Discovery of novel 5,6,7,8-tetrahydro[1,2,4]triazolo[4,3-a]pyridine derivatives as γ-secretase modulators

Article abstract of DOI:10.1016/j.bmcl.2015.07.101

Novel 5,6,7,8-tetrahydro[1,2,4]triazolo[4,3-a]pyridine derivatives were designed, synthesized, and evaluated as γ-secretase modulators (GSMs). An optimization study of this series resulted in the identification of (R)-11j, which showed a potent Aβ₄₂-lowering effect, high bioavailability and good blood-brain barrier permeability in mice. Oral administration of (R)-11j significantly reduced brain Aβ₄₂ in mice at a dose of 10 mg/kg.

Full text of DOI:10.1016/j.bmcl.2015.07.101

Bioorganic & Medicinal Chemistry Letters 25 (2015) 4245–4249
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Discovery of novel 5,6,7,8-tetrahydro[1,2,4]triazolo[4,3-a]pyridine
derivatives as c-secretase modulators
⇑
Takafumi Takai , Yasutaka Hoashi, Yoshihide Tomata, Sachie Morimoto, Minoru Nakamura,
Tomomichi Watanabe, Tomoko Igari, Tatsuki Koike
Pharmaceutical Research Division, Takeda Pharmaceutical Company Ltd, 26-1, Muraokahigashi 2-chome, Fujisawa, Kanagawa 251-8555, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Novel 5,6,7,8-tetrahydro[1,2,4]triazolo[4,3-a]pyridine derivatives were designed, synthesized, and evalu-
Received 13 May 2015
Revised 27 July 2015
Accepted 30 July 2015
Available online 7 August 2015
ated as c-secretase modulators (GSMs). An optimization study of this series resulted in the identification
of (R)-11j, which showed a potent Ab42-lowering effect, high bioavailability and good blood–brain barrier
permeability in mice. Oral administration of (R)-11j significantly reduced brain Ab42 in mice at a dose of
10 mg/kg.
Ó 2015 Elsevier Ltd. All rights reserved.
Keywords:
c
-Secretase modulators
Alzheimer’s disease
Amyloid beta
Alzheimer’s disease (AD) is an age-associated neurodegenera-
tive disorder characterized by impaired memory and cognition.1
A characteristic pathology of AD is the formation of plaques with
the accumulation of amyloid beta (Ab), which is produced by step-
wise processing of the amyloid precursor protein (APP) by b-secre-
structure–activity relationship (SAR) of novel 5,6,7,8-tetrahydro
[1,2,4]triazolo[4,3-a]pyridines.
Monocyclic 1,2,4-triazole derivative
3 was synthesized as
shown in Scheme 1. Bromide 126 was converted to nitrile 13,
which was coupled with hydrazide 14 and subsequently cyclized
to afford 3.
tase and
c c-secretase also cleaves other
-secretase.2 Because
substrates in addition to APP, such as Notch,3 it was reported that
The synthetic route for the preparation of the tetrahydro[1,2,4]-
triazolo[1,5-a]pyridine derivative 4 is illustrated in Scheme 2.
Hydrolysis of the cyano group of 13 yielded benzoic acid 15.
Coupling 15 with t-butyl carbazate and subsequent N-Boc depro-
tection afforded hydrazide 16a as an HCl salt form. Hydrazide
16b was also directly obtained as a free form by condensation of
15 with hydrazine monohydrate. Imidate 19 was prepared by alky-
lation of phenylacetonitrile 17, followed by ethanolysis of 18 under
acidic conditions. Finally, ring formation using 16a and 19 in the
presence of imidazole gave the tetrahydro[1,2,4]triazolo[1,5-a]pyr-
idine derivative 4.
Tetrahydro[1,2,4]triazolo[4,3-a]pyridine and related bicyclic
analogs 5 and 7–10 were synthesized as shown in Scheme 3.
