Design and synthesis of highly selective, orally active Polo-like kinase-2 (Plk-2) inhibitors

Article abstract of DOI:10.1016/j.bmcl.2013.02.065

Polo-like kinase-2 (Plk-2) is a potential therapeutic target for Parkinson's disease and this Letter describes the SAR of a series of dihydropteridinone based Plk-2 inhibitors. By optimizing both the N-8 substituent and the biaryl region of the inhibitors we obtained single digit nanomolar compounds such as 37 with excellent selectivity for Plk-2 over Plk-1. When dosed orally in rats, compound 37 demonstrated a 41-45% reduction of pS129-α-synuclein levels in the cerebral cortex.

Full text of DOI:10.1016/j.bmcl.2013.02.065

Bioorganic & Medicinal Chemistry Letters 23 (2013) 2743–2749
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Design and synthesis of highly selective, orally active Polo-like
kinase-2 (Plk-2) inhibitors
a
a
a
a
a
Simeon Bowers a, , Anh P. Truong , Michael Ye , Danielle L. Aubele , Jennifer M. Sealy , R. Jeffrey Neitz ,
Roy K. Hom a, Wayman Chan a, Michael S. Dappen a, Robert A. Galemmo Jr. a, Andrei W. Konradi a,
Hing L. Sham a, Yong L. Zhu b, Paul Beroza b, George Tonn c, Heather Zhang c, Jennifer Hoffman c,
Ruth Motter c, Donald Fauss c, Pearl Tanaka c, Michael P. Bova c, Zhao Ren c, Danny Tam c, Lany Ruslim c,
Jeanne Baker c, Deepal Pandya c, Linnea Diep c, Kent Fitzgerald c, Dean R. Artis b, John P. Anderson c,
Marcelle Bergeron c
⇑
a Department of Chemical Sciences, Elan Pharmaceuticals, 180 Oyster Point Boulevard, South San Francisco, CA 94080, USA
b Department of Molecular Design, Elan Pharmaceuticals, 180 Oyster Point Boulevard, South San Francisco, CA 94080, USA
c Department of Pharmacological Sciences, Elan Pharmaceuticals, 180 Oyster Point Boulevard, South San Francisco, CA 94080, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Polo-like kinase-2 (Plk-2) is a potential therapeutic target for Parkinson’s disease and this Letter describes
the SAR of a series of dihydropteridinone based Plk-2 inhibitors. By optimizing both the N-8 substituent
and the biaryl region of the inhibitors we obtained single digit nanomolar compounds such as 37 with
excellent selectivity for Plk-2 over Plk-1. When dosed orally in rats, compound 37 demonstrated a 41–
Received 19 November 2012
Revised 6 February 2013
Accepted 13 February 2013
Available online 24 February 2013
45% reduction of pS129-
a-synuclein levels in the cerebral cortex.
Ó 2013 Elsevier Ltd. All rights reserved.
Keywords:
Polo like kinase
PLK inhibitor
Parkinson’s disease
Alpha-synuclein
Parkinson disease (PD) is a devastating neurological disorder
characterized by a progressive loss of dopaminergic neurons with
the subsequent lack of dopamine causing the classical motor
symptoms of bradykinesia, rigidity and resting tremors.1 The major
histological marker of PD is the formation of neuronal intracellular
protein inclusions known as Lewy bodies.2 Lewy bodies are charac-
Plk-1 inhibitors for the treatment of cancer.6 Plk-2 is required for
centriole duplication near the G1 to S phase transition,7 while
Plk-3 directs centrosome localization.8 Plk-4 is the most structur-
ally divergent family member and, similar to Plk-2, also controls
centriole duplication.6 Plk-5 is expressed mainly in differentiated
tissues such as the cerebellum and lacks the kinase domain in
terized by the accumulation of a-synuclein possessing a number of
protein modifications with the major modification being phos-
phorylation at Ser-129.3 Consequently, the kinases responsible
for this phosphorylation have been the subject of intense investiga-
tion. Recent discoveries from both our laboratories4 and others5
have implicated Polo-like kinase 2 (Plk-2) as a major contributor
5
8
5
8
N
N
O
N
N
O
3
3
N
N
N
1
S
N
N
1
N
HN
N
to the formation of pS129 a-synuclein and thus, inhibitors of Plk-
N
2 have the potential to be useful therapeutics for the treatment
1
2
of PD and related Lewy body diseases.
