Design of noncompetitive interleukin-8 inhibitors acting on CXCR1 and CXCR2

3984

J. Med. Chem. 2007, 50, 3984-4002

Articles

Design of Noncompetitive Interleukin-8 Inhibitors Acting on CXCR1 and CXCR2

Alessio Moriconi,^†Maria Candida Cesta,^†Maria Neve Cervellera,^†Andrea Aramini,^†Silvia Coniglio,^†Sandro Colagioia,^†

Andrea Rosario Beccari,^†Cinzia Bizzarri,^†Michela Rita Cavicchia,^†Massimo Locati,^‡,§Emanuela Galliera,^‡

Paola Di Benedetto,^†Paolo Vigilante,^†Riccardo Bertini,^†and Marcello Allegretti*^,†

Research Centre, Dompe´ pha.r.ma s.p.a., Via Campo di Pile, 67100, L’Aquila, Italy, Institute of General Pathology, UniVersity of Milan,

20133 Milan, Italy, and Istituto Clinico Humanitas, 20089 Rozzano, Italy

ReceiVed December 22, 2006

Chemokines CXCL8 and CXCL1 play a key role in the recruitment of neutrophils at the site of inflammation.

CXCL8 binds two membrane receptors, CXCR1 and CXCR2, whereas CXCL1 is a selective agonist for

CXCR2. In the past decade, the physiopathological role of CXCL8 and CXCL1 has been investigated. A

novel class of small molecular weight allosteric CXCR1 inhibitors was identified, and reparixin, the first

drug candidate, is currently under clinical investigation in the prevention of ischemia/reperfusion injury in

organ transplantation. Reparixin binding mode to CXCR1 has been studied and used for a computer-assisted

design program of dual allosteric CXCR1 and CXCR2 inhibitors. In this paper, the results of modeling-

driven SAR studies for the identification of potent dual inhibitors are discussed, and three new compounds

(56, 67, and 79) sharing a common triflate moiety have been selected as potential leads with optimized

pharmacokinetic characteristics.

Introduction

CXCR2-selective agonist CXCL1. CXCR1 and CXCR2 are

mainly expressed on neutrophils but low expression is also

evident on keratinocytes, fibroblasts, endothelial, and melanoma

cells.¹⁴

The chemokine family is a group of low molecular weight,

multifunctional cytokines that play a primary role in the

inflammatory response, finely regulating the leukocytes recruit-

ment in the inflamed tissue.^1,2These molecules exert additional

functions in physiological and pathological conditions including

wound healing and tumorigenesis.^3,4

Disregulation of the chemokines network has been evoked

as a crucial event in the insurgence and progression of severe

chronic diseases including rheumatoid arthritis (RA^a), chronic

obstructive pulmonary disease (COPD), asthma, Alzheimer’s

disease, melanoma, ulcerative colitis (UC), multiple sclerosis,

and psoriasis.^5-13Hence, interference with chemokine function

is generally considered a promising approach for the develop-

ment of novel anti-inflammatory medications.

Among chemotactic factors, interleukin-8 (CXCL8) and

related chemokines, including GRO-R (CXCL1), belong to the

CXC chemokines family and play a major role in the activation

and recruitment of neutrophils.³Two membrane receptors, the

type A CXCL8 receptor (CXCR1) and type B CXCL8 receptor

(CXCR2), have been shown to bind CXCL8 with high affinity.¹⁴

CXCR1 and CXCR2 belong to the class A (rhodopsin-like) of

the 7-transmembrane G-protein coupled receptors (7-TM-

GPCRs) and share 78% amino acid identity.¹⁵CXCR1 is

selective for CXCL8, whereas CXCR2 also interacts with the

The role of CXCL8 in acute and chronic inflammatory

conditions characterized by a prominent polymorphonucleate

(PMN) infiltrate as, for instance, UC and COPD, has been

recently reviewed.¹⁶

Furthermore, recent data strongly support the hypothesis that

expression of CXCL8 and its receptors CXCR1 and CXCR2

contributes to aggressive growth and metastasis in human

malignant melanoma.

To date, a limited number of small molecular weight (SMW)

CXCL8 inhibitors have been disclosed in the literature and the

first drug candidates have only recently entered clinical studies

aimed at the assessment of the therapeutic potential of this

class.^17,18

The most comprehensively described class of SMW CXCL8

inhibitors is a class of bisaryl urea derivatives discovered and

characterized by GlaxoSmithKline (GSK) laboratories as selec-

tive CXCR2 antagonists.^19,20The first lead compound in this

series, N-(2-bromophenyl)-N′-(2-hydroxy-4-nitrophenyl)urea

(SB225002, Figure 1), is >150-fold selective for CXCR2 over

CXCR1 in a binding assay but, in vitro, it potently inhibited

human and rabbit neutrophil chemotaxis induced by both

CXCL8 and CXCL1. The in vivo efficacy of the compound

was proved in some relevant animal models such as CXCL8-

induced PMN margination in rabbits,¹⁹but protracted lead

optimization studies delayed the selection of appropriate clinical

candidates. To date, a novel candidate, N-(2-chloro-3-fluorophe-

nyl)-N′-[4-chloro-2-hydroxy-3-(piperazin-1-ylsulfonyl) phenyl]-

urea (SB656933, Figure 1) is in phase I clinical trials for the

treatment of COPD.

* To whom correspondence should be addressed. Phone: +39-0862-

338212. Fax: +39-0862-338215. E-mail: marcello.allegretti@dompe.it.

^†Research Centre, Dompe´ pha.r.ma s.p.a.

^‡University of Milan.

^§Istituto Clinico Humanitas.

^aAbbreviations: I/R, ischemia/reperfusion; RA, rheumatoid arthritis;

COPD, chronic obstructive pulmonary disease; UC, ulcerative colitis; DGF,

delayed graft function; PGD, primary graft dysfunction; PMN, polimor-

phonuclear neutrophil; LPS, lypopolysaccaride; 7-TM-GPCRs, 7-trans-

membrane G-protein coupled receptors; SMW, small molecular weight;

SAR, structure-activity relationship; COX, cyclooxygenase.

Published on Web 08/01/2007

CXCR1/CXCR2 Inhibitors

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17 3985

Figure 1. Chemical structures of SB225002, SB656933, and reparixin.

In recent years, a novel class of SMW noncompetitive

allosteric CXCR1 inhibitors has been widely investigated in our

laboratories.²¹

(2R)-2-[4-(2-methylpropyl)phenyl]-N-(methylsulfonyl)pro-

panamide (reparixin, Figure 1), the compound selected from

this class, is a potent and selective inhibitor of CXCL8-induced

chemotaxis and has been proved efficacious in several ischemia/

reperfusion (I/R) experimental models.^22-24On the basis of its

pharmacological characteristics, reparixin has been selected to

enter clinical development for the prevention and treatment of

the delayed graft function (DGF) in kidney transplant and

primary graft dysfunction (PGD) in lung transplant. Phase II

clinical trials are ongoing for both of these indications.

Reparixin is a potent functional inhibitor of CXCL8-induced

biological activities on human PMNs with a marked selectivity

(around 400-fold) for CXCR1, as shown in specific experiments

on CXCR1/L1.2 and CXCR2/L1.2 transfected cells and on

human PMNs.²²Reparixin mechanism of action has been deeply

investigated leading to the full characterization of a binding

pocket in the transmembrane (TM) region of CXCR1. The

binding of the inhibitor effectively blocks the agonist-induced

receptor signaling in the intracellular compartment, while it does

not directly affect the CXCL8 binding affinity or the number

of expressed receptors.

Figure 2. Migration of L1.2 transfectants expressing wild type CXCR1

(b) and Ile43Val CXCR1 (O) was induced by 10 nM CXCL8 in the

presence or absence of increasing concentrations of reparixin as

indicated. Results are expressed as percent of migration in the absence

of reparixin ((SD) at least in three independent experiments: (**) P

< 0.01 and (*) P < 0.05 vs cell migration in the absence of reparixin

(Mann-Whitney U test). Spontaneous migration was 4000 ( 3000

cells/mL; CXCL8-induced cell migration was 420 000 ( 20 000

cells/mL.

There is conflicting information as to whether neutrophil

chemotaxis, the best characterized biological effect of CXCL8,

is mediated by one or both the CXCL8 receptors.¹⁶

Results and Discussion

Ile43Val Replacement. Effect of Reparixin on hCXCR1

Ile43Val Mutant. The prediction of the above-described model

was that the hCXCR1 Ile43Val mutant still supported CXCL8-

induced chemotaxis, but with reduced sensitivity to reparixin

inhibition. The valine-replacement mutant was expressed tran-

siently in L1.2 cells and receptor expression was confirmed by

flow cytometry analysis (data not shown). The CXCR1 mutant

did not significantly differ from wild-type CXCR1 transfectant

in terms of receptor expression levels, ligand binding properties,

and chemotactic migration to CXCL8.

Reparixin was tested in a wide range of concentrations (1-

1000 nM). As expected, wild-type CXCR1/L1.2 transfectants

migration induced by CXCL8 (10 nM) was significantly

inhibited by reparixin (Figure 2), the inhibition being concentra-

tion-dependent and reaching the maximum (80%) at 0.1 µM of

reparixin. The efficacy of reparixin was significantly lower in

cells expressing Ile43Val CXCR1 mutant (IC₅₀values of 5.6

× 10^-9M and 8.0 × 10^-8M for CXCR1 wt and CXCR1

Ile43Val, respectively).

Chemistry. The results described in this study are shown in

Tables 1-3, and the related synthetic methods are outlined in

Schemes 1-8.

The synthesis of 2-arylpropionic acids was performed fol-

lowing several strategies depending on the different substitution

pattern of the aromatic ring. The choice of different methods

was guided by the commercial availability of starting materials

and reagents. Compounds 4 and 5 were obtained starting from

intermediate 80, prepared as described,²¹by direct coupling with

2-methyl-1-propenyltributyltin and cyclohexylzinc bromide,

As previously discussed, our data support a major role of

CXCR1 signaling in the regulation of human neutrophil

chemotaxis, which is the key pathophysiological event in post-

ischemia reperfusion conditions.

Starting from the above considerations, we attempted to

exploit the information on the reparixin binding mode to

rationally design a novel class of potent and selective CXCL8

inhibitors with comparable potency on CXCR1 and CXCR2 and

with appropriate pharmacokinetic (PK) profile for the treatment

of chronic diseases.

In fact, accumulating evidence indicates a predominant, if

not exclusive, role of the CXCR2 receptor subtype in complex

pathologies such as psoriasis, COPD, and melanoma.^25-27

Therefore, dual inhibitors of the CXCL8 activity, blocking

both CXCR1 and CXCR2 receptors, could be an appropriate

and a more complete therapeutic strategy in a number of

inflammatory diseases where the CXCR2 pathway is involved

specifically or in conjunction with the CXCR1 signaling.

Here we describe the results obtained by structure-activity

relationship (SAR) studies in the class of 2-arylpropionic

CXCL8 inhibitors that have led to the identification of novel,

potent, and selective noncompetitive dual inhibitors of CXCR1

and CXCR2 receptors.

The demonstration of in vivo efficacy for CXCR1/CXCR2

dual inhibitors could represent a major advance in the treatment

of severe pathological conditions ranging from COPD to

aggressive melanoma.

3986 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17

Moriconi et al.

Table 1. Effect of 3- and 4-Substituted 2-Arylpropionic Acids on

respectively, and following methyl ester basic hydrolysis

(Scheme 1). The direct alkylation of the commercial 2-[4-

(bromomethyl)phenyl] propanoic acid with the appropriate

organomagnesium reagents in Grignard reaction conditions

afforded the 4-alkylderivatives 8-10 (Scheme 2). Compound

20 was obtained by Heck coupling from the intermediate 82²¹

and R-styrene, followed by Pd/C hydrogenation and hydrolysis

(Scheme 3).

CXCL8- and CXCL1-Induced Human PMN Chemotaxis^a

The commercial 3-(1-cyanoethyl)benzoic acid was reacted

with thionyl chloride to give the corresponding acid chloride

that, by treatment with the appropriate Grignard reagent and

acidic hydrolysis, led to the compounds 23-26. Similarly, the

same intermediate was reacted with heteroaryl lithium to give,

after hydrolysis, compounds 27-30. Treatment of the above

acid chloride with sodium azide under Curtius reaction condi-

tions afforded the corresponding arylamine that was converted

under Sandmeyer conditions to the intermediate 81 that, by

reaction with benzenesulfinic acid sodium salt and hydrolysis,

was converted to the sulfone 37 (Scheme 4).

Following a typical Wittig procedure and basic hydrolysis,

the methyl ester of commercial ketoprofen was transformed into

the compound 31; parallely, the vinyl intermediate was hydro-

genated and hydrolyzed to give 35. The direct alkylation of

ketoprofen by methyl magnesium bromide afforded compound

36 (Scheme 5).

Compounds 39 and 40 were prepared starting from the

commercial 2-(4-hydroxyphenyl)propanoic acid and the ap-

propriate acid chloride following a classical Schotten-Baumann

procedure in aqueous solution. The methyl ester of the 2-(4-

hydroxyphenyl)propanoic acid was converted into 4-arylsul-

fonates 51-54 by reaction with selected sulfonyl chlorides and

following hydrolysis (Scheme 6).

The same procedure was used to obtain sulfonamides

44-50 from intermediate 83. Treatment of 83 with benzoyl

chloride and subsequent hydrolysis afforded compound 41, while

the condensation of 83 with 4-chlorobutyryl chloride, and

following basic hydrolysis, allowed to obtain compound 43

(Scheme 7).

(2R)-2-{4-[(Trifluoromethyl)sulfonyl]phenyl}propanoic acid

56 was transformed into compounds 58-62 by reaction with

the appropriate alkyl, aryl or heteroarylsulfonamide and into

compounds 68-79 by classical coupling procedures with

amines. For compound 79, the not commercially available amine

(2-amino-4-trifluoromethylthiazole) was prepared according to

a published procedure.²⁸Compound 56 was converted into the

hydroxamic acid 63 and into N- and O-alkyl derivatives 64-

66 by coupling with commercial N- or O-alkylhydroxylamines

(Scheme 8).

The synthetic methods for the preparation of compounds 1-3,

6, 7, 11-19, 21, 22, 32-34 and intermediates 80 and 82²¹and

of compounds 55, 56, 57, and 67²⁹have been described

before.

Structure-Activity Relationship (SAR) Studies. Reparixin,

a 2-phenylpropionic acid derivative, was previously demon-

strated to act as a noncompetitive allosteric blocker of CXCL8

receptors with a marked selectivity for CXCR1.²²

Alanine scanning mutagenesis studies, supported by molecular

modeling techniques, allowed us to define the overall pattern

of interactions engaged by reparixin in a pocket located in the

TM region of hCXCR1. For a better comprehension of amino

acid residues position in the TM region of the receptor, the

Weinstein-Ballesteros numbering will be used³⁰in the follow-

ing section and throughout the rest of the paper.