Alkylation of the benzyl position of phenylacetic acid 20 with
1-bromoalkanes 21a–c afforded 22a–c. Benzohydrazide 16a or
16b was acylated with 22a–d to give diacyl compounds 23a–d,
which were cyclized by treatment with CCl4 and PPh3 to yield oxa-
diazoles 24a–d. Alkylchloride 24a–d were subjected to nucle-
ophilic azidation, followed by reduction of the azide group to
give the primary amines 25a–d. Intracyclization of 25a–d pro-
ceeded smoothly under acidic conditions to provide the annulate
derivatives 5, 7, 8, and 10. Synthesis of dihydro[1,2,4]triazolo
c
c
-secretase inhibitor induced several toxicities in clinical trials.4
-Secretase modulators (GSMs) lower pathogenic Ab by cleavage
shift without affecting Notch signal and have been proposed as a
potential therapeutic agent for AD.5 We recently reported the
potent piperazine derivative 1 as a GSM, which selectively reduced
brain Ab42 levels in normal mice (Fig. 1).6 In the study, the oxa-
zolylphenyl moiety in 1 was validated as a suitable surrogate for
the imidazolylphenyl moiety,
a highly conserved component
across GSMs from several research laboratories.5 To identify a
novel class of GSM, the oxazolylphenyl moiety was introduced into
various right side fragments in reported GSMs, such as the 3-ben-
zyl-1,2,4-triazole in compound 2.7 As a result, we observed that the
1,2,4-triazole derivative 3 showed moderate Ab42-lowering activity
(Fig. 1). A recent analysis of GSMs indicated that their increased
lipophilicity enhanced their potency but reduced their drug-like-
ness.8 Here we describe lead generation from 3 using ligand-
lipophilicity efficiency (LLE) as a drug-likeness guideline, and the
⇑
Corresponding author. Tel.: +81 466 32 1133; fax: +81 466 29 4454.
E-mail address: takafumi.takai@takeda.com (T. Takai).
http://dx.doi.org/10.1016/j.bmcl.2015.07.101
0960-894X/Ó 2015 Elsevier Ltd. All rights reserved.

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T. Takai et al. / Bioorg. Med. Chem. Lett. 25 (2015) 4245–4249
Figure 1. Piperazine derivative 1, and 1,2,4-triazole derivatives 2 and 3.
Scheme 1. Reagents and conditions: (a) Zn(CN)2, Pd2(dba)3, DavePhos, DMF, 130 °C, 88%; (b) K2CO3, n-BuOH, 120 °C, 12%.
Scheme 2. Reagents and conditions: (a) n-BuOH, 8 M NaOH aq., reflux, 74%; (b) For 16a, (i) t-butyl carbazate, DEPC, Et3N, DMF, rt, 73%; (ii) 4 M HCl/EtOAc, EtOAc, rt, 96%; For
16b, hydrazine monohydrate, CDI, THF, 92%; (c) 1-bromo-3-chloropropane, t-BuOK, THF, À10 °C to rt, 35%; (d) AcCl, EtOH, 0 °C to rt, 76%; (e) 16a, imidazole, MeOH, rt to 65 °C,
46%.
Scheme 3. Reagents and conditions: (a) 1.6 M n-BuLi/hexane, THF; (b) 16a, HATU, Et3N, DMF, or 16b, DEPC, Et3N, DMF; (c) CCl4, PPh3, MeCN or Burgess reagent, THF;
(d) (i) NaN3, DMSO; (ii) PPh3, H2O, THF; (e) AcOH, reflux; (f) NaH, DMF, air, rt, 39%; (g) TsOHÁH2O, toluene, reflux, 94%.

T. Takai et al. / Bioorg. Med. Chem. Lett. 25 (2015) 4245–4249
4247
[4,3-a]pyridine 9 was achieved by oxidation of the benzyl position
of 5 under basic condition and subsequent dehydration of 26 with
p-toluene sulfonic acid.
replacing the 1,2,4-triazole ring of 5 with the 1,2,3-triazole ring of
6 retained the potency, the increased lipophilicity of 6 resulted in a
decrease in the LLE value. Expanding the tetrahydropyridine ring
gave the triazoloazepine analog 7 with a negative effect on LLE,
and ring contraction to a five-membered ring (8) also reduced
potency and the LLE value. Introduction of an olefin moiety (9) into
the tetrahydropyridine ring of 5 was unfavorable for the activity.
