Small, non-peptide C5a receptor antagonists: Part 1

Article abstract of DOI:10.1016/j.bmcl.2008.08.106

The optimisation of a series of amides for C5a receptor binding and functional activity, and physicochemical properties is described. The initial hit, 1 (IC₅₀ 1 μM), was discovered during high throughput screening, from which highly potent C5a receptor antagonists (e.g. 14, IC₅₀ 5 nM) were developed.

Full text of DOI:10.1016/j.bmcl.2008.08.106

Bioorganic & Medicinal Chemistry Letters 18 (2008) 5601–5604
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Small, non-peptide C5a receptor antagonists: Part 1
b
b
b
Julian Blagg a, Charles Mowbray a, David C. Pryde a, , Gary Salmon , Esther Schmid , David Fairman ,
*
Kevin Beaumont c
a Department of Discovery Chemistry, Pfizer Global Research and Development, Ramsgate Road, Sandwich, Kent CT13 9NJ, UK
b Department of Discovery Biology, Pfizer Global Research and Development, Ramsgate Road, Sandwich, Kent CT13 9NJ, UK
c Department of Pharmacokinetics, Distribution and Metabolism, Pfizer Global Research and Development, Ramsgate Road, Sandwich, Kent CT13 9NJ, UK
a r t i c l e i n f o
a b s t r a c t
Article history:
The optimisation of a series of amides for C5a receptor binding and functional activity, and physicochem-
Received 28 July 2008
Revised 26 August 2008
Accepted 28 August 2008
Available online 31 August 2008
ical properties is described. The initial hit, 1 (IC50
ing, from which highly potent C5a receptor antagonists (e.g. 14, IC50 5 nM) were developed.
Ó 2008 Elsevier Ltd. All rights reserved.
1 lM), was discovered during high throughput screen-
Keywords:
C5a receptor
Complement
Inflammation
Antagonist
C5a is a 74-amino acid peptide cleaved from C5 at sites of
inflammation or infection during activation of the complement
system.1 C5a has been implicated in several inflammatory condi-
tions. For example, C5a is a potent chemoattractant of leukocytes
and phagocytes and stimulates via cell surface receptors (C5aR)
both the upregulation of integrins on the cell surface, and cell
degranulation at the source of inflammation, leading to endothelial
damage.2 Blocking the action of C5a on its receptor should provide
an effective treatment for a variety of inflammatory diseases.
A C-terminal analogue of C5a has been reported to show potent
agonism of the C5aR,3 while conformationally restricted cyclic ana-
logues are antagonists.4 Few non-peptidic C5a receptor ligands
have been reported in the literature.5 Of these, both agonists,6
and isolated reports of antagonists7 are represented.
contains two potentially toxic functional groups,10 the replace-
ment of which was a priority.
This paper describes our attempts to improve the binding affin-
ity of 2, while also improving its physicochemical properties to-
wards
a compound suitable for oral dosing. The targeted
parameters were IC50 < 100 nM, LogD < 4.0 and a MWt. < 500, in
a series devoid of any potentially toxic functional groups.
Lead compound 2 and direct piperidine analogues were synthe-
sized in a straightforward manner according to Scheme 1. Cyclo-
hexane derivatives were synthesized from cyclohexan-1,4-dione
monoethylene ketal according to Scheme 2. After reductive amina-
tion and acylation, ketal deprotection revealed the ketone, which
Cl
Cl
O
Our own efforts towards a small, orally bioavailable C5a recep-
O
tor antagonist began with the hit, 1 (IC50
1 lM, Figure 1), from a
N
N
N
N
high throughput radiolabelled C5a receptor binding screen of the
corporate compound file. 1 was then elaborated using parallel
chemistry8 to the potent (IC50 31 nM), although lipophilic and
poorly soluble project lead CP-447,697, 2 which fails the Lipinski
rule-of-59 on two counts (MWt. > 500 and LogP > 5) and would
be expected to be poorly absorbed and rapidly metabolized (see
Table 4 below). In the benzothiophene and aniline groups, 2 also
S
1, CP-289,503
IC50 1μM
cLogP 5.7
2, CP-447,697
IC50 31nM
cLogP 5.7
MWt. 554
O
NH
F
F
OMe
* Corresponding author.
E-mail address: David.Pryde@pfizer.com (D.C. Pryde).
Figure 1. The HTS hit, 1, and the project lead 2, obtained following extensive library
modification of 1.
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.08.106

