Investigations of amide bond variation and biaryl modification in analogues of α7 nAChR agonist SEN12333

Article abstract of DOI:10.1016/j.ejmech.2014.07.029

Several lines of experimental evidence support the involvement of the α₇ nAChR in schizophrenia and Alzheimer's disease. Modulators of the α7 nAChR have been extensively reviewed for the treatment of the cognitive deficits associated with these pathologies. SEN12333 represents a novel α7 nAChR agonist chemotype with potential for reduced side effects but requiring further SAR exploration. The present work investigates the amide bond of SEN12333, specifically its connectivity and replacement with the tetrazole functionality, a known cis amide isostere. The results reveal the original amide bond connectivity of SEN12333 to be favorable for binding affinity and agonist activity at α7 nAChRs. The use of a tetrazole isostere completely abolishes affinity and functional activity and suggests that SEN12333 binds in a linear conformation. Results reported herein also suggest the pyridine nitrogen within the terminal aromatic ring of SEN12333 is not essential for binding affinity or functional activity. Further SAR investigations involving manipulation of other moieties contained within SEN12333 are warranted.

Full text of DOI:10.1016/j.ejmech.2014.07.029

European Journal of Medicinal Chemistry 84 (2014) 200e205
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Short communication
Investigations of amide bond variation and biaryl modification in
analogues of 7 nAChR agonist SEN12333
a
Corinne Beinat a, Tristan Reekie a, David Hibbs b, Teresa Xie c, Thao T. Olson c,
Yingxian Xiao c, Andrew Harvey d, Susan O'Connor d, Carolyn Coles d, John Tsanaktsidis e,
, f, g, *
Michael Kassiou a
a School of Chemistry, The University of Sydney, Sydney, NSW 2006, Australia
b School of Pharmacy, The University of Sydney, Sydney, NSW 2006, Australia
c Department of Pharmacology and Physiology, Georgetown University, Washington, DC 20057, USA
d Bionomics Limited, Thebarton, SA 5031, Australia
e CSIRO Materials Science & Engineering, Ian Wark Laboratory, Bayview Avenue, Clayton Victoria 3168, Australia
f Discipline of Medical Radiation Sciences, The University of Sydney, Sydney, NSW 2006, Australia
g Brain and Mind Research Institute, Sydney, NSW 2050, Australia
a r t i c l e i n f o
a b s t r a c t
Article history:
Several lines of experimental evidence support the involvement of the a7 nAChR in schizophrenia and
Received 12 May 2014
Received in revised form
2 July 2014
Accepted 3 July 2014
Available online 9 July 2014
Alzheimer's disease. Modulators of the
the cognitive deficits associated with these pathologies. SEN12333 represents a novel
a
7 nAChR have been extensively reviewed for the treatment of
7 nAChR agonist
a
chemotype with potential for reduced side effects but requiring further SAR exploration. The present
work investigates the amide bond of SEN12333, specifically its connectivity and replacement with the
tetrazole functionality, a known cis amide isostere. The results reveal the original amide bond connec-
tivity of SEN12333 to be favorable for binding affinity and agonist activity at a7 nAChRs. The use of a
Keywords:
tetrazole isostere completely abolishes affinity and functional activity and suggests that SEN12333 binds
in a linear conformation. Results reported herein also suggest the pyridine nitrogen within the terminal
aromatic ring of SEN12333 is not essential for binding affinity or functional activity. Further SAR in-
vestigations involving manipulation of other moieties contained within SEN12333 are warranted.
© 2014 Elsevier Masson SAS. All rights reserved.
a7 nicotinic receptors
Acetylcholine receptor
AChR
CNS
Structureeactivity relationships
Tetrazole
Isostere
1. Introduction
the
a
7 nAChR in schizophrenia and Alzheimer's disease (AD) [5].
