Design, synthesis and evaluation of peptidomimetics based on substituted bicyclic 2-pyridones-Targeting virulence of uropathogenic E. coli

Article abstract of DOI:10.1016/j.bmc.2006.07.017

Substituted bicyclic 2-pyridones, termed pilicides, are dipeptide mimetics that prevent pilus assembly in uropathogenic Escherichia coli. Here, we apply rational design to produce four classes of extended peptidomimetics based on two bioactive 2-pyridones

Full text of DOI:10.1016/j.bmc.2006.07.017

Bioorganic & Medicinal Chemistry 14 (2006) 7563–7581
Design, synthesis and evaluation of peptidomimetics based
on substituted bicyclic 2-pyridones—Targeting virulence
of uropathogenic E. coli
a,
Veronica Aberg, Magnus Sellstedt,a, Mattias Hedenstrom,a Jerome S. Pinkner,b
˚
¨
Scott J. Hultgrenb and Fredrik Almqvista,*
a
˚
˚
Organic Chemistry, Department of Chemistry, Umea University, SE-90187 Umea, Sweden
bMolecular Microbiology, Washington University in St. Louis School of Medicine, St. Louis, MO 63110, USA
Received 17 March 2006; accepted 3 July 2006
Available online 14 August 2006
Abstract—Substituted bicyclic 2-pyridones, termed pilicides, are dipeptide mimetics that prevent pilus assembly in uropathogenic
Escherichia coli. Here, we apply rational design to produce four classes of extended peptidomimetics based on two bioactive 2-pyri-
dones. The key intermediate in the synthesis was an amino-functionalised 2-pyridone scaffold, which could be obtained via a mild
and selective nitration and subsequent reduction. Procedures were then developed to further derivatize this amino-substituted core
and a total of 24 extended peptidomimetics were synthesised and evaluated for chaperone affinity and in vivo antivirulence activity
in P pili producing E. coli. Enhanced affinities for the target protein were observed within the generated set of compounds, while the
ability to prevent pilus assembly in vivo was significantly decreased compared to the parent lead compounds. The results suggest
that the limited in vivo potencies of the analogues are either uptake/distribution related or due to loss in binding specificity.
Ó 2006 Elsevier Ltd. All rights reserved.
1. Introduction
folds, have previously been described in the literature
(1–3, Fig. 1).1–4,12,13 The saturated counterparts to these
bicyclic systems are also useful as b-turn mimetics,13,14
and synthetic procedures to these derivatives have been
presented.15
Peptidomimetics can offer advantages as therapeutics in
terms of pharmacokinetic properties and may also ben-
efit from conformational constraints compared to natu-
ral peptides. Substituted 2-pyridone scaffolds have
previously been reported to serve as useful peptidomi-
metics for this purpose. For example, 2-pyridone based
peptidomimetics inhibit hepatitis C virus (HCV) NS3
serine protease in an extended b-sheet conformation.1
They also act as inhibitors of human rhinovirus
(HRV) 3C protease.2–4 In general, ring-fused 2-pyridone
frame-works are also present in numerous of com-
pounds with diverse biological application areas such
as ACE-inhibitors,5,6 anticancer agents7,8 and inhibitors
of Ab-peptide aggregation.9–11
We have earlier reported short and efficient syntheses of
the bicyclic 2-pyridones 516–18 and in this paper the
diversity of this scaffold is further increased via amino-
substitutions to 4 (Fig. 1).
When comparing our synthetic pathway to the ones
previously published, two important aspects should
be noted that can be advantageous. First, in 1–3 the sub-
stituent in position 6 is derived from the 2-pyridone
building blocks and the amine functionality is then
obtained by sequential substituent transformations.
Opposed to this, the amine-functionality in 4 is intro-
duced via substitution of the 2-pyridone scaffold 5
(Fig. 1). Hence, our pathway offers the possibility to
choose whether position 6 should be substituted or not
and, importantly, this position can be functionalised
via, for example, electrophilic aromatic substitutions.
Second, our synthesis to 5 provides a substituent in po-
sition 7 (R2, Fig. 1) directly in the ring formation step of
Synthetic routes to several amino-substituted bicyclic 2-
pyridones, which can function as peptidomimetic scaf-
Keywords: 2-Pyridone; Peptidomimetic; Antimicrobial; Pili.
*
Corresponding author. Tel.: +46 907866925; fax: +46 90138885;
e-mail: fredrik.almqvist@chem.umu.se
These two authors contributed equally to this work.
0968-0896/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2006.07.017

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
7564
1
1
R
O
R
R
R
1
8
2
2
7
9
R
R
S
S
S
2
5
N
N
N
R
N
N
4
6
3
H N
2
H N
2
H N
2
N
H
CO H
CO H
2
CO H
CO H
CO Me
2
2
2
2
O
O
O
O
1
2
3
4
5
Figure 1. Synthesis of 2-pyridone based dipeptide mimetics 1,1 2,2–4 and 312 have previously been reported and here we present syntheses to 4 from 5.
the 2-pyridone. This can be beneficial since position 7 is
not as easily accessible for substitutions as position 6
and 8. If desired, R1 (position 8) can be left unsubstitut-
ed and since R1 and R2 are derived from simple starting
material—nitriles and carboxylic acids, respectively—
our pathway offers great variability in the substitution
pattern. In comparison, amino-substituted 1–3 has so
far been reported with substituents only in position 6
and 8 of the 2-pyridone ring.
affinity for the chaperones PapD and FimC that are
involved in the formation of P pili and type 1 pili,
respectively.18,19 Furthermore, pilus biogenesis in E. coli
could be blocked by the pilicides.21 Importantly, and in
agreement with the theorized binding site, both a hydro-
phobic substituent in R2 and the carboxylate of these
bicyclic 2-pyridones have been shown to be important
for the activity (see Fig. 2).18,21
While this work was ongoing, binding studies with 15N
NMR spectroscopy pointed towards two plausible chap-
erone binding sites. The designed subunit binding site was
still valid but a second possible binding site near the so-
called F1-G1 loop of the chaperone was also revealed.31
Thus, the pilicides could be carboxylate-anchored in the
chaperone cleft according to the design, and/or bind in
the loop-region. In both cases pilus assembly could be
disrupted since the loop region is known to be involved
in the uncapping and delivery of the subunits at the outer
membrane assembly site, that is, the usher.32
We have previously applied the synthesis of 5 to produce
rigid dipeptide mimetics 7 (R3=H) of a C-terminal pep-
tide 6 (Fig. 2).16–21 This class of compounds interferes
with the formation of virulence associated organelles,
pili/fimbriae, in uropathogenic E. coli and is thus
referred to as pilicides. Uropathogenic E. coli (UPEC)
express two types of pili responsible for urinary tract
infections (UTIs). Type 1 pili are involved in bladder
infections (cystitis), whereas P pili mediate infection of
the upper urinary tract and kidneys (pyelonephritis).23–
25 The pathway by which pili are formed has been stud-
ied and described in great detail.26 Pili consist of a num-
ber of subunit proteins that are assembled via the highly
conserved chaperone–usher pathway. The subunits are
unstable in their monomeric forms and cannot be incor-
porated into the pilus rod without the aid of periplasmic
chaperones. Hence, by interfering with the vital chaper-
one function pilus assembly would be disrupted.22,27
In this paper, we investigate the possibilities to improve
the potency of the dipeptide mimicking pilicides by
developing synthetic pathways to highly substituted
extended peptidomimetics via R3 (Fig. 2). The intention
was also to establish structure–activity relationships that
could be used in future design and to shed light on the
two possible binding sites. The synthesised compounds
have been evaluated for chaperone affinity and in vivo
antivirulence activity in P pili producing E. coli.
The 2-pyridone pilicides target these chaperones and
were initially designed and synthesised to mimic the
C-terminus of the subunit PapG, which is illustrated in
Figure 2.19 We envisioned that they would competitively
inhibit the highly conserved subunit binding site of the
chaperones and thus prevent subunit binding29,30 and
thereby pilus assembly. The pilicides indeed displayed
2. Design and synthesis
Besides the general attention that is paid to the chemis-
try and biological applications of 2-pyridone containing
a
b
2
Anchored to Arg8 and Lys112
Conserved hydrophobic motif
Hydrophobic R and
carboxylate important for activity
Me
Me
1
R
2
3
R
R
S
O
H
O
H
N
N
N
H
N
N
N
H
CO H
2
CO H
O
O
O
2
O
Me Me
OH
3
R
provide extended mimetics
Lys110
Gln108
6
7
PapG C-terminus
Val310-Leu311-Ser312-Phe313-Pro314
2-pyridones as PapG mimetics
Figure 2. (a) Crystal structure28 of chaperone PapD and PapG peptide. (b) Bicyclic 2-pyridones 7 were designed to inhibit chaperones as C-terminal
mimetics of the pilus adhesin PapG 6. Hydrogen bonds are illustrated as ----.

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
7565
structures, we were particularly interested in obtaining
structure–activity relationships for the designed pepti-
domimetics as chaperone inhibitors and pilicides. Two
bioactive pilicides 8 and 9 (Fig. 3) were chosen for deriv-
atization into four classes of extended peptidomimetics.
A retrosynthetic analysis of the four classes A–D is sum-
marized in Figure 4.
Assuming that the pilicides share the binding mode of
the PapG C-terminus, we aimed for increased potency
by producing extended peptidomimetics with enhanced
resemblance to the PapG peptide (Fig. 2). Thus, the
obvious choice of tripeptidomimetics A was with serine
incorporated in the corresponding position to Ser312 in
PapG (Figs. 2 and 4).
Encouragingly, computational analysis using the soft-
ware MOE33 confirmed that a serine, substituted deriv-
ative of 8 was able to adopt the bound conformation of
the PapG peptide (Fig. 5 and Section 5).
Figure 5. The bioactive conformation of the C-terminus of PapG can
be adopted by a serine substituted derivative of 2-pyridone 8 in class A
(i.e., compound 34, Scheme 2).
Since the side chain of Ser312 is solvent exposed28 and
does not interact with PapD, glycine and alanine were
also included in these initial structure–activity studies
(A, Fig. 4). Alanine and glycine substituents would also
keep the molecular weight reasonably drug-like.34 Acet-
ylated amino acids were preferred since they best mimic
the features of a peptide sequence and also to avoid
zwitterionic compounds.
The crystal structure shows that the amide nitrogen in
Phe313 of the PapG peptide (corresponding to the aryl
amide nitrogen in A) hydrogen bonds to the carbonyl
oxygen of Lys110 in PapD. Moreover, the carbonyl oxy-
gen of Leu311 in PapG (corresponding to the acetyl car-
bonyl in A) is a hydrogen bond acceptor to the amide
nitrogen of Lys110 in PapD (Fig. 2).28
To further investigate the importance of the amide in R3
as well as its hydrogen bonding properties, classes B–D
were synthesised. The amides in C resemble the back-
bone of PapG, while in class D, the direction of the
amide bond is reversed. The sulfonamides B were con-
sidered attractive synthetic targets from a structure–
activity point of view, and also due to their increased
metabolic stability compared to amides. Classes B–D
comprised both isopropyl- and methyl-derivatives to
take steric effects into account and class C also includes
two additional formamide analogues.
According to the design, the amino acid substituents
would enhance binding to PapD by participating in
two hydrogen bond interactions to Lys110 (see Fig. 2).
S
S
N
N
CO H
2
CO H
2
O
O
8
9
The various amides in B–D could contribute to impor-
tant structure–activity relationships for future library
synthesis and perhaps also to clarify the binding mode
Figure 3. 2-Pyridones 8 and 9 prevent pilus assembly in uropathogenic
Escherichia coli and were chosen for further derivatization into
extended peptidomimetics.
O
1
1
R
R
H
N
3
3
S
S
N
A
B
R
R
=
=
N
H
O
R
A-C
N
H N
2
O N
2
6 compounds
CO Me
CO Me
2
2
O
O
1
1
O
O
R
R
E
F
S
R
N
S
S
H
N
N
3
4 compounds
O
R
CO Li
2
CO Me
2
O
O
1
1
3
3
1
R
N
R
R
C
D
R
R
=
=
10: R = cyclopropyl
H
1
D
11: R = phenyl
S
S
6 compounds
HO
N
H
O
N
H
N
CO Me
CO Me
2
2
R
O
O
O
O
G
H
4 compounds
Figure 4. Retrosynthetic analysis of the four classes of extended peptidomimetics A–D.

˚
V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
7566
of the pilicides. On this basis, the amino- and nitro-
substituted precursors E and F were also hydrolysed
and included in the biological evaluations.
of the nitrated compounds 20 and 21 with zinc in acetic
acid41 yielded amines 22 and 23 (81–90%, Scheme 1),
which served as precursors in the following backbone
elongations. The nitro- and amino-substituted methy-
lesters (20–23) were also hydrolysed to their correspond-
ing lithium carboxylates (24–27) and evaluated as
pilicides. The hydrolysis was performed with LiOH in
MeOH:THF.
Classes A–C could be synthesised from the amino-
substituted scaffolds E, which were derived from 10
and 11 via nitration followed by reduction to amine
(Fig. 4). Class D was synthesised via a three-step proce-
dure with formylation, oxidation and amine coupling.
Compounds 10 and 11, that is, the corresponding meth-
yl esters to bioactive 8 and 9 (Fig. 3) were synthesised in
four steps according to our previously published proce-
dures.16–18
2.2. Tripeptidomimetics via couplings to aminated
scaffold (A)
The amino-substituted intermediates 22 and 23 were
transformed into tripeptidomimetics 34–39 by couplings
of amino acids serine, glycine and alanine with DIC and
HOAt, followed by hydrolysis (Scheme 2). The cou-
plings proceeded smoothly in high yields (79–95%) in
spite of the relatively large naphthyl substituent on the
2-pyridone ring. Not surprisingly, the tert-butyl-protect-
ed serine being the most sterically demanding amino
acid also needed the longest reaction time (29, Scheme
2). In this case, some starting material still remained
after 48 h when work-up was performed. Cleavage of
the tert-butyl-protected 28 and 29 was carried out with
TFA and all methyl esters were hydrolysed as previously
described to yield the desired tripeptidomimetics 34–39
(Scheme 2).
2.1. Amino-substitution of the scaffold (E)
Amino-substituted 2-pyridones are often synthesised
using traditional nitration conditions with HNO3/
H2SO435 or HNO3/AcOH and heat36 followed by reduc-
tion of the obtained nitro-derivative to the amine with,
36,37
for example, SnCl2
or hydrogenation.35,38,39 To
avoid the harsh nitration conditions, an alternative pro-
cedure via a nitroso-derivative was planned. Thus, con-
ditions that had previously been used for nitrosation of
anisole40 with NaNO2 in CH2Cl2/TFA were slightly
modified and applied on a small-scale reaction of 10
(ꢀ10 mg). Unexpectedly, the main product turned out
to be the nitro-product 20 rather than the nitrosated
derivative (Scheme 1). Complete conversion into the ni-
tro-product could be obtained in larger scale (ꢀ500 mg),
provided that oxygen gas was supplied to the reaction.
Under nitrogen atmosphere the large-scale reaction
was sluggish and yielded byproducts. Based on this
observation, selective and mild nitrations into 20 and
21 (84–89%) could be performed under oxygen atmo-
sphere using NaNO2 in CH2Cl2/TFA.
2.3. Backbone extensions by acylations and sulfonylations
(B and C)
Formamide, acetamide and isobutyramide derivatives
40–45 were synthesised by direct acylations of the ami-
nated scaffolds 22 and 23 (Scheme 3). Both the formyl-
(40 and 41) and the acetyl derivatives (42 and 43) were
prepared in high yields (81–93%) using excess formyl-
acetic anhydride and acetic anhydride, respectively.
However, we were surprised to find that the same condi-
tions with isobutyryl anhydride or chloride mainly yield-
ed the diacylated product. The desired monoacylations
into 44 and 45 could be accomplished by using near
equimolar amounts of acid chloride.
It should be noted that the nitration was solvent sensi-
tive. CH2Cl2 worked well, while no reaction occurred
in MeOH or THF and only trace amounts of nitrated
product were observed in acetic acid. Facile reduction
Sulfonamides 46–49 that correspond to the acetamide
and isobutyramide derivatives 42–45 were synthesised
according to Scheme 3. The methyl-substituted sulfona-
mides 46 and 47 were easily prepared by reacting the
amino-substituted scaffolds 22 and 23 with excess meth-
ane sulfonyl chloride in pyridine. The isopropyl deriva-
tives on the other hand proved to be more of a
challenge. When repeating the procedure that worked
satisfactory with the methyl derivative, no conversion
was observed. Prolonged reaction times and/or
increased amounts of reagents were unfruitful and so
was also activation with 4-dimethylamino pyridine42 or
trimethyl silyl chloride.43 Other reaction conditions
using, for example, DMF/imidazole, MeCN/K2CO3 or
CH2Cl2/TEA44, did not lead to any progress and neither
did microwave heating. By replacing the naphthyl sub-
stituent in 23 for a methyl group without being able to
improve the reaction, it could be ruled out that the poor
reactivity was a result of steric hindrance alone. Gener-
ation of the anion would increase the reactivity but
1
1
1
R
R
R
a
b
S
S
S
N
N
N
O N
2
H N
2
CO Me
CO Me
2
CO Me
2
O
O
2
O
1
1
1
10: R = cyclopropyl
11: R = phenyl
20: R = cyclopropyl (84 %) 22: R = cyclopropyl (90 %)
21: R = phenyl (89 %)
1
1
1
23: R = phenyl (81 %)
c
c
1
1
R
R
S
N
S
N
O N
2
H N
2
CO Li
CO Li
2
2
O
O
1
1
24: R = cyclopropy (quant.) 26: R = cyclopropyl (quant.)
25: R = phenyl (quant.)
1
1
27:R = phenyl (quant.)
Scheme 1. Reagents and conditions: (a) NaNO2, O2(g), 4% (v/v) TFA,
CH2Cl2, rt; (b) Zn dust, AcOH, rt; (c) 0.1 M LiOH, THF/MeOH, rt.