^a(*) % of inhibition at 10^-8M. (**) % of inhibition at 10^-7M. n.t.: not

tested.

CXCR1/CXCR2 Inhibitors

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17 3987

Table 2. Effect of 3- and 4-substitued 2-Arylpropionic Acids on CXCL8- and CXCL1-Induced Human PMN Chemotaxis^a

^a(*) % of inhibition at 10^-8M. (**) % of inhibition at 10^-7M. n.t.: not tested.

3988 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17

Moriconi et al.

Table 3. Effect of (R)-2-Arylpropionic Derivatives on CXCL8- and CXCL1-Induced Human PMN Chemotaxis^a

^a(*) % of inhibition at 10^-8M. (**) % of inhibition at 10^-9M. n.t.: not tested.

According to the proposed binding mode, the acylmethane-

sulfonamide moiety engages strong polar interactions with five

This hypothesis was in further agreement with the previous

observation that Ile43Ala mutation in CXCR1 caused a 100-

fold reduction of the inhibitor potency. The significant decrease

of reparixin potency on CXCR1 Ile43Val mutant indirectly

suggests that the loss of hydrophobic interactions could account

for reparixin lower affinity toward CXCR2.

residues in the TM domain (Tyr46_1.39, Lys99_2.64, Asn120_3.35

,

Tyr258_6.51, and Glu291_7.39), whereas three aliphatic amino acids

(Val42_1.35, Val113_3.28, and Ile43_1.36) are putatively involved in

the recognition of the isobutyl residue of reparixin in a

hydrophobic cavity in the TM region of CXCR1.

Supported by these results, we decided to follow a molecular

modeling aided strategy to design dual CXCR1/CXCR2 allos-

teric inhibitors.

The chemotaxis experiments on this set of mutants clearly

showed that hydrophobic interactions strongly contribute to tune

the overall affinity of the inhibitor, since single replacement of

the three hydrophobic residues for Ala amino acid considerably

reduced the potency of reparixin.

In our previous work, SAR in the class of 2-arylpropionic

CXCR1 ligands were extensively investigated by using the

CXCL8-induced human PMN chemotaxis assay as primary

functional screening. Due to the specific objectives of the study,

the CXCR2 selectivity of the test compounds was not exten-

sively analyzed, and the CXCL1-induced chemotaxis assay was

performed on a limited number of molecules selected for the

lead optimization phase.

Reparixin binding site in CXCR2 was also studied by

molecular mechanics, molecular dynamics calculations (MM/

MD) and GRID analysis. In the model, as expected on the basis

of the high CXCR1 and CXCR2 homology in the TM region,

the pattern of polar interactions engaged by reparixin appeared

strictly conserved (Figure 3, panels A and B). In contrast, some

nonconserved residues in the helix bundle apparently account

for the larger dimensions of the hydrophobic cavity in the TM

of CXCR2. In particular, the replacement of Ile43 (CXCR1)

by Val52 (CXCR2) could, to some extent, explain the lower

affinity of the inhibitor.

With the objective to select novel scaffolds for the design of

equipotent CXCR1/2 inhibitors, and in strict analogy with the

approach followed in our previous paper,²¹we first investigated

the selectivity profile of a representative set of racemic

2-phenylpropionic acids in CXCL1-induced human PMN chemo-

CXCR1/CXCR2 Inhibitors

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17 3989

Scheme 1^a

Before proceeding with the lead optimization, we considered

a key factor for the success of the program the selection of

several compounds with different structural and physicochemical

properties. Two 3-substituted-2-phenylpropionic acids, previ-

ously selected by SAR studies as potent CXCL8 inhibitors,

3-isopropyl (13) and 3-benzoyl (21) phenylpropionic acids were

tested for CXCL1 biological activity.

Compound 13 showed a comparable potency in inhibiting

CXCL8- and CXCL1-induced chemotaxis. Binding of 13 in the

TM region of CXCR1 and CXCR2 was investigated by MM/

MD and GRID analysis. Interestingly, both polar and hydropho-

bic interactions engaged by the ligand are well conserved in

the two binding models. According to the model, the forced per-

pendicular orientation of the isopropyl group significantly mo-

difies, if compared to the 4-isobutyl derivatives, the hydrophobic

pattern of recognition that, in this case, involves conserved

residues in the two receptor subtypes (data not shown).

^aReagents and conditions: (a) Pd₂dba₃, AsPh₃, LiCl, CuI, 2-methyl-1-

propenyltributyltin, NMP, 90 °C; (b) KOH, MeOH, rt; (c) cyclohexylzinc

bromide, Pd(PPh₃)₄, LiCl, THF, reflux.

Scheme 2^a

The importance of the orientation of the hydrophobic chain/

group was confirmed by the observed SAR. In the series of

3-alkyl-substituted phenylpropionic acids (13-20), 14 showed

a good potency and, like 13, dual activity. Also the 3-sec-butyl

analogue 15 maintained a moderate activity, while the introduc-

tion of planar unsaturated groups (16) as well as flexible, linear,

or branched alkyl chains (17-20) invariably afforded a dramatic

loss of affinity.

The last phenylpropionic scaffold analyzed for the CXCR2

selectivity was R-ketoprofen (21), the first CXCL8 inhibitor ever

discovered in our laboratories. The polar interactions engaged

by 21 in the TM region of CXCR1 were previously discussed

in deep.²¹To allow a direct comparison of the binding pocket

in the receptor subtypes, a binding model of 21 in CXCR2 was

obtained by molecular modeling techniques. We found the

carboxylic group engaging a double ionic bond with Lys108_2.64

and Glu300_7.39in CXCR2, and in general, the hydrophilic

interactions pattern appeared strictly conserved in the two

receptor subtypes. Furthermore, the analysis shows that the

benzoyl moiety well accommodates (Figure 3, panels C and

D) in a conserved hydrophobic cavity spanned by Val122_3.28

(Val113_3.28in CXCR1), Ile292_7.31(Ile283_7.31), and the aliphatic

side chain of Lys126_3.32(Lys117_3.32). In both receptors, the

complex seems further stabilized by an electrostatic interaction

between the phenol group of Tyr55_1.39(Tyr46_1.39in CXCR1)

and the phenyl group of the ligand.

^aReagents and conditions: (a) RMgBr, CuI, THF, 0 °Cf rt.

Scheme 3^a

^aReagents and conditions: (a) Pd(PPh₃)₂Cl₂, TEA, R-methylstyrene,

DMF, 90 °C; (b) 10% Pd/C, EtOH, rt; (c) KOH, MeOH, rt.

taxis. Only potent CXCL8 inhibitors (>50% at c ) 10^-8M)

were selected for CXCL1 chemotaxis assay.

In agreement with the conserved binding mode, 21 potently

Starting from the above binding mode hypothesis, we

expected that, in the series of 4-substituted-phenylpropionic

acids, the introduction of larger hydrophobic substituents could

favorably influence the affinity to CXCR2.

inhibited CXCL1-induced PMN chemotaxis (Table 2).

A set of 2-(3-acylphenyl)propionic acids (22-30), ketoprofen

analogues, was designed and tested for activity toward CXCL8

and CXCL1.

In agreement with previous SAR studies, the replacement of

the 4-isobutyl residue with more rigid alkyl chains (1-4) or

cyclic aliphatic (5) substituents led to a dramatic loss of

biological activity and, similarly, the introduction of 4-aryl

substituents (6, 7) was not tolerated in CXCR1 binding pocket.

In the proposed binding model, the carbonyl group seems

not involved in direct interactions with polar residues in the

TM region even if, from a geometrical point of view, the amino

acid residue Lys117_3.32in CXCR1 (Lys126_3.32in CXCR2) could

be accessible for hydrogen-bond interaction. As a matter of fact,

a benzophenone carbonyl, due to the electron-withdrawing effect

of the two phenyl groups, is a weak hydrogen-bond acceptor.

Thus, we attempted to reinforce the acceptor characteristics of

the carbonyl group, introducing hydrophobic substituents with

electron-donor properties. Data reported in Table 2 show that

the replacement of the phenyl moiety with small and rigid alkyl

chains (22-24) was detrimental for the affinity of the ligands

and also that the inclusion of cycloalkyl residues (25, 26) did

not afford significant improvement of the receptor affinity.

Flexibility is required for a correct accommodation in the

cavity, and the size of the hydrophobic group was confirmed

as a key factor for the modulation of the affinity at CXCR2.

Nevertheless, as emerged in the series of 4-cycloalkylmethyl

substituted acids (8-12), a limited size increase was tolerated

and the cyclobutylmethyl residue was found as the best

compromise for high affinity to both receptors. Thus, in this

series, 9 was selected as the first potent dual inhibitor of

CXCL8- and CXCL1-induced chemotaxis.

3990 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17

Moriconi et al.

Scheme 4^a

^aReagents and conditions: (a) SOCl₂, reflux; (b) RMgCl, ZnCl₂, (dppf)PdCl₂, 0 °C f rt; (c) R′H (heterocycle), BuLi, THF, ZnCl₂, CuI, -78°f 0 °C;

(d) 37% HCl, 1,4-dioxane, 70 °C; (e) (1) NaN₃, CH₂Cl₂/H₂O, 0 °C; (2) TFA, reflux; (3) K₂CO₃, H₂O/MeOH, 60 °C; (f) concd HCl, NaNO₂, KI, 0°-5 °C;

(g) PhSO₂Na, CuI, L-proline sodium salt, DMSO, 80 °C.

Scheme 5^a

^aReagents and conditions: (a) CH₃MgBr (2 equiv), Et₂O, reflux; (b) concd H₂SO₄, MeOH, rt; (c) Ph₃CH₃P⁺Br^-, BuLi, THF, rt; (d) 1 N NaOH, MeOH,

rt; (e) 10% Pd/C, EtOH, rt.

The key role of aromatic hydrophobic interactions, anticipated

by the above observations, was further confirmed by the SAR

studies in a limited set of heteroaromatic analogues (27-30).

The thiophene derivative 27, a classical benzene bioisoster,

was a very potent dual inhibitor, while the introduction of the

more polar furan ring (28) was not well-tolerated. Analogous

SAR were found in the CXCL8-induced chemotaxis assay by

comparing the 2-oxazolyl (29) with the 2-thiazolyl (30) ana-

logue. Compound 30, as reported in Table 2, was also confirmed

as a potent CXCR2 inhibitor.

On the basis of these results, we argued that, in accordance

with the models, the second aromatic moiety is essential in this

series of derivatives and that the carbonyl group behaves as a

spacer for the correct orientation of the two aromatic rings.

To further support the above assumption, we synthesized the

phenylvinyl derivative (31) that, while loosing hydrogen-bond

CXCR1/CXCR2 Inhibitors

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17 3991

Scheme 7^a

Scheme 6^a

^aReagents and conditions: (a) conc. H₂SO₄, MeOH, rt; (b) RSO₂Cl,

Py, reflux; (c) 37% HCl, glacial AcOH, reflux; (d) 1 N NaOH, RCOCl, rt.

acceptor properties, retains a benzophenone-like spatial disposi-

tion. The obtained results (Table 2) confirmed the scarce

importance of the carbonyl oxygen but the good selectivity of

31 for CXCR1 suggests that the hydrophobic pattern of inter-

actions is not fully conserved in the TM cavities of CXCR1/2.

In apparent contradiction with the above considerations, the

hydroxyl derivatives 32-34, obtained by partial reduction of

21-23, retained good potency, despite the substantial change

in hybridization and geometry of the molecule.

Compound 32 was a good dual inhibitor, but interestingly,

in this series, the replacement of the aromatic moiety (32) for

smaller alkyl chains (33 and 34) did not compromise the activity

of the inhibitors.

Compounds 35 and 36 were synthesized with the aim to gain

further information on the unexpected biological activity of 32-

34. The hydroxyl group seems not directly involved in the

receptor binding, since the substitution of hydroxyl for methyl

(35), but not the methylation of the hydroxyl-bearing carbon

(36), did not compromise the biological activity.

The above-reported results are reminiscent of SAR in the

series of the analogues of the 3-isopropyl derivative 13, thus

suggesting that the two classes of derivatives could share a

similar binding mode in which the hydroxyl moiety is not

directly involved in interactions with the receptor.

On the basis of these considerations, the lack of activity of

the phenylsulfonyl (37) and phenoxyphenyl (38) derivatives is

most likely due to the different orientation and mutual distances

of the aromatic moieties than to electronic effects.

As a further step of the project, the relatively rigid structure

of the ketoprofen scaffold was chosen as template for design

and synthesis of novel 4-substituted phenylpropionic acids with

dual CXCR1/2 activity. With the aim of directing a second

aromatic group in the hydrophobic pocket occupied by the

benzoyl group, several spacers (ester, amide, sulfonate, and

sulfonamide) were evaluated in parallel.

^aReagents and conditions: (a) RSO₂Cl, Py, acetone, rt; (b) 2 N NaOH,

EtOH, rt; (c) RCOCl, Py, acetone, rt; (d) 6% NaOH, i-PrOH, rt.

The benzoic ester (39) showed a moderate activity, as CXCL8

inhibitor, that was completely lost in the corresponding propionic

ester (40). These preliminary results partially supported the

validity of the approach, but the in vivo liability of the ester

group prompted us to shift to alternative scaffolds for the

prosecution of the SAR studies.

The corresponding benzoyl amide (41) was found completely

inactive, whereas the tertiary cyclic amide (42) in which the

aromatic residue is part of a rigid bicyclic structure was a good

and selective CXCL8 inhibitor. The marked rigidity of the

structure could explain the weak inhibitory potency of 42 as

CXCL1 inhibitor, but also in this case, the crucial relevance of

the second aromatic ring was confirmed by the complete activity

loss of the corresponding oxopyrrolidinyl derivative (43).

With the aim of refining the geometry and the physicochem-

ical properties of the spacer, the activity of 4-aryl and 4-alkyl-

sulfonylamino derivatives was investigated.

The simple 4-methanesulfonamide (44) was completely

devoid of biological activity, but the replacement of the methyl

group with the isopropyl (45) led to a significant potency

increase on CXCL8. As expected, the introduction of a phenyl

group (46) strongly enhanced the affinity at CXCR1 but not

yet at CXCR2.

By varying the nature and the position of the substituents of

the phenylsulfonyl moiety (47-49), a clear modulation of the

affinity for both the receptors resulted and the 2-ethyl derivative

(47) was identified as a dual and potent inhibitor.

Because the direct comparison between the benzoate (39)

derivative and the corresponding benzamide (41) suggested a

3992 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17

Moriconi et al.

