Succinyl hydroxamates as potent and selective non-peptidic inhibitors of procollagen C-proteinase: Design, synthesis, and evaluation as topically applied, dermal anti-scarring agents

Article abstract of DOI:10.1016/j.bmcl.2008.10.036

Succinyl hydroxamates 1 and 2 are disclosed as novel series of potent and selective inhibitors of procollagen C-proteinase (PCP) which may have potential as anti-fibrotic agents. Carboxamide 7 demonstrated good PCP inhibition and had excellent selectivity

Full text of DOI:10.1016/j.bmcl.2008.10.036

Bioorganic & Medicinal Chemistry Letters 18 (2008) 6562–6567
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Succinyl hydroxamates as potent and selective non-peptidic inhibitors of
procollagen C-proteinase: Design, synthesis, and evaluation as topically
applied, dermal anti-scarring agents
a,
a
e
b
a
Simon Bailey a, , , Paul V. Fish , Stephane Billotte , Jon Bordner , Doris Greiling , Kim James ,
*
*
Andrew McElroy a, James E. Mills a, Charlotte Reed c, Robert Webster d
a Department of Discovery Chemistry, Pfizer Global Research and Development, Sandwich Laboratories, Ramsgate Road, Sandwich, Kent CT13 9NJ, UK
b Department of Discovery Biology, Pfizer Global Research and Development, Sandwich Laboratories, Ramsgate Road, Sandwich, Kent CT13 9NJ, UK
c Department of Pharmaceutical Sciences, Pfizer Global Research and Development, Sandwich Laboratories, Ramsgate Road, Sandwich, Kent CT13 9NJ, UK
d Department of Pharmacokinetics, Dynamics & Metabolism, Pfizer Global Research and Development, Sandwich Laboratories, Ramsgate Road, Sandwich, Kent CT13 9NJ, UK
e Department of Materials Science, Pfizer Global Research and Development, Groton Laboratories, Eastern Point Road, Groton, CT 06340, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Succinyl hydroxamates 1 and 2 are disclosed as novel series of potent and selective inhibitors of procol-
lagen C-proteinase (PCP) which may have potential as anti-fibrotic agents. Carboxamide 7 demonstrated
good PCP inhibition and had excellent selectivity over MMPs involved in wound healing. In addition, 7
was effective in a cell-based model of collagen deposition (fibroplasia model) and was very effective at
penetrating human skin in vitro. Compound 7 (UK-383,367) was selected as a candidate for evaluation
in clinical studies as a topically applied, dermal anti-scarring agent.
Received 19 September 2008
Revised 7 October 2008
Accepted 8 October 2008
Available online 11 October 2008
Keywords:
Ó 2008 Elsevier Ltd. All rights reserved.
Succinyl hydroxamates
Procollagen C-proteinase
Transepidermal delivery
Anti-scarring
Conformational analysis
Fibrosis is a complex process which is characterized by exces-
sive accumulation of extracellular matrix (ECM), principally type
I collagen.1,2 In the case of wounds to the skin, dermal fibroblasts
respond to activation by transforming growth factor-b (TGF-b1)
by releasing soluble procollagens and procollagen C-proteinase
(PCP; EC 3.4.24.19 also known as BMP-1).3–5 Proteolytic cleavage
of the C-terminal propeptide of types I, II, and III procollagens by
PCP affords collagen, which undergoes self-assembly into fibrils
followed by cross-linking to give an insoluble collagen matrix.