These results suggest that the ring size of the bicyclic system and
the direction of the distal phenyl group affect GSM activity and
that the conformation of 5 is the most desirable for activity in this
series. Triazolooxazine analog 10, which is expected to have a con-
formation similar to that of 5, showed decreased potency and LLE,
indicating that the polarity of the oxygen atom of the triazoloox-
azine core in 10 exerted an adverse effect on the activity. We
accordingly selected the tetrahydro[1,2,4]triazolo[4,3-a]pyridine
ring as a core scaffold.
We next investigated the SAR of the distal phenyl moiety with
monosubstituted analogs 11a–g and disubstituted analogs 11h–j
(Table 2). Removal of the chlorine atom at the 3-position in the
3,4-dichlorobenzene of 4 resulted in a slight decrease of potency,
but the LLE of 11a was retained by this modification. Compound
11b containing a 4-trifluoromethyl group showed potency and
LLE similar to those of 11a. The less lipophilic 4-fluoro analog
11c decreased the potency while retaining the same LLE as that
of 11a. This tendency was observed when polar groups, such as a
methoxy group (11d) or morpholino group (11e), were introduced
at the 4-position of the phenyl group. The regioisomeric trifluo-
romethyl analogs 11f (3-CF3) and 11g (2-CF3) showed the potency
similar to that of 11b. A strong correlation between potency and
logD value was observed among monosubstituted analogs (Fig. 2,
indicated by black circles). This result suggests that GSM activity
is dominated by lipophilicity in the tetrahydro[1,2,4]triazolo
[4,3-a]pyridine series, and that various substituents on the distal
phenyl ring would be tolerable for LLE. We accordingly prepared
Tetrahydro[1,2,4]triazolo[4,3-a]pyridines 11a–d, 30, and 11f–j
were synthesized by a manner similar to that described for 5 as
shown in Scheme 4. Carboxylic acids 28e and 28h were obtained
from the corresponding esters 31e and 31h by alkylation and
hydrolysis. In the case of preparation of 28g, direct alkylation of
the benzyl position of phenylacetic acid 27g did not proceed well.
The desired carboxylic acid 28g was smoothly obtained by alkyla-
tion of the protected compound 32 and the following deprotection.
A palladium-catalyzed cross coupling reaction of morpholine with
bromide 30 afforded 4-morpholine derivative 11e.
The synthetic route to tetrahydro[1,2,3]triazolo[1,5-a]pyridine
6 is illustrated in Scheme 5. The Sonogashira coupling of the
bromide 12 with 5-hexyn-1-ol, followed by oxidation of primary
alcohol 33 yielded the aldehyde 34. A coupling reaction of 34 with
3,4-dichlorophenylmagnesium bromide gave the secondary alco-
hol 35, which was treated with mesylchloride to yield 36. The
4,5,6,7-tetrahydro[1,2,3]triazolo[1,5-a]pyridine ring of 6 was con-
structed by nucleophilic substitution of 36 with sodium azide
and a subsequent intramolecular [3+2] cycloaddition in one pot.
Triazole derivatives 3–10 were evaluated for inhibitory activity
against Ab42 production in rat primary neuronal cells, lipophilicity
(logD at pH 7.4),9 and LLE (=pIC50 À logD) as shown in Table 1. We
initially introduced an additional ring between the benzyl position
and the triazole ring of 3 to identify the active conformation.
Enhancement of potency was observed with both the bicyclic
derivatives 4 and 5. The LLE value of 4 was equal to that of the
monocyclic triazole 3. This suggests that the increase of lipophilic-
ity of 4 from 3 may simply result in the enhancement of potency.
However, a marked improvement in LLE was achieved in 5.