Plk2 IC50 = 0.005 uM
Plk1 IC50 = 0.008 uM
Plk3 IC50 = 94 uM
Plk1/Plk2 = 1.6
OxMet % = m 0, r 0, h 0
Perm. nm/s = 114
P-gp efflux = 3.2
Plk2 IC50 = 0.06 uM
Plk1 IC50 = 0.77 uM
Plk3 IC50 = 12.4 uM
Plk1/Plk2 = 14
OxMet % = m 33, r 38, h 38
Perm. nm/s = 183
Five mammalian Plk family members have been identified (Plk-
1–5)6 with Plk-1 being the most characterized kinase in the group.
Plk-1 plays an essential role during mitosis and is over expressed in
human cancer tissues which has lead to an extensive search for
P-gp efflux = 1.2
Solubility = 100 uM
Solubility = 100 uM
⇑
Corresponding author. Tel.: +1 650 616 2630; fax: +1 650 794 2387.
E-mail address: simeongbowers@gmail.com (S. Bowers).
Figure 1. Typical dihydropteridinone Plk inhibitors.11,12
0960-894X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2013.02.065

2744
S. Bowers et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2743–2749
humans.6b While Plk-1 knockout mouse phenotypes are embryon-
MeO
HN
O
NO2
CO2Me
MeO
H2N
O
N
ically lethal, Plk-2À/À mice are viable6 and therefore, to avoid po-
tential toxicity, Plk-2 inhibitors should possess exquisite
selectivity over Plk-1.
a
b
c
Cl
N
N
R1
R1
3
4
5
We have previously reported our efforts to improve the isoform
selectivity and pharmacokinetic properties of a series of dihydropte-
ridinone based Plk inhibitors which occupy the ATP-binding site of
Plk.9–11 Modification of the biaryl region of compounds such as 1 re-
sulted in compound 2 (Fig. 1), which has good Plk-2 potency (Plk-2
IC50 = 55 nM), good passive permeability (Papp = 183 nm/s), low P-
gp efflux (P-gp efflux ratio = 1.2) and improved microsomal stability
compared to compound 1. Unfortunately, the isoform selectivity of
this compound is less than desirable and we describe in this Letter
the continued optimization of these analogs with an emphasis on
improving the isoform selectivity and metabolic stability.
Metabolite identification studies identified both the substituent
at the N-8 position of the pteridine and the phenyl ring of the biar-
yl system as the major sites of metabolism. It was envisioned that
either the incorporation of fluorine substituents13 or lowering the
lipophilicity of the inhibitor would improve the metabolic stability
of the compounds.14 The first series of analogs that were prepared
were based on the imidazole biaryl scaffold as this series has his-
torically provided excellent Plk isoform selectivity.9 These analogs
were prepared using one of two methods outlined in Scheme 1. In
Method 1, reductive amination of the appropriate aldehyde or ke-
tone with amino ester 3, followed by reaction with 2,4-dichloro-5-
nitropyrimidine gave intermediate 5. One-pot nitro reduction and
H
N
N
O
O
N
O
N
N
N
e
d
N
N
N
R1
Cl
N
N
R1
Cl
N
N
R1
N
R2
6
7
8
Method 2
N
NO2
NO2
CO2CH3
N
CO2CH3
N
O
N
g
f
N
N
N
R1
N
N
N
R1
Cl
N
N
R1
N
N
R2
R2
5
9
10
Scheme 1. Reagents and conditions: (a) aldehyde or ketone, NaBH(OAc)3, NaOAc,
CH2Cl2, rt, 18 h; (b) 2,4-dichloro-5-nitropyrimidine, NaHCO3, hexane, Et2O, 70 °C,
18 h; (c) Fe, AcOH, 90 °C, 2 h; (d) trimethylphosphate, K2CO3, dioxane, 100 °C, 18 h;
(e) 2-(4-R2-phenyl)-1H-imidazol-1-yl, Pd2(dba)3, BINAP, Cs2CO3, 140 °C,
lwave,
1 h; (f) 2-(4-R2-phenyl)-1H-imidazol-1-yl, DMSO, 120 °C, 1 h; (g) (i) Fe, AcOH, 90 °C,
2 h; (ii) trimethylphosphate, K2CO3, dioxane, 100 °C, 18 h.