The excessive accumulation of ECM by these processes leads to
the formation of scar tissue.6 Hence, inhibition of PCP activity7
was identified as an attractive point at which to intervene in the
collagen deposition pathway.
candidate for clinical evaluation as a topically applied, dermal
anti-scarring agent.8
O
O
O
O
R
R
N
N
NR1R2
NR1R2
HOHN
HOHN
3
5
4
2
O
N
O
5
X
1
2
Previously, we have described the identification of the succinyl
hydroxamate template by a combination of file screening of li-
braries of matrix metalloproteinase (MMP) inhibitors and specific
structural modifications; these efforts furnished oxadiazole ester 6
as a starting point for optimization (Fig. 1).8,9 Although it was clear
that the lipophilic side chain of 6 played an important role in regu-
lating activity against PCP (compare 3 vs 4 and 5 vs 6), it was
decided to explore replacements for the chemically and metaboli-
cally vulnerable C3-ester group in the first phase of analogue
synthesis. In particular, the effect of simple carboxamide substitu-
In this letter, we now describe the detailed SAR which lead to
the identification and optimization of oxadiazoles 1 and oxazoles
2 as potent and selective non-peptidic inhibitors of PCP. Further-
more, one of these compounds, 7 (UK-383,367) was selected as a
* Corresponding authors. Tel.: +1 858 526 4692 (S.B.); tel.: +44 (0)1304 644589
(P.V.F.).
E-mail addresses: simon.bailey@pfizer.com (S.B.), paul.fish@pfizer.com (P.V.F.).
Present address: Department of Chemistry, Pfizer Global R & D, La Jolla Labs, San
Diego, CA 92121, USA.
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.10.036

S. Bailey et al. / Bioorg. Med. Chem. Lett. 18 (2008) 6562–6567
6563
Table 1
Targeted Screening of
Pfizer Compound File
PCP and MMP-2 inhibition by oxadiazole carboxamides 7–21a,b
(e.g. MMP-i; Zn2+ ligands)
O
O
IC50 < 10 μM
N
NR1R2
HOHN
O
N
R
7-21
Compound
NR1R2
PCP IC50 (nM)
MMP-2 IC50 (nM)
O
O
H
7
(S)-7
8
NH2
NH2
NHMe
NMe2
NH-nPr
NH-i-Pr
NHCH2-cPr
NHCH2Ph
NHCH2(2-Py)
NHCH2CO2H
Pyrrolidine
Piperidine
Morpholine
4-Methylpiperazine
N(Me)CH2Ph
N(Me)CH2(2-Py)
44
>2000
28
10
32
21
33
57
37
21
43
26
25
54
65
40
>60,000
NT
N
N
Me
HOHN
H
O
>30,000
>100,000
21,700
>100,000
>30,000
NT
>30,000
>100,000
NT
74,900
NT
NT
81,800
50,500
Me
Me
Me
9
10
11
12
13
14
15
16
17
18
19
20
21
3: R = Ph
4: R = c-Hex
PCP: IC50 820 nM
PCP: IC50 8765 nM
MMP-2: IC50 135 nM
MMP-2: IC50 1000 nM
Reduce peptidic
nature
R
a
See Ref. 9 for complete details of assay conditions. IC50 values are geometric
means of at least three experiments. Differences of <2-fold should not be considered
significant.
O
N
CO2Et
HOHN
b
NT denotes not tested.
O
N
5: R = Ph
PCP: IC50 6600 nM
MMP-2: IC50 5900 nM
6: R = c-Hex
PCP: IC50 36 nM
MMP-2: IC50 80,000 nM
to afford the alcohols 35 and 36, respectively. Reaction of ser-
ine-derived alcohol 35 with the Burgess reagent promoted cyc-
lodehydration to oxazoline 37, which then underwent
aromatization to the corresponding oxazole by treatment with
CuBr2/DBU.12 At this point, a protected hydroxamic acid group
was introduced to give 38 prior to hydrolysis of the ethyl ester
and amide formation under standard conditions. Finally, the
hydroxamate benzyl protecting group was cleaved by hydrogen-
olysis to give 22 and 23. A different order of chemical steps was
employed for the fully substituted 5-Me-oxazole targets 24–25.
Oxidation of the secondary alcohol 36 using the Dess–Martin re-
agent gave the keto-amide 39 which underwent ring closure by
the action of PPh3-I2 to afford 4013 Oxazole 40 was converted to
the targets 24–25 using transformations similar to those de-
scribed above.