Conformational constraint in 5 could effectively fix the active con-
formation of 3 and contribute to the increased LLE value. Although
Scheme 4. Reagents and conditions: (a) n-BuLi, 1-bromo-3-chloropropane; (b) (i) 16a or 16b, HATU or DEPC; (ii) CCl4 or CCl3CN, PPh3, (c) (i) NaN3; (ii) PPh3, H2O; (iii) AcOH;
(d) (i) NaH, 1-bromo-3-chloropropane, DMF; (ii) NaOH, H2O, MeOH, THF; (e) 1,1-di-tert-butoxytrimethylamine, toluene, 80 °C, 81%; (f) (i) NaH, 1-chloro-3-iodopropane, DMF,
rt, 91%; (ii) TFA, rt, 99%; (g) morpholine, DavePhos, Pd2(dba)3, t-BuONa, toluene, 100 °C, 20%.

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T. Takai et al. / Bioorg. Med. Chem. Lett. 25 (2015) 4245–4249
Scheme 5. Reagents and conditions: (a) 5-hexyn-1-ol, PdCl2(PPh3)2, CuI, Et3N, 70 °C, 83%; (b) Dess–Martin periodinane, MeCN, DMSO, rt, 83%; (c) 3,4-dichlorophenylmag-
nesium bromide, THF, À78 °C, 41%; (d), MsCl, Et3N, THF, 0 °C, 97%; (e) sodium azide, DMSO, 110 °C, 48%.
Table 1
circles), and the most lipophilic 4-chloro-2-trifluoromethyl analog
Ab42-lowering effects of 3–10, logD values and LLE
11j showed the most potent Ab42-lowering effects, as we expected.
Finally, we obtained each enantiomer of 11j.10 The chiral configu-
ration greatly affected the potency and the eutomer (R)-11j exhib-
ited 14-fold more potent activity than the distomer (S)-11j.
Compound (R)-11j exhibited the most potent Ab42-lowering effect
a
in this series and a moderate Ab40-lowering effect (IC50 = 490 nM)
in the same cellular assay. In addition, (R)-11j was inactive against
Notch signal (IC50 >10 lM). In vivo pharmacokinetic and pharma-
Compound
Ab42 IC50 (nM)
logDb
LLEc
1.6
3
2000
860
4.1
cological profiles of (R)-11j were evaluated in mice. Sufficient
exposure of (R)-11j in brain and plasma was confirmed at 3 h after
oral administration at a dose of 10 mg/kg (Table 3). Furthermore,
compound (R)-11j showed a statistically significant reduction of
Ab42 in both brain and plasma at the same dosage (Table 4).
In conclusion, we designed and synthesized bicyclic analogs to
fix the active conformation of 3. By screening various bicyclic sys-
tems for lead generation, the novel tetrahydro[1,2,4]triazolo[4,3-a]
pyridine core scaffold with enhanced potency and LLE was
4
5
4.4
3.2
1.7
3.5
210
210
6
7
4.2
3.5
2.5
3.2
Table 2
Ab42-lowering effects of 11a-j, (S)-11j and (R)-11j, their logD values and their LLE
220
8
9
630
600
3.3
3.6
2.9
2.6
logDb
LLEc
a
Compound
R
Ab42 IC50 (nM)
11a
11b
11c
11d
11e
11f
11g
11h
11i
11j
(S)-11j
(R)-11j
4-Cl
4-CF3
4-F
4-OMe
4-morpholine
3-CF3
2-CF3
3-CF3, 4-Cl
2-CF3, 5-CF3
2-CF3, 4-Cl
2-CF3, 4-Cl
2-CF3, 4-Cl
570
510
1500
1900
2300
370
380
220
300
84
2.8
2.9
2.4
2.4
2.3
2.8
3.0
3.2
3.3
3.5
3.5
3.5
3.4
3.4
3.4
3.3
3.3
3.6
3.4
3.5
3.2
3.6
2.6
3.7
10
530
3.1
3.2
a
b
c
IC50 values are means of triplicate measurements.
logD at pH 7.4.9
LLE = pIC50 À logD.
