadze, N. Y.;
Clawson, D. K.; Dressman, B. A.; Hatch, S. D.; Khalil, D. A.;
Kosa, M. B.; Lubbehusen, P. P.; Muesing, M. A.; Patick,
A. K.; Reich, S. H.; Su, K. S.; Tatlock, J. H. J. Med. Chem.
1997, 40, 3979. (b) Rieger, D. L. J. Org. Chem. 1997, 62, 8546.
(c) Inaba, T.; Birchler, A. G.; Yamada, Y.; Sagawa, S.;
Yokota, K.; Ando, K.; Uchida, I. J. Org. Chem. 1998, 63,
7582. (d) Zook, S. E.; Busse, J. K.; Borer, B. C. Tetrahedron
Lett. 2000, 41, 7017. (e) Inaba, T.; Yamada, Y.; Abe, H.;
Sagawa, S.; Cho, H. J. Org. Chem. 2000, 65, 1623. (f) Albizati,
K. F.; Babu, S.; Birchler, A.; Busse, J. K.; Fugett, M.; Grubbs,
A.; Haddach, A.; Pagan, M.; Potts, B.; Remarchuk, T.; Rieger,
D.; Rodriguez, R.; Shanley, J.; Szendroi, R.; Tibbetts, T.;
Whitten, K.; Borer, B. V. Tetrahedron Lett. 2001, 42, 6481.
(g) Ma, D.; Zou, B.; Zhu, W.; Xu, H. Tetrahedron Lett. 2002,
43, 8511. (h) Ikunaka, M.; Matsumoto, J.; Fujima, Y.;
Hirayama, Y. Org. Proc. Res. Dev. 2002, 6, 49.
The initial geometries of the ligand-HIV PR complexes
were built by docking the ligand structures onto the
structures of nelfinavir (for compound 3) and saquinavir
(for compound 4) bound to HIV PR as in the optimized
reference complexes. The structure were first optimized by
˚
running an energy minimisation to a 0.05 Kcal/mol/A
energy gradient. The models were then subjected to a
molecular dynamics run in the NTV ensemble; the systems
were gradually heated to 300 K, in three steps, allowing a
25 psec interval per each 100 K, then equilibrated for
25 psec at 300 K, and finally, submitted to a 400 psec
collection run at 300 K. The lowest potential energy
equilibrium geometries were finally re optimized.
13. For reported syntheses of Saquinavir see: (a) Thompson, W. J.;
Ghosh, A. K.; Holloway, M. K.; Lee, H. Y.; Munson, P. M.;
Schwering, J. E.; Wai, J.; Darke, P. L.; Zugay, J.; Emini, E. A.;
Schleif, W. A.; Huff, J. R.; Anderson, P. S. J. Am. Chem. Soc.
1993, 115, 801. (b) Parkes, K. E. B.; Bushnell, D. J.; Crackett,
P. H.; Dunsdon, S. J.; Freeman, A. C.; Gunn, M. P.; Hopkins,
R. A.; Lambert, R. W.; Martin, J. A.; Merrett, J. H.; Redshaw,
S.; Spurden, W. C.; Thomas, G. J. J. Org. Chem. 1994, 59,
3656. (c) Fassler, A.; Bold, G.; Capraro, H.-G.; Cozens, R.;
Mestan, J.; Poncioni, B.; Rosel, J.; Tintelnot-Blomley, M.;
Lang, M. J. Med. Chem. 1996, 39, 3203. (d) Gohring, W.;
Gokhale, S.; Hilpert, H.; Roessler, F.; Schlageter, M.; Vogt, P.
Chimia 1996, 50, 532. (e) Ghosh, A. K.; Hussain, K. A.;
Fidanze, S. J. Org. Chem. 1997, 62, 6080.
Acknowledgements
The authors gratefully acknowledge MIUR (Ministry of
Instruction, University and Research-Rome, Project FIRB:
RBNE017F8N), University of Basilicata and ‘Centro di
Eccellenza in Biocristallografia’ of University of Trieste for
financial support.
14. (a) Tay, M. K.; About-Joudet, E.; Callignon, N.; Teulade,
M. P.; Savignac, P. Synth. Commun. 1988, 18, 1349.
(b) Kosakara, A.; Izumi, T.; Ogihara, T. J. Heterocycl.
Chem. 1989, 26, 597. (c) Rossi, R.; Carpita, A.; Ciofalo, M.;
Lippolis, V. Tetrahedron 1991, 47, 8443. (d) Guncheal, K.;
Don Gyu, L.; Sukbok, C. Bull. Korean Chem. Soc. 2001, 22,
943.
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