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
7567
1
1
1
R
R
R
a
b
S
S
S
O
O
H
N
H
N
N
N
N
H N
N
H
N
H
2
CO Me
CO Me
2
CO Li
2
2
R
O
O
O
R
O
O
1
1
1
22: R = cyclopropyl
28: R = cyclopropyl, R = CH O-t-Bu (85 %)
34: R = cyclopropyl, R = CH OH (66% two steps)
2
2
1
1
1
23: R = phenyl
29: R = phenyl, R = CH O-t-Bu (79 %)
35: R = phenyl, R = CH OH (72% two steps)
2
2
1
1
30: R = cyclopropyl, R = H (91 %)
36: R = cyclopropyl, R = H (quant.)
1
1
31: R = phenyl, R = H (95 %)
37: R = phenyl, R = H (quant.)
1
1
32: R = cyclopropyl, R = CH (90 %)
38: R = cyclopropyl, R = CH (quant.)
3
3
1
1
33: R = phenyl, R = CH (88 %)
39: R = phenyl, R = CH (quant.)
3
3
Scheme 2. Reagents and conditions: (a) DIC, HOAt, CH2Cl2, rt; (b) For 28 and 29: TFA/CH2Cl2, rt, and purification (66–72%), then 0.1 M LiOH,
THF/MeOH, rt. For 30–33: 0.1 M LiOH, THF/MeOH, rt.
1
1
1
R
R
R
a
d
S
S
S
O
O
N
N
N
R
N
H N
2
R
N
H
CO Li
2
H
CO Me
2
CO Me
2
O
O
O
1
1
40: R = cyclopropyl, R = H (93 %)
50: R = cyclopropyl, R = H (quant. )
1
22: R = cyclopropyl
1
1
41: R = phenyl, R = H (88 %)
51: R = phenyl, R = H (quant. )
1
23: R = phenyl
1
1
42: R = cyclopropyl, R = CH (92 %)
52: R = cyclopropyl, R = CH (quant. )
3
3
1
1
43: R = phenyl, R = CH (81 %)
53: R = phenyl, R = CH (quant. )
3
3
1
1
44: R = cyclopropyl, R = i-Pr (68 %)
54: R = cyclopropyl, R = i-Pr (quant. )
1
1
45: R = phenyl, R = i-Pr (81 %)
55: R = phenyl, R = i-Pr (quant. )
b
1
1
R
R
d
S
S
O
O
O O
S
N
S
N
R
N
R
N
H
2
CO Me
2
CO H
2
R
O
O
1
2
1
46: R = cyclopropyl, R = H, R = CH (88 %)
56: R = cyclopropyl, R = CH (quant. )
3
3
1
2
1
47: R = phenyl, R = H, R = CH (84 %)
57: R = phenyl, R = CH (quant. )
3
3
c
1
2
1
48: R = cyclopropyl, R = CHO, R = i-Pr (36 %)
58: R = cyclopropyl, R = i-Pr (91% )
1
2
1
49*: R = phenyl, R = CHO, R = i-Pr (41 %)
59: R = phenyl, R = i-Pr (quant.)
Scheme 3. Reagents and conditions: (a) formyl acetic anhydride (40 and 41) or acetic anhydride (42 and 43) or isobutyryl chloride (44 and 45),
pyridine/CH2Cl2, rt; (b) MeSO2Cl, pyridine, rt; (c) t-BuOK (* gave 49 as racemate) or K+i-PrNꢁCOOMe (gave 48), i-PrSO2Cl, THF, ꢁ42 °C ! rt;
(d) for 50–55: 0.1 M LiOH, MeOH/THF, rt. For 56–59: 0.1 M LiOH, MeOH/THF, rt, then AmberliteÒ IR120+.
problems encountered by the presence of an acidic a-
proton turned our attention to formamide formation
and subsequent deprotonation. Using this strategy, the
isopropyl derivatives 48 and 49 could finally be obtained
in modest yields (36–41%) from formamides 40 and 41,
which were deprotonated and reacted with isopropyl
sulfonyl chloride. Simultaneous cleavage of the activat-
ing formyl group and the methyl ester was possible un-
der the described saponification conditions (Scheme 3).
dium catalysed reactions have failed earlier on these sul-
fur containing 2-pyridone systems, we abandoned this
strategy and went for an alternative pathway (Scheme
4). This route based on formylation (60 and 61) and oxi-
dation (62 and 63) followed by a simple coupling reac-
tion with DCC and HOAt required one additional
reaction step to 64–67 compared to the direct amino-
carbonylation (Scheme 4). However, the two first steps
in the reaction sequence had previously been developed
within the group and were known to proceed in high
yields.47 The aminocarbonylated derivatives 64–67
could be synthesised in excellent overall yields of 68–
84% over three steps and were subsequently hydrolysed
to 68–71 (Scheme 4).
2.4. Aminocarbonylations (D)
Halogenation of 10 and 11 followed by one-pot amino-
carbonylations was first attempted to obtain the prod-
ucts with inverse direction of amide bonds compared
to the acylated compounds. Iodination was straightfor-
ward according to our previously published proce-
dures.20 Unfortunately, the reported microwave-
assisted aminocarbonylations employing Mo(CO)6 as
carbon monoxide source together with a Pd-catalyst
(Pd(OAc)2 or Pd(C)), base (K2CO3 or DBU) and
amine45,46 did not turn out to be successful in our case.
Trace amounts of product were observed, while more of
the dehalogenated product was found. Since other palla-
3. Biological evaluation
The produced 24 compounds were evaluated for their
ability to prevent pili formation in P pili producing E.
coli (HB101/pPAP5). Bacteria were cultured on agar
containing 3.5 mM pilicide and the level of piliation of
the bacteria was subsequently determined in a haemag-
glutination (HA) assay. The haemagglutinating ability

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
7568
1
1
1
R
R
1
R
R
c
d
a
S
S
S
S
H
H
N
N
N
N
R
N
N
R
R
CO Me
CO Li
2
2
CO Me
O
O
O
O
CO Me
2
O
O
2
O
1
1
1
1
10: R = cyclopropyl
60: R = cyclopropyl (87 %)
64: R = cyclopropyl, R = CH (96 %)
68: R = cyclopropyl, R = CH (quant. )
3
3
1
1
1
1
R = H
R = OH
11: R = phenyl
61: R = phenyl (79 %)
65: R = phenyl, R = CH (93 %)
69: R = phenyl, R =CH (quant. )
3
3
b
1
1
1
62: R = cyclopropyl (quant. )
66: R = cyclopropyl, R = i-Pr (91 %)
70: R = cyclopropyl, R = i-Pr (quant. )
1
1
1
63: R = phenyl (94 %)
67: R = phenyl, R = i-Pr (92 %)
71: R = phenyl, R = i-Pr (quant. )
Scheme 4. Reagents: (a) ClꢁMe2N+@CHCl, MeCN, reflux; (b) NaClO2, NaH2PO4, DMSO/H2O, rt; (c) MeNH2 or i-PrNH2, DCC, HOAt, CH2Cl2,
rt; (d) 0.1 M LiOH, THF/MeOH, rt.
reflects the amount of pili being expressed in the pres-
ence of pilicide. A low HA-titre demonstrates that a high
concentration of bacteria is required for agglutination to
occur. The non-pili producing strain E. coli (HB101/
pBR322) was included as a negative control and gave
the HA-titre 1, whereas fully piliated positive control
(HB101/pPAP5) gave the HA-titres 64 and 128 in dupli-
cate runs (Table 1, HA-titre, No pilicide). The lead com-
pounds 8 and 9 have a very clear effect at 3.5 mM and
their HA-titres of 1 denote that agglutination and thus
pilus assembly is completely, or almost completely,
blocked.
Table 1. Relative affinities for chaperone PapD were determined using
relaxation-edited 1H NMR spectroscopy at a 1:1 ratio of pilicide:chap-
erone (Relative affinity)
Compound
Class
HA-titrea
(duplicates)
Relative affinity
(%)b
8
—
—
F
1/1
1/1
46
—
53
—
86
—
0
9
24
32/64
64/128
8/16
25
26
F
E
27
34
E
64/128
32/64
64/128
64/64
64/64
64/128c
64/128
16/64
64/64
32/64
64/64
32/128c
32/64
64/64
64/64
64/128
64/128
64/128
64/128
64/128
64/64
64/128
A
A
A
A
A
A
C
C
C
C
C
C
B
35
36
—
27
—
0
The in vivo activity of the extended peptidomimetics as
pilicides proved to be limited and all 24 derivatives were
significantly less potent than the parent compounds 8
and 9 (Table 1, HA-titre). Within the new set of com-
pounds some pilicide activity was retained for the ami-
no-substituted analogue 26 and even smaller effects
were seen for compounds 24, 34, 50, 52 and 55 (Table
1, HA-titre). These compounds were all from the cyclo-
propyl serie of derivatives. The results clearly demon-
strate that among the 26 evaluated compounds, 8 and
9 were still superior as pilicides and could not be chal-
lenged as lead compounds by the new extended
peptidomimetics.
37
38
39
50
—
100
—
41
—
60
—
69
—
—
—
28
—
41
—
—
51
52
53
54
55
56
57
58
B
B
59
68
B
D
D
D
D
—
69
70
Still, the complexity of in vivo activity, especially in
Gram-negatives where uptake is a common problem,
prompted us to evaluate the cyclopropyl serie of ana-
logues for chaperone binding. Affinity ranking was per-
formed using relaxation-edited one-dimensional 1H
NMR spectroscopy48 and indeed, although being more
or less inactive in vivo, most of them did bind to the
chaperone PapD (Table 1, relative affinity). The relative
affinities of the pilicides are given as percentages com-
pared to the best binder, that is, the aminoformyl substi-
tuted derivative 50 (100% relative affinity, Table 1).
Compared to the parent compound 8, similar or even
enhanced affinities were observed for the analogues 24,
26, 50, 52, 54, 56 and 70. Thus, chaperone binding is
apparently dependent on the substituents in this posi-
tion and the nitro-, amine-, amide- and sulfonamide
derivatives are well tolerated. All compounds with in-
creased affinities have a hydrogen bond-donating substi-
tuent in R3 and the amino-substituted derivative 26 was
one of the best binders. The nitro-derivative 24, which
lacks a hydrogen-bond donor, displayed an affinity that
was comparable to that of the unsubstituted lead com-
pound 8. Moreover, the arrangement of the amide bond,
71
No pilicide
The ability to prevent P pilus assembly in E. coli HB101/pPAP5, cul-
tured with 3.5 mM pilicide in agar, was evaluated with haemaggluti-
nation (HA-titre).
a Inverse of dilution factor for duplicate runs (see Section 5 for details).
b Relative affinity compared to 50; 6% RSD from triplicate runs.
c Limited solubility: 38 tested at 1.8 mM and 54 tested at 1.2 mM.
and thereby the position of the hydrogen-bond donor,
appears to affect binding based on a comparison of the
aminocarbonylated derivatives 68 and 70 and the meth-
yl- and isopropyl amides 52 and 54 (Table 1, classes C
and D, relative affinity). The relative affinities were con-
sistently lower (ꢀ30%) for the aminocarbonylated com-
pounds in class D, arguing that an amide bond that best
resembles a peptide backbone is beneficial. Hence, a
properly located hydrogen bond donor in R3 appears
to be favourable for chaperone affinity. This is in agree-
ment with the design and the cleft binding site of the
chaperone but could also be valid for other binding
locations.

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
7569
In contradiction, amino acid-substituted tripeptidomi-
metics 34, 36 and 38, which also have a suitably posi-
tioned hydrogen bond donor, were poor binders and
the serine and alanine-bearing compounds exhibited
no affinity for PapD (Table 1, class A, relative affinity).
One would reason that the affinity of a cleft binder
would improve, or at least not become weaker, by the
enhanced PapG mimicking features provided by the
amino acids. Possibly the acceptance decreases with in-
creased bulk since the amino acids are indeed the most
sterically demanding substituents. This may be valid
both within the series of amino acid-substituted ana-
logues, where the tolerance for glycine is higher than
for alanine and serine, and also within the whole set of
derivatives. Large substituents in R3 should be accepted
in the cleft since it is a fairly wide and unrestricted bind-
ing site, given that the pilicide is carboxylate anchored
to Arg8 and Lys112 (see Fig. 2). Thus, the low tolerance
for amino acid substituents and steric bulk could instead
support a binding site near the F1-G1 loop (Fig. 2a) that
was suggested in a previous NMR study.31
lier SAR studies, which have supported the designed
cleft binding site, the new data rather suggested binding
near the F1-G1 loop of the chaperone. The binding site
could however not be unambiguously clarified. The in vi-
vo activity of the extended peptidomimetics proved to
be limited which presumably could be a consequence
of poor uptake and/or distribution. Still, insight has
been gained of structure–activity relationships that can
be used in future design of pilicides and studies are
ongoing to unravel their exact binding site and mecha-
nism of action.
5. Experimental
General synthesis. All reactions were carried out under
an inert atmosphere with dry solvents under anhydrous
conditions, unless otherwise stated. CH2Cl2 and 1,2-
dichlorethane were distilled from calcium hydride and
THF was distilled from potassium. DMF was distilled
˚
and dried over 3A molecular sieves. EtOH was dried
˚
over 3 A molecular sieves. Zinc dust was activated by
However, it is important to note that a large R3 substi-
tuent in combination with the already sterically demand-
ing R1 and R2 in some cases resulted in mixtures of
locked conformations, that is, atropisomers. The con-
formational constraints could prevent the compounds
from adopting a bioactive conformation, which compli-
cates the interpretation of the data. This was not further
investigated here but the observation has led to efforts to
isolate atropisomers to provide valuable information
about the binding mode and the bioactive conformation.
stirring it in 10% HCl for two minutes and then filtered
and washed with water and acetone. The activated zinc
was then dried under vacuum. Formyl acetic anhydride
was prepared as follows: Formic acid (0.40 ml,
10.6 mmol) and acetic acid anhydride (0.90 ml,
9.3 mmol) were stirred at rt for 5 h and used directly
without purification. HCl(g) was passed through con-
centrated H2SO4 prior to use. TLC was performed on
Silica Gel 60 F254 (Merck) using UV light detection.
Flash column chromatography (eluents given in brack-
˚
ets) employed normal phase silica gel (Matrex, 60 A,
The presented results suggest the poor in vivo activity of
the extended peptide mimetics to be related to uptake-
and/or distribution rather than affinity. An alternative
explanation could be that the increased affinities are
due to unspecific binding to PapD and it should be
35–70 lm, Grace Amicon). The 1H and 13C NMR spec-
tra were recorded at 298 K with a Bruker DRX-400
spectrometer in CDCl3 [residual CHCl3 (dH 7.26 ppm)
or CDCl3 (dC 77.0 ppm) as internal standard], or
MeOD-d4 [residual CD2HOD (dH 3.30 ppm) or CD3OD
(dC 49.0 ppm) as internal standard], or DMSO-d6 [resid-
ual DMSO (dH 2.49 ppm) or DMSO (dC 40.0 ppm) as
internal standard]. In the case of rotameric/diastereo-
meric mixtures the NMR signals and integrals are as-
signed as corresponding to either the major (maj) or
the minor (min) rotamer/diastereomer.
1
pointed out that relaxation-edited H NMR spectrosco-
py does not provide any information about the location
of binding. Overall, this study concludes a limited use of
the compounds in the four classes A–D as pilicides. Still,
it is clear that chaperone affinities in the range of what is
required for in vivo activity can be retained or even im-
proved with substituents in R3 and this acceptance is
encouraging for future synthetic work.
IR spectra were recorded on an ATI Mattson Genesis
Series FTIRTM spectrometer. Optical rotations were mea-
sured with a Perkin-Elmer 343 polarimeter at 20 °C.
HRMS data were recorded with fast atom bombard-
ment (FAB+) ionization on a JEOL JMS-SX 102
spectrometer.
4. Conclusions
Mild and efficient synthetic procedures to amino-substi-
tuted bicyclic 2-pyridone scaffolds have been developed.
These highly substituted core structures can serve as
building blocks in peptide mimetic related research.
Based on this new scaffold a set of 24 extended deriva-
tives was synthesised and biologically evaluated as
inhibitors of pilus assembly in E. coli. It could be con-
cluded that binding to the target chaperone PapD could
be improved with appropriate substituents in the inves-
tigated position. Specifically, a small, hydrogen-bond-
donating substituent appears to be favoured. In addi-
tion, it was beneficial if the hydrogen-bond donor was
located as in a peptide sequence. In contradiction to ear-
5.1. Synthesis of methyl esters 10–11, 20–23, 28–33,
40–49 and 60–67
Compounds 10 and 11 were synthesised according to
published procedures. Data in agreement with published
data.16,17
5.1.1. (3R)-8-Cyclopropyl-7-naphthalen-1-ylmethyl-6-ni-
tro-5-oxo-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-3-car-
boxylic acid methyl ester (20). To a mixture of 10
(750 mg, 1.92 mmol) and NaNO2 (139 mg, 2.0 mmol)