Figure 1. Summary of key structures in the identification of succinyl hydroxamate
lead series by a combination of file screening and specific structural modifications.
ents on the oxadiazole ring was assessed, and, as an initial venture –
CONH2 (7), –CONHMe (8), and –CONMe2 (9) were prepared and
screened for inhibition of PCP activity.
These carboxamides furnished promising leads as 7–9 all had
PCP inhibitory activity equivalent or superior to ester 6 and no sig-
nificant MMP-2 activity (Table 1). tert-Amide 9 was the most po-
tent inhibitor (IC50 10 nM) of this set and gave encouragement
that potent, selective inhibitors of PCP could be achieved. Hence,
a more detailed investigation of the SAR of the carboxamide substi-
tuent was undertaken in both the oxadiazole (1) and oxazole (2)
templates.
The synthesis of oxadiazole target compounds 7–21 (Table 1)
were prepared as described in Scheme 1. N-Hydroxy amidine 27
was coupled with (R)-succinic acid mono-ester 2610 to give the
O-acyl derivative 28 and then thermal cyclodehydration of 28 in
xylene created the 1,2,4-oxadiazoles 29. Two general methods
were used to install the 3-carboxamide: either selective deprotec-
tion of the tert-butyl esters with TFA gave carboxylic acid 30 and
then treatment with amines HNR1R2 gave amides 31, or alterna-
tively, these two steps were reversed (i.e., 29 ? 32 ? 31). Finally,
activation of the carboxylic acid 31 with i-BuOCOCl followed by
reaction with TMSONH2 and methanolic work-up afforded
hydroxamates 7–21. The (S)-enantiomer (S)-7 was prepared by
an identical sequence but starting with (S)-26.
All target compounds (Tables 1 and 2) were tested for their abil-
ity to inhibit the PCP mediated cleavage of a fluorogenic peptide
substrate.14 Selected compounds were then routinely assessed
for physicochemical properties consistent with transepidermal
delivery (TED)15 (mp > 110 °C; aq solubility > 50
lg/ml; stability
under autoclave conditions), and for selectivity over MMPs in-
volved in wound healing.16 Inhibition of PCP activity was con-
firmed for selected compounds with the endogenous substrate
procollagen. Finally, preferred compounds were evaluated in an
in vitro cell-based model of collagen deposition (fibroplasia mod-
el)17 and for their ability to penetrate human skin in vitro.18
A wide variety of amide substituents were tolerated at the C-3
position of the oxadiazole (Table 1). Thus, the primary amide 7,
n-alkyl secondary amides 8 and 10, as well as those with branching
at the a- or b-carbon, 11 and 12, respectively, all inhibit PCP with
IC50s in the 20–45 nM range. The benzylamide, 13, does show a
small, 2-fold, drop in potency compared with the methyl amide
8. However, activity is largely restored when a heteroaryl ring is
featured as in compound 14. The presence of a carboxylic acid
group, as in 15, did not detrimentally influence activity.
The oxazoles target compounds 22–25 (Table 2) were prepared
using a similar overall synthetic strategy as for the oxadiazoles but
employed the methods of Wipf11 for the creation of the oxazole
ring (Scheme 2). (R)-Succinic acid mono-ester 26 was esterified
with an appropriate L-serine (33) or L-threonine (34) derivative

6564
S. Bailey et al. / Bioorg. Med. Chem. Lett. 18 (2008) 6562–6567
O
R
O
R
O
R
a
b
R
H2N
HO
CO2Et
O
N
CO2Et
OH
+
tBuO
N
tBuO
tBuO
N
O
O
N
O
CO2Et
H2N
27
28
29
26
O
O
R
c
d
N
N
CO2Et
HO
O
O
O
R
O
O
R
O
30
e
N
NR1R2
N
HO
HOHN
NR1R2
O
N
R
O
O
N
c
d
31
N
N
NR1R2
7-21
tBuO
32
R = -CH2CH2CH2-cHex
Scheme 1. Synthesis of oxadiazole 3-carboxamides 7–21. Reagents and conditions: (a) WSCDI, HOBt, 1,4-dioxane, 0 °C ? rt; (b) xylene, 130 °C; (c) TFA, CH2Cl2, 0 °C ? rt; (d)
HNR1R2, EtOH; (e) i-BuOCOCl, N-methylmorpholine, THF or DMF, 0 °C, then H2NOSiMe3, rt, then MeOH, rt.