Scheme 8^a

^aReagents and conditions: (a) RSO₂NH₂, CDI, DBU, CH₂Cl₂, rt; (b) SOCl₂, RONH₂, NaHCO₃, CH₂Cl₂, rt; (c) oxalyl chloride, DMF, (CH₃)NHOH,

CH₂Cl₂, rt; (d) SOCl₂, RNH₂, CH₂Cl₂, rt.

detrimental effect of the amide hydrogen on the activity, a series

of alkyl and aryl sulfonates was synthesized and tested.

As expected on the basis of the pharmacophore hypothesis,

the simple mesylate (51) showed no activity, although this result

has to be prudently considered due to the intrinsic reactivity of

the mesylate group that could compromise the SAR interpreta-

tion. The gradual size increase of the hydrophobic group was

paralleled by a potency increase in the series of sulfonate

compounds (52-54) and, consistent with the premises, the

phenylsulfonate ester (53) exhibited higher potency than the

corresponding phenylsulfonamide (46).

Compounds 52-54 were all three potent and dual CXCR1/2

inhibitors and the sulfonate spacer was confirmed as optimal

from a spatial point of view for the correct orientation in the

hydrophobic pocket. Nevertheless, sulfonic esters, still more

clearly than carboxylic esters, suffer the problem of liability

due to their inherent sensitivity to hydrolytic conditions and

high reactivity toward nucleophilic groups.

To overcome this issue, we focused on the trifluoromethane-

sulfonyl (triflate) group, because this moiety, even if scarcely

used in medicinal chemistry programs, is considerably more

stable than aliphatic and aromatic sulfonates versus hydrolysis

and nucleophilic attack. Furthermore, due to the marked

electron-withdrawing effect of the substituent, aryltriflates are

expected to show a good resistance to in vivo metabolic

oxidation of the aromatic ring.^31-33

(56), which is to date the most potent dual inhibitor in the series

of 2-phenylpropionic acid derivatives. Nevertheless, the pro-

nounced receptor affinity of 56 does not obviously fit with the

above-described SAR that, along this series of compounds,

repeatedly confirmed the importance of strong hydrophobic

interactions for a favorable ligand binding.

The paucity of medicinal chemistry studies related to triflate

derivatives, hardly limited so far the comprehension of specific

properties, and the potential usefulness of this group for

bioisosteric replacement purposes remains unexplored.

The marked potency increase, observed substituting methyl

(51) for trifluoromethyl (55) group in the sulfonate esters series,

suggests a key role of this residue, but an adequate contribution

to the establishment of a network of hydrophobic interactions

seems hardly plausible due to the high polarity and relatively

small size of the group. In fact, logP/logD value calculations

(ACD Labs/logD Suite release 9.0; data not shown) do not

evidence a strong influence of the trifluoromethyl group on the

overall lipophilicity of the molecule. By contrast, the hydrogen-

bonding capability of the trifluoromethyl group is well-

documented in the literature, and several structural studies

highlight the evident hydrogen-bond acceptor capability in

protein-ligand complexes, as assessed by PDB analysis.³⁴

To gain insight into this topic, the binding mode of 56 in

CXCR1 and CXCR2 was studied by means of exhaustive MM/

MD and GRID analysis.

Starting from these considerations, particular attention was

paid to the results obtained with 2-(4-trifluoromethanesulfony-

loxy)phenylpropionic acid (55) and the related 2R-enantiomer

Like the other CXCL8 inhibitors, the acid carboxylic group

of 56 was found to directly engage strong polar interactions

with the common binding motif of the class (the couple Lys99_2.64

CXCR1/CXCR2 Inhibitors

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17 3993

Figure 3. (Panels A and B) Comparison of the 3D MIFs calculated on the allosteric site of reparixin in CXCR1 and CXCR2 by using the DRY

(contoured in yellow at -0.4 kcal/mol) and OH2 (contoured in cyan at -8.0 kcal/mol) probes. From a visual inspection it can be seen that CXCR2

does not have an extended favorable hydrophobic region as in CXCR1 cavity and this leads to a partial loss of reparixin activity in CXCR2. (Panels

C and D) GRID maps determined on the allosteric site of 21 in CXCR1 and CXCR2 by using the DRY and OH2 (cyan) probes. The hydrophobic

regions (contoured in yellow at -0.4 kcal/mol), generated from the residues of a conserved pocket of CXCR1 and CXCR2, closely map the

benzoyl moiety; furthermore, the hydrophilic (contoured in cyan at -8.0 kcal/mol) contours superimpose the carboxylic group of 21. For clarity,

only a part of the entire GRID maps is shown.

and Glu291_7.39) in both the receptors. However, in both the

derived binding models, the trifluoromethyl residue does not

seem to be directly involved in polar interactions but, as assumed

for the other phenylpropionic acids, it appears comfortably

accommodated in the hydrophobic cavity spanned by the helices

1, 3, and 7 (Figure 4, panels A and B). Molecular interaction

fields (MIFs) show that, in CXCR1 and CXCR2, the triflate

group well occupies the hydrophobic region generated by the

same residues previously described.

The experimental confirmation of the proposed binding model

would suggest an unexpected ability of the small trifluoro-

methyl group in establishing a network of strong hydrophobic

interactions.

At the end of this preliminary phase aimed at the SAR

investigation and the assessment of the binding hypotheses,

three phenylpropionic acids 9, 13, and 56 were selected as

potential candidates for the following lead optimization

phase.

As widely discussed, the lack of cyclooxygenases (COXs)

inhibitory activity and the selectivity toward other chemokine/

chemotactic factors were considered the main criteria for the

selection of candidates for in vivo studies.²¹

(data not shown). Among the tested compounds, only 56 was

completely inactive in the PGE₂production assay at the

concentration of 10^-5M (% inhibition: 17 ( 8).

Starting from the binding mode of 56, a set of phenylpropi-

onamide derivatives, ranging from weakly acidic to neutral

aliphatic or aromatic amides, was built up. We first evaluated

a series of acylsulfonamides as potential bioisosteres of the

carboxylic group. As shown in Table 3, along this class (57-

61), the increase of the hydrophobic group size led to a

significant potency loss toward CXCL8 and the substitution of

trifluoromethyl for methyl (62) led to an absolutely inactive

compound. This effect seemed still more pronounced in CXCL1

inhibition, and the methanesulfonamide 57 was the only potent

and dual inhibitor of the series. The results obtained in CXCL8-

induced chemotaxis assay are in fair agreement with the SAR

reported for reparixin analogues, and this observation further

supports the validity of the binding mode hypothesis. Following

the same approach as reported for reparixin derivatives,²¹we

investigated the possibility of replacing the ionic interaction

between the inhibitor and the Lys99_2.64/Glu291_7.39couple with

a double hydrogen-bond interaction. A set of hydroxamate

derivatives was evaluated as alternative bioisosteres of the

carboxylic acid functionality. The hydroxamic acid (63) was,

as expected, a dual and potent inhibitor, and both O- and

N-alkylation (64-66) were compatible with binding to CXCR1

and CXCR2.

The R- and S-enantiomers of 9, 13, and 56 were evaluated at

the maximal concentration for inhibition of the LPS-induced

PGE₂production in human macrophages and for the effect on

human PMN chemotaxis induced by fMLP, C5a, and MCP1

3994 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17

Moriconi et al.

and that the biologically validated reparixin/CXCR1 binding

mode can be used as a reliable tool for the interpretation of

SAR of this class of allosteric inhibitors.

Among the selected compounds, a set of dual inhibitors

has been further filtered on the basis of the selectivity

(related GPCRs and COX activity), PK profile, and oral

biovailability in rodent species (data not shown). Interestingly,

PK profile of 56, 67, and 79 confirmed the hypothesized in

vitro and in vivo metabolic stability of tri-

flate moiety, supporting further investigations on this group

up to date not yet duly explored for medicinal chemistry

purposes. Compounds 56, 67, and 79 are very stable in

plasma and corresponding half-life time ranges from 3 to 24 h.

In addition, the related metabolic pathways have been com-

pletely characterized and will be the subject of a specific

publication.

Compounds 56, 67, and 79, unlike reparixin, show a PK

profile and oral bioavailability from suitable to optimal,

amenable for long-term treatments. They are currently under

preclinical investigation, not only for I/R prevention, but also

for the treatment of several chronic diseases including UC and

metastatic melanoma.

Conclusion

In the present work, we report a computer-assisted design

approach for the identification of novel allosteric CXCL8

inhibitors acting with comparable potency at both CXCR1 and

CXCR2. The reparixin binding model in the CXCR1 TM region,

validated by extensive mutagenesis experiments, guided the

analysis of the main determinants for CXCR1/CXCR2 selectiv-

ity. Molecular modeling studies on selected inhibitors provided

useful information for the interpretation of the SAR results.

A series of 2-arylpropionic acids has been investigated for

the comprehension of SAR and for the selection of ring

substituents optimized in terms of dual CXCR1/CXCR2 activity.

The 4-triflate derivatives have been selected because of their

interesting pharmacophoric features, and a set of 56 analogues

has been further investigated. Among the selected dual inhibi-

tors, three potential novel lead compounds, 56, 67, and 79, share

the triflate moiety and exhibit optimized PK and disposition

characteristics.

Figure 4. (Panels A and B) MIFs calculated on the allosteric site of

56 in CXCR1 and CXCR2 by using the DRY (contoured in yellow at

-0.5 kcal/mol) and OH2 (contoured in cyan at -9.0 kcal/mol) probes.

The figure shows a common and favorable hydrophobic cavity spanned

by the elices 1, 3, and 7 of CXCR1 and CXCR2. The triflate group of

56 fits this hydrophobic pocket. For the sake of clarity, only a part of

the entire GRID map is shown.

Finally, several alkyl and aryl amides derived from 56 were

synthesized and tested. The primary amide (67) preserved a

potent inhibitory effect toward CXCL8 and CXCL1, whereas,

in strict analogy with reparixin derivatives, relatively small alkyl

substituents on the amido nitrogen were poorly tolerated (68,

70-72) in the receptor pocket. Nevertheless, the N-isopropyl

derivative (69) was identified as a potent and dual inhibitor of

CXCL8 and CXCL1 up to the concentration of 10^-9M.

Among the N-aryl derivatives (73-79), the N-phenyl amide

73 did not show any activity, but the insertion of a heteroatom

in the 2-position of the aromatic ring favorably influenced the

biological activity and, in fact, compounds 74, 77-79 were all

effective dual and potent inhibitors of CXCR1 and CXCR2,

whereas the shift of the heteroatom in the 3- and 4-position led

to inactive compounds (75, 76). As previously discussed, the

good activity of heteroaryl derivatives seems coherent with the

formation of an intramolecular hydrogen bond involving the

heteroatom as acceptor, able to determine a significant dextral

shift of the amido/imido equilibrium, thus tuning the binding

ability of the amido group.

The obtained data suggest an unexpected ability of the small

triflate group in establishing a network of strong hydrophobic

interactions.

The proposed binding site and the allosteric mechanism of

action have been confirmed by testing 57 on CXCR1/L1.2

transfected cells (paper in preparation), and specific point

mutagenesis studies will allow in the near future to definitively

clarify the peculiar pattern of interactions engaged by the triflate

group in the binding pocket. If confirmed, these results, together

with the in vivo PK and metabolic profile of the leads, could

offer new insights into the peculiar characteristic of the triflate

group and in its potential relevance in medicinal chemistry

research programs.

Several studies^35-38converge in indicating CXCL8 inhibition

as a promising pharmacological target for the treatment of still

unmet medical needs. Although CXCR1 activation has been

reported to play a major role in CXCL8-induced human PMN

chemotaxis, CXCR2 signaling seems directly and specifically

involved in the pathogenesis of demanding chronic diseases such

as psoriasis and COPD.

The overall data obtained with derivatives of 56 are in

excellent agreement with the SAR observed for reparixin

analogues and, in general, the modification/functionalization of

the carboxylic group parallely affects CXCL8 and CXCL1

inhibition.

Furthermore, there is accumulating evidence that the patho-

physiological role of CXCL8 in melanoma progression and

metastases could be mediated by CXCR2 activation.

This observation is a further confirmation that hydrophobic

interactions are key determinants for CXCR1/CXCR2 selectivity

CXCR1/CXCR2 Inhibitors

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17 3995

and 30 min later, 1 µg/mL LPS was added. Control wells received

saline at the appropriate dilution. Culture supernatants were

harvested 12 h after LPS stimulation. PGE₂production was

measured with an EIA kit (Amersham; sensitivity 2.5 pg/well).

Mutation Analysis of CXCR1. The human CXCR1 open

reading frame was PCR amplified from a CXCR1/pCEP4 plasmid

(kindly provided by Dr. P. M. Murphy, NIH, Bethesda) using the

high fidelity Pfx DNA polymerase. Point mutants were obtained

by two-step PCR, as previously described.⁴⁴Wild type and CXCR1

I43V were cloned in the mammalian expression vector pcDNA3.

The nucleotide sequence of each construct was confirmed by

double-stranded DNA sequencing. The mouse L1.2 lymphoma cell

line was grown and transfected transiently by electroporation, as

previously described.⁴⁵After 48 h from transfection, expression of

CXCR1 was assessed by flow cytometry using an antihuman

CXCR1 monoclonal antibody and cells were used for chemotaxis

experiments.

B. Molecular Modeling. All the calculations were performed

using an IBM eServer 326 Cluster under the Red Hat Enterprise

WS3 operating system. The conformational analysis of the ligands

was performed with the systematic torsional sampling method

implemented in Macromodel 8.6⁴⁶software. Atomic charges and

potentials were fixed using the standard OPLS2001 force-field, and

the energy level was fixed at 50 kJ/mol.

The availability of potent and selective dual inhibitors

blocking both CXCR1 and CXCR2 subtypes could hence

represent a powerful tool for the treatment of those pathologies

characterized by an abnormal activation of the CXCL8/CXCL1

pathway.

Finally, the results herein reported confirm the valuable

contribution of homology models to SAR program when MM/

MD analysis is matched with information coming from point

mutagenesis studies. Following the same approach, a novel class

of CXCR2 selective inhibitors has been recently identified, and

the pharmacological characterization of selected compounds is

currently in progress.

Experimental Section

A. Biology. Reagents. Compounds were routinely dissolved at

the indicated final concentrations in saline (Bieffe Medital, Sondrio,

Italy). All chemokines were from PeproTech (London, United

Kingdom). All chemicals, cell culture reagents, and protease

inhibitors were from Sigma (St. Louis, Missouri). Ficol/Hipaque,

Percoll, and dextran were from Pharmacia LKB. Diff-Quik was

from Harleco. Boyden chambers and polycarbonate filter were from

Neuroprobe, Inc. Transwell filters were from Costar (Cambridge,

Massachusetts). Thioglycolate and LPS (from E. coli 055:B5) were

from Difco (Detroit, MI). Pfx DNA polymerase and pcDNA3 were

from Invitrogen (Carlbad, California). Human CXCR1 monoclonal

antibody (clone 5A12) was from BD PharMingen, (San Diego,

California).