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J. Blagg et al. / Bioorg. Med. Chem. Lett. 18 (2008) 5601–5604
Table 1
O
N
R
O
C5a receptor binding and functional activity for a series of piperidine analogues of
compound 2
HN
N
O
R
a
b, c
R
N
Cl
O
then b, d or e
O
O
N
R
O
N
R
S
Compound
R
125I Binding
affinity IC50
(nM)
Functional
activity
IC50 (nM)
cLogP
(LogD)b
Scheme 1. Preparation of 2 and analogues. Reagents and conditions: (a) RNH2,
Na(AcO)3BH, DCM, rt, 18 h. (b) RCO2H, (COCl)2, 1 drop DMF, DCM, rt, 1 h. (c) TFA,
DCM, rt, 6 h. (d) RNCO, Et3N, DCM, rt, 5 h. (e) RHal, K2CO3, MeCN, reflux, 16 h.
a
a
H
N
F
2
3
31
250
nd
5.6
5.5
N
N
O
F
O
O
O
R
R
O
R
N
R
N
F
330
a, b
c-f
O
F
F
O
O
F
O
O
4
200
>1000
350
70
3000
nd
6.7 (4.1)
3.6
N
Scheme 2. Preparation of cyclohexane derivatives of compound 2. Reagents and
conditions: (a) RNH2, Na(AcO)3BH, DCM, rt, 18 h. (b) RCO2H, (COCl)2, 1 drop DMF,
DCM, rt, 1 h. (c) 2 N HCl, THF, 60 °C, 3 h. (d) AllylMgBr, THF, 0 °C, 4 h. (e) 9-BBN, THF,
reflux, 4 h then H2O2, NaOH, rt, 3 h. (f) PPh3, CBr4, DCM, rt, 16 h.
5
NH
6
nd
4.0
O
was reacted with a hydride source or a Grignard reagent.11 In the
case illustrated in Scheme 2, allylation of this ketone, followed
by olefin hydroboration/oxidation and cyclisation led to the tetra-
hydrofurans. Heterocyclic derivatives of the piperidine/cyclohex-
ane were prepared by simple displacement of halo-substituted
heterocycles with an amine (according to step (e) in Scheme 1 be-
low), followed by acylation.
This simple, modular chemistry was especially amenable to
parallel chemistry in 96-well plate format, allowing rapid modifi-
cation of all 3 substituents around the central amide N atom
through simple variation of the amine, acyl or cycloalkyl-/hetero-
aryl component.
7
1000
nd
4.4 (4.9)
4.4
OH
OH
8
200
75
O
9
300
nd
4.9 (5)
5.7
O
N
10
11
12
13
14
>1000
>1000
175
370
5
NH2
Biological activity was assessed initially using a radiolabelled
C5a binding assay in U937 cells to generate compound IC50s.12
A
secondary, functional screen assessed the inhibition of C5a-in-
duced elastase release from human neutrophils.13 Table 1 summa-
rises the activity and lipophilicity of some piperidine analogues of
2. Simple amide and amine derivatives, 3–5, were significantly less
potent than the parent urea. An early breakthrough came with the
discovery that the entire piperidine substituent could be excised,
with the ketone 6 showing good activity at 350 nM. Further
encouragement came with the trans-alcohol 7, some threefold
more active than its cis counterpart, and with the synthetically
very accessible ketal 9, equipotent with 7. Heterocyclic derivatives
10 and 11 were inactive, suggesting lipophilic groups to be fa-
voured. This was borne out by the good activity of the cyclohexyl
12 and benzyl 13 analogues. The most potent cyclohexyl substitu-
ent proved to be the cis tetrahydrofuran (THF) 14. When tested in
the functional screen, both 7 and 14 lost some 15-fold activity,
with the ketal 9 losing some fourfold in potency. The origin of
these drop offs in functional potency was unclear, but could be
due to non-specific binding of these lipophilic compounds to mem-
branes or variability in neutrophil sensitivity to a C5a challenge.
Due to ease of synthetic accessibility, the ketal was chosen as
the most suitable group around which to explore further SAR’s, de-
spite its obvious acid lability (see Table 4 below). The tetrahydro-
furan and trans alcohols offered more robust functionality to
swap at a later stage.
nd
4.0
O
Cyclohexyl
Benzyl
nd
6.5
nd
6.2
77
5.6
O
15
200
nd
5.6
O
a
Values are means of at least two experiments.
Measured at pH 7.4 in octanol/neutral buffer. nd = not determined.
b
teins,10 and were keen to replace it. As can be seen in Table 2, ini-
tial attempts to do this, even in very close analogues, for example,
16 and 17, were disappointing. It quickly became apparent that an
o-substituted aryl amide was a crucial pharmacophoric element.
This was particularly true if the o-substituent were non-polar.
Hence, while m-benzoyl, polar o-substituents or heterocyclic
analogues (18, 20–22) were weaker than benzothiophene, 1-naph-
thyl 19, was of equivalent potency, and the o-alkyl benzoyl deriv-
The next area we looked at was the acyl benzothiophene group.
We were particularly concerned about this group being S-oxidised
in vivo and resulting toxicity through addition of plasma pro-