A similar pattern was observed with tertiary amides 16–21,
with acyclic and cyclic moieties all having relatively little influence
on enzyme inhibition. The presence of heteroatoms 18, and basic
centers 19 were well tolerated with only minor variations in the
observed potencies. Again, the incorporation of a benzyl group 20
gives rise to slightly weaker activity than that seen with other moi-
eties. Hence, it has proved difficult to improve PCP inhibitory activ-
ity below a plateau of IC50 < 20–45 nM.
oxazoles 22–25 were prepared and screened for inhibition of PCP
activity (Table 2).
Comparison of PCP activity of C5-H oxazoles, 22 and 23, with
the corresponding oxadiazole analogues, 7 and 9, shows that activ-
ity is largely retained in the oxazole series but with a slight loss in
activity. In contrast, the C5-Me substituted oxazoles, 24 and 25, are
amongst the most potent PCP inhibitors identified in the carbox-
amide series.
It is of note that the absolute stereochemistry of chiral center
had a profound effect on PCP activity with the (R)-enantiomer
being superior (e.g., (R)-7 vs (S)-7). Furthermore, no compound
had any significant inhibition of MMP-2 activity providing evi-
dence that the 6-cyclohexyl-N-hydroxyhexanamide backbone con-
fers good affinity for PCP and selectivity over MMP-2.
For the PCP inhibitors in Table 1, it was expected that the C3
amide group and the 1,2,4-oxadiazole ring would lie co-planar
due to favorable mesomeric stabilization. As a consequence, there
would be barriers to free rotation of the amide substituent about
the carbonyl-heterocycle bond. In contrast, it was expected that a
carboxamide directly attached at C4 of a 1,3-oxazole ring might al-
low additional favorable binding modes to become available. The
oxazole ring would allow the incorporation of a H- or Me-substitu-
ent at C5 and promote an out-of-plane conformation of the amide
through alleviation of steric congestion. Consequently, the
These results prompted a more detailed analysis of the pre-
ferred conformation of the heterocycle-amide bond. Torsion angle
calculations predicted 7, 22, and 24 would all have a preference for
an in-plane conformation (Fig. 2) and these calculations were sup-
ported by a single crystal X-ray structure determination of 7 mono-
hydrate which also showed
a planar arrangement of the
carboxamide and oxadiazole ring (Fig. 3).19,20 These results were
somewhat unexpected, and the observations/principles may prove
to have general utility in the design of azole derivatives. Hence we
concluded that the slight increase in affinity for 24 (and 25) was
due to an additional positive binding interaction between the C5-
Me and the protein structure rather than conformational effects.
Overall the data presented in Tables 1 and 2 indicated that the
contribution to PCP binding affinity provided by the heterocycle
and the carboxamide substituent was relatively minor and, conse-
quently, this amide position proved to be a good handle for tuning
the physicochemical properties.
From these experiments, primary amide 7 emerged as having a
superior combination of PCP activity (IC50 44 nM) combined with
selectivity over MMP-2 (>1000-fold) and physicochemical proper-
ties thought suitable for TED delivery, and so was selected for fur-
ther assessment.
Table 2
PCP inhibition by oxazole carboxamides 22–25
Inhibition of PCP enzymatic activity was confirmed as amide 7
was found to inhibit the turnover of procollagen with a slightly en-
hanced potency (IC50 26 nM) (Table 3). Additional screening of 7
O
O
N
NR1R2
HOHN
against
a panel of MMPs showed no significant activity
O
(IC50 > 10,000 nM) (Table 3).