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
7570
was added 60 ml CH2Cl2. A balloon filled with oxygen
gas was connected to the flask via a rubber septum
and 2.5 ml TFA was added dropwise at rt. After 5 h
the brown solution was neutralized with NaHCO3(aq)
and then extracted with CH2Cl2. The organic phase
was dried over Na2SO4(s), filtered and concentrated.
Purification by silica gel chromatography (heptane/
EtOAc, 1:1 ! 1:2) gave 20 as a yellow foam (703 mg,
84%): [a]D ꢁ324 (c 0.5, CHCl3); IR m/cmꢁ1 1753, 1657,
1589, 1525, 1485, 1437, 1371, 1346, 1207, 1167, 1030,
(m, 1H), 7.50–7.44 (m, 1H), 7.29–7.23 (m, 1H), 6.96
(d, J = 7.0 Hz, 1H), 5.59 (dd, J = 2.2, 8.2 Hz, 1H),
4.55–4.39 (m, 2H), 3.86 (br s, 2H), 3.76 (s, 3H), 3.57
(dd, J = 8.4, 11.8 Hz, 1H), 3.42 (dd, J = 2.3, 11.7 Hz,
1H), 1.52–1.44 (m, 1H), 0.66–0.58 (m, 2H), 0.54–0.45
(m, 2H); 13C NMR (100 MHz, CDCl3) d 168.35,
156.25, 133.41, 132.22, 131.90, 131.76, 131.70, 128.42,
127.55, 126.73, 125.75, 125.33, 125.27, 122.96, 122.72,
114.27, 62.69, 52.71, 31.29, 30.30, 11.36, 6.54, 6.51;
HRMS (FAB+) calcd for [M]+ C23H22N2O3S
406.1351, obsd 406.1429.
1
1010, 961, 797, 769, 734; H NMR (400 MHz, CDCl3)
d 8.04 (d, J = 8.3 Hz, 1H), 7.85 (d, J = 8.0 Hz, 1H),
7.72 (d, J = 8.2 Hz, 1H), 7.58–7.47 (m, 2H), 7.37–7.31
(m, 1H), 7.04 (d, J = 7.1 Hz, 1H), 5.75–5.70 (m, 1H),
4.69–4.50 (m, 2H), 3.82 (s, 3H), 3.75–3.66 (m, 1H),
3.52 (dd, J = 2.0, 12.0 Hz, 1H), 1.23–1.13 (m, 1H),
0.66–0.48 (m, 4H); 13C NMR (100 MHz, CDCl3) d
167.51, 153.18, 152.24 (splitted), 148.30 (splitted),
139.51, 133.40, 131.99, 131.30, 128.66, 127.25, 126.21,
125.66, 125.31, 124.44, 122.52, 112.10, 63.37, 53.32,
31.46, 31.11, 11.11, 7.54, 7.10; HRMS (FAB+) calcd
for [M+H]+ C23H21N2O5S 437.1171, obsd 437.1180.
5.1.4. (3R)-6-Amino-7-naphthalen-1-ylmethyl-5-oxo-8-
phenyl-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-3-carbox-
ylic acid methyl ester (23). By following the procedure
described for the preparation of 22 from 20, 21
(623 mg, 1.32 mmol) and Zn dust (404 mg, 6.18 mmol)
in 9 ml acetic acid gave 23 (474 mg, 81%) after purifica-
tion with silica gel chromatography (heptane/EtOAc,
1:1 ! 1:2). [a]D ꢁ167 (c0.5, CHCl3); IR m/cmꢁ1 1740,
1633, 1572, 1488, 1440, 1315, 1207, 1155, 1073, 977,
906, 793, 731, 702; 1H NMR (400 MHz, CDCl3) d
7.89–7.83 (m, 2H), 7.75 (d, J = 8.2 Hz, 1H), 7.50–7.42
(m, 2H), 7.41–7.36 (m, 1H), 7.26–7.11 (m, 6H), 5.74
(dd, J = 2.4, 8.2 Hz, 1H), 4.07 (s, 2H), 3.86 (s, 5H),
3.65 (dd, J = 8.2, 11.7 Hz, 1H), 3.47 (dd, J = 2.4,
11.8 Hz, 1H); 13C NMR (100 MHz, CDCl3) d 168.44,
156.51, 136.97, 133.63, 132.55, 131.98, 131.70, 131.08,
129.51 (broad), 128.91 (broad), 128.57, 128.39 (2C,
broad), 127.77, 127.17, 125.90, 125.57, 125.50, 125.27,
123.66, 122.92, 117.52, 63.77, 53.09, 31.69, 31.52;
HRMS (FAB+) calcd for [M]+ C26H22N2O3S
442.1351, obsd 442.1358.
5.1.2.
(3R)-7-Naphthalen-1-ylmethyl-6-nitro-5-oxo-8-
phenyl-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-3-carbox-
ylic acid methyl ester (21). By following the procedure
described for the preparation of 20 from 10, 11
(750 mg, 1.75 mmol), NaNO2 (127 mg, 1.84 mmol),
50 ml CH2Cl2 and 2.0 ml TFA gave 21 as a yellow foam
(737 mg, 89%) after purification with silica gel chroma-
tography (heptane/EtOAc, 1:1 ! 1:4). [a]D ꢁ258 (c
0.5, CHCl3); IR m/cmꢁ1 1747, 1657, 1583, 1523, 1475,
1
1438, 1352, 1214, 1152, 1006, 780, 734, 700; H NMR
(400 MHz, CDCl3) d 7.77 (d, J = 8.1 Hz, 1H), 7.69 (d,
J = 8.3 Hz, 1H), 7.55 (d, J = 8.3 Hz, 1H), 7.43–7.31
(m, 3H), 7.16–7.09 (m, 3H), 7.09–7.02 (m, 1H), 7.00–
6.96 (m, 1H), 6.90 (d, J = 7.5 Hz, 1H), 5.80 (dd,
J = 2.4, 8.7 Hz, 1H), 4.30–4.12 (m, 2H), 3.90 (s, 3H),
3.75 (dd, J = 8.8, 11.9 Hz, 1H), 3.54 (dd, J = 2.4,
12.0 Hz, 1H); 13C NMR (100 MHz, CDCl3) d 167.48,
153.57, 151.55, 146.83, 139.43, 134.29, 133.43, 132.09,
131.29, 129.90, 129.34, 128.85, 128.80, 128.76, 128.58,
127.55, 126.07, 125.64, 125.58, 125.28, 122.54, 114.93,
64.43, 53.74, 31.91, 31.87; HRMS (FAB+) calcd for
[M+H]+ C26H21N2O5S 473.1171, obsd 473.1180.
5.1.5.
amino)-8-cyclopropyl-7-naphthalen-1-ylmethyl-5-oxo-2,3-
dihydro-5H-thiazolo[3,2-a]pyridine-3-carboxylic acid
(3R)-6-(2-Acetylamino-3-tert-butoxy-propionyl-
methyl ester (28). By following the procedure described
for the preparation of 30 from 22, 22 (60 mg,
0.15 mmol), HOAt (24 mg, 0.18 mmol), Ac-Ser(t-Bu)-
OH (39 mg, 0.19 mmol) and DIC (28 ll, 0.19 mmol) in
0.8 ml CH2Cl2:THF (1:1) (24 h) gave 28 (74 mg, 85%)
after purification with silica gel chromatography (hep-
tane/EtOAc/MeOH, 5:20:1 ! 5:40:4). [a]D ꢁ184 (c 0.5,
CHCl3); IR m/cmꢁ1 1747, 1639, 1587, 1496, 1363, 1207,
1081, 1022, 792, 773; As a mixture of diastereomers
5.1.3. (3R)-6-Amino-8-cyclopropyl-7-naphthalen-1-ylm-
ethyl-5-oxo-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-3-car-
boxylic acid methyl ester (22). Compound 20 (685 mg,
1.57 mmol) was dissolved in 11 ml acetic acid and then
activated Zn dust (472 mg, 7.22 mmol) was added in
portions in order to control the temperature of the reac-
tion. After 5 h of stirring at rt the solvent was removed
under vacuum. The residue was dissolved in CH2Cl2,
neutralized with Na2CO3(aq) and extracted with
CH2Cl2. The organic phase was dried over Na2SO4(s),
filtered and concentrated. Purification by silica gel chro-
matography (heptane/EtOAc, 1:1 ! 1:9) gave 22 as a
foam (571 mg, 90%): [a]D ꢁ190 (c 0.5, CHCl3); IR m/
cmꢁ1 1742, 1638, 1575, 1510, 1434, 1353, 1287, 1215,
1179, 1149, 1010, 960, 793, 772, 731; 1H NMR
(400 MHz, CDCl3) d 8.14 (d, J = 8.4 Hz, 1H), 7.82 (d,
J = 8.0 Hz, 1H), 7.67 (d, J = 8.2 Hz, 1H), 7.57–7.50
ꢀ5:4 1H NMR (400 MHz, CDCl3)
d 8.09 (d,
J = 8.2 Hz, 1H), 8.06 (s, 0.5H), 7.98 (s, 0.4H), 7.84 (d,
J = 8.1 Hz, 1H), 7.69 (d, J = 8.2 Hz, 1H), 7.58–7.46
(m, 2H), 7.33–7.26 (m, 1H), 6.89 (d, J = 7.0 Hz, 1H),
6.43 (d, J = 6.1 Hz, 0.4H), 6.35 (d, J = 6.3 Hz, 0.5H),
5.67 (dd, J = 2.2, 8.6 Hz, 1H), 4.62–4.49 (m, 2H),
4.43–4.32 (m, 1H), 3.84–3.80 (m, 3H), 3.73–3.61 (m,
2H), 3.51 (dd, J = 2.2, 11.9 Hz, 1H), 3.22 (t,
J = 8.4 Hz, 0.6H), 3.05 (t, J = 8.5 Hz, 0.5H), 1.86 (s,
1.4H), 1.84 (s, 1.5H), 1.35–1.25 (m, 1H), 0.88 (s, 5H),
0.82 (s, 4H), 0.69–0.60 (m, 2H), 0.60–0.51 (m, 2H); 13C
NMR (100 MHz, CDCl3) d 169.97 (splitted), 169.89,
168.39, 168.34, 157.84, 157.79, 150.86, 145.97, 145.78,
133.98, 133.78, 133.69, 131.77, 131.73, 128.73, 126.95,
126.93, 126.06, 125.68, 125.49, 125.46, 123.93, 11.89,
123.11, 122.21, 122.18, 113.60, 113.54, 74.14, 74.10,
63.31, 63.28, 60.98, 60.95, 53.35, 53.27, 53.23, 53.17,

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
7571
31.92, 31.86, 31.64, 31.58, 27.05–26.85 (3C), 22.95,
22.93, 11.73, 11.68, 7.49–7.24 (m, 2C), 7.34, 7.30;
HRMS (FAB+) calcd for [M+Na]+ C32H37N3NaO6S
614.2301, obsd 614.2310.
0.51 (m, 2H); 13C NMR (100 MHz, CDCl3) d 170.38,
168.79, 168.29, 158.20, 151.84, 146.19, 133.97, 133.62,
131.70, 128.72, 126.89, 126.13, 125.73, 125.47, 124.29,
123.21, 121.94, 113.92, 63.39, 53.28, 43.11 (broad),
31.82, 31.51, 22.50 (broad), 11.76, 7.50, 7.38; HRMS
(FAB+) calcd for [M+Na]+ C27H27N3NaO5S 528.1571,
obsd 528.1569.
5.1.6.
(3R)-6-(2-Acetylamino-3-tert-butoxy-propionyl-
amino)-7-naphthalen-1-ylmethyl-5-oxo-8-phenyl-2,3-dihy-
dro-5H-thiazolo[3,2-a]pyridine-3-carboxylic acid methyl
ester (29). By following the procedure described for the
preparation of 30 from 22, 23 (60 mg, 0.15 mmol),
HOAt (22 mg, 0.16 mmol), Ac-Ser(t-Bu)-OH (36 mg,
0.18 mmol) and DIC (28 ll, 0.19 mmol) in 0.8 ml
CH2Cl2:THF (1:1) (48 h) gave 29 (67 mg, 79%) after
purification with silica gel chromatography (heptane/
EtOAc/MeOH, 5:30:2 ! 20:40:1). [a]D ꢁ144 (c 0.5 in
CHCl3); IR m/cmꢁ1 2365, 2329, 1746, 1638, 1585, 1488,
1364, 1212, 1155, 1085, 746, 704; As a mixture of diaste-
5.1.8. (3R)-6-(2-Acetylamino-acetylamino)-7-naphthalen-
1-ylmethyl-5-oxo-8-phenyl-2,3-dihydro-5H-thiazolo[3,2-
a]pyridine-3-carboxylic acid methyl ester (31). By follow-
ing the procedure described for the preparation of 30
from 22, 23 (55 mg, 0.12 mmol), HOAt (24 mg,
0.18 mmol), N-acetyl glycine (19 mg, 0.16 mmol) and
DIC (25 ll, 0.16 mmol) in 0.6 ml CH2Cl2 (28 h) gave
31 (64 mg, 95%) after purification with silica gel chroma-
tography (heptane/EtOAc/MeOH, 1:10:1). [a]D ꢁ154 (c
0.5, CHCl3); IR m/cmꢁ1 2961, 1747, 1631, 1582, 1490,
1444, 1398, 1369, 1259, 1215, 1154, 1090, 1013, 739,
1
reomers ꢀ1:1 H NMR (400 MHz, CDCl3) d 8.06 (s,
0.5H), 8.00 (s, 0.5H), 7.80–7.75 (m, 1H), 7.73–7.62 (m,
2H), 7.44–7.34 (m, 2H), 7.31–7.25 (m, 1H), 7.19–6.97
(m, 5H), 6.97 (d, J = 7.0 Hz, 1H), 6.43 (d, J = 6.2 Hz,
0.5H), 6.29 (d, J = 6.3 Hz, 0.5H), 5.76–5.70 (m, 1H),
4.40–4.29 (m, 1H), 4.21–4.06 (m, 2H), 3.86 (s, 1.5H),
3.85 (s, 1.5H), 3.73–3.65 (m, 1H), 3.64–3.59 (m, 1H),
3.47 (dd, J = 2.2, 11.7 Hz, 1H), 3.18 (t, J = 8.3 Hz,
0.5H), 3.01–2.94 (t, J = 8.4 Hz, 0.5H), 1.85 (s, 1.5H),
1.82 (s, 1.5H), 0.92 (s, 4.5H), 0.85 (s, 4.5H); 13C NMR
(100 MHz, CDCl3) d 170.01, 169.80, 168.25, 168.23,
157.92, 157.86, 148.52, 145.32, 145.09, 136.10, 136.02,
133.88, 133.64, 133.54, 133.51, 131.48, 131.44, 129.75,
129.45, 129.29, 128.65–128.43 (m, 3C), 128.17, 126.95,
125.83, 125.81, 125.53, 125.50, 125.39, 125.32, 124.41,
124.62, 123.07, 123.05, 122.08, 122.05, 116.27, 116.25,
74.23, 74.18, 64.14, 34.11, 60.86, 60.82, 53.38, 53.34,
53.17, 32.78, 32.74, 31.71, 31.68, 27.09–26.84 (3C),
26.92, 22.95, 22.89; HRMS (FAB+) calcd for
[M+Na]+ C35H37N3NaO6S 650.2301, obsd 650.2295.
1
747, 703; H NMR (400 MHz, CDCl3) d 7.80–7.60 (m,
4H), 7.44–7.33 (m, 2H), 7.31–7.25 (m, 1H), 7.21–7.11
(m, 3H), 7.11–7.03 (m, 2H), 6.99 (d, J = 7.0 Hz, 1H),
6.45–6.36 (m, 1H), 5.71–5.53 (m, 1H), 4.20–4.06 (m,
2H), 3.89–3.68 (m, 5H), 3.68–3.53 (m, 1H), 3.50–3.34
(m, 1H), 1.72 (s, 3H); 13C NMR (100 MHz, CDCl3) d
170.38, 168.59, 168.14, 158.27, 149.51, 145.50, 135.95,
133.85, 133.45, 131.41, 129.75, 129.39, 128.60 (2C),
128.49, 128.24, 126.90, 125.87, 125.56, 125.34, 124.99,
123.15, 121.76, 116.55, 64.22, 53.37, 43.16, 32.69,
31.59, 22.45; HRMS (FAB+) calcd for [M+Na]+
C30H27N3NaO5S 564.1569, obsd 564.1571.
5.1.9. (3R)-6-(2-Acetylamino-propionylamino)-8-cyclo-
propyl-7-naphthalen-1-ylmethyl-5-oxo-2,3-dihydro-5H-
thiazolo[3,2-a]pyridine-3-carboxylic acid methyl ester
(32). By following the procedure described for the prep-
aration of 30 from 22, 22 (60 mg, 0.15 mmol), HOAt
(24 mg, 0.18 mmol), N-acetyl alanine (25 mg,
0.19 mmol) and DIC (30 ll, 0.19 mmol) in 0.8 ml
CH2Cl2:THF (1:1) (overnight) gave 32 (69 mg, 90%)
after purification with silica gel chromatography (hep-
tane/EtOAc/MeOH, 5:30:2 ! 1:10:1). [a]D ꢁ230 (c 0.5,
CHCl3); IR m/cmꢁ1 1748, 1632, 1582, 1497, 1436, 1258,
1211, 1014, 792, 747; As a mixture of diastereomers
5.1.7. (3R)-6-(2-Acetylamino-acetylamino)-8-cyclopro-
pyl-7-naphthalen-1-ylmethyl-5-oxo-2,3-dihydro-5H-thiaz-
olo[3,2-a]pyridine-3-carboxylic acid methyl ester (30). A
flask was charged with 22 (55 mg, 0.14 mmol), HOAt
(26 mg, 0.19 mmol) and N-acetyl glycine (21 mg,
0.18 mmol) and 0.6 ml CH2Cl2 was added. This was fol-
lowed by addition of DIC (27 ll, 0.17 mmol). The mix-
ture was stirred for 26 h and was then diluted with
CH2Cl2 and washed with NaHCO3 (aq). The organic
layer was washed with 10% citric acid (aq), and the com-
bined aqueous layers were reextracted with CH2Cl2. The
combined organic layers were dried over Na2SO4(s), fil-
tered and concentrated.
ꢀ7:2 1H NMR (400 MHz, CDCl3)
d 8.09 (d,
J = 8.2 Hz, 1H), 7.84 (d, J = 7.9 Hz, 1H), 7.71 (s, 1H),
7.68 (d, J = 8.3 Hz, 1H), 7.57–7.45 (m, 2H), 7.31–7.26
(m, 1H), 6.90 (d, J = 6.9 Hz, 1H), 6.38 (d, J = 7.1 Hz,
0.2H), 6.10 (d, J = 6.4 Hz, 0.7H), 5.63 (dd, J = 1.1,
8.4 Hz, 0.8H), 5.58 (dd, J = 1.4, 8.8 Hz, 0.2H), 4.62–
4.48 (m, 2H), 4.44–4.32 (m, 1H), 3.81–3.73 (m, 3H),
3.72–3.59 (m, 1H), 3.52–3.42 (m, 1H), 1.70–1.55 (m,
3H), 1.37–1.28 (m, 1H), 1.15 (d, J = 7.0 Hz, 2.5H),
1.06–0.99 (m, 0.7H), 0.70–0.61 (m, 1H), 0.61–0.51 (m,
1H); 13C NMR (100 MHz, CDCl3) d 171.66, 169.81
(min), 169.70 (maj), 168.34 (maj), 168.28 (min), 158.07,
151.48 (min), 151.27 (maj), 145.97, 133.95 (maj),
133.85 (min), 133.58, 131.70, 128.70, 126.81, 126.06,
125.65, 125.51, 124.12, 123.15, 122.08, 113.90 (min),
113.78 (maj), 63.29, 53.25, 48.65, 31.76, 31.58, 22.62
(min), 22.57 (maj), 17.86 (maj), 17.73 (min), 11.74,
7.46, 7.40; HRMS (FAB+) calcd for [M+Na]+
C28H29N3NaO5S 542.1726, obsd 542.1720.
Purification by silica gel chromatography (heptane/
EtOAc/MeOH, 5:30:2) gave 30 (62 mg, 91%): [a]D
ꢁ158 (c 0.5, CHCl3); IR m/cmꢁ1 1746, 1631, 1582,
1496, 1344, 1267, 1215, 1174, 1068, 1025, 965, 775,
729; 1H NMR (400 MHz, CDCl3)
d
8.08 (d,
J = 8.2 Hz, 1H), 7.84 (d, J = 7.8 Hz, 1H), 7.76 (s, 1H),
7.68 (d, J = 8.1 Hz, 1H), 7.58–7.45 (m, 2H), 7.31–7.25
(m, 1H), 6.91 (d, J = 5.3 Hz, 1H), 6.54–6.41 (m, 1H),
5.64–5.46 (m, 1H), 4.66–4.46 (m, 2H), 3.90–3.68 (m,
5H), 3.68–3.54 (m, 1H), 3.47 (d, J = 10.9 Hz, 1H), 1.73
(s, 3H), 1.39–1.22 (m, 1H), 0.72–0.61 (m, 2H), 0.61–