J. Blagg et al. / Bioorg. Med. Chem. Lett. 18 (2008) 5601–5604
5603
Table 2
Table 3
C5a receptor binding and functional activity for a series of benzothiophene analogues
of compound 9
C5a receptor binding and functional activity for a series of p-chloro-phenethylamine
analogues of compound 9
Cl
O
O
R
N
R
N
S
O
O
O
O
Compound
R
125I Binding
affinity IC50
(nM)
Functional
activity IC50
(nM)
cLogP
Compound
R
125I Binding
affinity IC50 (nM)
Functional
activity IC50
(nM)
cLogP
(LogD)b
a
a
(LogD)b
a
a
9
75
300
nd
4.9 (5)
4.9
9
75
300
nd
4.9 (5)
3.7
Cl
S
S
O
27
300
Cl
Me
16
17
18
19
20
21
22
23
24
25
>1000
>1000
1000
80
N
N
28
29
30
31
32
33
34
35
150
nd
5.3
2.7
4.0
5.7
5.4
5.6
6.0
4.9
nd
3.6
Cl
>1000
395
90
N
nd
4.0
Benzyl
Me Me
nd
OMe
nd
4.9
300
nd
Cl
O
NHMe
>1000
835
nd
3.2
435
5
N
N
Cl
Me
30
O
nd
3.9
EtO
O
Cl
Me
89
nd
MeO
MeO
O
175
nd
3.5
OMe
Me
Me
35
290
nd
O
Me
1.5
29
5.7
a
Me
Values are means of at least two experiments.
Measured at pH 7.4 in octanol/neutral buffer. nd = not determined.
b
18
nd
4.8
Me
The final area of SAR exploration around 2 was the chlorophen-
ethylamine group (Table 3). Conformational locks in the ethyl
chain provided modest gains in potency (28 and 31) while short-
ened/extended linkers (30 and 32), chlorophenyl isomers (27) or
heterocyclic analogues (29) of the phenyl group all lost potency.
400
1600
1360
3.8
N
26
110
3.7 (4.0)
Table 4
Human liver microsome stability and pharmacokinetic data for selected compounds
N
a
Compound cLogP
(LogDa)
HLM T1/2
(mins)
Pharmacokinetic parameters
Values are means of at least two experiments.
Measured at pH 7.4 in octanol/neutral buffer. nd = not determined.
b
2
7
5.6
11
nd
4.4 (4.9) 14
Rat: 0.5 mg/kg iv, 10 mg/kg po T1/2 0.5 h, Cl
67 ml/min/kg, Vd 3 L/kg, F 0.5%
nd
Dog: 0.5 mg/kg iv, T1/2 1.4 h, Cl 29 ml/min/kg,
Vd 3.5 L/kg
atives 23 and 24 were much more active. The latter two com-
pounds were of particularly poor metabolic stability and solubility
and were not pursued further. An investigation into introducing
polarity into the naphthyl ring through insertion of N atoms was
made. The best of these were 25 and particularly 26, of comparable
potency to naphthyl and a log unit more polar.
26
36
3.7 (4.0) 10b
4.3 (>5)
20
a
Measured at pH 7.4 in octanol/neutral buffer.
Primary metabolite was the alcohol, via hydrolysis and reduction of the ketal.
b
nd, not determined.