830
60
disubstituted analogs 11h–j to further enhance potency by increas-
ing lipophilicity. Definite correlation between potency and logD
value was also observed with 11h–j (Fig. 2, indicated by white
a
b
c
IC50 values are means of triplicate measurements.
logD at pH 7.4.9
LLE = pIC50 À logD.

T. Takai et al. / Bioorg. Med. Chem. Lett. 25 (2015) 4245–4249
4249
discussion. We are indebted to Mr. Motoo Iida for structural deter-
mination by single-crystal X-ray analysis. We express our gratitude
to Dr. Makoto Kamata and Dr. Takanobu Kuroita for helpful
discussion during the preparation of the manuscript. We also
thank Mr. Tomohiro Onishi, Dr. Yasuko Takahashi, Dr. Koji
Murakami, Ms. Yayoi Doken, Dr. Tatsuya Hayama and Dr. Shigeru
Morita for their contribution in the biological evaluation.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2015.07.
101. These data include MOL files and InChiKeys of the most
important compounds described in this article.
References and notes
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Figure 2. Plot of pIC50 against logD for 3–5 and 11a–j.
3. Beel, A. J.; Sanders, C. R. Cell Mol. Life Sci. 2008, 65, 1311.
Table 3
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9. logD at pH7.4 was measured by the retention time of reversed phase
chromatographic procedure whose condition was developed based on the
following published methods. (a) Nakashima, S.; Yamamoto, K.; Arai, Y.; Ikeda,
Y. Chem. Pharm. Bull. (Tokyo). 2013, 61, 1228;; (b) Masumoto, K.; Takeyasu, A.;
Oizumi, K.; Kobayashi, T. Yakugaku Zasshi 1995, 115, 213.
Cortex and plasma concentration of (R)-11j in C57BL/6 J
mice (male, 9 w.o.)
Cortexa
2.332
(l
g/g)
Plasmaa
6.450
(lg/mL)
a
Means of concentrations at 3 h after p.o. adminis-
tration at dose of 10 mg/kg, n = 6 (cortex), 5 (plasma).
Table 4
In vivo profile of (R)-11j in C57BL/6 J mice (male, 9 w.o.)
Brain Aba (%)
Plasmaa Ab (%)
Ab42
Ab40
Ab42
Ab40
50d
75b
31c
63c
a
b
c
Expressed as % vehicle at 3 h after p.o. administration at dose of 10 mg/kg, n = 6.
p <0.05,
p <0.01,
d
p <0.001 Welch test.
discovered. In subsequent SAR studies of the tetrahydro[1,2,4]tria-
zolo[4,3-a]pyridines, strong correlation between potency and logD
value was observed, and increasing lipophilicity of compounds
helped in identification of (R)-11j as a potent GSM. Compound
(R)-11j exhibited a significant Ab42-lowering effect with high
plasma and brain exposures by oral administration in mice.
These results show that the tetrahydro[1,2,4]triazolo[4,3-a]pyri-
dine derivative (R)-11j has the potential of being a therapeutic
agent for AD. Further profiling of (R)-11j is in progress and will
be reported in due course.
10. Compound 11j was separated by HPLC (column: CHIRALPAK AD 50 mmID x
500 mmL, manufactured by DAICEL CHEMICAL INDUSTRIES, LTD., mobile
phase: hexane/isopropyl alcohol 500/500) to give (S)-11j with
retention time and (R)-11j with a longer retention time. The specific rotations
of (S)-11j and (R)-11j were [ ] 25D +127.0 (c 0.41, MeOH) and [ ] 25D–123.8 (c
a shorter
a
a
0.47, MeOH), respectively. The absolute configuration of (S)-11j was
determined by single crystal X-ray crystallography. The crystal structure has
been deposited at the Cambridge Crystallographic Data Centre and the CCDC
deposition number is 1400477.
Acknowledgements
We thank Mr. Yuichi Kajita, Mr. Hitoaki Nishikawa, and Mr.
Tetsuya Tsukamoto for compound synthesis and helpful