X
22-25
Amide 7 was then assessed for its potential to block collagen
deposition in an in vitro fibroplasia model (Table 3).17 After cultur-
ing human dermal fibroblasts for 8 days, 7 showed a dose-depen-
dant reduction in collagen deposition, both by histological
analysis and by quantification of hydroxyproline content. From
the quantitative hydroxyproline analysis, 7 inhibited collagen
Compound
NR1R2
X
PCP IC50 (nM)
22
23
24
25
NH2
H
H
Me
Me
73
45
17
17
NMe2
NH2
NMe2

S. Bailey et al. / Bioorg. Med. Chem. Lett. 18 (2008) 6562–6567
6565
O
R
O
R
R
a
H
N
H2N
HO
CO2E
X
CO2E
X
OH
+
tBuO
tBuO
O
O
HO
26
33: X = H; E = Et
34: X = Me; E = Me
35: X = H; E = Et
36: X = Me; E = Me
O
R
O
R
O
R
O
b
c, d, e
f, g, h
N
CO2Et
N
CO2Et
N
BnOHN
HOHN
NR1R2
35
tBuO
O
O
O
37
38
22, 23
O
R
O
R
O
R
O
H
N
i
j
d-h
CO2Me
Me
N
CO2Me
Me
N
NR1R2
36
tBuO
tBuO
HOHN
O
O
O
O
Me
39
40
24, 25
R = -CH2CH2CH2-cHex
Scheme 2. Synthesis of oxazole 4-carboxamides 22–25. Reagents and conditions: (a) WSCDI, HOBt, i-Pr2NEt, CH2Cl2, 0 °C ? rt; (b) Burgess reagent, THF, 80 °C; (c) CuBr2, DBU,
HMTA, CH2Cl2, 0 °C ? rt; (d) TFA, CH2Cl2, 0 °C ? rt; (e) BnONH2, CDI, i-Pr2NEt, THF, rt; (f) LiOH, 1,4-dioxane-H2O, 0 °C; (g) HNR1R2, WSCDI, HOBt, N-methylmorpholine,
CH2Cl2, rt; (h) 5% Pd-BaSO4, H2 or NH4+HCO2À, EtOH; (i) Dess–Martin periodinane, CH2Cl2, rt; (j) I2, PPh3, NEt3, THF, À78 °C ? 0 °C.
deposition by >70% at 10
M.
The ability of 7 to penetrate human skin in vitro was measured
using standard transdermal assays. Human cadaver stratum cor-
neum was mounted in a Franz diffusion cell and exposed to a solu-
l
M and with an IC50 of approximately
favorably with values for agents which are known to penetrate skin
well in vivo.21 A more detailed analysis of the ability of 7 to pene-
trate human skin was then performed. When propylene glycol–cit-
rate buffer–ethanol (20:30:50) was employed as the TED vehicle,
2
l
the saturated solubility increased to 13,800
lg/ml and the TED flux
tion of
7
in 50% aqueous propylene glycol for 24 h. The
increased to 1.59 0.56
l
g/cm2/h.
concentration of 7 passing through the skin to a receptor chamber
was measured and used to calculate a TED flux rate (Table 3).
Encouragingly, in this system 7 permeated through the skin with
Compound 7 was screened for off-target pharmacology against
a panel of receptors, enzymes and ion channels and was found to
have modest binding affinity for only the PDE-4 enzyme (69% inhi-
a calculated flux value of 0.30 0.09
l
g/cm2/h which compared
bition at 10
lM; non-subtype specific). Further evaluation showed
7 to have modest affinity for all the PDE-4 subtypes [IC50
(lM):
PDE-4a = 1.8; 4b = 1.5; 4c = 2.4; 4d = 0.9]. The consequences of this
weak PDE-4 activity was not clear although PDE-4 inhibition has
been shown to attenuate TGF-b mediated pro-collagen synthesis
and can also be considered a potential anti-fibrotic target.22
In the context of treating dermal scarring by TED delivery, it
was felt that a compound with low systemic exposure would
-294995
-294996
*
-294997
-294998
-294999
-295000
-295001
-295002
-295003
-295004
-295005
O
*
N
O
NH2
5.5 kcal mol-1
N
R
7
-2284944
-2284946
*
O
-2284948
*
-2284950
O
NH2
-284952
N
9.6 kcal mol-1
-284954
R
-284956
-2284958
-2284960
22
-309620
-309622
-309624
-309626
-309628
-309630
-309632
-309634
-309636
-309638
-309640
Me
*
O
*
O
NH2
11.4 kcal mol-1
N
R
24
Figure 2. Torsion angle scan energy calculations of 7, 22, and 24 to illustrate the
lowest energy conformations of the carboxamide wrt the heterocycle. The lowest
*
energy structures are shown; the indicating the atoms for which the torsion angle
was measured. The calculations were carried out using Density Functional Theory at
Figure 3. Single crystal X-ray structure determination of 7 monohydrate which
**
B3LYP/6-31G level of theory (Jaguar v6.5).