7 nAChRs have been extensively reviewed for the
Modulators of
a
Neuronal nicotinic acetylcholine receptors (nAChRs) belong to
the superfamily of ligand-gated cation channels and have recently
generated extensive interest as potential targets of therapeutics in
the treatment of cognitive disorders [1e3]. These receptors exist as
pentameric transmembrane ion channels where subtype is formed
from either a homo- or heteropentameric combination of twelve
treatment of the cognitive deficits associated with these pathol-
ogies [6e8].
Multiple
due to unfavorable side effects and pharmacokinetic issues the
majority of these studies have been discontinued [9]. The known
a7 agonists have entered clinical trials. Unfortunately,
a7
ligands currently and previously in clinical trials display relatively
little structural diversity and are primarily centered on anabaseine,
quinuclidine, or diazabicyclic scaffolds [10]. Consequently, the
majority of them possess the same cross-reactivity profiles with
other sites and similar unwanted side effects. There is a strong
possible subunits: a2ea10 and b2eb4. The homomeric a7 nAChR is
one of the most commonly expressed nicotinic receptors in the
human brain, found in high levels in regions associated with
learning and memory, such as the cerebral cortex and the hippo-
campus [4]. Experimental evidence supports the involvement of
demand for structurally differentiated
a7 nAChR agonists with
more desirable drug-like properties and improved cross-reactivity
profiles for research in this field to further progress.
High-throughput screening by Siena Biotech and Wyeth has
identified piperazine 1 (Fig. 1) as a novel chemotype of weak,
* Corresponding author. School of Chemistry, The University of Sydney, Sydney,
NSW 2006, Australia.
partial agonist activity at a7 nAChRs. Preliminary SAR investigation
E-mail address: michael.kassiou@sydney.edu.au (M. Kassiou).
http://dx.doi.org/10.1016/j.ejmech.2014.07.029
0223-5234/© 2014 Elsevier Masson SAS. All rights reserved.

C. Beinat et al. / European Journal of Medicinal Chemistry 84 (2014) 200e205
201
Fig. 1. Agonists at a7 nAChRs identified by Siena Biotech and Wyeth.
Fig. 2. The incorporation of the urea functionality with pharmacophoric elements: basic center (blue), carbon chain (black), central linker (green) and biaryl system (red). (For
interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)
Fig. 3. Amido-varied SEN12333 analogues.
of the piperazine, biaryl and amide regions of compound 1 resulted
in the development of SEN12333 (2) [11]. SEN12333 is an 7 nAChR
enhanced potency at
selectivity, particularly against receptors containing
a
7 nAChRs, they also demonstrated low
a
a
3 subunit,
agonist, with selectivity over other nAChR subtypes, 5-HT3 re-
ceptors, and hERG channels [11,12]. Additionally, it displayed
favorable bioavailability and brain permeation in vivo [12]. Initial
investigations of SEN12333 (2) in animal models of episodic
memory confirmed its ability to reverse both scopolamine- and
MK-801-induced amnesia [11,12].
unfavorable hERG activity and substantial CYP450 inhibition [14].
The present investigations were aimed at exploring the con-
nectivity of the amide bond. The transposition of amide to reverse-
amide is a concept well-known to medicinal chemistry [15e17].
Additionally, the effect of the pyridine nitrogen within the terminal
aromatic ring of SEN12333 (2) was simultaneously investigated.
The lack of a clear rationale why the pyridine nitrogen was moved
from the 2-position in original hit 1 to the 3-position in SEN12333
(2), prompted us to question if it was required at all. Accordingly,
compounds 3e5 were designed to address this question (Fig. 3).