Cells. Human mononuclear cells and PMNs were obtained from

buffy coats of heparinized blood from normal volunteers through

the courtesy of Centro Trasfusionale, Ospedale S. Salvatore,

L’Aquila, Italy. Mononuclear cells were obtained by centrifugation

on Ficoll/Hipaque. Monocytes were separated by Percoll gradient

centrifugation.³⁹Human PMNs were prepared to 95% purity by

dextran sedimentation followed by hypotonic lysis of contaminating

red blood cells.⁴⁰Cellular viability was >95% in all experiments,

as measured by trypan blue dye exclusion.

Migration Assay. Cell migration of human PMNs, human

monocytes, was evaluated using a 48-well micro-chemotaxis

chamber, as previously described.⁴¹Cell suspension (1.5 - 3 ×

10⁶cells/mL) was incubated at 37 °C for 15 min in the presence

of vehicle or of indicated concentrations of compounds and next

seeded in triplicates in the upper compartment of the chemotactic

chamber. Different agonists were seeded in the lower compartment

of the chamber at the following concentrations: 1nM CXCL8, 0.03

nM fMLP, 10 nM CXCL1, 2.5 nM CCL2, 30 nM C5a. The

chemotactic chamber was incubated at 37 °C in air with 5% CO₂

for 45 min (human PMNs) or 2 h (monocytes). At the end of

incubation, the filter was removed, fixed, and stained and five oil

immersion fields at high magnification (100×) were counted for

each migration well after sample coding. L1.2 migration was

evaluated using 5 µm pore size Transwell filters, as previously

described.⁴²Briefly, cells were preincubated overnight in growth

medium containing 5 mM sodium butyrate to maximize receptor

expression, resuspended at 10⁷cells/mL in culture medium, and

pretreated for 30 min at 37 °C with indicated concentrations of

compound. After treatment, 100 µL of cell suspension were placed

onto each Transwell filter, which was transferred into wells

containing 600 µL of culture medium supplemented with 1 nM

CXCL8 and compound (at the same concentration of the corre-

sponding upper chamber). After 4 h of incubation at 37 °C, the

upper chamber was removed and the number of migrated cells was

evaluated in a Bu¨rker chamber.

The minimum-energy conformer of each ligand was manually

docked in the putative binding cavity of CXCL8 receptor homology

models. With the aim to relax nonbonding interactions in the system,

the complexes were energy-refined using steepest descent and

conjugate gradient algorithms, as implemented in the DISCOVER

package, using the CVFF force field.⁴⁷After, the complexes were

submitted to molecular dynamics calculations by using a previously

described protocol.²¹

The interaction energies were carried out by using GRID.⁴⁸The

grid spacing was fixed at 0.5 Å and the directive MOVE was set

to 0. The water (OH2) and DRY probes were used to calculate the

MIFs in the putative CXCR1 and CXCR2 allosteric sites of CXCL8

inhibitors. The water probe (OH2) simulates the solvation-

desolvation process, whereas the hydrophobic probe (DRY) com-

putes the hydrophobic energy.

All the figures were depicted by using the BODIL package.⁴⁹

C. Chemistry. General Experimental Procedures. Optical

rotations were measured on a Perkin-Elmer 241 polarimeter and

25

1

the [R]_Dvalues are given in 10^-1deg cm²g^-1. H NMR spectra

were recorded on a Bruker ARX 300 spectrometer. Melting points

were determined using a Bu¨chi capillary melting point apparatus

and are uncorrected. Elemental analyses were within (0.4% of the

theoretical values calculated for C, H, and N and are reported only

with symbols.

All reagents and solvents were purchased from Sigma-Aldrich

or Lancaster and used without further purification. Reaction courses

and product mixtures were monitored by thin-layer chromatography

on silica gel (precoated F₂₅₄Macherey-Nagel plates); the spots

were examined with UV light and visualized with I₂.

The 2-arylpropionic acids 38 (Fenoprofen) and 42 (Indoprofen)

are commercially available (Sigma).

Synthesis of Racemic 2-Arylpropanoic Acids (Schemes 1-7).

2-{4[(2-Methylprop-1-enyl]phenyl}propanoic Acid (4). To a

solution of intermediate 80 (0.35 g, 1.12 mmol) in anhydrous

1-methyl-2-pyrrolidinone (3 mL) under nitrogen and vigorous

stirring, LiCl (0.142 g, 3.37 mmol), CuI (11 mg, 0.056 mmol),

AsPh₃(27 mg, 0.09 mmol), and Pd₂dba₃(21 mg, 0.02 mmol) were

added. After stirring 10 min, 2-methyl-1-propenyltributyltin (pre-

pared as described;⁵⁰0.46 g, 1.34 mmol) was added, and the reaction

mixture was left stirring 5 h at 90 °C. After cooling at room

temperature, a saturated solution of KF (10 mL) was added and,

after stirring 15 min, water was added (5 mL). The aqueous solution

was extracted with Et₂O (3 × 10 mL) and the collected organic

extracts were washed back with water (2 × 10 mL), dried over

Na₂SO₄, and evaporated under reduced pressure to give a crude

residue that, after purification by flash chromatography (n-hexane/

EtOAc 9:1), afforded methyl 2-{4[(2-methylprop-1-enyl)phenyl]-

Macrophage Preparation and LPS-Induced PGE₂Produc-

tion. Peritoneal exudates cells were collected from peritoneal

washing 5 days after i.p. inoculum of 3% thioglycolate in saline

(1.5 mL for mouse), as previously described.⁴³Cells were then

plated at the density of 6 × 10⁵/well in 96-well plates in RPMI

1640 medium, and nonadherent cells were removed by repeated

washing 2 h later. Compounds were then added to the macrophages,

3996 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17

Moriconi et al.

propanoate as a yellow oil (0.18 g, 74% yield). The methyl ester

was dissolved in 95% EtOH (3 mL), a 20% alcoholic solution of

KOH (3 mL) was added, and the resulting mixture was left stirring

4 h at room temperature. After solvent evaporation, the residue

was diluted with water (5 mL) and washed with Et₂O (3 × 5 mL);

the aqueous phase was acidified with 1 N HCl (2 × 5 mL) to pH

1 and extracted with EtOAc (2 × 5 mL). The organic collected

extracts were dried over Na₂SO₄and evaporated under reduced

pressure to give a crude residue that, after purification by flash

chromatography (petroleum ether/EtOAc 1:1), afforded pure 4 as

an off-white wax (0.143 g, 85% yield). ¹H NMR (CDCl₃): δ 7.47

(d, 2H, J ) 7 Hz), 7.18 (d, 2H, J ) 7 Hz), 6.24 (m, 1H), 3.74 (q,

1H, J ) 7 Hz), 1.88 (d, 6H, J ) 8 Hz), 1.53 (d, 3H, J ) 7 Hz).

Anal. (C₁₃H₁₆O₂) C, H.

ladium(II) chloride (0.14 g, 0.2 mmol) and triethylamine (0.28 mL,

2 mmol) were added. After stirring for 10 min, R-methylstyrene

(0.29 mL, 2.2 mmol) was added and the reaction was left stirring

for 24 h at 90 °C. After cooling at room temperature, the reaction

mixture was diluted with toluene (10 mL) and washed with water

(10 mL) and 6 N HCl. The aqueous layers were extracted back

with toluene (2 × 10 mL), and the collected organic extracts were

washed with water (10 mL) and brine (10 mL), dried over Na₂-

SO₄, and evaporated under reduced pressure to give a crude residue

that, after purification by flash chromatography (n-hexane/EtOAc

95:5), afforded methyl 2-{3-[2-phenylprop-1-en-1-yl]phenyl}-

propanoate as yellow oil (0.42 g, 75% yield). To palladium on

charcoal (10% Pd, 50 mg), a solution of the methyl ester (0.4 g,

1.42 mmol) in EtOH (10 mL) was added, and the mixture was

stirred under hydrogen until complete disappearance (6 h) of the

starting material (TLC). After filtration on a Celite pad, concentra-

tion of the mother liquors gave pure methyl 2-[3-(2-phenylpropyl)-

phenyl]propanoate (0.39 g, 96% yield). Hydrolysis of the methyl

ester was performed as described for 4. Pure 20 was isolated by

flash chromatography (n-hexane/EtOAc 9:1) as a pale yellow oil

(0.31 g, 85% yield). ¹H NMR (CDCl₃): δ 7.65 (s, 1H), 7.45-7.12

(m, 7H), 7.05 (t, 1H, J ) 3 Hz), 3.60 (q, 1H, J ) 7 Hz), 3.35 (q,

1H, J ) 7 Hz), 2.85 (dd, 2H, J₁) 7 Hz, J₂) 3 Hz), 1.55 (d, 3H,

J ) 7 Hz), 1.25 (d, 3H, J ) 7 Hz). Anal. (C₁₈H₂₀O₂) C, H.

2-(3-Propionylphenyl)propanoic Acid (23). Compound 23 was

prepared following a described procedure⁵²with minor modifica-

tions. To a suspension of ZnCl₂(0.390 g, 2.85 mmol) in dry THF

(5 mL) at 0 °C under a nitrogen atmosphere commercial ethyl-

magnesium chloride (2M in Et₂O, 2.85 mL, 5.70 mmol) was added.

After stirring for 20 min, dichloro[1,1′-bis(diphenylphosphino)-

ferrocene] palladium(II) dichloromethane adduct (PdCl₂(dppf); 1

mol %, 46 mg, 0.057 mmol) was added and then a solution of

3-(1-cyanoethyl)benzoyl chloride (1.11 g, 5.72 mmol) in dry THF

(5 mL) was dripped. The mixture was stirred for 1 h at 0 °C and

for 3 h at room temperature. After cooling to 0 °C, 3 N HCl (10

mL) and Et₂O (30 mL) were added. The two phases were separated

and the organic one was washed sequentially with a saturated

solution of NaHCO₃(2 × 30 mL) and brine (30 mL), dried over

Na₂SO₄, and evaporated under vacuum to a crude oil. The crude

was purified by flash chromatography (n-hexane/EtOAc 95:5) to

afford the intermediate 2-(3-propionylphenyl) propionitrile as pale

yellow oil (0.87 g, 81% yield). ¹H NMR (CDCl₃): δ 7.86 (s, 1H),

7.76 (d, 1H, J ) 7 Hz), 7.45-7.35 (m, 2H), 3.84 (q, 1H, J ) 7

Hz), 3.45 (m, 1H), 1.68 (d, 3H, J ) 7 Hz), 1.1 (d, 6H, J ) 7 Hz).

To a solution of 2-(3-propionylphenyl)propionitrile (0.85 g, 4.54

mmol) in dioxane (10 mL), 37% HCl (10 mL) was added. The

resulting mixture was left stirring at 70 °C for 4 h. After cooling

at room temperature, the solvent was evaporated and the residue

was diluted with cold water (10 mL) and extracted with EtOAc

(2× 8 mL). The organic layer was extracted back with 1 N NaOH

(2 × 5 mL), and the basic aqueous phase was acidified with 1 N

HCl to pH 1 and extracted with EtOAc (2 × 5 mL). The collected

organic extracts were dried over Na₂SO₄and evaporated to give

pure 23 as a pale yellow oil (0.81 g, 85% yield). ¹H NMR

(CDCl₃): δ 10.50 (bs, 1H), 8.05 (s, 1H), 7.92 (d, 1H, J ) 7 Hz),

7.65-7.48 (m, 2H), 3.95 (q, 1H, J ) 7 Hz), 3.10 (q, 2H, J ) 7

Hz), 1.70 (d, 3H, J ) 7 Hz), 1.35 (t, 3H, J ) 7 Hz). Anal.

(C₁₄H₁₄O₃) C, H.

2-[4-(Cyclohexylphenyl)propanoic Acid (5). To a solution of

80 (5 g, 16.01 mmol) in dry THF (20 mL) under nitrogen

atmosphere, LiCl (2.035 g, 48.03 mmol) and Pd(PPh₃)₄(0.74 g,

0.64 mmol) were added. After stirring 10 min at room temperature,

cyclohexylzinc bromide (0.5 M in THF; 5.82 g, 24.02 mmol) was

added, and the resulting mixture was refluxed overnight. After

cooling at room temperature, the solvent was evaporated and the

residue was diluted with EtOAc (15 mL), washed with water (2 ×

15 mL), dried over Na₂SO₄, and evaporated to give an oily crude

that was purified by flash chromatography (n-hexane/EtOAc 8:2)

to give methyl 2-[4-(cyclohexyl)phenyl]propanoate (3.04 g, 77%

yield) as a colorless oil. The methyl ester hydrolysis was performed

as described for 4. Pure 5 was isolated as a white powder (2.64 g,

1

92% yield), mp 72-74 °C. H NMR (CDCl₃): δ 10.20 (bs, 1H),

7.18 (d, 2H, J ) 7 Hz), 7.10 (d, 2H, J ) 7 Hz), 3.75 (q, 1H, J )

7 Hz), 2.50 (m, 1H), 1.90-1.70 (m, 6H), 1.52 (d, 3H, J ) 7 Hz),

1.45-1.28 (m, 4H). Anal. (C₁₅H₂₀O₂) C, H.

2-[4-(Cyclopropylmethyl)phenyl]propanoic Acid (8). A cata-

lytic amount of CuI was added to commercial cyclopropylmagne-

sium bromide (0.5 M in THF) (0.61 g, 4.21 mmol) under a nitrogen

atmosphere at 0-5 °C, and after stirring 30 min at the same

temperature, a solution of commercial 2-[4-(bromomethyl)phenyl]-

propanoic acid (0.5 g, 2.05 mmol) in dry THF (5 mL) was added

by dripping. After stirring overnight at room temperature, the

resulting dark green solution was quenched by adding a KH₂PO₄

buffer solution (pH 2.5, 20 mL). CH₂Cl₂(20 mL) was added and

the two phases were separated. The organic phase was extracted

with a saturated solution of NaHCO₃(3 × 5 mL); the collected

aqueous extracts were acidified with 2 N HCl to pH 2 and extracted

back with CH₂Cl₂(3 × 10 mL); the collected organic extracts were

dried over Na₂SO₄and, after solvent removal under reduced

pressure, afforded the pure compound 8 (0.31 g, yield 75%) as

1

pale yellow oil. H NMR (CDCl₃): δ 7.25 (s, 4H), 3.75 (q, 1H, J

) 7 Hz), 2.55 (d, 2H, J ) 7 Hz), 1.55 (d, 3H, J ) 7 Hz), 1.05 (m,

1H), 0.60-0.48 (m, 2H), 0.15 (m, 2H). Anal. (C₁₃H₁₆O₂) C, H.