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
7572
5.1.10. (3R)-6-(2-Acetylamino-propionylamino)-7-naph-
thalen-1-ylmethyl-5-oxo-8-phenyl-2,3-dihydro-5H-thiaz-
olo[3,2-a]pyridine-3-carboxylic acid methyl ester (33). By
following the procedure described for the preparation of
30 from 22, 23 (60 mg, 0.14 mmol), HOAt (23 mg,
0.17 mmol), N-acetyl alanine (23 mg, 0.18 mmol) and
DIC (28 ll, 0.18 mmol) in 0.8 ml CH2Cl2:THF (1:1)
(23 h) gave 33 (66 mg, 88%) after purification with silica
(min), 128.74 (maj), 127.79 (min), 126.99 (maj), 126.57
(min), 126.20, 125.79 (maj), 125.45, 123.96 (maj),
123.27, 122.85 (min), 122.37 (min), 120.97 (maj),
114.17 (maj), 113.76 (min), 63.45 (splitted), 53.44
(min), 53.36 (maj), 32.24 (maj), 31.75 (min), 31.66
(maj), 31.60 (min), 11.83 (maj), 11.64 (min), 7.60
(maj), 7.46, 7.06 (min); HRMS (FAB+) calcd for
[M+H]+ C24H23N2O4S 435.1379, obsd 435.1386.
gel
chromatography
(heptane/EtOAc/MeOH,
5:30:2 ! 1:10:1). [a]D ꢁ174 (c 0.5, CHCl3); IR m/cmꢁ1
3283, 3225, 2955, 2925, 28 50, 1747, 1631, 1490, 1439,
1369, 1260, 1212, 1155, 1012, 977, 906, 792, 725, 702;
As a mixture of diastereomers ꢀ10:3 1H NMR
(400 MHz, CDCl3) d 7.77 (d, J = 7.6 Hz, 1H), 7.71 (d,
J = 7.8 Hz, 1H), 7.67–7.61 (m, 2H), 7.44–7.34 (m, 2H),
7.31–7.25 (m, 1H), 7.22–7.05 (m, 4.7H), 7.05–7.01 (m,
0.3H), 7.00–7.94 (m, 1H), 6.23–6.15 (m, 0.3H), 5.94–
5.85 (m, 0.6H), 5.75–5.65 (m, 1H), 4.41–4.30 (m, 1H),
4.20–4.08 (m, 2H), 3.83 (s, 2H), 3.81 (s, 1H), 3.72–3.61
(m, 1H), 3.49–3.41 (m, 1H), 1.70–1.66 (m, 0.9H), 1.59
(s, 2H), 1.11 (d, J = 6.8 Hz, 2H), 1.03–0.96 (m, 1H);
13C NMR (100 MHz, CDCl3) d 171.25, 169.76 (min),
169.70 (maj), 168.24, 158.18, 148.82 (min), 148.71
(maj), 145.26, 136.16 (maj), 136.12 (min), 133.97 (maj),
133.89 (min), 133.53, 131.52, 129.82, 129.51 (splitted),
128.70 (2C), 128.62, 128.32, 126.98 (min), 126.94
(maj), 125.95, 125.63, 125.49 (maj), 125.45 (min),
124.75 (maj), 124.63 (min), 123.19, 121.93 (maj),
121.87 (min), 116.64 (min), 116.56 (maj), 64.18, 53.46,
48.84 (min), 48.70 (maj), 32.87 (maj), 32.78 (min),
31.78, 22.77 (min), 22.62 (maj), 17.72; HRMS (FAB+)
calcd for [M+Na]+ C31H29N3O5S 578.1726, obsd
578.1722.
5.1.12. (3R)-6-Formylamino-7-naphthalen-1-ylmethyl-5-
oxo-8-phenyl-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-3-
carboxylic acid methyl ester (41). By following the pro-
cedure described for the preparation of 40 from 22, 23
(250 mg, 0.56 mmol) in 1.0 ml pyridine/CH2Cl2 (1:1),
and formyl acetic anhydride solution (0.35 ml,
4.4 mmol) gave 41 (243 mg, 88%) after purification
with silica gel chromatography (heptane/EtOAc/
MeOH, 20:40:1 ! 5:20:1). [a]D ꢁ158 (c 0.5 in CHCl3);
IR m/cmꢁ1 1745, 1681, 1633, 1584, 1486, 1433, 1216,
1155, 770, 696; As mixture of rotamers ꢀ7:4 1H
NMR (400 MHz, CDCl3) d 8.54 (d, J = 11.4 Hz,
0.3H), 8.1 (s, 0.6H), 7.86–7.60 (m, 3H), 7.59–7.31 (m,
3H), 7.31–6.91 (m, 7H), 5.77 (d, J = 8.2 Hz, 0.4H),
5.65 (dd, J = 8.5, 2.0 Hz, 0.6H), 4.28–4.08 (m, 2H),
3.86 (s, 1H), 3.76 (s, 2H), 3.76–3.57 (m, 1H), 3.54–
3.39 (m, 1H); 13C NMR (100 MHz, CDCl3) d 168.04,
164.41 (min), 160.25 (maj), 158.09, 148.84 (maj),
145.27 (maj), 144.17 (min), 143.67 (min), 135.92
(maj), 135.59 (min), 133.80 (maj), 133.57 (min),
133.35 (maj), 132.29 (min), 131.41 (maj), 131.26
(min), 129.64, 129.33 (maj), 128.98 (min), 128.74–
128.28 (m, 3C + 1 min C), 128.12 (maj), 127.65
(min), 126.76 (maj), 126.21 (min), 125.88 (min),
125.78 (maj), 125.43 (maj), 125.23, 124.72 (maj),
123.85 (min), 123.09 (maj), 122.69 (min), 122.18
(min), 120.91 (maj), 116.58 (maj), 116.36 (min), 64.27
(min), 64.12 (maj), 53.38 (min), 53.25 (maj), 32.91
(maj), 32.33 (min), 31.53; HRMS (FAB+) calcd for
[M+H]+ C27H23N2O4S 471.1379, obsd 471.1382.
5.1.11. (3R)-8-Cyclopropyl-6-formylamino-7-naphthalen-
1-ylmethyl-5-oxo-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-
3-carboxylic acid methyl ester (40). Compound 22
(250 mg, 0.62 mmol) was dissolved in 1.0 ml pyridine/
CH2Cl2 (1:1). Then formyl acetic anhydride solution
(0.4 ml, 5.0 mmol) was added (prepared as described in
general experimentals). The solution was stirred at rt
overnight and then 10% citric acid (aq). was added
and the aqueous layer was extracted with CH2Cl2. The
combined organic layers were dried over Na2SO4(s), fil-
tered, and concentrated. Purification with silica gel chro-
matography (heptane/EtOAc/MeOH, 50:50:1 ! 5:20:1)
gave 40 (248 mg, 93%). [a]D ꢁ200 (c 0.5 in CHCl3); IR
m/cmꢁ1 2360, 2329, 1743, 1674, 1633, 1581, 1501, 1435,
1398, 1342, 1248, 1215, 1170, 1025, 960, 792, 755, 665;
5.1.13. (3R)-6-Acetylamino-8-cyclopropyl-7-naphthalen-
1-ylmethyl-5-oxo-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-
3-carboxylic acid methyl ester (42). Compound 22
(60 mg, 0.15 mmol) was dissolved in 0.2 ml pyridine
and acetic anhydride (28 ll, 0.30 mmol) was added.
After 5.5 h of stirring at rt the solution was diluted with
CH2Cl2 and washed with 10% citric acid (aq). The com-
bined aqueous layers were reextracted with CH2Cl2. The
combined organic layers were dried over Na2SO4(s), fil-
tered and concentrated. Purification with silica gel chro-
matography (heptane/EtOAc/MeOH, 1:9:1) gave 42
(61 mg, 92%). [a]D ꢁ213 (c 0.5, CHCl3); IR m/cmꢁ1
1
As a mixture of rotamers ꢀ7:5 H NMR (400 MHz,
CDCl3) d 8.46 (d, J = 11.3 Hz, 0.4H), 8.17–8.06 (m,
1.5H), 7.92–7.83 (m, 1H), 7.79–7.68 (m, 1H), 7.64–7.48
(m, 2H), 7.35–7.28 (m, 1H), 7.10 (s, 0.5H), 6.88 (d,
J = 7.0 Hz, 0.6H), 6.82 (d, J = 6.8 Hz, 0.4H), 6.77 (d,
J = 11.2 Hz, 0.3H), 5.75–5.69 (m, 0.4H), 5.67–5.61 (m,
0.6H), 4.71–4.52 (m, 2H), 3.86 (s, 1.2H), 3.81 (s,
1.8H), 3.77–3.65 (m, 1H), 3.60–3.50 (m, 1H), 1.49–1.40
(m, 0.4H), 1.40–1.31 (m, 0.6H), 0.76–0.64 (m, 2H),
0.64–0.51 (m, 2H); 13C NMR (100 MHz, CDCl3) d
168.32 (maj), 168.21 (min), 164.35 (min), 160.22 (maj),
158.08, 151.14 (maj), 146.15 (min/maj), 145.99 (maj/
min), 144.71 (min), 134.05 (maj), 133.87 (min), 133.67
(maj), 132.38 (min), 131.79 (maj), 131.69 (min), 128.94
1
1749, 1635, 1581, 1500, 1213, 1027, 964, 792, 773; H
NMR (400 MHz, CDCl3) d 8.09 (d, J = 8.1 Hz, 1H),
7.86 (d, J = 7.8 Hz, 1H), 7.70 (d, J = 8.1 Hz, 1H),
7.58–7.46 (m, 2H), 7.34–7.26 (m, 1H), 7.01 (s, 1H),
6.92 (d, J = 6.5 Hz, 1H), 5.61 (d, J = 8.0 Hz, 1H),
4.67–4.51 (m, 2H), 3.8 (s, 3H), 3.73–3.63 (m, 1H),
3.55–3.47 (m, 1H), 1.90 (s, 3H), 1.39–1.29 (m, 1H),
0.75–0.54 (m, 4H); 13C NMR (100 MHz, CDCl3) d
169.69, 168.40, 158.48, 151.23, 145.45, 134.52, 133.65,
131.79, 128.72, 126.86, 126.08, 125.70, 125.42, 124.25,
123.32, 122.42, 114.07, 63.40, 53.29, 32.19, 31.58,