Aside from investigating the amide connectivity, the amide
linker was also replaced with a tetrazole group. The selection of a
tetrazole linker is based upon the knowledge that tetrazoles are cis-
amide isosteres [18e21]. It was envisioned that this would provide
us further mechanistic detail into the binding mode of SEN12333 (2)
Limited SAR data is reported for this unique class of a7 nAChR
ligands, with brief studies conducted into modification of the
morpholine ring and variation of the arylanilide groups [11]. Pre-
vious work within our laboratory has identified the 4-carbon chain
linker of compound 2 to be optimal for binding and functional
activity at a7 nAChRs [13]. Minimal SAR investigation is available
for the amide bond of SEN12333 (2). Haydaar and co-workers have
described replacement of the amide moiety with the urea func-
tionality (Fig. 2) [14]. Though these compounds generally exhibited
Scheme 1. Reagents and conditions: (a) (COCl)2, rt, 45 min; (b) 4-bromoaniline, Et3N, CH2Cl2, e78 ꢀC to rt, 4 h, 91% yield over 2 steps; (c) morpholine, NaI, Et3N, DMF, reflux, 16 h,
85%; (d) Pd(PPh3)4, appropriate boronic acid, MeCN/sat. aq. Na2CO3 (1:1), reflux, 18 h, 68 or 83%.

202
C. Beinat et al. / European Journal of Medicinal Chemistry 84 (2014) 200e205
Scheme 2. Reagents and conditions: (a) Morpholine, NaI, Et3N, THF, reflux, 16 h, 66%; (b) NH2NH2$H2O, EtOH, reflux, 1 h; (c) 4-bromobenzoyl chloride, Et3N, CH2CH2, e78 ꢀC to rt,
2 h, 64% over two steps; (d) Pd(PPh3)4, appropriate boronic acid, MeCN/sat. aq. Na2CO3 (1:1), reflux, 18 h, 70 or 91%.
Scheme 3. Reagents and conditions. (a) (i) PCl5, CH2Cl2, rt, 2 h; (ii) TMSN3, 0 ꢀC, 16 h, 40e71%.
at a7 nAChRs and potentially reduce the number of rotatable bonds
within the molecule. Tetrazoles are resistant to certain metabolic
pathways, and their incorporation can often result in increased
bioavailability of a drug [18]. There are two regioisomeric tetrazoles
to be studied. One series would be accessed from SEN12333 (2) and
analogues containing this original amide bond connectivity (3);
whereas the other tetrazole series could be accessed from the
related ‘reverse-amide’ bond analogues (4 and 5).
acid gave the desired retro-amide ligands 4 and 5 (70 or 91%)
respectively.
The corresponding tetrazole analogues 13e16 were readily
synthesized from amides 2e5, a representative example is shown
in Scheme 3. The amides 2e5 were treated with phosphorus pen-
tachloride to form the presumed corresponding imidoyl chloride.
However this was not isolated as the addition of trimethylsilylazide
furnished the desired tetrazoles 13e16 following basic workup
(40e71%).
Single crystals of SEN12333 (2) and tetrazole 13 were grown by
slow recrystallization in CH2Cl2 and n-hexane and subjected to X-
Ray crystal analysis. In addition to structure confirmation, it was
thought that by identifying their solid-state conformations it would
help to further predict the mode of binding of the amide and tet-
razole analogues. The Oak Ridge Thermal Ellipsoid Plots (ORTEP) of
SEN12333 (2) and tetrazole 13 are shown in Figs. 4 and 5 respec-
tively. SEN12333 (2, Fig. 4) has a large degree of conformational
flexibility and it is immediately apparent that its lowest energy
configuration state adopts a trans amide conformation to linearize
the structure and minimize steric interactions. In contrast, the
tetrazole ring present within compound 13 locks the amide bond
2. Chemistry
The syntheses of analogues 2 and 3 are depicted in Scheme 1
and involved the initial conversion of 5-bromovaleric acid (6) to
the corresponding acid chloride. Subsequent treatment with 4-
bromoaniline in the presence of triethylamine gave desired ani-
lide 7 (91%). Alkylation of morpholine with bromide 7 in the
presence of catalytic iodide afford compound 8 in 85% yield. Sub-
jecting bromoarene 8 to Suzuki cross-coupling with either 3-
pyridylboronic acid or phenylboronic acid gave the desired li-
gands SEN12333 (2) and 3 (83 or 68%) respectively. SEN12333 (2)
was synthesized for direct comparison with the new analogues.