2-[4-(Cyclobutylmethyl)phenyl]propanoic Acid (9). Following

the same procedure described for the preparation of 8 and starting

from 2-[4-(bromomethyl)phenyl]propanoic acid (0.5 g, 2.05 mmol)

and cyclobutylmagnesium bromide (prepared by treatment of

cyclobutyl bromide with magnesium turnings;⁵¹0.57 g, 4.21 mmol),

after workup, 9 was isolated as a colorless oil (0.35 g, yield 79%).

¹H NMR (CDCl₃): δ 7.24 (d, 2H, J ) 7 Hz), 7.10 (d, 2H, J ) 7

Hz), 3.75 (q, 1H, J ) 7 Hz), 2.75 (d, 2H, J ) 7 Hz), 2.55 (m, 1H),

2.11-2.01 (m, 2H), 1.91-1.80 (m, 2H), 1.77-1.68 (m, 2H), 1.52

(d, 3H, J ) 7 Hz). Anal. (C₁₄H₁₈O₂) C, H.

2-(3-Isobutyrylphenyl)propanoic Acid (24). Following the

same procedure described for 23 and starting from 3-(1-cyanoethyl)-

benzoyl chloride (1.10 g, 5.70 mmol) and commercial isopropyl-

magnesium chloride (2 M in Et₂O; 0.58 g, 5.70 mmol), after workup

and following hydrolysis of the nitrile intermediate, 24 was isolated

as a colorless oil (0.86 g, 85% yield). ¹H NMR (CDCl₃): δ 10.60

(bs, 1H), 7.86 (s, 1H), 7.76 (d, 1H, J ) 7 Hz), 7.45-7.35 (m, 2H),

3.79 (q, 1H, J ) 7 Hz), 3.45 (m, 1H), 1.45 (d, 3H, J ) 7 Hz), 1.15

(d, 6H, J ) 7 Hz). Anal. (C₁₃H₁₆O₃) C, H.

2-[3-(Cyclopentylcarbonyl)phenyl]propanoic Acid (25). Fol-

lowing the same procedure described for 23 and starting from 3-(1-

cyanoethyl)benzoyl chloride (1.10 g, 5.70 mmol) and commercial

cyclopentylmagnesium chloride (2 M in Et₂O; 0.73 g, 5.70 mmol),

2-[4-(Cyclopentylmethyl)phenyl]propanoic Acid (10). Fol-

lowing the same procedure described for the preparation of 8 and

starting from 2-[4-(bromomethyl)phenyl]propanoic acid (0.5 g, 2.05

mmol) and cyclopentylmagnesium bromide (0.63 g, 4.21 mmol),

after workup, 10 was isolated as a colorless oil (0.35 g, yield 73%).

¹H NMR (CDCl₃): δ 7.25 (d, 2H, J ) 7 Hz), 7.10 (d, 2H, J ) 7

Hz), 3.72 (q, 1H, J ) 7 Hz), 2.60 (d, 2H, J ) 7 Hz), 2.05 (m, 1H),

1.81-1.60 (m, 6H), 1.55 (d, 3H, J ) 7 Hz), 1.32-1.12 (m, 2H).

Anal. (C₁₅H₂₀O₂) C, H.

2-[3-(2-Phenylpropyl)phenyl]propanoic Acid (20). To a sus-

pension of intermediate 82 (0.63 g, 2 mmol) in dry DMF (5 mL)

under nitrogen and vigorous stirring, bis(triphenylphosphine) pal-

CXCR1/CXCR2 Inhibitors

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17 3997

after workup and following hydrolysis of the nitrile intermediate,

25 was isolated as a pale yellow oil (1.04 g, 74% yield). ¹H NMR

(CDCl₃): δ 7.86 (m, 1H), 7.79 (d, 1H, J ) 7 Hz), 7.52 (d, 1H, J

) 7 Hz), 7.37 (m, 1H), 3.82 (q, 1H, J ) 7 Hz), 3.71 (m, 1H), 2.22

(s, 2H), 2.01 (m, 3H), 1.82 (m, 3H), 1.58 (d, 3H, J ) 7 Hz). Anal.

(C₁₅H₁₈O₃) C, H.

2-[3-(Cyclohexylcarbonyl)phenyl]propanoic Acid (26). Fol-

lowing the same procedure described for 23 and starting from 3-(1-

cyanoethyl)benzoyl chloride (1.10 g, 5.70 mmol) and commercial

cyclohexylmagnesium chloride (2 M in Et₂O; 0.81 g, 5.70 mmol),

after workup and following hydrolysis of the nitrile intermediate,

26 was isolated as a pale yellow oil (1.19 g, 80% yield). ¹H NMR

(CDCl₃): δ 7.86 (m, 1H), 7.79 (d, 1H, J ) 7 Hz), 7.52 (d, 1H, J

) 7 Hz), 7.37 (m, 1H), 3.80 (q, 1H, J ) 7 Hz), 3.15 (m, 1H),

1.95-1.80 (m, 4H), 1.52 (d, 3H, J ) 7 Hz), 1.42-1.18 (m, 6H).

Anal. (C₁₆H₂₀O₃) C, H.

hydrolysis of the nitrile intermediate, 30 was isolated as a white

glassy solid (1.43 g, 72% yield). H NMR (CDCl₃): δ 8.40 (m,

2H), 8.10 (s, 1H), 7.75 (s, 1H), 7.60 (d, 1H, J ) 7 Hz), 7.50

(t, 1H, J ) 7 Hz), 3.90 (q, 1H, J ) 7 Hz), 1.60 (d, 3H, J ) 7 Hz).

Anal. (C₁₃H₁₁NO₃S) C, H, N, S.

2-[3-(1-Phenylvinyl)phenyl]propanoic Acid (31). To a solution

of commercial 2-(3-benzoylphenyl)propanoic acid (ketoprofen; 1

g, 3.93 mmol) in methanol (100 mL), a few drops of concentrated

H₂SO₄were added, and the resulting mixture was left stirring at

room temperature overnight. After solvent evaporation at reduced

pressure, the crude was diluted with CH₂Cl₂(25 mL), and the

organic phase was washed with 1 N NaOH (2 × 20 mL), dried

over Na₂SO₄, and evaporated under vacuum to give 2-(3-ben-

zoylphenyl)propanoic acid methyl ester (0.95 g, 3.54 mmol) as a

colorless oil.

1

To a solution of methyltriphenylphosphonium bromide (1.26 g,

3.54 mmol) in dry THF (10 mL) at room temperature under a

nitrogen atmosphere, n-BuLi (1.6 M in hexanes, 2.23 mL, 3.54

mmol) was added. After stirring for 3 h, a solution of 2-(3-

benzoylphenyl)propanoic acid methyl ester (0.95 g, 3.54 mmol) in

dry THF (10 mL) was added by dripping, and the resulting mixture

was left stirring overnight at room temperature. After solvent

evaporation, the crude was diluted with EtOAc (20 mL) and washed

with a 40% Na₂S₂O₅solution (2 × 15 mL) and with water (2 × 15

mL), dried over Na₂SO₄, and evaporated under vacuum to give a

crude residue. The crude was purified by flash chromatography

(n-hexane/EtOAc 95:5) to give methyl 2-[3-(1-phenylvinyl)phenyl]-

propanoate (0.56 g, 2.12 mmol) as a colorless oil. The ester (0.31

g, 1.16 mmol) was dissolved in methanol (8 mL) and 1 N NaOH

(2 mL) was added. The solution was left stirring overnight at room

temperature. After solvent evaporation under reduced pressure, the

residue was diluted with CH₂Cl₂(10 mL) and the organic phase

was extracted with H₂O (2 × 5 mL); the collected aqueous extracts

were acidified to pH 1 with 1 N HCl and extracted back with

CH₂Cl₂(2 × 10 mL). After drying over Na₂SO₄and solvent

evaporation under vacuum, pure 31 was obtained (0.29 g, quantita-

tive yield) as colorless oil. ¹H NMR (CDCl₃): δ 7.30-7.15

(m, 9H), 5.50 (m, 2H), 3.80 (q, 1H, J ) 7 Hz), 1.55 (d, 3H, J )

7 Hz). Anal. (C₁₇H₁₆O₂) C, H.

2-[3-(Thien-2-ylcarbonyl)phenyl]propanoic Acid (27). Com-

pound 27 was prepared essentially following a described procedure⁵³

but with some modifications. To a solution of thiophene (0.61 mL,

7.60 mmol) in dry THF (50 mL) at -78 °C under nitrogen

atmosphere n-BuLi (1.6 M in hexanes, 4.7 mL, 7.60 mmol) was

added. After stirring for 20 min, ZnCl₂(2.071 g, 15.20 mmol) was

added and the mixture was warmed to 0 °C and stirred for 45 min.

CuI (1.45 g, 7.60 mmol) was added, and after stirring for 20 min,

a solution of 3-(1-cyanoethyl)benzoyl chloride (2.94 g, 15.2 mmol)

in dry THF (10 mL) was added by dripping. After stirring for 2 h,

the mixture was diluted with EtOAc (30 mL) and washed

sequentially with a saturated solution of NaHCO₃(2 × 50 mL)

and brine (50 mL), dried over Na₂SO₄, and evaporated under

vacuum to a crude oil. The crude was purified by flash chroma-

tography (n-hexane/EtOAc 95:5) to afford the intermediate 2-[3-

(thien-2-ylcarbonyl)phenyl)propanenitrile as pale yellow oil (1.36

1

g, 74% yield). H NMR (CDCl₃): δ 7.85 (m, 2H), 7.70 (m, 1H),

7.60 (d, 1H, J ) 7 Hz), 7.52 (m, 1H), 7.46 (t, 1H, J ) 7 Hz), 7.09

(t, 1H, J ) 7 Hz), 3.95 (q, 1H, J ) 7 Hz), 1.65 (d, 3H, J ) 7 Hz).

To a solution of 2-[3-(thien-2-ylcarbonyl)phenyl)propanenitrile

(1.07 g, 4.43 mmol) in dioxane (10 mL), 37% HCl (10 mL) was

added. The resulting mixture was left stirring at 70 °C for 4 h.

After cooling at room temperature, the solvent was evaporated under

vacuum and the residue was diluted with EtOAc (15 mL) and

washed with cold water (10 mL). The organic layer was extracted

back with 1 N NaOH (2 × 5 mL). The aqueous phase was acidified

with 1 N HCl to pH 1 and extracted with EtOAc (2 × 10 mL).

The organic collected extracts were dried over Na₂SO₄and

evaporated under vacuum to give pure 27 as a brown oil (0.83 g,

72% yield). ¹H NMR (CDCl₃): δ 7.85 (m, 2H), 7.70 (m, 1H), 7.60

(d, 1H, J ) 7 Hz), 7.52 (m, 1H), 7.46 (t, 1H, J ) 7 Hz), 7.15

(t, 1H, J ) 7 Hz), 3.85 (q, 1H, J ) 7 Hz), 1.60 (d, 3H, J ) 7 Hz).

Anal. (C₁₄H₁₂O₃S) C, H, S.

2-[3-(1-Phenethylethyl)phenyl]propanoic Acid (35). The syn-

thesis of 35 was performed following a described procedure,²¹using

racemate methyl 2-(3-benzoylphenyl)propanoate as starting material.

35 was isolated pure as pale yellow oil. ¹H NMR (CDCl₃): δ 7.35-

7.12 (m, 9H), 4.15 (q, 1H, J ) 7 Hz), 3.75 (q, 1H, J ) 7 Hz), 1.70

(d, 3H, J ) 7 Hz), 1.48 (d, 3H, J ) 7 Hz). Anal. (C₁₇H₁₈O₂) C, H.

2-[3-(1-Hydroxy-1-phenethylethyl)phenyl]propanoic Acid (36).

To a solution of 2-(3-benzoylphenyl)propanoic acid (0.33 g, 1.30

mmol) in dry Et₂O (10 mL) at room temperature under a nitrogen

atmosphere, methylmagnesium bromide (3 M in Et₂O, 0.9 mL, 2.7

mmol) was added. The resulting mixture was refluxed for 3 h and,

after cooling at room temperature, diluted with Et₂O (10 mL),

washed with 2 N HCl (3 × 10 mL) and with water (2 × 10 mL),

dried over Na₂SO₄, and evaporated under vacuum to give a crude.

After purification by flash chromatography (eluent: CHCl₃/CH₃-

OH 98:2), pure 36 was obtained as a white powder (0.26 g, 74%

2-[3-(Furan-2-carbonyl)phenyl]propanoic Acid (28). Follow-

ing the same procedure described for 27 and starting from

commercial furan (0.55 mL, 7.60 mmol) and 3-(1-cyanoethyl)-

benzoyl chloride (2.94 g, 15.2 mmol), after workup and following

hydrolysis of the nitrile intermediate, 28 was isolated as a pale

yellow oil (1.48 g, 80% yield). ¹H NMR (CDCl₃): δ 7.95 (m, 1H),

7.85 (m, 1H), 7.72 (s, 1H); 7.60 (m, 1H), 7.48 (t, 1H, J ) 7 Hz),

7.20 (d, 1H, J ) 3 Hz), 6.60 (m, 1H), 3.85 (q, 1H, J ) 7 Hz), 1.65

(d, 3H, J ) 7 Hz). Anal. (C₁₄H₁₂O₄) C, H.

1

yield): mp 93-95 °C. H NMR (CDCl₃): δ 7.45-7.38 (m, 4H),

7.30-7.15 (m, 5H), 3.65 (q, 1H, J ) 7 Hz), 1.95 (s, 3H), 1.55

(d, 3H, J ) 7 Hz). Anal. (C₁₇H₁₈O₃) C, H.

2-[3-(1,3-Oxazol-2-ylcarbonyl)phenyl]propanoic Acid (29).

Following the same procedure described for 27 and starting from

commercial oxazole (0.54 g, 7.60 mmol) and 3-(1-cyanoethyl)-

benzoyl chloride (2.94 g, 15.2 mmol), after workup and following

hydrolysis of the nitrile intermediate, 29 was isolated as a white

solid (1.21 g, 65% yield): mp 121-122 °C. ¹H NMR (CDCl₃): δ

8.45 (m, 2H), 7.90 (s, 1H), 7.60 (d, 1H, J ) 7 Hz), 7.45 (t, 1H, J

) 7 Hz), 7.35 (s, 1H), 3.85 (q, 1H, J ) 7 Hz), 1.60 (d, 3H, J ) 7

Hz). Anal. (C₁₃H₁₁NO₄) C, H, N.

2-[3-(Phenylsulfonyl)phenyl]propanoic Acid (37). Compound

37 was prepared following a described procedure⁵⁴starting from

intermediate 81 (0.5 g, 1.94 mmol) and commercial benzenesulfinic

acid sodium salt (0.38 g, 2.33 mmol). The following hydrolysis of

the nitrile intermediate 2-[3-(phenylsulfonyl)phenyl]propanenitrile

allowed to obtain pure 37 as a white powder (75% yield from 81):

mp 95-98 °C. H NMR (CDCl₃): δ 8.00 (s, 1H), 7.95-7.90 (m,

3H), 7.50-7.42 (m, 3H), 7.30 (m, 2H), 3.90 (q, J ) 7 Hz, 1H),

1.55 (d, 3H, J ) 7 Hz). Anal. (C₁₅H₁₄O₄S) C, H, S.

