Inhalation by design: Dual pharmacology β-2 agonists/M3 antagonists for the treatment of COPD

Article abstract of DOI:10.1016/j.bmcl.2010.10.132

This paper describes the successful design and development of dual pharmacology β-2 agonists-M3 antagonists, for the treatment of chronic obstructive pulmonary disorder using the principles of 'inhalation by design'. A key feature of this work is the comb

Full text of DOI:10.1016/j.bmcl.2010.10.132

Bioorganic & Medicinal Chemistry Letters 21 (2011) 2759–2763
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Inhalation by design: Dual pharmacology b-2 agonists/M3 antagonists
for the treatment of COPD
Lyn H. Jones a, , Helen Baldock a, Mark E. Bunnage a, Jane Burrows b, Nick Clarke c, Michele Coghlan c,
⇑
David Entwistle b, David Fairman d, Neil Feeder b, Craig Fulton b, Laura Hilton a, Kim James a, Rhys M. Jones d,
Amy S. Kenyon a, Stuart Marshall a,b,c, Sandra D. Newman a, Rachel Osborne a, Sheena Patel c,
Matthew D. Selby a, Emilio F. Stuart c, Michael A. Trevethick c, Karen N. Wright c, David A. Price a
a Sandwich Chemistry, World Wide Medicinal Chemistry, Pfizer, Ramsgate Road, Sandwich CT13 9NJ, UK
b Pharmaceutical Sciences, Pfizer, Ramsgate Road, Sandwich CT13 9NJ, UK
c Allergy and Respiratory Biology, Pfizer, Ramsgate Road, Sandwich CT13 9NJ, UK
d Pharmacokinetics, Dynamics and Metabolism, Pfizer, Ramsgate Road, Sandwich CT13 9NJ, UK
a r t i c l e i n f o
a b s t r a c t
Article history:
This paper describes the successful design and development of dual pharmacology b-2 agonists-M3
antagonists, for the treatment of chronic obstructive pulmonary disorder using the principles of ‘inhala-
tion by design’. A key feature of this work is the combination of balanced potency and pharmacodynamic
duration with desirable pharmacokinetic and material properties, whilst keeping synthetic complexity to
a minimum.
Received 3 September 2010
Revised 25 October 2010
Accepted 26 October 2010
Available online 31 October 2010
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
COPD
Inhalation
Dual pharmacology
MABA
Crystallinity
Chronic bronchitis and emphysema are components of chronic
obstructive pulmonary disease (COPD) that damages the airways
of the lungs leading to reduced air flow. COPD is a major cause of
disability and is predicted to be the third largest cause of death
by 2030.1 Treatments for COPD include the use of bronchodilators
that increase lung air flow, the two major classes being inhaled b-2
adrenergic agonists and muscarinic M3 antagonists. Long acting
b-2 agonists (LABAs) and long acting muscarinic antagonists
(LAMAs) have been, and continue to be, developed to enable
once-daily inhalation dosing. Significant efficacy improvements
can be seen from the combination of LABAs and LAMAs.2 Moreover,
triple therapy, from the optimal synergistic combination of a LABA,
LAMA and an inhaled corticosteroid, represents an exciting new
treatment paradigm for this debilitating condition.3 However, the
complexity of combining three different drugs that operate via
three distinct mechanisms into a single device for inhalation dos-
ing, such that patient compliance is high, is considerable. To facil-
itate the triple therapy concept, we and others,4 have pursued a
strategy to incorporate muscarinic antagonism and b-2 agonism
into a single molecule (MABA), such that combination with an
ICS could be achieved in a dry powder inhaler. Indeed, a number
of dual combination inhalation therapies are already available for
the treatment of COPD.
A clear advantage of inhalation is the potential to target the
active drug directly to the site of action and, importantly, minimize
the potential for side effects associated with systemic drug
exposure. Effective inhaled drugs require a number of essential
attributes to be secured: very low dose and optimal material prop-
erties (to facilitate delivery), suitable solubility and pharmacoki-
netics, and the desired pharmacodynamic duration of action in
the lung. Although the medicinal chemistry associated with
inhaled drug discovery is distinct from classical oral drug design,
many of the key insights and learnings may be transferable into
more mainstream medicinal chemistry.
To design a dual pharmacology molecule5 we chose a strategy
to conjugate a b-2 agonist motif onto that of an M3 antagonist. This
would create larger molecules, with high lipophilicity, that are
likely to possess high metabolism and low oral absorption to min-
imize potential systemically-driven side effects from the swal-
lowed component following inhalation. A caveat of this strategy
is the potential to create a highly synthetically complex molecule,
that would prohibit rapid development, and therefore enabling
synthesis was a key component of our medicinal chemistry design.
⇑
Corresponding author. Tel.: +44 1304 644 256; fax: +44 1304 651 821.
E-mail address: lyn.jones@pfizer.com (L.H. Jones).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.10.132