An alternative synthesis was proposed for reverse-amide ana-
logues 4 and 5 (Scheme 2). Alkylation of morpholine with bromide
9 gave amine 10 (66%), subsequent hydrazinolysis and amide bond
formation yielded compound 12 (64% over two steps). Final Suzuki
cross-coupling with either 3-pyridylboronic acid or phenylboronic
Fig. 4. ORTEP representation of SEN12333 (2).
Fig. 5. ORTREP representation of tetrazole 13.

C. Beinat et al. / European Journal of Medicinal Chemistry 84 (2014) 200e205
203
Table 1
Binding affinities of newly synthesized compounds.
Compound
Ki (nM)a
Physicochemical properties
a
7
a
4
b2
a
3
b4
CLogPb
PSAb
LLEc
670 119
>10,000
>10,000
2.25
53.93
3.92
2300 1000
>10,000
>10,000
>10,000
>10,000
3.58
1.93
41.57
53.93
2.06
3.02
11,000 4000
1800 200
>10,000
>10,000
3.28
2.08
41.57
65.15
2.46
1.59
216,000
DNC
DNC
236,000
165,000
DNC
DNC
>1,000,000
>1,000,000
3.47
2.81
52.79
65.15
0.16
0.97
156,000
DNC
>100,000
4.31
52.79
ꢁ0.5
a
Competition curves were constructed in which 10 concentrations of a sample were included. DNC ¼ the fitting does not converge due to extremely low affinities. Ki values
are the Mean SEM of three to six independent measurements. For low binding affinity compounds, estimated Ki values or Ki value ranges were provided.
b
Calculated using ChemDraw12.0.
Calculated according to equation LLE ¼ pIC50 (or pKi) ꢁ cLogP.
c
into a cis conformation and as illustrated in Fig. 5. It is evident that
the compound 13 resides in a bent form. If the binding conforma-
tion of SEN12333 (2) is similar to the low energy, trans amide
conformation depicted in Fig. 4, these studies suggest the mode of
binding of SEN12333 (2) and tetrazole 13 would be vastly different.
It was envisioned that this would additionally provide greater
insight into the binding mode of SEN12333 (2), specifically, if the
amide bond was residing in either a cis or trans conformation. It was
thought that reverse-amide 4 would have a similar conformation to
SEN12333 (2) while the tetrazole 15 would mimic analogue 13,
however, X-Ray crystal analysis was only obtained for a represen-
tative example of each class of compounds.
determine their affinity and selectivity for the
sults are summarized in Table 1. All compounds were selective for
the 7 nAChR subtype; however, a large range of affinities was
evident. Removal of the pyridine nitrogen from reference
SEN12333 (2) to give biphenyl analogue 3 resulted in a decrease in
affinity. Reversal of the amide bond order of SEN12333 (2) to give
compound 4 led to a marked decrease in affinity, however, this
decrease in affinity was restored partially in retro-amide biphenyl
analogue 5. These results suggest the original amide bond order of
SEN12333 (2) to be favorable and that the pyridine nitrogen is not
a7 nAChR. The re-
a
essential for binding or selectivity at
compounds to their corresponding tetrazole resulted in a loss of
affinity at 7 receptors with Ki values over 150 M for all tetrazole
a7 receptors. Conversion of
a
m
3. Pharmacology
analogues. Considering the high concentrations required for the
tetrazole series, a clear determination of the effect of the pyridine
or biphenyl compounds in this series was not possible. In contrast
to compounds 2e5, tetrazoles synthesized from the original amide
All newly synthesized compounds and SEN12333 (2) as a
reference were evaluated in competitive binding assays to

204
C. Beinat et al. / European Journal of Medicinal Chemistry 84 (2014) 200e205
important guide in the process of lead optimization, and also as a
flag for selection of preclinical candidates [30]. A typical oral drug
with cLogP of 2.5 and potency within the range of 1e10 nM sug-
gests a target LLE of 5e7 or greater, a value >5 is considered highly
optimized for drug discovery. The advantages of high in vivo po-
tency are low total doses in humans and decreased adventitious
compound related toxicity [31].