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
7573
23.26, 11.87, 7.65, 7.47; HRMS (FAB+) calcd for
[M+H]+ C25H25N2O4S 449.1535, obsd 449.1541.
chloride (13 ll, 0.12 mmol) and after 6 h adding more
isobutyryl chloride (2 ll) together with 0.5 ml CH2Cl2
(overnight) gave 45 (51 mg, 81%) after purification with
silica gel chromatography (heptane/EtOAc/MeOH,
20:40:1 ! 10:30:1). [a]D ꢁ140 (c 0.5, CHCl3); IR m/
cmꢁ1 2969, 2933, 1746, 1679, 1633, 1585, 1488, 1442,
5.1.14. (3R)-6-Acetylamino-7-naphthalen-1-ylmethyl-5-
oxo-8-phenyl-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-3-
carboxylic acid methyl ester (43). By following the pro-
cedure described for the preparation of 42 from 22, 23
(60 mg, 0.14 mmol) in pyridine, with acetic anhydride
(26 ll, 0.28 mmol) and after 7 h adding more acetic
anhydride (26 ll, 0.28 mmol) (overnight) gave 43
(53 mg, 81%) after purification with silica gel chroma-
tography (heptane/EtOAc/MeOH, 5:15:1). [a]D ꢁ180 (c
0.5, CHCl3); IR m/cmꢁ1 1751, 1682, 1631, 1582, 1514,
1
1398, 1366, 1211, 1156, 746, 705; H NMR (400 MHz,
CDCl3) d 7.80–7.70 (m, 2H), 7.64 (d, J = 8.1 Hz, 1H),
7.44–7.34 (m, 2H), 7.29–7.16 (m, 4H), 7.16–7.08 (m,
1H), 7.01 (d, J = 7.1 Hz, 1H), 6.92 (d, J = 7.0 Hz, 1H),
6.87 (s, 1H), 5.69 (dd, J = 2.1, 8.4 Hz, 1H), 4.32–4.09
(m, 2H), 3.86 (s, 3H), 3.67 (dd, J = 8.7, 11.7 Hz, 1H),
3.47 (dd, J = 2.3, 11.8 Hz, 1H), 2.36–2.24 (m, 1H),
0.96 (d, J = 7.0 Hz, 3H), 0.91 (d, J = 7.0 Hz, 3H); 13C
NMR (100 MHz, CDCl3) d 176.11, 168.28, 158.52,
148.19, 144.07, 136.34, 134.38, 133.46, 131.59, 129.76,
129.56, 128.62 (2C, splitted), 128.44, 128.15, 126.72,
125.76, 125.44, 125.24, 124.84, 123.31, 122.30, 116.84,
64.11, 53.38, 35.64, 33.10, 31.68, 19.22 (2C); HRMS
(FAB+) calcd for [M+H]+ C30H29N2O4S 513.1848, obsd
513.1857.
1
1487, 1437, 1366, 1254, 1212, 1154, 1005, 780, 698; H
NMR (400 MHz, CDCl3) d 7.78 (d, J = 7.6 Hz, 1H),
7.72 (d, J = 8.1 Hz, 1H), 7.63 (d, J = 8.1 Hz, 1H),
7.45–7.34 (m, 2H), 7.30–7.24 (m, 1H), 7.25–7.11 (m,
4H), 7.10–7.03 (m, 1H), 6.99–6.89 (m, 2H), 5.66 (d,
J = 8.0 Hz, 1H), 4.28–4.10 (m, 2H), 3.83 (s, 3H), 3.71–
3.61 (m, 1H), 3.47 (d, J = 11.6 Hz, 1H), 1.87 (s, 3H);
13C NMR (100 MHz, CDCl3) d 169.40, 168.21, 158.49,
148.81, 144.56, 136.21, 134.35, 133.46, 131.51, 129.79,
129.56, 128.58 (2C), 128.46, 128.17, 126.78, 125.77,
125.46, 125.23, 125.02, 123.27, 122.29, 116.57, 64.16,
53.32, 33.08, 31.62, 23.23; HRMS (FAB+) calcd for
[M+H]+ C28H25N2O4S 485.1535, obsd 485.1520.
5.1.17. (3R)-8-Cyclopropyl-6-methanesulfonylamino-7-
naphthalen-1-ylmethyl-5-oxo-2,3-dihydro-5H-thiazolo[3,2-
a]pyridine-3-carboxylic acid methyl ester (46). Com-
pound 22 (60 mg, 0.15 mmol) was dissolved in 0.2 ml
pyridine and methane sulfonyl chloride (23 ll,
0.30 mmol) was added. After 5 h CH2Cl2 was added
and the organic layer was washed with 10% citric acid
(aq). The combined aqueous layers were reextracted
with CH2Cl2. The combined organic layers were dried
over Na2SO4(s), filtered and concentrated. Purification
with silica gel chromatography (heptane/EtOAc/MeOH,
50:50:1 ! 20:40:1) gave 46 (63 mg, 88%). [a]D ꢁ160 (c
0.5, CHCl3); IR m/cmꢁ1 1745, 1631, 1581, 1498, 1434,
1392, 1317, 1243, 1218, 1149, 973, 779; 1H NMR
(400 MHz, CDCl3) d 8.20 (d, J = 8.9 Hz, 1H), 7.85 (d,
J = 8.1 Hz, 1H), 7.70 (d, J = 8.2 Hz, 1H), 7.60–7.47
(m, 2H), 7.32–7.26 (m, 1H), 6.83 (d, J = 7.1 Hz, 1H),
6.47 (s, 1H), 5.70 (dd, J = 1.5, 8.5 Hz, 1H), 5.00–4.76
(m, 2H), 3.82 (s, 3H), 3.70 (dd, J = 8.9, 12.1 Hz, 1H),
3.53 (dd, J = 1.6, 12.0 Hz, 1H), 3.03 (s, 3H), 1.22–1.13
(m, 1H), 0.71–0.52 (m, 4H); 13C NMR (100 MHz,
CDCl3) d 167.99, 159.09, 155.40, 147.29, 134.21,
133.68, 131.85, 128.64, 126.93, 126.16, 125.74, 125.24,
124.30, 123.49, 121.43, 114.73, 63.46, 53.41, 41.08,
31.69, 31.54, 12.03, 7.80, 7.39; HRMS (FAB+) calcd
for [M+H]+ C24H25N2O5S2 485.1205, obsd 485.1209.
5.1.15. (3R)-8-Cyclopropyl-6-isobutyrylamino-7-naphtha-
len-1-ylmethyl-5-oxo-2,3-dihydro-5H-thiazolo[3,2-a]pyri-
dine-3-carboxylic acid methyl ester (44). Compound 22
(55 mg, 0.14 mmol) was dissolved in 0.2 ml pyridine.
Then isobutyryl chloride (15 ll, 0.14 mmol) was added
and the solution was stirred at rt for 5 h before heating
it to 45 °C overnight. Additional isobutyryl chloride
(3 ll) was added together with 0.5 ml CH2Cl2 and stir-
ring at rt was continued for 6 h. CH2Cl2 was added
and the organic layer was washed with 10% citric acid
(aq). The combined aqueous layers were reextracted
with CH2Cl2. The combined organic layers were dried
over Na2SO4(s), filtered and concentrated. Purification
with silica gel chromatography (heptane/EtOAc/MeOH,
5:20:1) gave 44 (44 mg, 68%). [a]D ꢁ219 (c 0.5, CHCl3);
IR m/cmꢁ1 1745, 1681, 1631, 1587, 1504, 1267, 1209, 790;
1H NMR (400 MHz, CDCl3) d 8.10 (d, J = 8.2 Hz, 1H),
7.54 (d, J = 7.9 Hz, 1H), 7.70 (d, J = 8.2 Hz, 1H), 7.57–
7.43 (m, 2H), 7.33–7.27 (m, 1H), 6.91 (d, J = 7.1 Hz,
1H), 6.87 (s, 1H), 5.63 (dd, J = 8.3, 1.7 Hz, 1H), 4.69–
4.50 (m, 2H), 3.83 (s, 3H), 3.69 (dd, J = 8.8, 11.7 Hz,
1H), 3.52 (dd, J = 2.0, 11.7 Hz, 1H), 2.38–2.26 (m,
1H), 1.39–1.30 (m, 1H), 0.99 (d, J = 6.9 Hz, 3H), 0.94
(d, J = 6.9 Hz, 3H), 0.78–0.56 (m, 4H); 13C NMR
(100 MHz, CDCl3) d 176.36, 168.42, 158.48, 150.69,
145.02, 134.56, 133.62, 131.81, 128.66, 126.78, 126.03,
125.64, 125.40, 124.10, 123.32, 122.35, 114.32, 63.33,
53.29, 35.60, 32.14, 31.59, 19.25 (2C), 11.86, 7.80, 7.45;
HRMS (FAB+) calcd for [M+H]+ C27H29N2O4S
477.1848, obsd 477.1853.
5.1.18.
(3R)-6-Methanesulfonylamino-7-naphthalen-1-
ylmethyl-5-oxo-8-phenyl-2,3-dihydro-5H-thiazolo[3,2-a]pyr-
idine-3-carboxylic acid methyl ester (47). By following
the procedure described for the preparation of 46 from
22, 23 (60 mg, 0.14 mmol) in pyridine (0.2 ml), and
methane sulfonyl chloride (21 ll, 0.27 mmol) gave 47
(58 mg, 84%) after purification with silica gel chroma-
tography (heptane/EtOAc, 1:3). [a]D ꢁ125 (c 0.5,
CHCl3); IR m/cmꢁ1 1751, 1631, 1577, 1494, 1442, 1390,
1
5.1.16. (3R)-6-Isobutyrylamino-7-naphthalen-1-ylmethyl-
5-oxo-8-phenyl-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-3-
carboxylic acid methyl ester (45). By following the proce-
dure described for the preparation of 44 from 22, 23
(50 mg, 0.11 mmol) in pyridine (0.2 ml), and isobutyryl
1319, 1216, 1145, 975, 777, 700; H NMR (400 MHz,
CDCl3) d 7.78–7.71 (m, 2H), 7.63 (d, J = 8.1 Hz, 1H),
7.43–7.32 (m, 2H), 7.23–7.17 (m, 1H), 7.16–7.09 (m,
2H), 7.08–6.98 (m, 2H), 6.87 (d, J = 7.9 Hz, 1H), 6.79
(d, J = 7.0 Hz, 1H), 6.55 (s, 1H), 5.74 (dd, J = 8.7,

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
7574
2.2 Hz, 1H), 4.65–4.33 (m, 2H), 3.83 (s, 3H), 3.67 (dd,
J = 8.7, 11.8 Hz, 1H), 3.45 (dd, J = 2.1, 11.8 Hz, 1H),
3.11 (s, 3H); 13C NMR (100 MHz, CDCl3) d 167.88,
159.20, 153.03, 146.74, 135.79, 133.97, 133.48, 131.60,
129.77, 129.31, 128.54, 128.47, 128.35, 128.23, 126.90,
125.86, 125.47, 125.44, 125.06, 123.41, 121.27, 117.10,
64.31, 53.48, 41.31, 32.60, 31.57; HRMS (FAB+) calcd
for [M+H]+ C27H25N2O5S2 521.1205, obsd 521.1212.
were dried over Na2SO4(s), filtered and concentrated.
Purification with silica gel chromatography (heptane/
EtOAc/MeOH, 130:70:1 ! 20:40:1) gave 49 (30 mg,
41%): [a]D 0 (c 0.5, CHCl3); IR m/cmꢁ1 2923, 1750,
1710, 1644, 1582, 1488, 1441, 1398, 1346, 1261, 1214,
1145, 1114, 1016, 941, 793, 748, 695; As a mixture of
1
rotamers ꢀ3:2 H NMR (400 MHz, CDCl3) d 8.75 (s,
0.3H), 8.70 (s, 0.6H), 7.74–7.57 (m, 3H), 7.39–7.08 (m,
6H), 7.02–6.92 (m, 1H), 6.80–6.74 (m, 1H), 6.62–6.53
(m, 1H), 5.76–5.70 (m, 1H), 4.58–4.06 (m, 2H), 3.89 (s,
1.3H), 3.84 (s, 1.8H), 3.78–3.67 (m, 1.5H), 3.62–3.53
(m, 0.6H), 3.54–3.46 (m, 1.1H), 1.60–1.47 (m, 6H); 13C
NMR (100 MHz, CDCl3) d 167.75, 161.62 (maj),
161.14 (min), 158.03 (min), 157.67 (min), 157.52 (maj),
157.44 (maj), 150.05 (maj), 149.81 (min), 135.38 (min),
135.31 (maj), 133.34 (maj), 133.26 (maj), 133.19 (2 min
C), 131.41 (maj), 131.34 (min), 129.92 (maj), 129.86
(min), 129.42 (maj), 129.16 (min), 128.48 (maj), 128.42
(min), 128.28 (maj), 128.25 (min), 128.17 (maj), 128.12
(splitted), 127.99 (min), 126.93 (maj), 126.88 (min),
126.72, 125.68 (maj), 125.54 (min), 125.26, 125.19
(min), 125.10 (maj), 123.05 (maj), 123.00 (min), 118.88
(min), 118.60 (maj), 116.85 (maj), 116.69 (min), 64.42
(maj), 64.22 (min), 56.76 (min), 56.46 (maj), 53.63
(min), 53.38 (maj), 32.50 (maj), 32.44 (min), 31.75
(min), 31.66 (maj), 16.87 (maj), 16.76 (min), 15.82
(maj), 15.73 (min); HRMS (FAB+) calcd for [M+H]+
C30H29N2O6S2 577.1467, obsd 577.1475.
5.1.19. (3R)-8-Cyclopropyl-6-[formyl-(propane-2-sulfo-
nyl)-amino]-7-naphthalen-1-ylmethyl-5-oxo-2,3-dihydro-
5H-thiazolo[3,2-a]pyridine-3-carboxylic acid methyl ester
(48). Isopropyl carbamic acid methyl ester (18 mg,
0.15 mmol) was dissolved in 1 ml THF and cooled to
ꢁ42 °C, then t-BuOK (0.95 M in THF, 115 ll,
0.11 mmol) was added. After 40 min, a solution of 40
(45 mg, 0.10 mmol) in 1 ml THF was added. Additional
THF (1 ml) was used to transfer remains of 40 into the
reaction vessel. After 30 min, isopropyl sulfonyl chloride
(14 ll, 0.13 mmol) was added and after another 70 min
the temperature was allowed to reach rt. After 3 h,
10% citric acid (aq) was added and the product was
extracted with CH2Cl2. The combined organic layers
were dried over Na2SO4(s), filtered and concentrated.
Purification with centrifugal preparative silica gel chro-
matography
(heptane/EtOAc/MeOH,
130:70:1 !
20:40:1) gave 48 (20 mg, 36%): [a]D ꢁ31 (c 0.5, CHCl3);
IR m/cmꢁ1 2929, 2849, 1749, 1709, 1641, 1581, 1492,
1436, 1347, 1267, 1214, 1141, 1110, 1033, 963, 793,
749, 693; As a mixture of rotamers ꢀ3:2 1H NMR
(400 MHz, CDCl3) d 8.77 (s, 1H), 8.20–8.14 (m, 1H),
7.87–7.82 (m, 1H), 7.72–7.67 (m, 1H), 7.59–7.53 (m,
1H), 7.52–7.53 (m, 1H), 7.52–7.47 (m, 1H), 7.37–7.29
(m, 1H), 7.06 (d, J = 7.1 Hz, 1H), 5.73–5.56 (m, 1H),
4.81–4.55 (m, 2H), 3.87 (s, 1.1H), 3.82 (s, 1.8H), 3.81–
3.70 (m, 1H), 3.70–3.48 (m, 2H), 3.57–3.51 (m, 3H),
3.50–3.43 (m, 3H), 1.08–0.91 (m, 1H), 0.68–0.45 (m,
4H); 13C NMR (100 MHz, CDCl3) d 167.92 (min),
167.89 (maj), 161.73 (maj), 161.30 (min), 159.47 (maj),
159.38 (min), 158.03 (min), 157.51 (maj), 150.33 (broad),
133.62 (maj), 133.59 (min), 133.48 (maj), 133.33 (min),
131.74, 128.78, 126.95, 126.11 (maj), 126.09 (min),
125.72–125.43 (m, 3C), 123.23 (maj), 123.12 (min),
119.19 (min), 118.81 (maj), 114.40 (maj), 114.06 (min),
63.56 (min), 63.50 (maj), 56.75 (min), 56.42 (maj),
53.57 (min), 53.35 (maj), 32.00 (min), 31.98 (maj),
31.69 (broad), 16.87 (maj), 16.77 (min), 15.75 (maj),
15.69 (min), 12.11 (maj), 11.97 (min), 8.53 (min), 8.49
(maj), 7.25 (min), 6.74 (maj); HRMS (FAB+) calcd for
[M+H]+ C27H29N2O6S2 541.1467, obsd 541.1478.
Compounds 60–63 were synthesized according to pub-
lished procedures. Data in agreement with published
data.47
5.1.21. (3R)-8-Cyclopropyl-6-methylcarbamoyl-7-naph-
thalen-1-ylmethyl-5-oxo-2,3-dihydro-5H-thiazolo[3,2-a]-
pyridine-3-carboxylic acid methyl ester (64). By following
the procedure described for the preparation of 66 from
62, 62 (60 mg, 0.14 mmol) and methylamine (2 M in
THF, 140 ll, 0.28 mmol) gave 64 as a white foam
(59 mg, 96%): [a]D ꢁ162 (c 0.5, CHCl3); IR m/cmꢁ1
3256, 2992, 2949, 1750, 1649, 1531, 1483, 1436, 1214,
1168 cmꢁ1 1H NMR (400 MHz, CDCl3) d 8.41 (d,
;
J = 4.76, 1H), 8.19 (d, J = 8.42, 1H), 7.84 (d, J = 7.96,
1H), 7.67 (d, J = 8.14, 1H), 7.59–7.45 (m, 2H), 7.30
(m, 1H), 6.86 (d, J = 7.04, 1H), 5.70 (dd, J = 8.96,
2.47, 1H), 5.35–5.15 (m, 2H), 3.87 (s, 3H), 3.73 (dd,
J = 11.89, 8.96, 1H), 3.53 (dd, J = 11.89, 2.47, 1H),
2.78 (d, J = 4.76, 3H), 1.24 (m, 1H), 0.73–0.51 (m,
4H); 13C NMR (100 MHz, CDCl3) d 168.33, 165.90,
160.80, 160.18, 149.92, 135.63, 133.70, 132.05, 128.65,
126.42, 125.91, 125.53, 125.50, 123.94, 123.39, 119.18,
116.18, 63.81, 53.45, 33.04, 31.17, 26.10, 11.87, 8.16,
7.53; HRMS (FAB+) calcd for [M+H]+ C25H25N2O4S
449.1535, obsd 449.1540.
5.1.20. 6-[Formyl-(propane-2-sulfonyl)-amino]-7-naphtha-
len-1-ylmethyl-5-oxo-8-phenyl-2,3-dihydro-5H-thiazolo-
[3,2-a]pyridine-3-carboxylic acid methyl ester (49). Com-
pound 41 (60 mg, 0.13 mmol) was dissolved in 1.5 ml
THF and the solution was cooled to ꢁ42 °C. Then
t-BuOK (0.95 M in THF, 135 ll, 0.13 mmol) was added
dropwise. After stirring at ꢁ42 °C for 45 min, isopropyl
sulfonyl chloride (43 ll, 0.38 mmol) was added. After
1.5 h the temperature was allowed to reach rt and the
reaction mixture was stirred overnight. Saturated NaH-
CO3(aq.) and brine was added and the product was
extracted with CH2Cl2. The combined organic layers
5.1.22. (3R)-6-Methylcarbamoyl-7-naphthalen-1-ylmeth-
yl-5-oxo-8-phenyl-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-
3-carboxylic acid methyl ester (65). By following the pro-
cedure described for the preparation of 64 from 62, 63
(65 mg, 0.14 mmol) and methylamine gave 65 as a white
foam (65 mg, 93%): [a]D ꢁ154 (c 0.5, CHCl3); IR m/cmꢁ1
3283, 3000, 2937, 1751, 1651, 1534, 1483, 1441, 1214,
1152 cmꢁ1 1H NMR (400 MHz, CDCl3) d 8.51 (d,
;