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L. H. Jones et al. / Bioorg. Med. Chem. Lett. 21 (2011) 2759–2763
OH
Cl
Cl
OH
H
N
H
N
N
O
HO
HO
Tolterodine
LABA 1
OH
OH
H
N
H
N
O
HO
HO
Figure 1. b-2 Agonists and the M3 antagonist tolterodine.
O
HO
Salmeterol
HN
Formoterol
CHO
OBn
OBn
i.
ref. 10
ii.
N
N
tolterodine
NC
O
12
13
OH
OH
OH
H
N
H
N
N
iii.
N
H2N
O
HO
HO
2
14
Scheme 1. Synthesis of the amide-tethered series. Reagents: (i) MeCN, KOH aq; (ii) Pd/C, H2, AcOH; (iii) Acid, WSCDI, pyridine.
Novel saligenin LABAs, exemplified by compound 1 (Fig. 1),
were described previously, that possess impressive pharmacologi-
cal profiles.6 An in vitro b-2 wash-off assay was developed to pro-
vide evidence of a long duration of action (a small shift in potency
when the receptor is washed),7 that was confirmed via prolonged
functional antagonism of electrically field stimulated guinea-pig
trachea. Low membrane permeability was achieved through the
introduction of hydrogen-bonding groups,8 particularly through
the benzylamide, that also ensures high first-pass metabolism.
Moreover, the availability of bulk material for this template and
the ability to analog the benzylamide provided an attractive syn-
thetic starting point to explore conjugation of the M3 motif.
We decided the potent M3 antagonist tolterodine (Fig. 1) was an
ideal fragment to incorporate into our design of a dual pharmacol-
ogy agent. We have previously described the ability to functionalise
the toluene methyl group to create a fluorometric antimuscarinic
for assay development.9 Consequently, the structure–activity
relationships also suggest this position of the tolterodine template
may be resilient to conjugation to a b-2 motif.
measuring offset kinetics using a dilution-offset methodology
whereby the offset is inferred from the on rate of co-administered
3H-NMS (N-methyl scopolamine) and is expressed as the time ta-
ken to reach 50% of total 3H-NMS for solvent treated membranes.11
Interestingly, although the offset kinetics were considerably
improved over tolterodine itself (Table 1), we believed that further
improvements could be made to the M3 off rate of saligenin 2.
We explored the SAR of this amide series by investigating the
effect of the b-2 agonist trigger head group, and prepared the sul-
fonamide 3 and resorcinol 4 derivatives.12 These compounds pos-
sessed similar profiles to the saligenin 2 and disappointingly
retained the mediocre M3 kinetics (Table 1). Similarly, the trun-
cated derivative 5 achieved potent antimuscarinic and b-2 effects,
but lacked pharmacological duration against both receptors in this
case. When a long aliphatic chain was used to link the b-2 and M3
motifs, there was a significant loss of b-2 potency (compound 6).
All derivatives possessed rapid metabolic turnover, as assessed in
human liver microsomes predicting for high first pass metabolism
and low systemic exposure.
The synthesis commenced with benzyl protection of the toltero-
dine phenol and subsequent oxidation of the toluene group to the
benzaldehyde 12 (Scheme 1).10 Homologation with deprotonated
acetonitrile provided the unsaturated nitrile 13 that was reduced
and benzyl-deprotected to the amine 14 in one step. Amidation
yielded the saligenin 2 in a very efficient total synthesis, in just
four steps from an antimuscarinic drug. Other amide-linked
MABAs were prepared in an analogs manner.
We next decided to explore structural and physicochemical-
kinetics relationships of the linker on M3 offset and focused on
replacing the polar amide moiety with an ether linkage. In partic-
ular, we concentrated on the aryl–alkyl and alkyl–alkyl ether con-
nectivities present in the potent b-2 agonists formoterol and
salmeterol (Fig. 1).13 The functional tether from the tolterodine
motif was created using ‘bromotolterodine’ 15,14 that was homol-
ogated using Heck reaction conditions to provide 16 (Scheme 2).
Hydrogenation of the olefin and benzyl deprotection, followed by
acid-mediated cleavage of the Boc group, revealed amine 17, that
was subsequently coupled to bromide 18.14 Debenzylation and
Amide 2 possessed remarkable b-2 and M3 potency and the b-2
duration, as assessed in the wash-off assay, was comparable to the
lead saligenin LABA 1 (Table 1). We then assessed M3 duration by