1

2-[3-(1,3-Thiazol-2-ylcarbonyl)phenyl]propanoic Acid (30).

Following the same procedure described for 27 and starting from

commercial thiazole (0.54 mL, 7.60 mmol) and 3-(1-cyanoethyl)-

benzoyl chloride (2.94 g, 15.2 mmol), after workup and following

2-[4-(Benzoyloxy)phenyl]propanoic Acid (39). 2-(4-hydrox-

yphenyl)propanoic acid (1 g, 6.01 mmol) was dissolved in 1 N

NaOH (18 mL). Benzoyl chloride (1.74 mL, 15.02 mmol) was

dripped into the solution and, at the same time, 2 N NaOH was

3998 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17

Moriconi et al.

dripped in the flask to basic pH. At the end of the additions, the

reaction mixture was left stirring at room temperature for 90 min,

3 N HCl was added to pH 2, and pure 39 was isolated by filtration

as a white powder (1.35 g, 83% yield): mp 150-153 °C. ¹H NMR

(CDCl₃): δ 8.25 (d, 2H, J ) 7 Hz), 7.68 (t, 1H, J ) 7 Hz), 7.55

(t, 2H, J ) 7 Hz), 7.40 (d, 2H, J ) 7 Hz), 7.21 (d, 2H, J ) 7 Hz),

3.82 (q, 1H, J ) 7 Hz), 1.55 (d, 3H, J ) 7 Hz). Anal. (C₁₆H₁₄O₄)

C, H.

2-{4-[(Isopropylsulfonyl)amino]phenyl}propanoic Acid (45).

Following the same procedure described for 44 and starting from

83 (1.94 g, 10.82 mmol) and isopropylsulfonyl chloride (1.88 mL,

13 mmol), after workup and purification, 45 was obtained as a pale

1

brown powder (1.7 g, 87% yield): mp 168-172 °C. H NMR

(CDCl₃): δ 7.40 (bs, 1H), 7.28 (d, 2H, J ) 7 Hz), 7.17 (d, 2H, J

) 7 Hz), 3.70 (q, 1H, J ) 7 Hz), 3.25 (m, 1H), 1.52 (d, 3H, J )

7 Hz), 1.35 (d, 6H, J ) 7 Hz). Anal. (C₁₂H₁₇NO₄S) C, H,

N, S.

2-[4-(Propionyloxy)phenyl]propanoic Acid (40). Following the

same procedure described for 39 and starting from 2-(4-hydrox-

yphenyl)propanoic acid (1 g, 6.01 mmol) and propionyl chloride

(1.3 mL, 15.02 mmol), after workup, 40 was obtained as a waxy

2-{4-[(Phenylsulfonyl)amino]phenyl}propanoic Acid (46). Fol-

lowing the same procedure described for 44 and starting from 83

(1.94 g, 10.82 mmol) and benzenesulfonyl chloride (1.66 mL, 13

mmol), after workup and purification, 46 was obtained as a white

1

solid (1.00 g, 75% yield). H NMR (CDCl₃): δ 7.38 (d, 2H, J )

1

powder (2.64 g, 80% yield), mp 161-164 °C. H NMR (CDCl₃):

7 Hz), 7.15 (d, 2H, J ) 7 Hz), 3.75 (q, 1H, J ) 7 Hz), 2.62 (q, 2H,

J ) 7 Hz), 1.52 (d, 3H, J ) 7 Hz), 1.27 (t, 3H, J ) 7 Hz). Anal.

(C₁₂H₁₄O₄) C, H.

δ 9.38 (bs, 1H), 7.73 (m, 2H), 7.40-7.25 (m, 3H), 7.05 (d, 2H, J

) 7 Hz), 6.92 (d, 2H, J ) 7 Hz), 3.45 (q, 1H, J ) 7 Hz), 1.27

(d, 3H, J ) 7 Hz). Anal. (C₁₅H₁₅NO₄S) C, H, N, S.

2-[4-(Benzoylamino)phenyl]propanoic Acid (41). To a cooled

(0-5 °C) solution of benzoyl chloride (0.38 mL, 3.3 mmol) in

acetone (5 mL), pyridine (0.64 mL, 7.92 mmol) and methyl 2-(4-

aminophenyl)propanoate 83 (0.71 g, 3.3 mmol) were added. The

resulting solution was left stirring at room temperature overnight.

The solvent was evaporated, and the oily crude was diluted with

CH₂Cl₂(15 mL) and washed with 1 N HCl (2 × 15 mL), water

(2 × 10 mL), and brine (20 mL), dried over Na₂SO₄, and evaporated

under vacuum to give methyl 2-[4-(benzoylamino)phenyl]propanoic

acid (0.67 g, 72% yield) as a yellow solid. To a solution of the

methyl ester (0.67 g, 2.36 mmol) in EtOH (10 mL), 2 N NaOH

(1.18 mL, 3.54 mmol) was added, and the resulting solution was

stirred overnight at room temperature. HCl (1 N) was added by

dripping until the precipitation of 41 was complete. Compound 41

was isolated by filtration as a pale yellow powder (0.635 g,

quantitative yield): mp > 200 °C. ¹H NMR (DMSO-d₆): δ 10.20

(bs, 1H), 8.05 (d, 2H, J ) 7 Hz), 7.75 (d, 2H, J ) 7 Hz), 7.60-

7.50 (m, 3H), 7.32 (d, 2H, J ) 7 Hz), 3.65 (q, 1H, J ) 7 Hz), 1.52

(d, 3H, J ) 7 Hz). Anal. (C₁₆H₁₅NO₃) C, H, N.

2-(4-{[(2-Ethylphenyl)sulfonyl]amino}phenyl)propanoic Acid

(47). Following the same procedure described for 44 and starting

from 83 (1.94 g, 10.82 mmol) and 2-ethyl benzenesulfonyl chloride

(prepared by treatment of 2-ethylbenzene sulfonic acid with thionyl

chloride; 2.66 g, 13 mmol), after workup and purification, 47 was

1

obtained as a glassy solid (2.81 g, 65% yield). H NMR (CDCl₃):

δ 8.15 (d, 1H, J ) 7 Hz), 7.67 (m, 1H), 7.52 (d, 1H, J ) 7 Hz),

7.40 (bs, 1H), 7.35 (d, 2H, J ) 7 Hz), 7.10 (d, 2H, J ) 7 Hz), 3.80

(q, 1H, J ) 7 Hz), 3.18 (q, 2H, J ) 7 Hz), 1.57 (d, 3H, J ) 7 Hz),

1.40 (t, 3H, J ) 7 Hz). Anal. (C₁₇H₁₉NO₄S) C, H, N, S.

2-(4-{[(4-Methylphenyl)sulfonyl]amino}phenyl)propanoic Acid

(48). Following the same procedure described for 44 and starting

from 83 (1.94 g, 10.82 mmol) and p-toluenesulfonyl chloride (2.47

g, 13 mmol), after workup and purification, 48 was obtained as a

1

white powder (2.56 g, 74% yield): mp 150-152 °C. H NMR

(DMSO-d₆): δ 12.23 (bs, 1H), 10.19 (bs, 1H), 7.65 (d, 2H, J ) 8

Hz), 7.34 (d, 2H, J ) 8 Hz), 7.12 (d, 2H, J ) 7 Hz), 7.02 (d, 2H,

J ) 7 Hz), 3.54 (q, 1H, J ) 7 Hz), 2.33 (s, 3H), 1.27 (d, 3H, J )

7 Hz). Anal. (C₁₆H₁₇NO₄S) C, H, N, S.

2-[4-(2-Oxopyrrolidin-1-yl)phenyl]propanoic Acid (43). To a

solution of 83 (0.6 g, 3.84 mmol) in CH₂Cl₂(8 mL), triethylamine

(0.54 mL, 3.84 mmoL) was added, and after stirring for 5 min,

4-chlorobutyryl chloride (0.54 g, 3.84 mmol) was dripped into the

solution. After stirring for 20 min at room temperature, the solution

was diluted with water (15 mL) and extracted with CH₂Cl₂(3 ×

10 mL). The collected organic extracts were washed with water,

dried over Na₂SO₄, and evaporated under vacuum to give methyl

2-{4-[(4-chlorobutanoyl)amino]phenyl}propanoate pure enough to

be used in the next step. The ester was dissolved in 2-propanol

(6 mL), and 6% NaOH was added by dripping until the complete

conversion to 43 (TLC). The mixture was diluted with water (10

mL) and washed with Et₂O (20 mL). The aqueous layer was

acidified with 1 N HCl and extracted with EtOAc (2 × 10 mL).

The organic extracts were dried over Na₂SO₄and evaporated under

2-(4-{[(2-Chlorophenyl)sulfonyl]amino}phenyl)propanoic Acid

(49). Following the same procedure described for 44 and starting

from 83 (1.94 g, 10.82 mmol) and 2-chlorobenzenesulfonyl chloride

(prepared by treatment of 2-chlorobenzene sulfonic acid with thionyl

chloride; 2.74 g, 13 mmol), after workup and purification, 49 was

obtained as a wax (2.28 g, 62% yield). ¹H NMR (CDCl₃): δ 8.05

(d, 1H, J ) 7 Hz), 7.40 (q, 2H, J ) 3 Hz), 7.25 (m, 1H), 7.10 (d,

2H, J ) 7 Hz), 6.95 (d, 2H, J ) 7 Hz), 3.58 (q, 1H, J ) 7 Hz),

1.45 (d, 3H, J ) 7 Hz). Anal. (C₁₅H₁₄ClNO₄S) C, H, Cl,

N, S.

2-(4-{[(Trifluoromethyl)sulfonyl]amino}phenyl)propanoic Acid

(50). Following the same procedure described for 44 and starting

from 83 (1.94 g, 10.82 mmol) and trifluoromethanesulfonyl chloride

(1.38 mL, 13 mmol), after workup and purification, 50 was obtained

1

as a white solid (1.41 g, 44% yield): mp 132-135 °C. H NMR

1

vacuum to give pure 43 as a waxy solid (0.58 g, 65% yield). H

(CDCl₃): δ 7.25 (d, 2H, J ) 7 Hz), 7.12 (d, 2H, J ) 7 Hz), 3.60

(q, 1H, J ) 7 Hz), 1.45 (d, 3H, J ) 7 Hz). Anal. (C₁₀H₁₀F₃NO₄S)

C, H, F, N, S.

NMR (CDCl₃): δ 7.59 (d, 2H, J ) 7 Hz), 7.34 (d, 2H, J ) 7 Hz),

3.84 (t, 2H, J ) 7 Hz), 3.74 (q, 1H, J ) 7 Hz), 2.62 (t, 2H, J ) 7

Hz), 2.14 (dd, 2H, J₁) 7 Hz, J₂) 4 Hz), 1.51 (d, 3H, J ) 7 Hz).

Anal. (C₁₃H₁₅NO₃) C, H, N.

2-{4-[(Methylsulfonyl)oxy]phenyl}propanoic Acid (51). To a

solution of commercial 2-(4-hydroxy)phenylpropanoic acid (50 g,

0.29 mol) in CH₃OH (50 mL), concentrated H₂SO₄(0.4 mL) was

added. The resulting solution was left stirring overnight at room

temperature. After solvent evaporation, the oily residue was diluted

with CH₂Cl₂and washed with a saturated solution of NaHCO₃

(2 × 100 mL) and water (3 × 150 mL), dried over Na₂SO₄, and

evaporated to give methyl 2-(4-hydroxy)phenylpropanoate (51.6 g,

0.28 mol). To a solution of the methyl ester (3.4 g, 18.9 mmol) in

pyridine (25 mL), methanesulfonyl chloride (1.46 mL, 18.9 mmol)

was added by dripping. The reaction mixture was refluxed for 24

h. After cooling at room temperature, the solvent was evaporated,

and the oily crude was diluted with CH₂Cl₂(50 mL), washed with

1 N HCl (2 × 50 mL), 1 N NaOH (2 × 35 mL), and water (2 ×

100 mL), dried over Na₂SO₄, and evaporated under vacuum to give

methyl 2-{4-[(methylsulfonyl)oxy]phenyl}propanoate (3.48 g, 71%

yield) as an oil. The methyl ester was dissolved in glacial AcOH

2-{4-[(Methylsulfonyl)amino]phenyl}propanoic Acid (44). To

a solution of 83 (1.85 g, 10.35 mmol) in acetone (20 mL), pyridine

(1.24 mL, 15.48 mmol) and methanesulfonyl chloride (0.81 g, 10.4

mmol) were added. The resulting solution was left stirring at room

temperature overnight. The solvent was evaporated and the oily

crude was diluted with CH₂Cl₂(50 mL) and washed with water (3

× 100 mL), dried over Na₂SO₄, and evaporated under vacuum to

give methyl 2-{4-[(methylsulfonyl)amino]phenyl}propanoate as a

slightly pink powder by pulping in diisopropyl ether (2.15 g, 81%

yield). The methyl ester hydrolysis was performed as described

for 41. Pure 44 was isolated as a white powder (1.87 g, 92%

yield): mp 102-105 °C. ¹H NMR (CDCl₃): δ 7.32 (d, 2H, J ) 7

Hz), 7.24 (d, 2H, J ) 7 Hz), 6.52 (bs, 1H), 3.75 (q, 1H, J ) 7 Hz),

3.05 (s, 3H), 1.52 (d, 3H, J ) 7 Hz). Anal. (C₁₀H₁₃NO₄S) C, H, N,

S.

CXCR1/CXCR2 Inhibitors

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17 3999

(15 mL), 37% HCl (2.7 mL) was added, and the resulting solution

was refluxed overnight. After cooling at room temperature, the

residue was dissolved in CH₂Cl₂(30 mL) and washed with water

(2 × 50 mL), dried over Na₂SO₄, and evaporated under vacuum to

give 51 (3.11 g, 95% yield) as a pale yellow powder, mp 80-

8.10 (d, 1H, J ) 7 Hz), 7.68-7.62 (m, 1H), 7.58-7.43 (m, 3H),

7.45 (d, 2H, J ) 7 Hz), 7.38 (d, 2H, J ) 7 Hz), 3.85 (q, 1H, J )

7 Hz), 1.30 (d, 3H, J ) 7 Hz). Anal. (C₁₆H₁₃ClF₃NO₆S₂) C, H, S,

N.