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
7575
J = 4.67, 1H), 7.74 (d, J = 8.05, 1H), 7.67–7.59 (m, 2H),
7.42–7.25 (m, 3H), 7.14–6.92 (m, 5H), 6.87 (d, J = 7.50,
1H), 5.74 (dd, J = 8.96, 2.74, 1H), 4.87–4.67 (m, 2H),
3.90 (s, 3H), 3.71 (dd, J = 11.98, 8.96, 1H), 3.46 (dd,
J = 11.98, 2.74, 1H), 2.77 (d, J = 4.67, 3H); 13C NMR
(100 MHz, CDCl3) d 168.18, 165.76, 161.03, 157.94,
149.85, 135.83, 135.74, 133.41, 131.68, 129.90, 129.45,
128.50, 128.44, 128.31, 128.26, 126.39, 125.55, 125.19
(2C), 124.89, 123.15, 118.98, 118.48, 64.70, 53.51,
33.78, 31.17, 26.09; HRMS (FAB+) calcd for [M+H]+
C28H25N2O4S 485.1535, obsd 485.1534.
allowed to attain rt while stirring overnight and was
then concentrated and lyophilized from MeCN/H2O
(ꢀ1:2) to give quantitative yields of the lithium
carboxylates.
Data in agreement with published data for 8 and 9.18,19
5.2.1. (3R)-8-Cyclopropyl-7-naphthalen-1-ylmethyl-6-ni-
tro-5-oxo-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-3-lith-
ium carboxylate (24). [a]D ꢁ216 (c 0.5, MeOH); IR m/
cmꢁ1 1632, 1614, 1485, 1386, 1333, 1217, 1168; 1H
NMR (400 MHz, MeOD-d4) d 8.14 (d, J = 7.8 Hz,
1H), 7.88 (d, J = 7.4 Hz, 1H), 7.75 (d, J = 7.9 Hz, 1H),
7.62–7.47 (m, 2H), 7.38–7.30 (m, 1H), 7.08 (d,
J = 6.5 Hz, 1H), 5.55 (d, J = 8.4 Hz, 1H), 4.75–4.50
(m, 2H), 3.88–3.78 (m, 1H), 3.67 (d, J = 11.3 Hz, 1H),
1.35–1.20 (m, 1H), 0.68–0.51 (m, 4H); 13C NMR
(100 MHz, MeOD-d4) d 173.10, 155.86, 155.52, 149.09,
140.77, 135.18, 134.07, 133.07, 129.86, 128.36, 127.43,
126.88, 126.50, 126.04, 124.00, 114.05, 68.15, 34.21,
32.37, 12.34, 8.40, 7.77.
5.1.23. (3R)-8-Cyclopropyl-6-isopropylcarbamoyl-7-naph-
thalen-1-ylmethyl-5-oxo-2,3-dihydro-5H-thiazolo[3,2-a]pyri-
dine-3-carboxylic acid methyl ester (66). Compound 62
(60 mg, 0.14 mmol), HOAt (24 mg, 0.18 mmol) and
DCC (28 mg, 0.14 mmol) were dissolved in CH2Cl2
(1.5 ml) and isopropylamine (24 ll, 0.28 mmol) was add-
ed at rt. After stirring overnight the reaction mixture was
concentrated. Purification by silica gel chromatography
(heptane/EtOAc, 1:9) gave 66 as a white foam (60 mg,
91%): [a]D ꢁ188 (c 0.5, CHCl3); IR m/cmꢁ1 3007, 2976,
1753, 1652, 1525, 1483, 1214, 1168 cmꢁ1
;
1H NMR
5.2.2.
(3R)-7-Naphthalen-1-ylmethyl-6-nitro-5-oxo-8-
(400 MHz, CDCl3) d 8.18 (d, J = 8.32, 1H), 7.84 (d,
J = 7.87, 1H), 7.78 (d, J = 6.95, 1H), 7.68 (d, J = 8.32,
1H), 7.59–7.45 (m, 2H), 7.31 (m, 1H), 6.89 (d, J = 6.77,
1H), 5.71 (dd, J = 8.78, 2.01, 1H), 5.22–5.07 (m, 2H),
4.01 (m, 1H), 3.86 (s, 3H), 3.71 (dd, J = 11.89, 8.78,
1H), 3.52 (dd, J = 11.89, 2.01, 1H), 1.26 (m, 1H), 1.08–
0.96 (m, 6H), 0.73–0.50 (m, 4H); 13C NMR (100 MHz,
CDCl3) d 168.30, 164.32, 160.36, 158.99, 149.49, 135.56,
133.67, 132.03, 128.63, 126.46, 125.93, 125.52, 125.47,
124.06, 123.35, 120.28, 115.93, 63.57, 53.42, 41.14,
32.98, 31.38, 22.49 (2C, splitted), 11.80, 8.10, 7.38;
HRMS (FAB+) calcd for [M+H]+ C27H29N2O4S
477.1848, obsd 477.1843.
phenyl-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-3-lithium
carboxylate (25). The hydrolysis of 21 required a total of
2.0 equiv LiOH (0.1 M aq) and the obtained lithium car-
boxylate 25 was therefore treated with AmberliteÒ
IR120+ ion-exchange resin to yield the corresponding
carboxylic acid of 25. Characterization was performed
on the carboxylic acid. [a]D ꢁ83 (c 0.5, DMSO); IR m/
cmꢁ1 1744, 1649, 1477, 1342, 1220, 1155, 800, 778,
1
703; H NMR (400 MHz, DMSO-d6) d 13.85 (bs, 1H),
7.88–7.83 (m, 1H), 7.78–7.72 (m, 2H), 7.49–7.37 (m,
3H), 7.26–7.06 (m, 6H), 5.74 (dd, J = 1.9, 9.3 Hz, 1H),
4.26–4.05 (m, 2H), 3.95 (dd, J = 9.3, 12.0 Hz, 1H),
3.62 (dd, J = 1.9, 12.0 Hz, 1H); 13C NMR (100 MHz,
DMSO-d6) d 169.14, 154.22, 153.21, 145.64, 138.79,
134.97, 133.40, 132.69, 131.23, 130.35, 130.07, 129.16,
129.09 (2C), 128.88, 127.64, 126.71, 126.25, 125.73,
125.44, 123.25, 113.84, 64.99, 32.39, 31.94.
5.1.24.
(3R)-6-Isopropylcarbamoyl-7-naphthalen-1-yl-
methyl-5-oxo-8-phenyl-2,3-dihydro-5H-thiazolo[3,2-a]pyri-
dine-3-carboxylic acid methyl ester (67). By following the
procedure described for the preparation of 66 from 62, 63
(65 mg, 0.14 mmol) and isopropylamine gave 67 as a
white foam (65 mg, 92%): [a]D ꢁ152 (c 0.5, CHCl3); IR
m/cmꢁ1 2970, 2953, 1753, 1649, 1526, 1484, 1442, 1216,
5.2.3. (3R)-6-Amino-8-cyclopropyl-7-naphthalen-1-yl-
methyl-5-oxo-2,3--5H-thiazolo[3,2-a]pyridine-3-lithium
carboxylate (26). [a]D ꢁ179 (c 0.5, MeOH); IR m/cmꢁ1
1612, 1557, 1505, 1396, 1274, 1024, 782; 1H NMR
(400 MHz, MeOD-d4) d 8.25 (d, J = 8.5 Hz, 1H), 7.88
(d, J = 8.1 Hz, 1H), 7.72 (d, J = 8.2 Hz, 1H), 7.62–7.53
(m, 1H), 7.53–7.48 (m, 1H), 7.34–7.27 (m, 1H), 7.03
(dd, J = 7.1, 0.7 Hz, 1H), 5.48 (dd, J = 1.5, 8.3 Hz,
1H), 4.62–4.51 (m, 2H), 3.72 (dd, J = 8.4, 11.3 Hz,
1H), 3.59 (dd, J = 1.5, 11.3 Hz, 1H), 1.52–1.43 (m,
1H), 0.64–0.57 (m, 2H), 0.57–0.50 (m, 1H), 0.48–0.40
(m, 1H); 13C NMR (100 MHz, MeOD-d4) d 174.35,
158.30, 136.15, 135.42, 134.25, 133.67, 133.42, 131.24,
129.81, 128.02, 127.15, 126.70, 126.65, 124.78, 124.32,
116.42, 67.67, 34.02, 31.67, 12.75, 7.59, 7.36.
1156 cmꢁ1 1H NMR (400 MHz, CDCl3) d 7.79 (d,
;
J = 7.14, 1H), 7.74 (d, J = 7.96, 1H), 7.71–7.59 (m, 2H),
7.43–7.27 (m, 3H), 7.16–6.96 (m, 5H), 6.91 (d, J = 7.50,
1H), 5.75 (dd, J = 8.96, 2.01, 1H), 4.76–4.57 (m, 2H),
3.89 (s, 3H), 3.69 (dd, J = 11.98, 8.78, 1H), 3.46 (dd,
J = 11.89, 2.01, 1H), 1.02–0.95 (m, 6H); 13C NMR
(100 MHz, CDCl3) d 168.14, 164.18, 160.52, 156.68,
149.35, 135.89, 135.68, 133.39, 131.70, 129.91, 129.46,
128.53, 128.47, 128.29, 128.25, 126.43, 125.62, 125.25
125.15, 124.92, 123.15, 119.72, 118.68, 64.44, 53.49,
41.11, 33.53, 31.37, 22.45, 22.38; HRMS (FAB+) calcd
for [M+H]+ C30H29N2O4S 513.1848, obsd 513.1841.
5.2. General procedure for hydrolysis into lithium
carboxylates 8–9, 24–27, 34–39, 50–59 and 68–71
5.2.4. (3R)-6-Amino-7-naphthalen-1-ylmethyl-5-oxo-8-
phenyl-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-3-lithium
carboxylate (27). [a]D ꢁ165 (c 0.5, MeOH); IR m/cmꢁ1
1621, 1566, 1487, 1440, 1395, 1372, 1315, 790, 766,
685; H NMR (400 MHz, MeOD-d4) d 7.85–7.78 (m,
2H), 7.69 (d, J = 8.2 Hz, 1H), 7.44–7.36 (m, 2H),
The methyl ester was dissolved in THF/MeOH (3:7) to a
concentration of 50 mM then 1.0 equiv of 0.1 M LiOH
(aq) was added dropwise at 0 °C. The solution was
1

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
7576
7.36–7.31 (m, 1H), 7.22–7.02 (m, 6H), 5.54 (dd,
J = 1.5, 8.3 Hz, 1H), 4.11–4.01 (m, 2H), 3.70 (dd,
J = 8.4, 11.3 Hz, 1H), 3.53 (dd, J = 1.6, 11.3 Hz,
5.2.6. (3R)-6-(2-Acetylamino-3-hydroxy-propionylami-
no)-7-naphthalen-1-ylmethyl-5-oxo-8-phenyl-2,3-dihydro-
5H-thiazolo[3,2-a]pyridine-3-lithium carboxylate (35). (i)
By following the procedure described for the preparation
of 34 from 28, 29 (38 mg, 0.06 mmol) gave the tert-butyl-
deprotected methyl ester (25 mg, 72%) after purification
with silica gel chromatography (heptane/EtOAc/MeOH,
5:40:4 ! 5:60:8). [a]D ꢁ124 (c 0.5 in CHCl3); IR m/cmꢁ1
1744, 1641, 1630, 1581, 1493, 1440, 1371, 1260, 1201,
1128, 1073, 1012, 979, 794, 749, 703; As a mixture of dia-
1H); 13C NMR (100 MHz, MeOD-d4)
d 174.23,
158.34, 138.93, 135.26, 134.80, 134.07, 133.60,
133.28, 131.11 (broad), 130.53 (broad), 129.64,
129.28 (2C, broad), 128.75, 128.22, 128.06, 126.97,
126.58 (2C, splitted), 125.35, 124.09, 119.16, 68.47,
34.04, 32.36.
1
5.2.5. (3R)-6-(2-Acetylamino-3-hydroxy-propionylami-
no)-8-cyclopropyl-7-naphthalen-1-ylmethyl-5-oxo-2,3-di-
hydro-5H-thiazolo[3,2-a]pyridine-3-lithium carboxylate
(34). (i) Compound 28 (42 mg, 0.071 mmol) was dis-
solved in 2 ml CH2Cl2/TFA (9:1) and stirred at rt over-
night. The solution was then concentrated, first from
CH2Cl2/TFA 9:1, then from THF/MeOH/H2O and
finally from toluene. Purification with silica gel
chromatography (heptane/EtOAc/MeOH, 5:40:4 !
5:60:8) gave the tert-butyl-deprotected methyl ester
(25 mg, 66%). [a]D ꢁ155 (c 0.5 in CHCl3); IR m/cmꢁ1
2960, 1748, 1629, 1579, 1500, 1436, 1344, 1258, 1200,
1175, 1126, 1064, 1018, 965, 793, 747; As a mixture
stereomers ꢀ1:1 H NMR (400 MHz, CDCl3) d 8.14–
8.04 (m, 1H), 7.80–7.75 (m, 1H), 7.74–7.68 (m, 1H),
7.66 (d, J = 8.3 Hz, 1H), 7.45–7.36 (m, 2H), 7.33–7.27
(m, 1H), 7.20–6.95 (m, 6.7H), 6.83–6.77 (d, J = 6.9 Hz,
0.4H), 5.82–5.73 (m, 1H), 4.38–4.28 (m, 1H), 4.22–4.07
(m, 2H), 3.84–3.76 (m, 4H), 3.74–3.39 (m, 4H), 1.62–
1.44 (m, 3H); 13C NMR (100 MHz, CDCl3) d 171.12,
171.06, 171.02, 170.86, 168.34, 168.30, 158.59, 150.59,
150.52, 146.62, 135.66, 133.49, 133.40, 131.42, 129.68,
129.32, 128.70 (2C), 128.53, 128.47, 128.43 (2C),
127.10, 126.08, 125.75, 125.71, 125.44, 124.62, 124.53,
123.16, 123.07, 122.09, 122.02, 117.25, 117.19, 64.47,
62.51, 62.33, 55.27, 54.87, 53.56, 53.54, 32.41, 31.67,
22.23, 22.13; HRMS (FAB+) calcd for [M+Na]+
C28H30N3NaO5S 594.1675, obsd 594.1676.
1
of diastereomers ꢀ5:4 H NMR (400 MHz, CDCl3) d
8.11 (dd, J = 8.4, 2.7 Hz, 1H), 8.04–7.94 (m, 1H),
7.85 (dd, J = 8.0 Hz, 1H), 7.69 (dd, J = 8.1 Hz, 1H),
7.61–7.48 (m, 2H), 7.32–7.25 (m, 1H), 7.05–6.97 (m,
0.5H), 6.90–6.77 (m, 1.5H), 5.74–5.66 (m, 1H), 4.62–
4.51 (m, 2H), 4.40–4.26 (m, 1H), 3.85–3.40 (m, 8H),
1.58 (s, 1.7H), 1.52 (s, 1.3H), 1.40–1.30 (m, 1H),
0.71–0.61 (m, 2H), 0.59–0.49 (m, 2H); 13C NMR
(100 MHz, CDCl3) d 171.34, 171.21, 170.94, 170.72,
168.44, 168.40, 158.53, 153.10, 147.32, 133.67, 133.47,
133.42, 131.72, 128.77, 128.70, 127.08, 126.30 (splitted),
125.91, 125.86, 125.56 (splitted), 123.94, 123.80, 123.25,
123.14, 122.24, 122.14, 114.69, 114.60, 63.63, 62.65,
62.37, 55.31, 54.87, 53.47, 53.45, 31.60 (2C), 22.32,
22.13, 11.71, 7.37, 7.23, 7.15; HRMS (FAB+) calcd
for [M+Na]+ C28H29N3NaO6S 558.1675, obsd
558.1678.
(ii) The purified product from the tert-butyl deprotec-
tion of 29 was then subjected to alkaline ester hydrolysis
according to the general procedure to yield 35.
[a]D ꢁ114 (c 0.5, MeOH); IR m/cmꢁ1 1621, 1490, 1371,
1
791, 701; As a mixture of diastereomers ꢀ1:1 H NMR
(400 MHz, MeOD-d4) d 7.78–7.74 (m, 1H), 7.72–7.67
(m, 1H), 7.64 (d, J = 8.2 Hz, 1H), 7.41–7.32 (m, 2H),
7.32–7.26 (m, 1H), 7.15–6.95 (m, 6H), 5.56–5.51 (m,
1H), 4.40 (t, J = 5.3 Hz, 0.5H), 4.36 (t, J = 5.1 Hz,
0.5H), 4.19–4.07 (m, 2H), 3.80–3.72 (m, 1H), 3.72–3.65
(m, 1.4H), 3.60–3.53 (m, 1.6H), 1.75 (s, 1.6H), 1.71 (s,
1.4H); 13C NMR (100 MHz, MeOD-d4) d 173.86,
173.37, 173.26, 172.78, 172.69, 160.54, 160.48, 151.48,
151.40, 149.73, 149.63, 137.88, 135.17, 134.97, 132.94,
131.33 (splitted, broad), 130.80 (splitted, broad), 129.45
(splitted), 129.39 (2C), 129.02, 127.80 (splitted), 126.90
(splitted), 126.65–126.41 (m, 3C), 124.15, 124.10,
122.79, 122.56, 117.80 (splitted), 68.83, 63.18, 62.96,
57.31, 56.85, 34.02, 33.03, 33.00, 22.28, 22.25.
(ii) The purified product from the tert-butyl deprotec-
tion was then subjected to alkaline ester hydrolysis
according to the general procedure to yield 34.
[a]D ꢁ190 (c 0.5, MeOH); IR m/cmꢁ1 1617, 1570, 1503,
1435, 1377, 1269, 1201, 1179, 1137, 1059, 1034, 792;
As
a
mixture of diastereomers ꢀ5:4 1H NMR
5.2.7. (3R)-6-(2-Acetylamino-acetylamino)-8-cyclopro-
pyl-7-naphthalen-1-ylmethyl-5-oxo-2,3-dihydro-5H-thiaz-
olo[3,2-a]pyridine-3-lithium carboxylate (36). [a]D ꢁ198
(c 0.5, MeOH); IR m/cmꢁ1 1621, 1498, 1371, 1262,
1029, 791, 663; 1H NMR (400 MHz, MeOD-d4) d
8.18 (d, J = 8.3 Hz, 1H), 7.86 (d, J = 8.0 Hz, 1H),
7.70 (d, J = 8.2 Hz, 1H), 7.59–7.53 (m, 1H), 7.52–
7.46 (m, 1H), 7.34–7.28 (m, 1H), 7.04 (d, J = 7.1 Hz,
1H), 5.50–5.44 (m, 1H), 4.62–4.49 (m, 2H), 3.86–3.75
(m, 2H), 3.76–3.69 (m, 1H), 3.63–3.57 (m, 1H), 1.77
(s, 3H), 1.31-1.22 (m, 1H), 0.64–0.45 (m, 4H); 13C
(400 MHz, MeOD-d4) d 8.23–8.16 (m, 1H), 7.86 (d,
J = 8.1 Hz, 1H), 7.71 (d, J = 8.2 Hz, 1H), 7.59–7.54
(m, 1H), 7.53–7.47 (m, 1H), 7.34–7.28 (m, 1H),
7.06–7.01 (m, 1H), 5.51–5.46 (m, 1H), 4.65–4.50 (m,
2H), 4.39 (t, J = 5.5 Hz, 0.6H), 4.33 (t, J = 5.0 Hz,
0.5H), 3.80–3.55 (m, 4H), 1.74 (s, 1.7H), 1.69 (s,
1.3H), 1.32–1.24 (m, 1H), 0.63–0.44 (m, 4H); 13C
NMR (100 MHz, MeOD-d4)
d
173.99, 173.39,
173.26, 172.76, 172.70, 160.36, 160.33, 153.87,
153.83, 150.54, 150.40, 135.45 (splitted), 135.22,
133.31, 129.75, 129.73, 127.85 (splitted), 127.82,
127.17, 126.73, 126.70, 126.64, 125.76, 125.62,
124.40, 124.34, 122.93, 122.68, 115.25, 115.19, 68.02,
63.18, 62.96, 57.33, 56.79, 33.96, 32.53, 32.46, 22.28,
22.22, 12.70 (splitted), 8.11, 8.07, 7.80, 7.71.
NMR (100 MHz, MeOD-d4)
d
173.96, 173.65,
171.65, 160.36, 153.80, 150.32, 135.54, 135.20,
133.26, 129.75, 127.82, 127.17, 126.70, 126.63,
125.78, 124.34, 122.50, 115.09, 67.95, 43.73, 33.94,
32.51, 22.21, 12.74, 8.17, 7.81.