All compounds display acceptable PSA values and cLogP < 5.
Whilst biphenyl compound 5 displays a binding affinity similar to
that of SEN12333 (2) (Ki values as 670 vs 1800 nM, respectively), its
decreased LLE value of 2.46 suggests that it may not be as successful
in vivo. While the cLogP and PSA values of compounds 4 and 5 are
acceptable, their LLE values (3.02 and 2.46 respectively) also suggest
their limited potential in vivo. Due to the extremely low binding
affinities of tetrazole containing analogues 13e16, they all display
unacceptable LLE values, further confirming the limited use of tet-
razoles as an amide isostere in SEN12333 (2) and related analogues.
Fig. 6. Dose response curve of SEN12333 (2) on rat a7 GH4C1 cells; EC50 ¼ 0.86
mM.
To determine theiragonistic activityat
all amide-varied and tetrazole analogues were evaluated in a fluo-
rescent calcium assay using a GH4C1 cell line expressing rat 7 re-
ceptors. Compounds were tested at concentrations of 0.3, 1, 3 and
10
a7 nAChRs, SEN12333 and
bond order exhibited decreased affinity compared to tetrazoles
synthesized from analogues containing the reversed amide bond
a
order (Ki values 13 and 14 approximately 220
mM vs. those of 15 and
m
M (n ¼ 3). Fluo4 Direct Dye from Molecular Probes was used and
16 approximately 130 M). These findings illustrate that the tet-
m
the assay was performed on a Perkin Elmer CellLux high throughput
fluorescent reader. Peak/Base ratios were calculated to show activity
where a peak/base values >1.00 was indicative of agonist activity.
Unfortunately, aside from SEN12333 (2), only compounds 3 and
razole motif is not tolerated as an amide isostere in this series of
compounds. This could be attributed to one or more of a combi-
nation of effects including the electronics of the tetrazole ring,
increased steric bulk, or the cis conformation of the tetrazole iso-
stere as illustrated in Fig. 5.
5 showed agonism at a7 nAChRs (Figs. 6 and 7). This was to be
expected given the concentration range tested and the Ki values
from the binding studies (Table 1). The EC50 of SEN12333 (2) was
Table 1 also describes the physicochemical properties cLogP,
polar surface area (PSA) and ligand lipophilicity efficiency (LLE) for
all newly synthesized compounds. It has been recognized for
numerous years that the success of a lead in drug development is
largely affected by its lipophilicity and the influence of LogP on drug
potency, pharmacokinetics and toxicity [22]. Lipophilic bases can
potentially trigger cardiovascular toxicological effects through
interaction with hERG channels and tissue toxicity through pro-
motion of cellular phospholipidosis [23]. In terms of absorption and
permeability, the ‘rule of five’ guidelines define the optimum for
cLogP to be less than 5. In line with this, the average clogP value of
marketed CNS drugs is 3.43 [24e27]. PSA is commonly utilized in
drug design in the optimization of a drug's capability to permeate
cell membranes such as the BBB [28]. PSA of approximately 120 Å2
is deemed necessary to permeate cell membranes and 40e70 Å2 is
required to cross the BBB [28,29]. The development of a novel index
known as ligand lipophilicity efficiency (LLE) has served as an
observed to be 0.86
activity at 7 receptors with an EC50 of 1.7
biphenyl analogue 5 similarly maintained agonistic activity with
an EC50 of 2.0 M. The lack of activity of the tetrazole containing
compounds 13e16 highlights incorporation of this moiety is also
m
M. The biphenyl analogue 3, retained agonistic
a
mM. Retro-amide
m
detrimental for agonistic activity at a7 nAChRs.