Table 1
SAR of N-8 aliphatic modifications
N
O
N
N
N
N
N
R1
R2
R1
R2
Plk-2 IC50
(lM)
Plk-1 IC50 (lM)
Plk-1/Plk-2
logP
OxMet %b (m, r, h)
a
a
Compds
11
12
13
iPr
H
H
F
0.025
0.009
0.042
0.87
0.25
3.55
36
30
85
na
12, 0, 14
2, 0, 0
CF3
CF3
3.1
3.1
32, 30, 27
14c
F
F
0.013
0.012
0.32
9.39
25
80
3.1
3.0
15, 8, 6
F
F
15
30, 14, 19
F
F
16
17
18
F
1.04
>50
—
1.6
1.9
1.8
2.0
74, 80, 74
51, 13, 17
27, 0, 10
O
H
H
F
0.008
0.021
0.033
0.38
0.48
7.53
48
23
230
O
O
O
19
63, 67, 58
a
b
c
See Ref. 12.
See Ref. 12. Percent remaining after 30 min incubation with liver microsomes (m = mouse, r = rat and h = human).
Compound is racemic.

S. Bowers et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2743–2749
2745
Method 1
O
cyclization gave compound 6, which was methylated with trimeth-
ylphosphate to give the key intermediate, chloropteridine 7. Inter-
mediate 7 could then be coupled to imidazoles11 under palladium
catalysis to give the analogs shown in Table 1. In some cases, the
final imidazole coupling proceeded in low yield, and the alterna-
tive synthesis, Method 2, was used. Chloropyrimidine 5 was re-
acted with the imidazole and this intermediate, 9, was then
elaborated into the pteridine using similar conditions to those in
Method 1.
NO2
N
R1
23
MeO
HN
O
b
a
N
CO2CH3
R1NH2
21
OMe
Cl
N
Br
R1
20
22
N
O
N
O
N
N
c
d
N
N
N
R1
Cl
N
N
R1
Our efforts to replace the N-8 substituent are outlined in Table 1.
Replacement of the isopropyl N-8 substituent (11) with a 3,3,3-tri-
fluoropropyl group (12) resulted in a threefold improvement in
Plk-2 potency but provided no significant change in the com-
pounds in vitro metabolic stability. Interestingly, the addition of
a fluorine substituent in the biaryl phenyl region of the inhibitor
(13) resulted in an increase in metabolic stability along with an in-
crease in the Plk-2 selectivity over Plk-1. The addition of gem-di-
fluoro substituents to the cyclopentyl (14) and cyclobutyl rings
(15) provided similar microsomal stability compared to 13 but of-
fered no clear advantage. Lowering the logP of the inhibitors by
incorporating heteroatoms into the N-8 substituent may also pro-
vide an increase in the stability of the compounds,14 and a series of
tetrahydrofurans and oxetanes were prepared. Oxetane 16
(logP = 1.6) has excellent metabolic stability but loses Plk-2 activ-
ity compared to the alkyl and fluoroalkyl analogs. Tetrahydrofuran
17 retains the excellent Plk-2 potency of the alkyl analogs and has
a slightly improved metabolic stability, but when combined with
the 4-fluorophenyl imidazole (19) a significant improvement in
metabolic stability was achieved. Consistent with previous results,
the additional fluorine substituent provided a significant increase
in the isoform selectivity of the inhibitor compared to hydrogen
at the R2-position.