L. H. Jones et al. / Bioorg. Med. Chem. Lett. 21 (2011) 2759–2763
2761
Table 1
Pharmacological and metabolic profiles of tolterodine, LABA 1 and MABAs 2-11a
b
Compound
b-2 EC50
(nM)
b-2-fold
M3 Ki
(nM)
M3 offsetc
(min)
MW
325
c Log P
HLMd
l/min/mg)
shiftb
(l
OH
N
n/a
n/a
2.9
3.6
n/a
<15
n/a
5.2
163
tolterodine
Cl
Cl
OH
H
N
H
N
1.1
17
517
723
3.1
5.3
>440
O
HO
HO
1
OH
OH
OH
OH
H
N
H
N
N
N
2.8
1.1
0..7
0.091
0.276
0.397
355
166
135
263
O
HO
HO
2
H
N
H
N
8.1
15
787
695
5.2
5.3
109
O
HO
HO
HN
O
S
3
O
OH
OH
H
N
H
N
N
>440
O
OH
4
OH
O
H
N
N
10
19
6.10
<15
583
3.4
nd
OH
HO
HN
S
O
5
OH
OH
H
N
H
N
N
116
7.3
1.9
2.6
nd
1.45
0.634
0.765
4.7
nd
781
732
669
654
712
6.3
6.3
6.4
6.9
5.6
87
67
54
nd
57
O
HO
HO
HN
HN
O
S
6
O
OH
H
OH
OH
OH
N
N
13
>1200
>1200
nd
O
S
O
7
O
OH
H
N
N
129
173
5.7
O
HO
HO
8
OH
H
N
HO
N
O
OH
9
OH
OH
O
H
N
N
O
0.7
0.99
>1200
HO
HN
10
S
O
(continued on next page)

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L. H. Jones et al. / Bioorg. Med. Chem. Lett. 21 (2011) 2759–2763
Table 1 (continued)
b
Compound
b-2 EC50
(nM)
b-2-fold
M3 Ki
(nM)
M3 offsetc
(min)
MW
706
c Log P
HLMd
l/min/mg)
shiftb
(l
OH
H
N
OH
N
2.4
1.1
0.305
>1200
6.6
74
O
HO
HN
11
O
a
nd = not determined; n/a = not applicable.
b
c
Washed and unwashed agonist potency of compounds at the b-2 adrenoceptor, by measuring their ability to stimulate intracellular cAMP production in CHO-hb-2 cells.
Offset kinetics from human cloned M3 receptor using [3H] N-methyl scopolamine (dilution method).
Clearance values in human liver microsomes.
d
OBn
BocHN
OBn
ii, iii
N
i.
N
O
Br
16
15
Si
O
O
OH
H2N
Br
iv, v, vi
N
11
+
O
BnO
18
HN
17
Scheme 2. Synthesis of the ether-tethered series. Reagents: (i) Olefin14, P(o-toluene)3, Pd(OAc)2, Hünigs base, MeCN; (ii) H2, Pd/C, EtOH; (iii) 2 N HCl/Et2O, EtOH; (iv) NaHCO3,
MeCN; (v) ammonium formate, Pd(OH)2, MeOH; (vi) Et3NÁ3HF, MeOH, THF.
Figure 2. Relationship between M3 offset kinetics and lipophilicity (compounds 2, 3, 4, 5, 7, 8, 10, 11 and tolterodine).
silyl ether deprotection furnished MABA 11 in an efficient manner.
Other ether-linked MABAs were prepared using an analogs
methodology.
All derivatives in the ether series were potent dual pharmacol-
ogy MABAs, although b-2 potency for the saligenin 8 and resorcinol
9 was lower than expected (Table 1). Other b-2 head groups inves-
tigated were sulfonamide (6 and 10) and quinolinone (11).12 Pleas-
ingly, the M3 offset kinetics for this series were very impressive
and a link with lipophilicity appears possible, (Figure 2). Again,
the desired metabolic instability has been retained across the
We then assessed the potency of our key compounds in the gui-
nea-pig trachea (GPT) model using electric field stimulation to re-
lease endogenous acetylcholine.6 The drive for bronchoconstriction
in this model operates through both b-2 and M3 mechanisms and
therefore values here reflect combined bronchodilatory effects. A
‘duration of action’ was defined as the time taken for the muscle
tone at a submaximal concentration of the compound to recover
by 50% of the inhibition induced. As expected the amide 3 and
ether 11 are potent bronchodilators in this model with respectable
duration of action (Table 2).15 To delineate the M3 drive, the b-2
antagonist propranolol was added to the EFS model to block the
series (HLM >50 ll/min/mg).