4. Conclusions
In summary, seven new analogues of SEN12333 (2) have been
synthesized to examine the effects of amide bond orientation, the
use of a tetrazole as an amide bond isostere and the importance of
the pyridine nitrogen within the biaryl system. The original amide
bond orientation of SEN12333 (2) was found to be favored; reversal
of the amide bond resulted in a significant loss of binding and
agonistic activity at a7 nAChRs (2 vs 4). Biphenyl analogues of the
reverse amide series were found to be more potent than their
corresponding phenyl-pyridine analogues (5 vs 4) and importantly
maintained agonistic activity at
a7 nAChRs. Furthermore, the
biphenyl motif of compound 5 was found to restore the potency lost
in the reversed amide pyridine compound 4. The use of a tetrazole
functionality as an amide bond isostere resulted in a dramatic loss
of potency and a complete loss of agonist activity for all types of
analogues. The most potent compound of the series was still
SEN12333 (2, Ki ¼ 670 nM) where this amide bond orientation
appears to be optimal for binding and activity at a7 nAChRs.
Acknowledgments
CB acknowledges receipt of an Australian Postgraduate Award
and a John A. Lamberton research scholarship.
Appendix A. Supplementary data
Supplementary data related to this article can be found at http://
dx.doi.org/10.1016/j.ejmech.2014.07.029.
Fig. 7. Dose response curve of compound
EC50 ¼ 1.7 M, 5: EC50 ¼ 2 M.
3 and 5 on rat a7 GH4C1 cells; 3:
m
m

C. Beinat et al. / European Journal of Medicinal Chemistry 84 (2014) 200e205
205
[15] A. Reichelt, C. Gaul, R.R. Frey, A. Kennedy, S.F. Martin, J. Org. Chem. 67 (2002)
4062e4075.
References
[16] B. Hu, J. Ellingboe, S. Han, E. Largis, R. Mulvey, A. Oliphant, F.-W. Sum, J. Tillett,
J. Med. Chem. 44 (2001) 1456e1466.
[17] S.L. Roderick, M.C. Fournie-Zaluski, B.P. Roques, B.W. Matthews, Biochemistry
28 (1989) 1493e1497.
[18] R.J. Herr, Bioorg. Med. Chem. 10 (2002) 3379e3393.
[19] K.L. Yu, R.L. Johnson, J. Org. Chem. 52 (1987) 2051e2059.
[20] J. Zabrocki, J.B. Dunbar, K.W. Marshall, M.V. Toth, G.R. Marshall, J. Org. Chem.
57 (1992) 202e209.
[21] J. Zabrocki, G.D. Smith, J.B. Dunbar, H. Iijima, G.R. Marshall, J. Am. Chem. Soc.
110 (1988) 5875e5880.
[22] H. van de Waterbeemd, D.A. Smith, K. Beaumont, D.K. Walker, J. Med. Chem.
44 (2001) 1313e1333.
[23] M.J. Waring, C. Johnstone, Bioorg. Med. Chem. Lett. 17 (2007) 1759e1764.
[24] C.A. Lipinski, F. Lombardo, B.W. Dominy, P.J. Feeney, Adv. Drug Deliv. Rev. 23
(1997) 3e25.
[25] C.A. Lipinski, Drug Discov. Today Technol. 1 (2004) 337e341.
[26] T. Fichert, M. Yazdanian, J.R. Proudfoot, Bioorg. Med. Chem. Lett. 13 (2003)
719e722.
[27] H.L. Pajouhesh, G.R. Lenz, NeuroRx 2 (2005) 541e553.