N
24
25
R2
Method 2
N
NO2
NO2
CO2CH3
N
CO2CH3
N
O
N
f
e
N
N
N
R1
N
N
N
R1
Cl
N
N
R1
N
N
23
26
27
R2
R2
Method 3
N
O
N
O
N
N
g
N
N
N
N
N
N
N
H
N
N
N
33
28
Scheme 2. Reagents and conditions: (a) K2CO3, KI, acetonitrile, 90 °C, 18 h; (b) 2,4-
dichloro-5-nitropyrimidine, NaHCO3, hexane, Et2O, 70 °C, 18 h; (c) (i) Fe, AcOH,
90 °C, 2 h, (ii) trimethylphosphate, K2CO3, dioxane, 100 °C, 18 h; (d) 2-(4-R2-
phenyl)-1H-imidazol-1-yl, Pd2(dba)3, BINAP, Cs2CO3, 140 °C,
lwave, 1 h; (e) 2-(4-
R2-phenyl)-1H-imidazol-1-yl, DMSO, 120 °C, 1 h; (f) (i) Fe, AcOH, 90 °C, 2 h, (ii)
trimethylphosphate, K2CO3, dioxane, 100 °C, 18 h; (g) 5-bromopyrimidine, Pd2dba3,
Xantphos, NaOtBu, toluene, 110 °C, 18 h.
Metabolite identification studies with compound 19 indicated
that N-dealkylation of the N-8 substituent is a major metabolic
Table 2
SAR of N-8 aryl modifications
N
O
N
N
N
N
N
R1
R2
R1
R2
H
Plk-2 IC50
0.017
(lM)
Plk-1 IC50
2.83
(lM)
Plk-1/Plk-2
164
logP
OxMet %b (m, r, h)
28, 5, 21
a
a
Compds
30c
Ph
3.3
31c
32c
F
F
0.015
0.031
2.34
3.76
156
122
3.4
2.7
41, 11, 47
49, 48, 63
F
CN
33d
34
H
F
0.044
0.012
0.009
7.76
8.67
3.42
175
744
380
1.3
1.8
2.2
60, 76, 55
66, 82, 73
N
N
N
NH
35c
F
67, 65, 72
N
NH
(continued on next page)

2746
S. Bowers et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2743–2749
Table 2 (continued)
a
a
Compds
R1
R2
F
Plk-2 IC50
(lM)
Plk-1 IC50
(lM)
Plk-1/Plk-2
155
logP
OxMet %b (m, r, h)
50, 86, 69
36
0.021
3.29
2.0
N
N
Me
Me
N
37
F
F
0.007
0.532
1.35
28.8
193
54
2.4
1.9
25, 26, 63
72, 80, 74
N
38b
Me
N
N
a
See Ref. 12.
b
See Ref. 12. Percent remaining after 30 min incubation with liver microsomes (m = mouse, r = rat and h = human).
Compound is racemic.
Prepared using Method 3.
c
d
Table 3
SAR of the imidazole replacements
N
O
N
R1
N
N
R2
R1
R2
Plk-2 IC50
0.036
(lM)
Plk-1 IC50
0.25
(lM)
Plk-1/Plk-2
7
OxMet %b (m, r, h)
44, 39, 47
a
a
Compds
S
N
41
N
N
H
O
S
N
42
0.039
0.092
0.130
1.41
3.03
1.82
37
33
14
47, 39, 51
57, 48, 84
na
N
N
N
N
H
O
N
N
S
N
43
N
H
NH
N
S
N
44c
N
H
Me
F
45
0.103
0.168
1.26
1.82
12
11
5, 13, 48
N
N
N
N
N
N
N
H
Me
F
46c
64, 87, 88
N
H
N
Me
a
b
c
See Ref. 12.
See Ref. 12. Percent remaining after 30 min incubation with liver microsomes (m = mouse, r = rat and h = human).