4-{(1R)-1-Methyl-2-oxo-2-[(pyridin-3-ylsulfonyl)amino]ethyl}-

phenyl trifluoromethanesulfonate (61). Following the same

procedure described for 58 and starting from 56 (0.2 g, 0.67 mmol)

and pyridine-2-sulfonamide (0.1 g, 0.67 mmol), after workup and

purification by flash chromatography (CH₂Cl₂/CH₃OH 9:1), 61 was

obtained as a white powder (0.194 g, 66% yield): mp 157-

1

82 °C. H NMR (CDCl₃): δ 7.45 (d, 2H, J ) 7 Hz), 7.25 (d, 2H,

J ) 7 Hz), 3.82 (q, 1H, J ) 7 Hz), 3.15 (s, 3H), 1.55 (d, 3H, J )

7 Hz). Anal. (C₁₀H₁₂O₅S) C, H, S.

2-{4-[(Isopropylsulfonyl)oxy]phenyl}propanoic Acid (52). Fol-

lowing the same procedure described for 51 and starting from

methyl 2-(4-hydroxy)phenyl propanoate (3.4 g, 18.9 mmol) and

2-propanesulfonyl chloride (2.12 mL, 18.9 mmol), after workup

and following hydrolysis of the methyl ester intermediate, 52 was

obtained as a pale yellow powder (3.76 g, 73% yield): mp 62-

25

1

160 °C. [R]_D-82° (c 0.5, CH₃OH). H NMR (CDCl₃): δ 9.20

(m, 1H), 8.92 (m, 1H), 8.40 (m, 1H), 7.55 (m, 1H), 7.25 (s, 4H),

3.65 (q, 1H, J ) 7 Hz), 1.48 (d, 3H, J ) 7 Hz). Anal. (C₁₅H₁₃-

ClF₃N₂O₆S₂) C, H, S, N.

1

64 °C. H NMR (CDCl₃): δ 7.37 (d, 2H, J ) 7 Hz), 7.25 (d, 2H,

4-[(1R)-1-Methyl-2-oxo-2-{[(trifluoromethyl)sulfonyl]amino}-

ethyl]phenyl trifluoromethanesulfonate (62). Following the same

procedure described for 58 and starting from 56 (0.2 g, 0.67 mmol)

and trifluoromethanesulfonamide (0.1 g, 0.67 mmol), after workup

and purification by flash chromatography (CH₂Cl₂/CH₃OH 9:1),

62 was obtained as a white powder (0.155 g, 54% yield), mp 102-

J ) 7 Hz), 3.71 (q, 1H, J ) 7 Hz), 3.45 (m, 1H), 1.55 (d, 3H, J )

7 Hz). Anal. (C₁₂H₁₆O₅S) C, H, S.

2-{4-[(Phenylsulfonyl)oxy]phenyl}propanoic Acid (53). Fol-

lowing the same procedure described for 51 and starting from

methyl 2-(4-hydroxy)phenyl propanoate (3.4 g, 18.9 mmol) and

benzenesulfonyl chloride (2.41 mL, 18.9 mmol), after workup and

following hydrolysis of the methyl ester intermediate, 53 was

obtained as a pale brown powder (4.74 g, 82% yield): mp 68-

25

1

104 °C. [R]_D-11° (c 0.5, CH₃OH). H NMR (CDCl₃): δ 7.85

(bs, 1H), 7.45 (d, 2H, J ) 7 Hz), 7.38 (d, 2H, J ) 7 Hz), 3.85 (q,

1H, J ) 7 Hz), 1.60 (d, 3H, J ) 7 Hz). Anal. (C₁₁H₉F₆NO₆S₂) C,

H, S, N.

1

70 °C. H NMR (CDCl₃): δ 7.86 (d, 2H, J ) 7 Hz), 7.68 (t, 1H,

J ) 7 Hz), 7.54 (t, 2H, J ) 7 Hz), 7.24 (d, 2H, J ) 7 Hz), 6.96

(d, 2H, J ) 7 Hz), 3.71 (q, 1H, J ) 7 Hz), 1.43 (d, 3H, J ) 7 Hz).

Anal. (C₁₅H₁₄O₅S) C, H, S.

Synthesis of N- and O-Alkyl Derivatives (Scheme 8). 4-[(1R)-

2-(Hydroxyamino)-1-methyl-2-oxoethyl]phenyl trifluoromethane-

sulfonate (63). Compound 56 (1.00 g, 3.3 mmol) was dissolved in

toluene (10 mL), and SOCl₂(3 mL) was added. The resulting

solution was left stirring at reflux for 3 h. After cooling at room

temperature, the mixture was evaporated under reduced pressure.

The crude acyl chloride was diluted with dry CH₂Cl₂(10 mL) and

added by dripping to a suspension of hydroxylamine hydrochloride

(0.23 g, 3.3 mmol) and NaHCO₃(0.28 g, 3.3 mmol) in dry CH₂Cl₂

(5 mL). The resulting mixture was left under stirring at room

temperature overnight. The mixture was diluted with CH₂Cl₂(10

mL), and the organic layer was washed with 1 N HCl (2 × 10

mL) and brine (3 × 10 mL), dried over Na₂SO₄, and concentrated

under reduced pressure to give a crude residue that was purified

by pulping in n-hexane to give, after filtration, pure 63 as a pale

2-(4-{[(4-Methylphenyl)sulfonyl]oxy}phenyl)propanoic Acid

(54). Following the same procedure described for 51 and starting

from methyl 2-(4-hydroxy)phenyl propanoate (3.4 g, 18.9 mmol)

and p-toluenesulfonyl chloride (3.6 g, 18.9 mmol), after workup

and following hydrolysis of the methyl ester intermediate, 54 was

obtained as a pale yellow oil (4.72 g, 78% yield). ¹H NMR

(CDCl₃): δ 7.73 (d, 2H, J ) 7 Hz), 7.33 (d, 2H, J ) 7 Hz), 7.25

(d, 2H, J ) 7 Hz), 6.96 (d, 2H, J ) 7 Hz), 3.71 (q, 1H, J ) 7 Hz),

2.47 (s, 3H), 1.43 (d, 3H, J ) 7 Hz). Anal. (C₁₆H₁₆O₅S) C, H, S.

Synthesis of Acylsulfonamides (Scheme 8). 4-{(1R)-2-[(Iso-

propylsulfonyl)amino]-1-methyl-2-oxoethyl}phenyl

trifluo-

romethanesulfonate (58). To a cooled mixture (0-5 °C) of 56

(0.5 g, 1.68 mmol) in CH₂Cl₂(3 mL), 1,1′-carbonyldiimidazole

(CDI) (0.27 g, 1.70 mmol) was added. After stirring 2 h at 0-

5 °C, isopropylsulfonamide (0.21 g, 1.68 mmol) and diazobicyclo-

[5.4.0]undec-7-ene (DBU; 0.26 mL, 1.68 mmol) were added, and

the mixture was left stirring for 4 h. Glacial AcOH (0.2 mL, 3.5

mmol) was added, the reaction mixture was diluted with CH₂Cl₂

(10 mL), and the organic layer was washed with 10% buffer NaH₂-

PO₄(pH 4) (4 × 10 mL) and water (3 × 10 mL), dried over Na₂-

SO₄, and concentrated under reduced pressure to give a crude

residue that was purified by flash chromatography (CH₂Cl₂/CH₃-

OH 95:5) to afford 58 as a pale yellow oil (1.24 g, 74% yield).

[R]_D²⁵-20° (c 0.72, CH₃OH). ¹H NMR (CDCl₃): δ 7.85 (bs, 1H),

7.52 (d, 2H, J ) 7 Hz), 7.32 (d, 2H, J ) 7 Hz), 3.75 (m, 2H), 1.57

(d, 3H, J ) 7 Hz), 1.40 (d, 3H, J ) 7 Hz), 1.25 (d, 3H, J ) 7 Hz).

Anal. (C₁₃H₁₆F₃NO₆S₂) C, H, S, N.

25

brown powder (0.9 g, 87% yield): mp 82-84 °C. [R]_D-25°

(c 0.7, CH₃OH). ¹H NMR (CDCl₃): δ 8.20 (bs, 2H), 7.35 (d, 2H,

J ) 7 Hz), 7.15 (d, 2H, J ) 7 Hz), 3.48 (q, 1H, J ) 7 Hz), 1.47

(d, 3H, J ) 7 Hz). Anal. (C₁₀H₁₀F₃NO₅S) C, H, S, N.

4-{(1R)-2-[Hydroxy(methyl)amino]-1-methyl-2-oxoethyl}-

phenyl trifluoromethanesulfonate (64). To a cooled (-20 °C)

solution of DMF (0.42 mL, 5.42 mmol) in CH₂Cl₂(2 mL), a

solution of oxalyl chloride (0.16 mL, 1.83 mmol) in CH₂Cl₂

(5 mL) was added by dripping. The temperature was raised up to

0 °C, and after stirring for 30 min, 56 (0.5 g, 1.67 mmol) and

4-methylmorpholine (0.185 mL, 1.67 mmol) were added. After

stirring for an additional 30 min, N-methylhydroxylamine hydro-

chloride (0.27 g, 3.3 mmol) and 4-methylmorpholine (0.73 mL,

6.6 mmol) were added. The resulting mixture was left stirring

overnight at room temperature. After filtration of the formed

precipitate, the mother liquors were evaporated, the crude was

dissolved in CH₂Cl₂(10 mL), and the organic layer was washed

with 1 N HCl (2 × 10 mL), a saturated solution of NaHCO₃(2 ×

10 mL), and water (10 mL), dried over Na₂SO₄, filtered, and

concentrated under reduced pressure to give a crude oily residue

that, after flash chromatography (CH₂Cl₂/CH₃OH 95:5), afforded

4-{(1R)-1-Methyl-2-oxo-2-[(phenylsulfonyl)amino]ethyl}-

phenyl trifluoromethanesulfonate (59). Following the same

procedure described for 58 and starting from 56 (0.5 g, 1.68 mmol)

and benzenesulfonamide (0.26 g, 1.68 mmol), after workup and

purification by flash chromatography (CHCl₃/CH₃OH 95:5), 59 was

25

obtained as a pale yellow oil (0.602 g, 82% yield). [R]_D-60°

25

(c 0.62, CH₃OH). ¹H NMR (CDCl₃): δ 8.05 (bs, 1H), 7.92 (d, 2H,

J ) 7 Hz), 7.65 (t, 1H, J ) 7 Hz), 7.52 (t, 2H, J ) 7 Hz), 7.18

(s, 4H), 3.60 (q, 1H, J ) 7 Hz), 1.45 (d, 3H, J ) 7 Hz). Anal.

(C₁₆H₁₄F₃NO₆S₂) C, H, S, N.

pure 64 as a colorless oil (0.27 g, 50% yield). [R]_D-23° (c 1,

1

CH₃OH). H NMR (DMSO-d₆): δ 9.95 (bs, 1H), 7.45 (m, 4H),

4.35 (q, 1H, J ) 7 Hz), 3.10 (s, 3H), 1.32 (d, 3H, J ) 7 Hz). Anal.

(C₁₁H₁₂F₃NO₅S) C, H, S, N.

4-[(1R)-2-{[(2-Chlorophenyl)sulfonyl]amino}-1-methyl-2-oxo-

ethyl]phenyl trifluoromethanesulfonate (60). Following the same

procedure described for 58 and starting from 56 (0.5 g, 1.68 mmol)

and 2-chlorobenzenesulfonamide (0.32 g, 1.68 mmol), after workup

and purification by flash chromatography (CHCl₃/CH₃OH 95:5),

60 was obtained as a white powder (0.602 g, 76% yield): mp 120-

4-[(1R)-2-(Methoxyamino)-1-methyl-2-oxoethyl]phenyl trif-

luoromethanesulfonate (65). Following the same procedure de-

scribed for 63 and starting from 56 (0.5 g, 1.67 mmol) and

methoxyamine hydrochloride (0.14 g, 1.67 mmol), after workup

and purification by flash chromatography (CH₂Cl₂/CH₃OH 98:2),

25

pure 65 was obtained as a colorless oil (0.37 g, 63% yield). [R]_D

25

1

125 °C. [R]_D-62° (c 0.4, CH₃OH). H NMR (DMSO-d₆): δ

-18° (c 0.5, CH₃OH). H NMR (DMSO-d₆): δ 10.95 (bs, 1H),

4000 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17

Moriconi et al.

7.50-7.30 (m, 4H), 3.85 (m, 1H), 3.45 (q, 1H, J ) 7 Hz), 1.30 (d,

3H, J ) 7 Hz), 1.05 (d, 6H, J ) 7 Hz). Anal. (C₁₃H₁₆F₃NO₅S) C,

H, S, N.

J ) 7 Hz), 2.05-1.95 (m, 2H), 1.68-1.60 (m, 4H), 1.52 (d, 3H, J

) 7 Hz), 1.35-1.20 (m, 2H). Anal. (C₁₅H₁₈F₃NO₄S) C, H, S, N.

4-[(1R)-2-Anilino-1-methyl-2-oxoethyl]phenyl

trifluoro-

4-[(1R)-2-(Isopropoxyamino)-1-methyl-2-oxoethyl]phenyl tri-

fluoromethanesulfonate (66). Following the same procedure

described for 63 and starting from 56 (0.5 g, 1.67 mmol) and

O-isopropylhydroxylamine hydrochloride (0.19 g, 1.67 mmol), after

workup and purification by pulping in n-hexane, pure 66 was

methanesulfonate (73). Following the same procedure described

for 68 and starting from 4-[(1R)-2-chloro-1-methyl-2-oxoethyl]-

phenyl trifluoromethanesulfonate (0.54 g, 1.68 mmol) and aniline

(0.46 mL, 5.04 mmol), after workup and purification by flash

chromatography (CH₂Cl₂/CH₃OH 99:1), pure 73 was obtained as

a colorless oil (0.31 g, 50% yield). [R]_D²⁵-30° (c 1, CH₃OH). ¹H

NMR (CDCl₃): δ 7.45 (d, 2H, J ) 7 Hz), 7.32 (m, 2H), 7.25 (d,

2H, J ) 7 Hz), 7.05 (t, 1H, J ) 7 Hz), 6.95-6.80 (m, 2H), 5.30

(bs, 1H), 3.55 (q, 1H, J ) 7 Hz), 1.52 (d, 3H, J ) 7 Hz). Anal.

(C₁₆H₁₄F₃NO₄S) C, H, S, N.

25

obtained as a waxy solid (0.44 g, 81% yield). [R]_D-24° (c 0.5,

1

CH₃OH). H NMR (CDCl₃): δ 8.20 (bs, 1H), 7.48 (d, 2H, J ) 7

Hz), 7.22 (d, 2H, J ) 7 Hz), 3.72 (s, 3H), 3.45 (q, 1H, J ) 7 Hz),

1.55 (d, 3H, J ) 7 Hz). Anal. (C₁₁H₁₂F₃NO₅S) C, H, S, N.