[28] J.J. Lu, K. Crimin, J.T. Goodwin, P. Crivori, C. Orrenius, L. Xing, P.J. Tandler,
T.J. Vidmar, B.M. Amore, A.G.E. Wilson, P.F.W. Stouten, P.S. Burton, J. Med.
Chem. 47 (2004) 6104e6107.
[1] J.A. Dani, D. Bertrand, Annu. Rev. Pharmacol. Toxicol. 47 (2007) 699e729.
[2] L.M. Broad, E. Sher, P.C. Astles, R. Zwart, M.J. O'Neill, Drug Future 32 (2007)
161e170.
[3] D. D'Hoedt, D. Bertrand, Expert Opin. Ther. Targets 13 (2009) 395e411.
[4] P. Seguela, J. Wadiche, K. Dineley-Miller, J.A. Dani, J.W. Patrick, J. Neurosci. 13
(1993) 596e604.
[5] A. Taly, P.-J. Corringer, D. Guedin, P. Lestage, J.-P. Changeux, Nat. Rev. Drug
Discov. 8 (2009) 733e750.
[6] P. Lippiello, M. Bencherif, T. Hauser, K. Jordan, S. Letchworth, A. Mazurov,
Expert Opin. Drug Discov. 2 (2007) 1185e1203.
[7] R. Freedman, C.E. Adams, S. Leonard, J. Chem. Neuroanat. 20 (2000) 299e306.
[8] W.R. Kem, Behav. Brain Res. 113 (2000) 169e181.
[9] J. Toyohara, K. Hashimoto, Open Med. Chem. J. 4 (2010) 37e56.
[10] A. Mazurov, T. Hauser, C.H. Miller, Curr. Med. Chem. 13 (2006) 1567e1584.
[11] S.N. Haydar, C. Ghiron, L. Bettinetti, H. Bothmann, T.A. Comery, J. Dunlop, S. La
Rosa, I. Micco, M. Pollastrini, J. Quinn, R. Roncarati, C. Scali, M. Valacchi,
M. Varrone, R. Zanaletti, Bioorg. Med. Chem. 17 (2009) 5247e5258.
[12] R. Roncarati, C. Scali, T.A. Comery, S.M. Grauer, S. Aschmi, H. Bothmann, B. Jow,
D. Kowal, M. Gianfriddo, C. Kelley, U. Zanelli, C. Ghiron, S. Haydar, J. Dunlop,
G.C. Terstappen, J. Pharmacol. Exp. Ther. 329 (2009) 459e468.
[13] C. Beinat, S.D. Banister, P.S. van, M.R. Doddareddy, D. Hibbs, M. Sako,
M. Chebib, T. Tran, N. Al-Muhtasib, Y. Xiao, M. Kassiou, Bioorg. Med. Chem.
Lett. 22 (2012) 2380e2384.
[14] C. Ghiron, S.N. Haydar, S. Aschmies, H. Bothmann, C. Castaldo, G. Cocconcelli,
T.A. Comery, L. Di, J. Dunlop, T. Lock, A. Kramer, D. Kowal, F. Jow, S. Grauer,
B. Harrison, S. La Rosa, L. Maccari, K.L. Marquis, I. Micco, A. Nencini, J. Quinn,
A.J. Robichaud, R. Roncarati, C. Scali, G.C. Terstappen, E. Turlizzi, M. Valacchi,
M. Varrone, R. Zanaletti, U. Zanelli, J. Med. Chem. 53 (2010) 4379e4389.
[29] J. Kelder, P.J. Grootenhuis, D. Bayada, L.C. Delbressine, J.-P. Ploemen, Pharm.
Res. 16 (1999) 1514e1519.
[30] A.R. Leach, M.M. Hann, J.N. Burrows, E.J. Griffen, Mol. Biosyst.
2 (2006)
429e446.
[31] P.D. Leeson, B. Springthorpe, Nat. Rev. Drug Discov. 6 (2007) 881e890.