Compound is racemic.
pathway for this series of compounds. In an effort to stabilize these
analogs, the alkyl N-8 substituents were replaced with aromatic
rings. These analogs were prepared using one of three methods
outlined in Scheme 2. Anilines or aminoheterocycles were reacted
with methyl 2-bromobutanoate (20) to give 22 which was elabo-
rated into the pteridines in a similar manner to that described in
Scheme 1. Alternatively, chloropyrimidine 23 was reacted with
an appropriate 2-phenyl imidazole to give nitropyrimidine 26,
which was then converted into pteridine 27 in a two-step proce-
dure. Chiral HPLC separation of the racemic products provided
the analogs describef in Tables 2 and 3. Pyrimidine analog 33
was prepared by palladium catalyzed N-arylation of 28 (Method
3). Unfortunately, this method was not general and rapid access
to a wide variety of heterocycles using this methodology was not
possible.
The initial analog in the series, aniline 30 (Table 2), demon-
strates that aromatic substituents at the N-8 position of the pteri-
dine are tolerated (Plk-2 IC50 = 17 nM) and provide excellent

S. Bowers et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2743–2749
2747
selectivity over Plk-1 (164-fold). The addition of fluorine to both
the N-8 substituent and the phenyl imidazole (31) provided a mod-
est increase in metabolic stability while maintaining excellent Plk-
2 potency (Plk-2 IC50 = 15 nM). In order to increase the microsomal
stability further, previous examples (Table 1) suggested that the
logP of the inhibitor would need to be below 3. The addition of a
nitrile to the N-8 phenyl ring reduced the logP to 2.7 (32) but only
offered a slight increase in the microsomal stability. Replacement
of the phenyl group at the N-8 position with heterocycles should
further reduce the logP of the inhibitors and, in turn, increase their
metabolic stability. Pyrimidine 33 (logP = 1.3), possessed excellent
metabolic stability and good Plk-2 potency (Plk-2 IC50 = 44 nM).
Encouraged by this result with a 6-membered heterocycle, the
SAR of a series of 5-membered heterocycles was explored with
pyrazoles 34–38 (Table 2). Both 4-aminopyrazole (34) and 3-ami-
nopyrazole (35) were among the most potent Plk-2 inhibitors in
this series and both had excellent in vitro microsomal stability.
Furthermore, these compounds had excellent isoform selectivity,
with 34 being 744-fold selective for Plk-2 over Plk-1. N-Methyl
pyrazoles 36 and 37 had similar potency to the NH pyrazoles but
with slightly attenuated isoform selectivity and, as expected,
slightly lower metabolic stability.
ilar to that of the imidazoles, and it appeared that the N-8 substi-
tuent had negligible benefits in this series.
Metabolite identification studies also indicated that the ethyl
group attached to the 7-position of the pteridine core was a meta-
bolic liability, and a small set of analogs was prepared in an at-
tempt to prevent this metabolic pathway (Table 4). These
analogs were prepared in a manner analogous to Scheme 1.
Replacement of the ethyl group with a 2,2,2-trifluoroethyl sub-
stituent (47) had little effect on the metabolic stability of the com-
pound compared to ethyl analog 13 (Table 1). Similarly,
replacement of the hydrogen at the 7-position of the pteridine with
a methyl group (48) had no effect on the metabolic stability of the
inhibitors. Both spirocyclobutyl (49) and gem-dimethyl (50) also
did not provide any advantages in terms of potency or metabolic
stability compared to ethyl at this position, and no further analogs
in this area were prepared.
In order for these compounds to reach their target they must be
able to pass through the blood–brain barrier which requires a high
rate of permeability and a low P-gp efflux. Table 5 contains the
in vitro permeability and P-gp efflux for a select group of com-
pounds that displayed good in vitro metabolic stability and good
Plk-2 potency. For the C-2 imidazole analogs with alkyl N-8 sub-
stituents (13 and 19) the P-gp efflux ratios were low to moderate.
Generally, the aromatic N-8 substituents (33–37) had greater P-gp
efflux compared to the alkyl substituents, but, interestingly, re-
moval of the hydrogen bond donor in pyrazoles 34 and 35 by
methylation (36 and 37) resulted in a significant reduction in the
P-gp efflux, with 37 having excellent permeability and essentially
not being a substrate for P-gp (efflux ratio = 1.1). Previously, we
have reported that the replacement of the imidazole with a pyra-
zole results in lower P-gp efflux ratios,10 and for the alkyl N-8
(41–42) substituents this was the case. Unfortunately, this effect
was not observed with the aryl N-8 analogs, with 46 possessing
As we have reported previously,10 by replacing the imidazole at
the 2-position of the pteridinone with pyrazoles, analogs were ob-
tained with much improved P-gp efflux and permeability (2).10 In
order to determine if these improvements could be combined with
the improved N-8 substituents, which had more favorable in vitro
metabolic properties a series of analogs were prepared. These com-
pounds could be prepared by coupling chloropteridine 24 with an
appropriate boronic acid 3910 (Scheme 3).