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
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5.2.8. (3R)-6-(2-Acetylamino-acetylamino)-7-naphthalen-
1-ylmethyl-5-oxo-8-phenyl-2,3-dihydro-5H-thiazolo[3,2-
a]pyridine-3-lithium carboxylate (37). [a]D ꢁ138 (c 0.5,
MeOH); IR m/cmꢁ1 1617, 1565, 1494, 1384, 1370, 1283,
50.59 (min), 50.48 (maj), 34.06, 33.05, 22.18 (min),
22.09 (maj), 17.69 (min), 17.49 (maj).
5.2.11. (3R)-8-Cyclopropyl-6-formylamino-7-naphthalen-
1-ylmethyl-5-oxo-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-
3-lithium carboxylate (50). [a]D ꢁ138 (c 0.3, DMSO);
IR m/cmꢁ1 1610, 1562, 1501, 1472, 1395, 1378, 1273,
1250, 775, 661; As a mixture of rotamers ꢀ7:3 1H
NMR (400 MHz, MeOD-d4) d 8.22–8.14 (m, 1H),
8.07 (s, 0.3H), 8.02 (s, 0.6H), 7.90–7.84 (m, 1H),
7.75–7.68 (m, 1H), 7.61–7.47 (m, 2H), 7.36–7.28 (m,
1H), 7.02 (dd, J = 0.7, 7.1 Hz, 0.7H), 7.00–6.96 (m,
0.3H), 5.53–5.48 (m, 1H), 4.72–4.52 (m, 2H), 3.82–
3.73 (m, 1H), 3.66–3.59 (m, 1H), 1.43–1.27 (m, 1H),
0.66–0.47 (m, 4H); 13C NMR (100 MHz, MeOD-d4)
d 173.93 (maj), 173.88 (min), 168.19 (min), 163.38
(maj), 160.98 (min), 160.05 (maj), 153.24 (maj),
151.38 (min), 150.55 (maj), 150.16 (min), 135.27,
135.21 (maj), 134.97 (min), 133.29 (splitted), 129.83
(min), 129.79 (maj), 128.17 (min), 127.86 (maj),
127.33 (min), 127.18 (maj), 126.89 (min), 126.71
(maj), 126.64 (maj), 126.55 (min), 125.60 (maj),
125.27 (min), 124.12, 122.49 (min), 121.68 (maj),
115.14 (min), 115.08 (maj), 68.07 (min), 67.96 (maj),
34.03, 32.65 (maj), 32.36 (min), 12.70 (maj), 12.63
(min), 8.11 (broad), 7.75 (maj), 7.63 (min).
1
1262, 773, 702; H NMR (400 MHz, MeOD-d4) d 7.78–
7.73 (m, 1H), 7.69 (d, J = 8.0 Hz, 1H), 7.64 (d,
J = 8.1 Hz, 1H), 7.41–7.32 (m, 2H), 7.32–7.26 (m, 1H),
7.15–6.98 (m, 6H), 5.53 (dd, J = 1.2, 8.6 Hz, 1H), 4.12
(s, 2H), 3.81 (d, J = 5.6 Hz, 2H), 3.74 (dd, J = 8.7,
11.2 Hz, 1H), 3.55 (dd, J = 1.2, 11.3 Hz, 1H), 1.78
(s, 3H); 13C NMR (100 MHz, MeOD-d4) d 173.84,
173.62, 171.64, 160.54, 151.40, 149.53, 137.95, 135.29,
134.97, 132.91, 131.33, 130.86, 129.47, 129.41 (2C,
broad), 128.99, 127.78, 126.89, 126.65, 126.46 (2C,
broad), 124.13, 122.37, 117.69, 68.75, 43.75, 34.03,
33.07, 22.21.
5.2.9. (3R)-6-(2-Acetylamino-propionylamino)-8-cyclo-
propyl-7-naphthalen-1-ylmethyl-5-oxo-2,3-dihydro-5H-
thiazolo[3,2-a]pyridine-3-lithium carboxylate (38). [a]D
ꢁ190 (c 0.5, DMSO); IR m/cmꢁ1 1620, 1565, 1487,
1408, 1381, 1264, 1155, 670; As a mixture of diaster-
eomers ꢀ10:3; 1H NMR (400 MHz, MeOD-d4) d
8.21–8.15 (m, 1H), 7.86 (d, J = 8.1 Hz, 1H), 7.70 (d,
J = 8.3 Hz, 1H), 7.59–7.53 (m, 1H), 7.52–7.46 (m,
1H), 7.33–7.28 (m, 1H), 7.01 (d, J = 7.1 Hz, 1H),
5.48 (dd, J = 1.6, 8.6 Hz, 1H), 4.63–4.48 (m, 2H),
4.34–4.21 (m, 1H), 3.79–3.71 (m, 1H), 3.61 (dd,
J = 1.6, 11.3 Hz, 1H), 1.70 (s, 0.7H), 1.62 (s, 2.3H),
1.37–1.26 (m, 1H), 1.15 (d, J = 7.1 Hz, 2.4H), 1.00
(d, J = 7.1 Hz, 0.7H), 0.64–0.44 (m, 4H); 13C NMR
5.2.12. (3R)-6-Formylamino-7-naphthalen-1-ylmethyl-5-
oxo-8-phenyl-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-3-
lithium carboxylate (51). [a]D ꢁ136 (c 0.5, MeOH); IR
m/cmꢁ1 1674, 1610, 1565, 1493, 1381, 1286, 1259, 766,
700; As a mixture of rotamers ꢀ7:3; 1H NMR
(400 MHz, MeOD-d4) d 8.15 (s, 0.3H), 8.04 (s,
0.6H), 7.82–7.62 (m, 3H), 7.42–7.28 (m, 3H), 7.17–
7.05 (m, 6H), 5.59–5.53 (m, 1H), 4.18 (s, 0.6H), 4.14
(s, 1.4H), 3.82–3.73 (m, 1H), 3.61 (m, 1H); 13C
NMR (100 MHz, MeOD-d4) d 173.79 (maj), 173.74
(min), 168.17 (min), 163.33 (maj), 161.11 (min),
160.19 (maj), 150.76 (maj), 149.73 (maj), 149.38
(min), 148.80 (min), 137.93 (maj), 137.76 (min),
135.07 (min), 135.03 (maj), 135.00 (maj), 134.74
(min), 132.94 (maj), 132.89 (min), 131.34, 130.85
(maj), 130.74 (min), 129.62 (min), 129.59–129.36 (m),
129.22 (min), 129.08 (maj), 128.17 (min), 127.83
(maj), 127.07 (min), 126.92 (maj), 126.69 (min),
126.49 (2 maj C), 126.41 (min), 126.37 (maj), 126.05
(min), 123.89, 122.44 (min), 121.58 (maj), 117.74
(min), 117.66 (maj), 68.89 (min), 68.77 (maj), 34.12,
33.22 (maj), 32.88 (min).
(100 MHz, MeOD-d4)
d
174.66, 173.96, 173.04
(maj), 172.95 (min), 160.29, 153.58 (min), 153.55
(maj), 150.44 (min), 150.25 (maj), 135.42 (maj),
135.33 (min), 135.19, 133.28, 129.76, 127.81 (min),
127.78 (maj), 127.16 (splitted), 126.70 (2C, splitted),
125.61 (maj), 125.52 (min), 124.29 (maj), 124.25
(min), 122.66 (maj), 122.46 (min), 115.26 (min),
115.12 (maj), 67.95 (splitted), 50.60 (min), 50.44
(maj), 33.97 (broad), 32.48 (splitted), 22.16 (min),
22.09 (maj), 17.70 (min), 17.46 (maj), 12.69, 8.09,
7.81.
5.2.10. (3R)-6-(2-Acetylamino-propionylamino)-7-naph-
thalen-1-ylmethyl-5-oxo-8-phenyl-2,3-dihydro-5H-thia-
zolo[3,2-a]pyridine-3-lithium carboxylate (39). [a]D ꢁ163
(c 0.5, MeOH); IR m/cmꢁ1 1613, 1493, 1375, 1289, 1258,
770, 689; As a mixture of diastereomers ꢀ2:1; 1H NMR
(400 MHz, MeOD-d4) d 7.79–7.74 (m, 1H), 7.71 (d,
J = 8.0 Hz, 1H), 7.67–7.62 (m, 1H), 7.41–7.27 (m, 3H),
7.14–7.01 (m, 6H), 5.56–5.51 (m, 1H), 4.35–4.22 (m,
1H), 4.17–4.05 (m, 2H), 3.79–3.71 (m, 1H), 3.59–3.53
(m, 1H), 1.71 (s, 1H), 1.63 (s, 2H), 1.14 (d, J = 7.1 Hz,
2H), 0.96 (d, J = 7.1 Hz, 1H); 13C NMR (100 MHz,
MeOD-d4) d 174.66 (maj), 174.61 (min), 173.83, 173.04
(maj), 172.94 (min), 160.44 (min), 160.40 (maj), 151.07,
149.59 (min), 149.41 (maj), 137.96 (maj), 137.92 (min),
135.17 (maj), 135.06 (min), 134.95, 132.92, 131.29 (split-
ted), 130.76, 129.58-129.32 (m, 3C), 129.08 (min), 129.02
(maj), 127.78 (min), 127.73 (maj), 126.93 (min), 126.89
(maj), 126.57–126.35 (m, 2C+ 1 maj C), 126.27 (min),
124.06 (maj), 123.99 (min), 122.54 (maj), 122.37 (min),
117.81 (min), 117.73 (maj), 68.77 (min), 68.72 (maj),
5.2.13. (3R)-6-Acetylamino-8-cyclopropyl-7-naphthalen-
1-ylmethyl-5-oxo-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-
3-lithium carboxylate (52). [a]D ꢁ240 (c 0.5, MeOH);
IR m/cmꢁ1 1615, 1565, 1497, 1391, 1342, 1265, 1016,
791, 770; 1H NMR (400 MHz, MeOD-d4) d 8.16 (d,
J = 8.3 Hz, 1H), 7.86 (d, J = 7.8 Hz, 1H), 7.70 (d,
J = 8.2 Hz, 1H), 7.59–7.52 (m, 1H), 7.52–7.46 (m,
1H), 7.34–7.28 (m, 1H), 7.04 (d, J = 7.1 Hz, 1H),
5.48 (dd, J = 1.6, 8.6 Hz, 1H), 4.62–4.50 (m, 2H),
3.74 (dd, J = 8.6, 11.3 Hz, 1H), 3.6 (dd, J = 1.6,
11.3 Hz, 1H), 1.85 (s, 3H), 1.37–1.26 (m, 1H), 0.68–
0.49 (m, 4H); 13C NMR (100 MHz, MeOD-d4) d
173.99, 173.30, 160.52, 126.64, 150.10, 135.57,