shows a planar arrangement of the 3-carboxamide and 1,2,4-oxadiazole ring.

6566
S. Bailey et al. / Bioorg. Med. Chem. Lett. 18 (2008) 6562–6567
Table 3
Summary of physicochemical properties, MMP selectivity, efficacy in the fibroplasia model and ability to penetrate human skin for compounds 7 and 9a,b
7
9
Physicochemical properties:
Mw
Log D7.4
pKa
Mp (°C)
324
2.6
9.0
136–138
35
352
3.6
NTc
Oil
Satd aqueous solubility in PBS (pH 7.4) (
lg/ml)
900
PCP activity:
Peptide substrate, IC50 (nM)
44
26
10
9.3
[3H]-procollagen, IC50 (nM)
MMP selectivity:
MMP-1, IC50 (nM)
MMP-2, IC50 (nM)
MMP-3, IC50 (nM)
MMP-9, IC50 (nM)
MMP-14, IC50 (nM)
>10,000
>60,000
>10,000
>10,000
>10,000
NT
>100,000
28,000
NT
NT
Efficacy in the fibroplasia model:d
% I @ 10
IC50 (nM)
l
M
76
5
67 14
NT
2100 500
In vitro penetration through human skin in 50% aqueous propylene glycol (pH 6.8):e
Solubility, Co g/ml)
g/cm2/h)
(l
670
0.30 0.09
8260
2.6 1.6
TED flux (
l
a
See Ref. 9 for complete details of assay conditions.
NT denotes not tested.
b
c
pKa ꢀ 9.0 by analogy with the measured value for 7.
d
e
Mean SEM (n P 4).
Mean SEM (n = 6–10).
Table 4
Topical pharmacokinetics in vivo confirmed 7, dosed as a solu-
tion of 0.9 mg/ml in 50% aqueous propylene glycol, penetrated in-
tact pig skin with tissue levels in the upper dermis reaching a
Pharmacokinetic parameters of 7 after single intravenous and oral administration to
rat and doga
Male rat (n = 2)
Male dog (n = 2)
steady state concentration of 10 lM after 24 h. Furthermore, no
systemic exposure of 7 was detected following topical administra-
tion in this model following repeat dosing over 7 days.
Intravenous dose (mg/kg)
2
0.5
1.5
35
Elimination half-life, T1/2 (h)
Plasma clearance, Cl (ml/min/kg)
Volume of distribution, Vd (l/kg)
0.8
157
12
N,N-Dimethyl amide 9 was the most potent PCP inhibitor
(IC5010 nM) of this set and was also selected for further character-
ization (Table 3). Amide 9 was comparable to 7 in nearly all param-
eters except that it lacked a suitable crystalline solid form.
Although not appropriate for potential clinical use, 9 proved to
be valuable tool in the development of preclinical models of scar-
ring. Evaluation of 9 in a rabbit model of cutaneous scarring estab-
lished that inhibition of PCP in vivo reduces scar hypertrophy.24
These results support the potential use of PCP inhibitors to miti-
gate hypertrophic scarring.