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J. Blagg et al. / Bioorg. Med. Chem. Lett. 18 (2008) 5601–5604
Amino-acid ester derived side chains (33), showed good pri-
proach wherein ionisable groups were targeted, is detailed in the
second paper in this series.
mary and functional potency, but attempts to then saturate/reduce
the chlorophenyl group in compounds 34 and 35 were not success-
ful. Overall, the chlorophenyl group appeared to offer the best bal-
ance of potency in a relatively small side chain.
Acknowledgments
At regular intervals throughout the above SAR explorations,
progress towards our goal of a compound suitable for oral dosing
was assessed. Table 4 summarises some in vitro and in vivo data
for a selection of compounds. The benzothiophene analogues 2
and 7 suffered from very short hepatic microsomal half-lives,
the latter exhibiting low oral bioavailability in the rat due to high
Cl with respect to liver blood flow (67 vs 80 ml/min/kg),
combined with a potential for incomplete oral absorption due
to poor aqueous solubility. Both of these issues relate to the high
lipophilicity. Metabolite identification with the quinolinyl ketal
26 confirmed our expectations of the dioxolanyl group being
readily metabolized. The naphthyl group had emerged as one of
the more effective benzothiophene replacements, which was
combined with a trans cyclohexyl alcohol in compound 36, which
at IC50 125 nM, demonstrated comparable potency to its dioxola-
nyl direct analogue.
The authors gratefully acknowledge the technical support of
Chris Carr, Kevin Coote, Katherine Grant, Vicky Stubbs, Alan MacIn-
nes, Tony Chuck, Christopher Luckhurst, Paul Glossop, David Beal,
Lindsey Sprigens, Fidelma Atkinson and Alison Harper.
References and notes
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O
N
7. (a) Lanza, T. J.; Durette, P. L.; Rollins, T.; Siciliano, S. J.; Cianciarulo, D. N.;
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36
IC50 125nM
Function. IC50 380nM
cLogP 4.4
OH
Following iv administration to dog, 36 exhibited moderate clear-
ance with respect to liver blood flow (29 vs 50 ml/min/kg). How-
ever, when combined with its moderate volume of distribution
this led to a short elimination T1/2. The microsomal half-life sug-
gested that clearance in man would also be moderate to high, with
respect to liver blood flow, indicating that 36 would be likely to ex-
hibit moderate to low oral bioavaliability and a short elimination
T1/2 in man.
In conclusion, no compounds from this investigation had the
appropriate combination of functional potency and physicochemi-
cal properties to be therapeutically useful oral agents. The data in
Table 4 clearly shows a need to reduce lipophilicity and clearance
in the series to provide a compound with a reasonable oral phar-
macokinetic profile. The above SAR indicates the difficulty in
achieving this goal through introduction of polar functionality or
downsizing without compromising potency. An alternative ap-
2544;
g Chemocentryx Inc. WO Patent 07051062, 2007.; h Jerini AG,
WO06128670, 2006.
8. Details of this work will be reported elsewhere.
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1997, 23, 3.
10. Treiber, A.; Dansette, P. M.; Amri, H. E.; Girault, J.-P.; Ginderow, D.; Mornon, J.-
P.; Mansuy, D. J. Am. Chem. Soc. 1997, 119, 1565.
11. Hydride reduction of cyclohexanones forms predominantly trans alcohols,
while larger nucleophiles favour cis. Isomers were separable by
chromatography.
12. Bound C5a was measured at 4 °C using dibutyryl cAMP-differentiated U937
cells and a filtration assay using 160pM 125I-labelled r-hC5a (NEN) to measure
bound radioligand.
13. Neutrophil degranulation was measured in cytochalasin B-primed isolated
human neutrophils at 37 °C by quantification of released elastase by cleavage
of the chromogenic MeOSuc-Ala-Ala-Pro-Val-pNA. C5aR antagonists were pre-
incubated with neutrophils for 5 min at 37 °C prior to a 1 nM r-hC5a (Sigma)
challenge.