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
7578
135.19, 133.24, 129.80, 127.78, 127.13, 126.66, 126.64,
125.83, 124.14, 123.00, 115.05, 67.91, 33.97, 32.44,
22.46, 12.74, 8.19, 7.82.
J = 1.4, 8.8 Hz, 1H), 4.94–4.77 (m, 2H), 3.85 (dd,
J = 8.9, 12.0 Hz, 1H), 3.60 (dd, J = 1.5, 12.0 Hz, 1H),
3.02 (s, 3H), 1.17–1.08 (m, 1H), 0.64–0.48 (m, 4H); 13C
NMR (100 MHz, MeOD-d4) d 170.86, 160.95, 157.46,
150.11, 135.77, 135.23, 133.31, 129.74, 127.85, 127.16,
126.73, 126.41, 125.74, 124.40, 122.54, 115.62, 65.23,
41.95, 32.73, 32.56, 13.01, 8.57, 8.14.
5.2.14. (3R)-6-Acetylamino-7-naphthalen-1-ylmethyl-5-
oxo-8-phenyl-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-3-
lithium carboxylate (53). [a]D ꢁ165 (c 0.5, MeOH); IR m/
cmꢁ1 1615, 1570, 1491, 1439, 1372, 1285, 1259, 772, 696;
1H NMR (400 MHz, MeOD-d4) d 7.79–7.75 (m, H),
7.74–7.69 (m, H), 7.65 (d, J = 8.2 Hz, H), 7.41–7.33
(m, H), 7.33–7.27 (m, H), 7.20–7.06 (m, H), 5.53 (dd,
J = 1.6, 8.6 Hz, H), 4.16–4.06 (m, H), 3.74 (dd, J = 8.7,
11.4 Hz, H), 3.55 (dd, J = 1.6, 11.4 Hz, H), 1.84 (s, H);
13C NMR (100 MHz, MeOD-d4) d 173.86, 173.26,
160.67, 151.18, 149.28, 138.06, 135.34, 134.97, 132.90,
131.36, 130.89, 129.55, 129.44 (2C), 129.04, 127.75,
126.87, 126.58, 126.48, 126.43, 123.94, 122.86, 117.68,
68.71, 34.06, 33.03, 22.49.
5.2.18.
(3R)-6-Methanesulfonylamino-7-naphthalen-1-
ylmethyl-5-oxo-8-phenyl-2,3-dihydro-5H-thiazolo[3,2-a]pyr-
idine-3-carboxylic acid (57). Hydrolysis of 47 required
additional 0.5 equiv LiOH and the residue was therefore
treated with AmberliteÒ IR120+ ion-exchange resin to
yield 57. [a]D ꢁ106 (c 0.5, DMSO); IR m/cmꢁ1 1749,
1624, 1497, 1385, 1311, 1161, 1112, 775; 1H NMR
(400 MHz, MeOD-d4) d 7.77–7.73 (m, 1H), 7.69 (d,
J = 8.3 Hz, 1H), 7.62 (d, J = 8.1 Hz, 1H), 7.40–7.30
(m, 2H), 7.25–7.19 (m, 1H), 7.11–7.05 (m, 2H), 7.04–
6.97 (m, 1H), 6.96–6.90 (m, 1H), 6.87–6.81 (m, 2H),
5.76 (dd, J = 1.7, 8.9 Hz, 1H), 4.55–4.33 (m, 2H),
3.90–3.82 (m, 1H), 3.56 (dd, J = 1.7, 11.9 Hz, 1H),
3.09 (s, 3H); 13C NMR (100 MHz, MeOD-d4) d
170.82, 161.11, 155.17, 149.52, 137.66, 135.48, 135.00,
133.00, 131.07, 128.80, 129.51, 129.47, 129.43, 129.12,
127.75, 126.83 (2C, broad), 126.42, 126.22, 124.23,
122.40, 118.10, 66.06, 42.03, 33.42, 32.65.
5.2.15. (3R)-8-Cyclopropyl-6-isobutyrylamino-7-naphtha-
len-1-ylmethyl-5-oxo-2,3-dihydro-5H-thiazolo[3,2-a]pyri-
dine-3-lithium carboxylate (54). [a]D ꢁ223 (c 0.3, DMSO);
IR m/cmꢁ1 1615, 1565, 1497, 1391, 1342, 1265, 1016, 791,
770; 1H NMR (400 MHz, DMSO-d6) d 8.72 (s, 0.9H), 8.30
(s, 0.2H), 8.16 (d, J = 8.3 Hz, 1H), 7.9 (d, J = 8.1 Hz, 1H),
7.72 (d, J = 8.2 Hz, 1H), 7.59–7.48 (m, 2H), 7.36–7.30 (m,
1H), 6.93 (d, J = 7.0 Hz, 1H), 5.11 (dd, J = 1.5, 7.9 Hz,
1H), 4.44–4.34 (m, 2H), 3.63–3.52 (m, 2H), 2.41–2.33
(m, 1H), 1.24–1.16 (m, 1H), 0.81 (d, J = 6.8 Hz, 3H),
0.72 (d, J = 6.8 Hz, 3H), 0.60–0.49 (m, 2H), 0.44–0.33
(m, 2H); 13C NMR (100 MHz, DMSO-d6) d 176.14,
168.74, 158.48, 148.81, 146.96, 135.24, 133.65, 131.98,
129.01, 126.68, 126.50, 126.02 (2C), 124.74, 123.68,
122.95, 110.85, 66.87, 34.14, 33.76, 31.27, 19.84, 19.75,
11.99, 7.82, 7.31.
5.2.19. (3R)-8-Cyclopropyl-7-naphthalen-1-ylmethyl-5-
oxo-6-(propane-2-sulfonylamino)-2,3-dihydro-5H-thiazolo-
[3,2-a]pyridine-3-carboxylic acid (58). Compound 48
(19 mg, 0.035 mmol) was dissolved in 0.7 ml THF/
MeOH (3:7) and 0.1 M LiOH (aq.) (705 ll, 0.071 mmol)
was added dropwise at 0 °C. The solution was allowed
to attain rt while stirred overnight and was then cocon-
centrated, first from EtOH and then MeOH. This gave
the lithium carboxylate of 58 together with the lithium
formate from the deprotection. To remove the lithium
formate, the residue was dissolved in 2.0 ml MeOH/
CH2Cl2 (3:1) and swirled with a small spoon of Amber-
liteÒ IR120+ ion-exchange resin. The resin was removed
by filtration and the filtrate was concentrated to give 58
(16 mg, 91%). [a]D ꢁ38 (c 0.1, DMSO); IR m/cmꢁ1 2923,
5.2.16. (3R)-6-Isobutyrylamino-7-naphthalen-1-ylmethyl-
5-oxo-8-phenyl-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-3-
lithium carboxylate (55). [a]D ꢁ132 (c 0.4, DMSO); IR m/
cmꢁ1 1617, 1568, 1492, 1440, 1393, 1291, 770; 1H NMR
(400 MHz, MeOD-d4) d 7.80–7.75 (m, 1H), 7.75–7.69
(m, 1H), 7.65 (d, J = 8.2 Hz, 1H), 7.42–7.33 (m, 2H),
7.33–7.28 (m, 1H), 7.18–7.07 (m, 6H), 5.55 (dd,
J = 1.6, 8.7 Hz, 1H), 4.15–4.04 (m, 2H), 3.77 (dd,
J = 8.7, 11.4 Hz, 1H), 3.56 (dd, J = 1.6, 11.4 Hz, 1H),
2.42–2.31 (m, 1H), 0.92 (d, J = 6.9 Hz, 3H), 0.74 (d,
J = 6.9 Hz, 3H); 13C NMR (100 MHz, MeOD-d4) d
179.65, 173.90, 160.69, 150.82, 149.10, 138.14, 135.18,
134.96, 132.92, 131.31, 130.83, 129.57, 129.48 (2C),
129.08, 127.70, 126.88, 126.56, 126.45, 126.25, 123.86,
122.88, 117.81, 68.71, 36.05, 34.11, 33.07, 19.80, 19.37.
1
2848, 1741, 1715, 1621, 1499, 1311, 1243, 1134, 772; H
NMR (400 MHz, MeOD-d4) d 8.23 (d, J = 8.3 Hz, 1H),
7.87 (d, J = 8.1 Hz, 1H), 7.71 (d, J = 8.3 Hz, 1H), 7.59–
7.54 (m, 1H), 7.52–7.47 (m, 1H), 7.33–7.28 (m, 1H), 6.92
(dd, J = 0.7, 7.1 Hz, 1H), 5.68 (dd, J = 1.6, 8.8 Hz, 1H),
4.92–4.80 (m, 2H), 3.85 (dd, J = 8.9, 11.9 Hz, 1H), 3.61
(dd, J = 1.7, 12.0 Hz, 1H), 3.43–3.32 (m, 1H), 1.36 (d,
J = 6.8 Hz, 6H), 1.16–1.08 (m, 1H), 0.65–0.50 (m, 4H);
13C NMR (100 MHz, MeOD-d4) d 170.87, 161.24,
157.57, 149.80, 135.78, 135.26, 133.35, 129.76, 127.88,
127.18, 126.74, 126.44, 125.85, 124.41, 122.99, 115.59,
65.18, 55.94, 32.97, 32.60, 17.08 (2C), 13.05, 8.55, 8.19;
HRMS (FAB+) calcd for [M+H]+ C25H27N2O5S2
499.1361, obsd 499.1378.
5.2.17.
(3R)-8-Cyclopropyl-6-methanesulfonylamino-
7-naphthalen-1-ylmethyl-5-oxo-2,3-dihydro-5H-thiazolo-
[3,2-a]pyridine-3-carboxylic acid (56). Hydrolysis of 46
required additional 0.5 equiv LiOH and the residue
was therefore treated with AmberliteÒ IR120+ ion-ex-
change resin to yield 56. [a]D ꢁ144 (c 0.5, DMSO); IR
m/cmꢁ1 1729, 1625, 1498, 1392, 1313, 1244, 1144, 970,
777; 1H NMR (400 MHz, MeOD-d4) d 8.21 (d,
J = 8.4 Hz, 1H), 7.85 (d, J = 8.1 Hz, 1H), 7.70 (d,
J = 8.2 Hz, 1H), 7.58–7.52 (m, 1H), 7.51–7.45 (m, 1H),
7.32–7.26 (m, 1H), 6.89 (d, J = 7.1 Hz, 1H), 5.70 (dd,
5.2.20. 7-Naphthalen-1-ylmethyl-5-oxo-8-phenyl-6-(pro-
pane-2-sulfonylamino)-2,3-dihydro-5H-thiazolo[3,2-a]pyri-
dine-3-carboxylic acid (59). By following the procedure
described for the preparation of 58 from 48, 49
(28 mg, 0.049 mmol) in 1 ml THF/MeOH (3:7), and
0.1 M LiOH (aq) (0.975 ml, 0.098 mmol) gave the lith-

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
7579
ium carboxylate of 59 together with the lithium for-
mate from the deprotection. The lithium formate
was removed using AmberliteÒ IR120+ ion-exchange
resin as described in the preparation of 58. This gave
59 (26 mg, quant.). [a]D 0 (c 0.5, MeOH); IR m/cmꢁ1
1736, 1625, 1493, 1256, 1132; 1H NMR (400 MHz,
MeOD-d4) d 7.76–7.73 (m, 1H), 7.69 (d, J = 8.2 Hz,
1H), 7.62 (d, J = 8.2 Hz, 1H), 7.39–7.29 (m, 2H),
7.25–7.19 (m, 1H), 7.09–6.98 (m, 3H), 6.91–6.85 (m,
3H), 5.72 (dd, J = 1.7, 8.9 Hz, 1H), 4.51–4.39 (m,
2H), 3.81 (dd, J = 8.9, 12.0 Hz, 1H), 3.53 (dd,
J = 1.7, 12.0 Hz, 1H), 3.49–3.38 (m, 1H), 1.44 (t,
J = 6.9 Hz, 6H); 13C NMR (100 MHz, MeOD-d4) d
170.78, 161.33, 155.25, 149.14, 137.72, 135.46,
134.96, 132.98, 131.07, 130.78, 129.44 (3C, splitted),
129.05, 127.74, 126.92, 126.81, 126.40, 126.23,
124.21, 122.81, 118.05, 65.94, 55.99, 33.65, 32.63,
17.16, 17.08; HRMS (FAB+) calcd for [M+H]+
C28H27N2O5S2 535.1361, obsd 535.1360.
154.69, 152.53, 135.90, 135.14, 133.34, 129.74, 127.75,
127.14, 126.64, 126.57, 126.26, 124.71, 124.12, 115.17,
67.69, 42.61, 34.00, 33.43, 22.23, 22.14, 12.40, 7.99, 7.92.
5.2.24. (3R)-6-Isopropylcarbamoyl-7-naphthalen-1-yl-
methyl-5-oxo-8-phenyl-2,3- dihydro-5H-thiazolo[3,2-a]pyr-
idine-3-lithium carboxylate (71). [a]D ꢁ42 (c 0.5, MeOH);
IR m/cmꢁ1 3341 (broad), 2965, 3058, 2971, 2922, 1613,
1491, 1387, 1238; 1H NMR (400 MHz, MeOD-d4) d
7.79–7.60 (m, 3H), 7.41–7.28 (m, 3H), 7.23
(dd,J = 8.32, 0.64, 1H), 7.18–6.93 (m, 5H), 5.57 (dd,
J = 8.69, 1.46, 1H), 4.41 (d, J = 16.01, 1H), 4.12 (d,
J = 16.01, 1H), 3.85–3.70 (m, 2H), 3.59 (dd, J = 11.43,
1.46, 1H), 1.02 (d, J = 6.50, 3H), 0.64 (d, J = 6.50,
3H); 13C NMR (100 MHz, MeOD-d4)
d 174.05,
167.87, 160.96, 152.56, 152.01, 137.73, 135.71, 134.89,
133.00, 131.35, 130.99, 129.45 (2C), 129.39, 129.09,
127.69, 127.11, 126.84 (2C), 126.38, 124.13, 123.88,
118.51, 68.54, 42.61, 34.09, 33.74, 22.19, 22.16.
5.2.21. (3R)-8-Cyclopropyl-6-methylcarbamoyl-7-naph-
thalen-1-ylmethyl-5-oxo-2,3-dihydro-5H-thiazolo[3,2-a]py-
ridine-3-lithium carboxylate (68). [a]D ꢁ131 (c 0.5,
MeOH); IR m/cmꢁ1 3320 (broad), 3062, 3003, 2938,
1611, 1501, 1388, 1243, 1224, 1170; 1H NMR
(400 MHz, MeOH-d4) d 8.18 (d, J = 8.32, 1H), 7.86 (d,
J = 7.59, 1H) 7.70 (d, J = 8.32, 1H), 7.61–7.45 (m, 2H),
7.32 (dd,J = 8.96, 2.47, 1H), 7.06 (dd,J = 7.04, 0.73,
1H), 5.52 (dd, J = 8.69, 1.56, 1H), 4.80–4.60 (m, 2H),
3.77 (dd, J = 11.34, 8.69, 1H), 3.63 (dd, J = 11.34, 1.56,
1H), 2.63 (s, 3H), 1.18 (m, 1H), 0.67–0.45 (m, 4H); 13C
NMR (100 MHz, MeOH-d4) d 174.13, 169.74, 161.01,
155.36, 152.83, 139.95, 135.16, 133.31, 129.75, 127.75,
127.12, 126.65, 126.58, 126.15, 124.20 (broad, 2C),
115.71, 67.79, 33.89, 33.67, 26.35, 12.44, 8.26, 8.03.
5.3. Computational procedures
Flexible alignment of compound 34 to the fixed bioac-
tive conformation of the PapG peptide was performed
using MOE2004.03.33 The PapG peptide coordinates
were obtained from the published crystal complex of
the 19-mer of PapG and PapD.28 The PapG peptide
was truncated to the corresponding length of the pepti-
domimetic 34. The starting conformation of compound
34 was achieved by first generating a 3D structure with
the software CORINA. CORINA can be tested on the
internet at http://www2.chemie.uni-erlangen.de/soft-
ware/corina/free_struct.html and is available from
Molecular Networks GmbH, Germany, info@mol-
net.de, http://www.mol-net.de. The 3D structures were
energy minimized in MOE with the MMFF94x force
field and default settings with preserved chirality. Flexi-
ble alignment was carried out with default settings in
MOE except for the addition of a partial charge similar-
ity term, which was set to 1. This was on the basis that
the carboxylate functionality is important for the
anchoring to the protein.
5.2.22. (3R)-6-Methylcarbamoyl-7-naphthalen-1-ylmeth-
yl-5-oxo-8-phenyl-2,3-dihydro-5H-thiazolo[3,2-a]pyridine-
3-lithium carboxylate (69). [a]D ꢁ56 (c 0.5, MeOH); IR m/
cmꢁ1 3328 (broad), 3055, 2935, 1612, 1492, 1392, 1243,
1155; 1H NMR (400 MHz, MeOD-d4) d 7.74 (d,
J = 7.96, 1H), 7.70–7.60 (m, 2H), 7.42–7.27 (m, 3H),
7.19 (d,J = 6.95, 1H), 7.11–7.63 (m, 5H), 5.57 (dd,
J = 8.78, 1.46, 1H), 4.38 (d, J = 15.64, 1H), 4.21 (d,
J = 15.64, 1H), 3.77 (dd, J = 11.43, 8.78, 1H), 3.57 (dd,
J = 11.43, 1.46, 1H), 2.62 (s, 3H); 13C NMR (100 MHz,
MeOD-d4) d 173.99, 169.55, 161.29, 153.36, 152.41,
137.68, 135.83, 134.88, 132.96, 131.40, 130.98, 129.42,
129.38, 129.35, 129.03, 127.69, 127.14, 126.76, 126.34
(2C), 123.97, 123.45, 118.53, 68.66, 34.01, 33.99, 26.33.
1
5.4. Affinity measurements using H relaxation-edited
NMR spectroscopy
Each sample contained PapD (95 lM), pilicide (95 lM)
and 1-naphthyl acetic acid (included as a non-binding ref-
erence, 95 lM) in phosphate buffer with 5% DMSO-d6.
The samples were prepared from a solution of PapD in
phosphate buffer (50 mM, pH 7.4) and stock solutions
of pilicide and 1-naphthyl acetic acid (2.5 mM in
DMSO-d6). Additional DMSO-d6 was added to obtain a
final 5% content. A 200 ms cpmg spin-lock filter was used
to efficiently removethe signalsfrom PapD and bound pil-
icide. Suppression of water signal was accomplished with
a WATERGATE sequence.49 Spectra were recorded at
25 °C on a Bruker DRX 600 MHz spectrometer.
5.2.23. (3R)-8-Cyclopropyl-6-methylcarbamoyl-7-naph-
thalen-1-ylmethyl-5-oxo-2,3-dihydro-5H-thiazolo[3,2-a]-
pyridine-3-lithium carboxylate (70). [a]D ꢁ83 (c 0.5,
MeOH); IR m/cmꢁ1 3327 (broad), 2965, 2937, 1617,
1501, 1408, 1217; 1H NMR (400 MHz, MeOD-d4) d
8.17 (d, J = 8.32, 1H), 7.86 (d, J = 7.87, 1H), 7.70
(d,J = 8.32, 1H), 7.60–7.44 (m, 2H), 7.33 (dd, J = 8.32,
7.23, 1H), 7.08 (d, J = 7.23, 1H), 5.52 (dd, J = 8.78,
1.37, 1H), 4.79–4.61 (m, 2H), 3.85–3.71 (m, 2H), 3.64
(dd, J = 11.43, 1.37, 1H), 1.28 (m, 1H), 1.01 (d, J = 6.59,
3H), 0.67 (d, J = 6.59, 3H), 0.66–0.47 (m, 4H); 13C
NMR (100 MHz, MeOD-d4) d 174.18, 168.03, 160.72,
5.5. In vivo activity evaluated with haemagglutination
Escherichia coli HB101/pPAP5 was grown in the
absence or presence of 3.5 mM compound in TSA

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V. Aberg et al. / Bioorg. Med. Chem. 14 (2006) 7563–7581
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(trypticase soy agar) plates for 24 h at 37 °C. The plates
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DMSO and the final DMSO content in the plate was
ꢀ5%. Positive controls were used from plates containing
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diluted in a V-bottomed 96-well microtitre plate prior
to addition of RBC and the plates were kept at 4 °C.
The last well with agglutination was visually determined.
The reported HA-titre is the inverse of the dilution fac-
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pili. The non-pili producing strain HB101/pBR322 was
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Acknowledgment
Almqvist, F. J. Comb. Chem. 2002, 4, 630–639.
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˚
We are grateful to Dr. Anna Linusson at Umea
University for helpful discussions. We thank the
Swedish Natural Science Research Council and the
Knut and Alice Wallenberg foundation for financial
support.
˚
20. Pemberton, N.; Aberg, V.; Almstedt, H.; Westermark, A.;
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22. Pinkner, J. S.; Pemberton, N.; Aberg, V.; Larsson, A.;
Supplementary data
Hedenstro¨m, M.; Emtena¨s, H.; Kihlberg, J.; Hultgren, S.
J.; Almqvist, F. Unpublished.
Supplementary data associated with this article can be
found, in the online version, at doi:10.1016/
j.bmc.2006.07.017.
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