Synthesis of Amides (Scheme 8). 4-[(1R)-1-Methyl-2-(methy-

lamino)-2-oxoethyl]phenyl trifluoromethanesulfonate (68). To

a solution of 4-[(1R)-2-chloro-1-methyl-2-oxoethyl]phenyl trifluo-

romethanesulfonate (prepared as described for 63; 0.54 g, 1.68

mmol) in CH₂Cl₂(5 mL), methylamine (2.0 M in THF; 2.52 mL,

5.04 mmol) was added by dripping. After stirring for 4 h, solvents

were evaporated, the crude was dissolved in CH₂Cl₂(5 mL), and

the organic layer was washed with 6 N HCl (2 × 10 mL), a

saturated solution of NaHCO₃(2 × 10 mL), and water (2 × 20

mL), dried over Na₂SO₄, and evaporated under reduced

pressure to give a crude residue. By flash chromatography

(CH₂Cl₂/CH₃OH 98:2), pure 68 was isolated as a pale yellow

4-[(1R)-1-Methyl-2-oxo-2-(pyridin-2-ylamino)ethyl]phenyl tri-

fluoromethanesulfonate (74). Following the same procedure

described for 68 and starting from 4-[(1R)-2-chloro-1-methyl-2-

oxoethyl]phenyl trifluoromethanesulfonate (0.54 g, 1.68 mmol),

2-aminopyridine (0.16 g, 1.68 mmol) and triethylamine (0.23 mL,

1.68 mmol), after workup and purification by pulping in isopropyl

ether, pure 74 was obtained as a waxy solid (0.49 g, 78% yield).

25

1

[R]_D-35° (c 1, CH₃OH). H NMR (CDCl₃): δ 8.28-8.20 (m,

2H), 8.10 (bs, 1H), 7.78-7.70 (m, 1H), 7.48 (d, 2H, J ) 7 Hz),

7.30 (d, 2H, J ) 7 Hz), 7.12-7.05 (m, 1H), 3.78 (q, 1H, J ) 7

Hz), 1.65 (d, 3H, J ) 7 Hz). Anal. (C₁₅H₁₃F₃N₂O₄S) C, H, S, N.

4-[(1R)-1-Methyl-2-oxo-2-(pyridin-3-ylamino)ethyl]phenyl tri-

fluoromethanesulfonate (75). Following the same procedure

described for 68 and starting from 4-[(1R)-2-chloro-1-methyl-2-

oxoethyl]phenyl trifluoromethanesulfonate (0.54 g, 1.68 mmol),

3-aminopyridine (0.16 g, 1.68 mmol) and triethylamine (0.23 mL,

1.68 mmol), after workup and purification by pulping in isopropyl

ether, pure 75 was obtained as a waxy solid (0.47 g, 75% yield).

25

oil (0.36 g, 69% yield). [R]_D-20° (c 1, MeOH). ¹H NMR

(CDCl₃): δ 7.42 (d, 2H, J ) 7 Hz), 7.25 (d, 2H, J ) 7 Hz), 5.35

(bs, 1H), 3.56 (q, 1H, J ) 7 Hz), 2.80 (s, 3H), 1.55 (d, 3H, J )

7 Hz). Anal. (C₁₁H₁₂F₃NO₄S) C, H, S, N.

4-[(1R)-2-(Isopropylamino)-1-methyl-2-oxoethyl]phenyl trif-

luoromethanesulfonate (69). Following the same procedure de-

scribed for 68 and starting from 4-[(1R)-2-chloro-1-methyl-2-

oxoethyl]phenyl trifluoromethanesulfonate (0.54 g, 1.68 mmol) and

isopropylamine (0.29 g, 5.04 mmol), after workup and purification

by pulping in n-hexane, pure 69 was obtained as a white powder

25

1

[R]_D-37° (c 1, CH₃OH). H NMR (CDCl₃): δ 8.20 (s, 1H),

8.15 (bs, 1H), 8.05 (m, 1H), 7.45 (d, 2H, J ) 7 Hz), 7.30 (d, 2H,

J ) 7 Hz), 7.26-7.20 (m, 1H), 7.12-7.05 (m, 1H), 3.75 (q, 1H, J

) 7 Hz), 1.60 (d, 3H, J ) 7 Hz). Anal. (C₁₅H₁₃F₃N₂O₄S) C, H, S,

N.

25

(0.34 g, 60% yield): mp 78-80 °C. [R]_D-2° (c 0.5, CH₃OH).

¹H NMR (CDCl₃): δ 7.40 (d, 2H, J ) 7 Hz), 7.25 (d, 2H, J ) 7

Hz), 5.15 (bs, 1H), 4.08 (m, 1H), 3.50 (q, 1H, J ) 7 Hz), 1.55

(d, 3H, J ) 7 Hz), 1.15 (d, 3H, J ) 7 Hz), 1.05 (d, 3H, J ) 7 Hz).

Anal. (C₁₃H₁₆F₃NO₄S) C, H, S, N.

4-[(1R)-1-Methyl-2-oxo-2-(pyridin-4-ylamino)ethyl]phenyl tri-

fluoromethanesulfonate (76). Following the same procedure

described for 68 and starting from 4-[(1R)-2-chloro-1-methyl-2-

oxoethyl]phenyl trifluoromethanesulfonate (0.54 g, 1.68 mmol),

4-aminopyridine (0.16 g, 1.68 mmol) and triethylamine (0.23 mL,

1.68 mmol), after workup and purification by flash chromatography

(CH₂Cl₂/CH₃OH 95:5), pure 76 was obtained as a colorless oil (0.51

4-[(1R)-2-(Cyclopropylamino)-1-methyl-2-oxoethyl]phenyl tri-

fluoromethanesulfonate (70). Following the same procedure

described for 68 and starting from 4-[(1R)-2-chloro-1-methyl-2-

oxoethyl]phenyl trifluoromethanesulfonate (0.54 g, 1.68 mmol) and

cyclopropylamine (0.35 mL, 5.04 mmol), after workup and

purification by pulping in n-hexane, pure 70 was obtained as a white

25

1

g, 82% yield). [R]_D-41° (c 1, CH₃OH). H NMR (CDCl₃): δ

8.15 (bs, 1H), 8.03 (d, 2H, J ) 5 Hz), 7.45 (d, 2H, J ) 7 Hz), 7.30

(d, 2H, J ) 7 Hz), 6.90 (d, 2H, J ) 5 Hz), 3.75 (q, 1H, J ) 7 Hz),

1.60 (d, 3H, J ) 7 Hz). Anal. (C₁₅H₁₃F₃N₂O₄S) C, H, S, N.

4-[(1R)-1-Methyl-2-oxo-2-(1,3-thiazol-2-ylamino)ethyl]phe-

nyl trifluoromethanesulfonate (77). Following the same procedure

described for 68 and starting from 4-[(1R)-2-chloro-1-methyl-2-

oxoethyl]phenyl trifluoromethanesulfonate (0.54 g, 1.68 mmol) and

2-aminothiazole (0.5 g, 5.04 mmol), after workup and purification

by pulping in isopropyl ether, pure 77 was obtained as a pale brown

powder (0.47 g, 73% yield): mp 128-130 °C. [R]_D²⁵-55° (c 0.5,

CH₃OH). ¹H NMR (CDCl₃): δ 10.65 (bs, 1H), 7.45 (m, 3H), 7.30

(d, 2H, J ) 7 Hz), 7.05 (d, 1H, J ) 3 Hz), 3.90 (q, 1H, J ) 7 Hz),

1.75 (d, 3H, J ) 7 Hz). Anal. (C₁₃H₁₁F₃N₂O₄S₂) C, H, S, N.

4-[(1R)-1-Methyl-2-(1,3-oxazol-2-ylamino)-2-oxoethyl]phe-

nyl trifluoromethanesulfonate (78). Following the same procedure

described for 68 and starting from 4-[(1R)-2-chloro-1-methyl-2-

oxoethyl]phenyl trifluoromethanesulfonate (0.54 g, 1.68 mmol) and

2-aminooxazole (0.42 g, 5.04 mmol), after workup and purification

by pulping in n-hexane, pure 78 was obtained as a white powder

25

powder (0.41 g, 73% yield): mp 58-61 °C. [R]_D-5° (c 0.5,

1

CH₃OH). H NMR (CDCl₃): δ 7.40 (d, 2H, J ) 7 Hz), 7.25 (d,

2H, J ) 7 Hz), 5.45 (bs, 1H), 3.50 (q, 1H, J ) 7 Hz), 2.80 (m,

1H), 1.52 (d, 3H, J ) 7 Hz), 0.85-0.75 (m, 2H), 0.50-0.38 (m,

2H). Anal. (C₁₃H₁₄F₃NO₄S) C, H, S, N.

4-{(1R)-2-[(Cyclopropylmethyl)amino]-1-methyl-2-oxoethyl}-

phenyl trifluoromethanesulfonate (71). Following the same

procedure described for 68 and starting from 4-[(1R)-2-chloro-1-

methyl-2-oxoethyl]phenyl trifluoromethanesulfonate (0.54 g, 1.68

mmol) and (aminomethyl)cyclopropane (0.44 mL, 5.04 mmol), after

workup and purification by flash chromatography (CH₂Cl₂/CH₃-

OH 99:1), pure 71 was obtained as a pale yellow oil (0.41 g, 70%

yield). [R]_D²⁵-15° (c 0.5, CH₃OH). ¹H NMR (CDCl₃): δ 7.45 (d,

2H, J ) 7 Hz), 7.25 (d, 2H, J ) 7 Hz), 5.50 (bs, 1H), 3.55 (q, 1H,

J ) 7 Hz), 3.10 (m, 2H), 1.52 (d, 3H, J ) 7 Hz), 0.90 (m, 1H),

0.45 (d, 2H, J ) 7 Hz), 0.15 (d, 2H, J ) 7 Hz). Anal. (C₁₄H₁₆F₃-

NO₄S) C, H, S, N.

4-[(1R)-2-(Cyclopentylamino)-1-methyl-2-oxoethyl]phenyl tri-

fluoromethanesulfonate (72). Following the same procedure

described for 68 and starting from 4-[(1R)-2-chloro-1-methyl-2-

oxoethyl]phenyl trifluoromethanesulfonate (0.54 g, 1.68 mmol) and

cyclopropylamine (0.35 mL, 5.04 mmol), after workup and

purification by pulping in isopropyl ether, pure 72 was obtained as

a white powder (0.34 g, 56% yield): mp 107-110 °C. [R]_D²⁵-2°

25

(0.24 g, 70% yield): mp 127-130 °C. [R]_D-8° (c 0.5, CH₃-

1

OH). H NMR (CDCl₃): δ 8.50 (bs, 1H), 7.75 (s, 1H), 7.45 (s,

1H), 7.40 (d, 2H, J ) 7 Hz), 7.27 (d, 2H, J ) 7 Hz), 4.15 (q, 1H,

J ) 7 Hz), 1.60 (d, 3H, J ) 7 Hz). Anal. (C₁₃H₁₁F₃N₂O₅S₂) C, H,

S, N.

4-[(1R)-1-Methyl-2-oxo-2-{[4-(trifluoromethyl)-1,3-thiazol-2-

yl]amino}ethyl]phenyl trifluoromethanesulfonate (79). Following

the same procedure described for 68 and starting from 4-[(1R)-2-

1

(c 0.5, CH₃OH). H NMR (CDCl₃): δ 7.45 (d, 2H, J ) 7 Hz),

7.25 (d, 2H, J ) 7 Hz), 5.30 (bs, 1H), 4.20 (m, 1H), 3.55 (q, 1H,

CXCR1/CXCR2 Inhibitors

Journal of Medicinal Chemistry, 2007, Vol. 50, No. 17 4001

chloro-1-methyl-2-oxoethyl]phenyl trifluoromethanesulfonate (0.54

g, 1.68 mmol) and 2-amino-4-trifluoromethylthiazole²⁸(0.85 g, 5.04

mmol), after workup and purification by pulping in n-hexane, pure

79 was obtained as a white powder (0.68 g, 90% yield): mp 150-

153 °C. [R]_D-21° (c 1, CH₃OH). H NMR (CDCl₃): δ 8.75

(bs, 1H), 7.50-7.40 (m, 3H), 7.28 (d, 2H, J ) 7 Hz), 3.85 (q, 1H,

J ) 7 Hz), 1.55 (d, 3H, J ) 7 Hz). Anal. (C₁₄H₁₀F₆N₂O₄S₂) C, H,

S, N.

Acknowledgment. This work was supported by the Euro-

pean Union FP6 (INNOCHEM, Grant Number LSHB-CT-2005-

518167).

25

1

Supporting Information Available: Results from elemental

analysis of all the listed compounds and ¹³C NMR spectra of some

representative compounds. This material is available free of charge

via the Internet at http://pubs.acs.org.

Synthesis of Intermediates. 2-(3-Iodophenyl)propanenitrile

(81). Commercial 3-(1-cyanoethyl)benzoic acid (2.5 g, 14.25 mmol)

was dissolved in SOCl₂(5 mL), and the resulting solution was left

stirring for 3 h at reflux. After cooling at room temperature, the

mixture was evaporated under reduced pressure. The crude acyl

chloride was diluted with CH₂Cl₂(15 mL), tetrabutylammonium

bromide (23 mg, 0.07 mmol) was added, and the mixture was

cooled to 0-5 °C. Under vigorous stirring, a solution of NaN₃

(1.275 g, 19.5 mmol) in H₂O (5 mL) was added, and the resulting

mixture was left stirring at 0-5 °C for 2 h. The formed precipitate

was filtered off, and the organic phase was washed with H₂O (3 ×

25 mL), dried over Na₂SO₄, treated with trifluoroacetic acid (TFA;

1.59 mL, 21.38 mmol), and refluxed for 48 h. After cooling at room

temperature, TFA was evaporated under reduced pressure and the

crude residue was diluted with CH₂Cl₂(50 mL) and washed

sequentially with a saturated solution of NaHCO₃(2 × 25 mL)

and H₂O (50 mL). After Na₂SO₄drying and solvent evaporation

under reduced pressure, 2-[(3-trifluoroacetylamino)phenyl]pro-

panenitrile was obtained pure enough (TLC) to be used for the

following step. A mixture of 2-[(3-trifluoroacetylamino) phenyl]-

propanenitrile (2.5 g, 9.25 mmol) and K₂CO₃(2.55 g, 17.6 mmol)

in H₂O/CH₃OH (3:1; 50 mL) was heated at 60 °C for 16 h. After

cooling at room temperature and solvent evaporation, the crude

was diluted with H₂O (25 mL) and extracted with CH₂Cl₂(3 × 25

mL). The collected organic extracts were dried over Na₂SO₄and

evaporated under reduced pressure to give pure 2-(3-aminophenyl)-

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JM061469T

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Design of noncompetitive interleukin-8 inhibitors acting on CXCR1 and CXCR2

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