In summary, succinyl hydroxamates 1 and 2 are disclosed as no-
vel series of potent and selective inhibitors of PCP which may have
potential as anti-fibrotic agents. Compound 7 demonstrated good
PCP inhibition and had excellent selectivity over MMPs involved
in wound healing. In addition, 7 was effective in a cell-based model
of collagen deposition (fibroplasia model) and was very effective at
penetrating human skin in vitro. Furthermore, compound 9 (UK-
369,930), a close relative to 7 in terms of both structure and profile,
reduced scar hypertrophy in a rabbit model of scarring. Based on
this information, 7 (UK-383,367) was selected as a candidate for
evaluation in clinical studies as a topically applied, dermal anti-
scarring agent. The results of these studies will be reported in
future publications.
4.6
Oral dose (mg/kg)
Cmax (ng/ml)
Tmax (h)
NT
—
—
2
110
0.5–1.5
13
Oral bioavailability, Fo (%)
—
a
NT denotes not tested.
minimize any potential safety concerns through any off-target
activity. Screening in ADME assays showed 7 to have low stability
in liver microsomes [T1/2 (min): HLM 19; DLM 9; RLM < 2] consis-
tent with high predicted clearance. Investigation of the primary
routes of metabolism in human, dog, and rat microsomes sug-
gested 7 was primarily metabolized at the hydroxamate group to
yield the corresponding carboxylic acid and primary carboxamide.
These metabolites were not inhibitors of PCP activity
(IC50 > 30 lM).
Pharmacokinetic data for 7 were generated in vivo in rat and
dog following intravenous and oral administration (Table 4).23 Fol-
lowing single intravenous administration to rat, the plasma clear-
ance of 7 was high (Cl 157 ml/min/kg) resulting in an elimination
half-life of 0.8 h. Concentrations were below the limit of quantifi-
cation (<0.1 lg/ml) after 2 h post-administration of the dose. Plas-
ma clearance in dog was lower than that observed in rat but still
high relative to liver blood flow. The oral bioavailability of 7 in
dog was low (13%) in keeping with the plasma clearance observed.
Following administration of 7 to the isolated perfused rat liver a
hepatic extraction rate of 75–86% was obtained, suggesting that
7 undergoes extensive hepatic clearance. These results would pre-
dict rapid systemic clearance following TED delivery.
Acknowledgments
We thank the following colleagues for their expert and enthusi-
astic technical assistance: Lyndsey Addison, Yvonne Ailwood,
Cathy Allan, Gillian Allan, Gerwyn Bish, Usa Datta, Vikki Dawe,
Samantha Gaboardi, John Harvey, Carol Loosley, Emma Newstead,
Julie Owen, Christelle Pasquinet, Clint Pereira, and Simon Planken.

S. Bailey et al. / Bioorg. Med. Chem. Lett. 18 (2008) 6562–6567
6567
15. (a) Kasting, G. B.; Smith, R. L.; Cooper, E. R.. In Pharmacol. Skin; Karger: Basel,
1987; Vol. 1. p 138; (b) Hadgraft, J.; Valenta, C. Int. J. Pharm. 2000, 200, 243; (c)
Potts, R. O.; Guy, R. H. Pharm. Res. 1995, 12, 1628.
In addition we acknowledge the excellent support of the Physical
Sciences group.
16. Lijnen, H. R.; Collen, D. Plasminogen 2003, 189.
References and notes
17. (a) Clark, R. A. F.; McCoy, G. A.; Folkvord, J. M.; McPherson, J. M. J. Cell. Physiol.
1997, 170, 69; (b) Newstead, E. L.; Gaboardi, S.; Huggins, J. P.; Greiling, D. J.
Dermatol. Sci. 1998, 16(Suppl. 1), S128.
18. (a) Barry, B. W. Dermatological Formulations in Drugs and the Pharmaceutical Sciences;
Marcel Dekker Inc.: NY, 1983. p 127; (b) Roberts, M. S. Structure–permeability
considerations in percutaneous absorption. In Prediction, of Percutaneous Penetration—
Methods, Measurements, and Modelling; Scott, R. C., Guy, R. H., Hadgraft, J., Bodde, H. E.,
Eds.; IBC Technical Services, 1991; Vol. 2, p 210.