As shown in Table 3, the Plk-2 activity was slightly lower in this
series of compounds compared to the imidazole C-2 pteridinones,
and, as observed with 2, the Plk- isoform selectivity was also much
lower. The metabolic stability for the pyrazoles was generally sim-
significant P-gp liability. The ability of these analogs to reduce
a-
synuclein phosphorylation in Plk-2 transfected HEK-293 cells is
shown in Table 5.12 Generally, the cellular IC50 tracks well with
the enzyme IC50, with an approximate 50–80 fold shift in the cel-
lular activity.
Compound 37 demonstrated a favorable combination of good
Plk-2 potency (Plk-2 IC50 = 7 nM), good selectivity over Plk-1
(193-fold), low P-gp efflux, good permeability and moderate
microsomal stability, and was chosen for further evaluation. Com-
pound 37 was screened against 308 kinases and had an exception-
N
O
N
O
B(OH)2
S
N
N
a, b
SEM
N
N
S
N
R1
N
Cl
N
N
R1
N
HN
N
N
24
39
40
Scheme 3. Reagents and conditions: (a) Pd(PPh3)4, Na2CO3, DME/H2O, 100 °C, 18 h;
(b) HCl/dioxane, MeOH, 70 °C.
Table 4
SAR of the 7-position of the pteridine
N
O
N
R2
R3
N
N
N
N
R1
F
a
a
Compds
R1
R2
R3
H
Plk-2 IC50
0.12
(lM)
Plk-1 IC50
3.48
(lM)
Plk-1/Plk-2
28
OxMet %b (m, h)
42, 30, 15
47c
48c
49
CH2CF3
CF3
CF3
Et
Me
0.03
1.15
2.71
7.16
25.3
>100
233
22
—
29, 34, 17
7, 1, 4
25, 10, 54
Isopropyl
Isopropyl
Spirocyclobutyl
Me
Spirocyclobutyl
Me
50
a
b
c
See Ref. 12.
See Ref. 12. Percent remaining after 30 min incubation with liver microsomes (m = mouse, r = rat and h = human).
Compound is racemic.

2748
S. Bowers et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2743–2749
Table 5
In vitro P-gp, permeability and metabolic stability data for select analogs
N
O
N
R1
N
N
R2
a
a
a
Compds
R1
R2
Plk-2 IC50
0.042
(l
M)
293 cell IC50
(lM)
Papp (nm/s)
P-gp effluxa
1.7
CF3
N
N
N
N
13
2.56
158
234
122
87
F
F
19
0.033
0.044
0.012
2.05
3.87
1.87
1.5
4.2
8.6
O
N
N
N
N
N
N
N
N
N
33b
34
N
F
F
NH
36
0.021
0.78
212
2
N
Me
N
N
N
N
35b
37
0.009
0.007
0.74
0.48
132
185
4.9
1.1
N
F
NH
N
N
Me
F
S
N
41
0.036
0.039
0.092
2.99
3.19
5.82
189
168
66
0.8
1.1
8.0
N
N
N
N
N
H
O
S
N
42c
43
N
H
O
N
N
S
N
N
H
NH
N
46c
0.168
—
101
3.4
N
H
N
Me
a
b
c
See Ref. 12.
Compound is racemic.
See Ref. 14.