L. H. Jones et al. / Bioorg. Med. Chem. Lett. 21 (2011) 2759–2763
2763
Table 2
Ex vivo pharmacological profiles of MABAsa
Compound
GPT EFS
EC50 (nM)
GPT EFS
duration T50% (h)
GPT EFS + propranolol
EC50 (nM)
GPT EFS + propranolol
durationc (h)
GPT histamine
EC50 (nM)
b
b
c
d
OH
OH
OH
H
N
H
N
N
O
1.3
4.1
>6.5
5.3
4.0
4.8
2.1
4.0
HO
HO
HN
O
S
3
O
OH
H
N
N
O
>7.7
>10
HN
11
O
a
All data are geometric means with 95% CI, n = 4.
GPT = Guinea pig trachea. EFS = Electric field stimulation. Potency and duration for b-2 and M3 components of bronchodilation. Submaximal concentration used for the
b
duration is that which gives 70% inhibition of the EFS response.
c
Potency and duration for M3 component of bronchodilation.
Potency for b-2 component of bronchodilation.
d
b-2 effect. Amide 3 was found to retain the potency, but lost phar-
macological duration, in line with the fast offset M3 kinetics. How-
ever, ether 11 retains both potency and duration of action,
resulting from the long M3 offset. To confirm the b-2 pharmacolog-
ical component, the GPT assay was modified to use histamine as
the bronchoconstricting agent.16 Both 3 and 11 are potent bron-
chodilators in this model as expected.
Acknowledgements
We thank Farhat Hussain for investigating the scale-up of the
quinolinone b-2 head group, and Paul Glossop, Graham Lunn and
John Harvey for useful discussions.
References and notes
Ether 11 effected a dose-related inhibition of methacholine-in-
duced bronchoconstriction in dogs17 when dosed intratracheally,
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index over cardiovascular side effects (heart rate and contractility)
threefold superior to salmeterol in this model, further supporting
the exciting potential of this novel MABA.
As the product profile of a MABA requires formulation into a
dry powder inhaler, either as a standalone agent, or as a combi-
nation therapy with an ICS, it is imperative to have the desired
material properties. Following an extensive sitting-drop salt
screen,18 the fumarate salt of 11 was identified as a crystalline
hit and converted on scale-up to a 1:1 fumarate hydrate (melting
point 151.8–153.3 °C), thus enabling further formulation develop-
ment of this compound. Additionally, ether 11 was clean in
in vitro genetic toxicity testing (AMES and micronucleus) and
wide-ligand profiling against a broad protein panel at CEREP,
highlighting the exquisite selectivity that can be achieved in this
dual pharmacological approach. Rat pharmacokinetics experi-
ments on 11, when combined with the poor membrane perme-
ability and high metabolic instability,19 predict negligible
human oral bioavailability (<5%) thus minimizing potential risks
from systemic exposure.
In conclusion, we have created novel MABAs using the princi-
ples of ‘inhalation by design’. Balanced potency and pharmacolog-
ical duration was combined with high metabolic clearance, low
oral bioavailability, desirable material properties and minimized
synthetic complexity, culminating in compound 11. Further elabo-
ration and development of the ether-tethered series will appear
elsewhere.
14. Jones, L. H.; James, K.; Price, D. A. WO2007107828.
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Clarke, N.; Trevethick, M.; Yeadon, M.; Perros-Huguet, C. Presented at the
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19. Rat intravenous pharmacokinetics (dose 2 mg/kg): AUC 2670 ng h/mL;
clearance 12.6 mL/min/kg.