1. Trojanowska, M.; LeRoy, E. C.; Eckes, B.; Krieg, T. J. Mol. Med. 1998, 76, 266.
2. Border, W. A.; Noble, N. A. NEJM 1994, 331, 1286.
3. Lee, S.; Solow-Corero, D. E.; Kessler, E.; Takahara, K.; Greenspan, D. S. J. Biol.
Chem. 1997, 272, 19059.
4. Kessler, E. In Procollagen C-Proteinase. In The Handbook of Proteolytic Enzymes;
Academic press, 1998. pp 1–5.
5. Kessler, E.; Takahara, T.; Biniamov, L.; Brusel, M.; Greenspan, D. S. Science 1996,
271, 360.
6. Rhett, J. M.; Ghatnekar, G. S.; Palatinus, J. A.; O’Quinn, M.; Yost, M. J.; Gourdie, R.
G. Trends Biotechnol. 2008, 26, 173.
7. For a review of PCP inhibitors, see: Turtle, E. D.; Ho, W.-B. Expert Opin. Ther.
Patents 2004, 14, 1185.
8. (a) Bailey, S.; Billotte, S.; Derrick, A. M.; Fish, P. V.; James, K.; Thomson, N.
M. WO 2001047901.; (b) Bailey, S.; Fish, P. V.; James, K.; McElroy, A.
Abstracts of Papers, 227th ACS Nation Meeting, Anaheim, CA, March 28–
April 1, 2004.
9. Fish, P. V.; Allan, G. A.; Bailey, S.; Blagg, J.; Butt, R.; Collis, M. G.; Greiling, D.;
James, K.; Kendall, J.; McElroy, A.; McCleverty, D.; Reed, C.; Webster, R.;
Whitlock, G. A. J. Med. Chem. 2007, 50, 3442.
10. Derrick, A. M.; Thomson, N. M. EP 1199301, 2002.
11. Wipf, P.; Miller, C. P. Tetrahedron Lett. 1992, 33, 907.
12. Barrish, J. C.; Singh, J.; Spergel, S. H.; Han, W. C.; Kissick, T. P.; Kronenthal, D. R.;
Mueller, R. H. J. Org. Chem. 1993, 58, 4494.
19. Crystallographic data (excluding structure factors) for 7ꢁH2O have been
deposited with the Cambridge Crystallographic Data Centre as
Supplementary Publication Number CCDC707773. Copies of the data can be
obtained free of charge, on application to CCDC, 12 Union Road, Cambridge,
CB2 1EZ, UK.
20. An analysis of crystal structures of related compounds available in the PDB and
CCDC also showed planar arrangements between the carboxamide and the
oxadiazole and oxazole rings.
21. To establish a baseline level of acceptable flux in our in vitro TED model, a set
of compounds with commercial precedence of successful transdermal delivery
were evaluated, utilizing the same vehicle and test conditions. For example, a
solution of clonidine at 1 mg/ml had a TED flux of 0.14 l
g/cm2/h.
22. (a) Grady, M. W.; Bloor, S. 1999, EP 930069; (b) Duncan, M. R.; Frazier, K. S.;
Abramson, S.; Williams, S.; Klapper, H.; Huang, X.; Grotendorst, G. R. FASEB J.
1999, 13, 1774.
23. Allan, G. A.; Gedge, J. I.; Nedderman, A. N. R.; Roffey, S. J.; Small, H. F.; Webster,
R. Xenobiotica 2006, 36, 399.
13. Wipf, P.; Miller, C. P. J. Org. Chem. 1993, 58, 3604.
14. Complete experimental procedures for all assays can be found in Ref. 9.
24. Reid, R. R.; Mogford, J. E.; Butt, R.; deGiorgio-Miller, A.; Mustoe, T. A. Wound
Rep. Reg. 2006, 14, 138.