A
B
0.06
0.04
0.02
0.00
100
80
60
40
20
0
***
***
Veh
100
200
Veh
100
200
Dose of Compound 37 (mg/kg)
Dose of Compound 37 (mg/kg)
Figure 2. A single oral dose of 37 significantly reduces pS129-
a
-synuclein levels in rat cerebral cortex (A) whereas total-
a
-synuclein levels are unchanged (B). The data
represents the mean SD with n = 6 for each group. Significant reduction (⁄⁄⁄p <0.001) versus vehicle control group (Veh) was determined by one way ANOVA with Dunnett’s
post hoc test.

S. Bowers et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2743–2749
2749
Table 6
Plasma and brain levels of 37 at 6 h after a single oral dose
Dose (mg/kg, PO)
Time (h)
Plasma levels mean SD (ng/ml)
Brain levels mean SD (ng/ml)
Brain/plasma (B/P)
100
200
6
6
6286 2594
4881 3239
2252 1051
1382 768
0.36
0.28
ally clean profile, with only two non-Plk- kinases inhibited at 50%
at 10
M compound concentration.12 The ability of 37 to reduce
Plk-2 mediated phosphorylation of -synuclein at Ser-129 in the
brain was evaluated after single oral doses (100 and 200 mg/kg)
in male Sprague–Dawley rats. A 41–45% reduction (p <0.001) of
References and notes
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pS129-a-synuclein levels was measured by ELISA in the cerebral
cortex 6 h after dosing (Fig. 2A). Similar reductions in pS129-
a
-
synuclein levels were also noted in other brain regions including
the olfactory bulb, striatum, ventral midbrain and hippocampus
(not shown). As expected in vehicle-treated rats (5% DMSO/1%
methylcellulose), total
a-synuclein reached 60–70 lg/g with
pS129- -synuclein representing only about 0.1% of the total. The
a
levels of total
a-synuclein were unaffected by treatment with 37
(Fig. 2B).
The plasma and brain exposures of 37 at 6 h after oral dosing in
rats showed good brain-to-plasma ratios (Table 6) in general
agreement with the in vitro PK properties of the compound (Ta-
ble 5). The 6-h brain concentrations of 37 at the 100 and
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200 mg/kg doses were 5.2 and 3.2 l
M,15 respectively, which were
well above the Plk-2 cellular IC50 value (IC50 = 0.485
lM) obtained
from Plk-2-transfected HEK-293 cells.
In conclusion, we have expanded the SAR of our Plk- inhibitors.
By optimizing both the N-8 substituent and the biaryl region of the
inhibitor, we obtained single digit nanomolar compounds with
excellent selectivity for Plk-2 over Plk-1, which, when dosed orally,
11. (a) Neitz, R. J.; Troung, A. P.; Galemmo, R. A.; Ye, X. M.; Sealy, J.; Adler, M.;
Bowers, S.; Beroza, P.; Anderson, J. P.; Aubele, D. L.; Artis, D. R.; Hom, R. K.; Zhu,
Y.-L. WO 2012048129, 2012.; (b) Galemmo, R. A., Jr.; Artis, D. R.; Ye, X. M.;
Aubele, D. L.; Truong, A. P.; Bowers, S.; Hom, R. K.; Zhu, Y.-L.; Neitz, R. J.; Sealy,
J.; Adler, M.; Beroza, P.; Anderson, J. P. WO 2011079118, 2011.; (c) Galemmo, R.
A., Jr.; A., Dean R.; Ye, X. M.; Aubele, D. L.; Truong, A. P.; Bowers, S.; Hom, R. K.;
Zhu, Y.-L.; Neitz, R. J.; Sealy, J.; Adler, M.; Beroza, P.; Anderson, J. P. WO
2011079114, 2011.
12. See Supplementary data.
13. Purser, S.; Moore, P. R.; Swallow, S.; Gouverneur, V. Chem. Soc. Rev. 2008, 37,
320.
14. Thompson, T. N. Med. Res. Rev. 2001, 21, 412.
15. The large variability in the individual data points are most likely due to the
poor solubility of 37 at high doses when dosed as a suspension.
demonstrated a robust reduction in pS129-
affecting total synuclein levels. Compound 37 will be a valuable
tool to help understand the role of -synuclein phosphorylation
a-synuclein without
a
in the pathophysiology of Parkinson-like symptoms.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2013.
02.065.