Discovery of VTP-27999, an alkyl amine renin inhibitor with potential for clinical utility

Article abstract of DOI:10.1021/ml200137x

Structure guided optimization of a series of nonpeptidic alkyl amine renin inhibitors allowed the rational incorporation of additional polar functionality. Replacement of the cyclohexylmethyl group occupying the S1 pocket with a (R)-(tetrahydropyran-3-yl)

Full text of DOI:10.1021/ml200137x

LETTER
pubs.acs.org/acsmedchemlett
Discovery of VTP-27999, an Alkyl Amine Renin Inhibitor with Potential
for Clinical Utility
Lanqi Jia,† Robert D. Simpson,† Jing Yuan,† Zhenrong Xu,† Wei Zhao,† Salvacion Cacatian,† Colin M. Tice,*,†
Joan Guo,† Alexey Ishchenko,† Suresh B. Singh,† Zhongren Wu,† Brian M. McKeever,† Yuri Bukhtiyarov,†
Judith A. Johnson,† Christopher P. Doe,‡ Richard K. Harrison,† Gerard M. McGeehan,†
Lawrence W. Dillard,† John J. Baldwin,† and David A. Claremon†
†Vitae Pharmaceuticals, 502 West Office Center Drive, Fort Washington, Pennsylvania 19034, United States
‡GlaxoSmithKline, 709 Swedeland Road, King of Prussia, Pennsylvania 19406, United States
S Supporting Information
b
ABSTRACT: Structure guided optimization of a series of nonpeptidic alkyl amine renin
inhibitors allowed the rational incorporation of additional polar functionality. Re-
placement of the cyclohexylmethyl group occupying the S1 pocket with a
(R)-(tetrahydropyran-3-yl)methyl group and utilization of a different attachment point
led to the identification of clinical candidate 9. This compound demonstrated excellent
selectivity over related and unrelated off-targets, >15% oral bioavailability in three species,
oral efficacy in a double transgenic rat model of hypertension, and good exposure in humans.
KEYWORDS: Renin, aspartyl protease, hypertension, structure-based drug design
he renin angiotensin aldosterone system (RAAS) plays a
major role in the control of vascular function and blood
Its oral bioavailability in rat and monkey was just below 10%, and its
clearance was medium in rat but low in cynomolgus monkey
(Table 2). However, 2 had an IC50 of 4.6 μM against CYP3A4,
which was considered undesirable. Reducing inhibition of CYP3A4,
while retaining or improving oral bioavailability and excellent
potency against plasma renin activity (PRA), became the major
goal for the further optimization of 2.
T
pressure.1 Chronic stimulation of the RAAS leads to tissue inflam-
mation and fibrosis with end organ dysfunction, particularly of the
kidney.2,3 Renin is a highly selective aspartyl protease secreted from
the kidneys, which cleaves a decapeptide, angiotensin (Ang) I, from
the N-terminus of angiotensinogen. Ang I is further processed by
angiotensin converting enzyme (ACE) to give an octapeptide, Ang
II, which binds to the angiotensin receptor type 1 (AT1), leading to
vasoconstriction and, with chronic stimulation, inflammation and
fibrosis. Recently, local RAASs have been shown to function in
various tissues, including kidney and heart.4 Two major classes of
antihypertensives, ACE inhibitors and AT1 receptor blockers
(ARBs), function by blocking the RAAS.5 The high selectivity of
renin and its activity at the top of the RAAS cascade suggest that a
direct renin inhibitor (DRI) may offer a superior therapeutic profile
to the ACE inhibitors and ARBs.6,7 RAAS blockers are also used
clinically for the treatment of chronic kidney disease (CKD).8
Preliminary studies indicate that DRIs may provide better end
organ protection in the treatment of CKD.9,10
’ RESULTS AND DISCUSSION
Our strategy to accomplish this goal was to introduce addi-
tional polar functionality into 2 at positions which would allow
favorable interactions with renin. We anticipated that the added
polar functionality would cause unfavorable interactions with the
largely hydrophobic binding pocket of CYP3A4.21 We used the 1.9
Å resolution X-ray structure of 2 bound to human renin (PDB code:
3KM4), combined with our proprietary structure based drug design
tool, Contour, to grow and score analogues of 2 within the renin
binding site.
Compounds 3 and 5, both of which vary in the group filling the S1
pocket,22 emerged from our design process as attractive candidates to
meet this goal. Despite the added polar groups, models of 3 and 5
bound to renin retained the key interactions and overall binding
mode observed in the X-ray structure of 2 (PDB code: 3KM4). The
tetrahydropyran ring of the (R)-(tetrahydropyran-3-yl)methyl com-
pound 3 was predicted to retain the same pucker observed for the
cyclohexane ring in the complex of 2 with renin and thus position the
ring ether oxygen to accept a hydrogen bond from the side chain
hydroxyl of Thr77 in the flap region of the protein. By contrast, the
Renin has proved to be a historically challenging target for
medicinal chemists. During the 1980s, intensive efforts that
focused on peptidomimetics led to the discovery of many potent
compounds;11,12 however, because of poor oral bioavailability,
none of them was successfully developed as a drug.13 Beginning
in the mid-1990s, several classes of nonpeptidic renin inhibitors
were reported.13ꢀ17 One line of investigation culminated in the
discovery of aliskiren (1, Figure 1), the first approved DRI, which
was brought to market in 2007.18
We recently described the structure based discovery of a new
class of nonpeptidic renin inhibitors represented by alkyl amine
2.19,20 Compound 2 had excellent potency against renin (Table 1).
Received: June 9, 2011
Accepted: August 9, 2011
Published: August 09, 2011
r
2011 American Chemical Society
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|

ACS Medicinal Chemistry Letters
LETTER
Figure 1. Aliskiren and alkyl amine renin inhibitors.
Table 1. Enzyme Inhibitiona,b
predicted to be less potent. The hydroxy group of R-hydroxy-
(cyclohexyl)methyl compound 5 was expected to donate a hydrogen
bond to Asp32, one of the catalytic aspartates.
compd
no.
IC50
low renin
PRAe
(nM)
CYP3A4
c
(nM)
IC50d (nM)
IC50f (nM)
The synthesis of the analogues in Figure 1 was accomplished
by formation of the central urea linkage from the appropriate left-
hand piperidine and a suitably protected right-hand diamine. The
final steps in the synthesis of 3 from piperidine 1019 and protected
diamine 11 are shown in Scheme 1. Complete synthetic schemes
and procedures are provided in the Supporting Information.
Compounds 3 and 5 both retained potency against renin in the
absence or presence of plasma comparable to that of 2 (Table 1).
Compound 3 was a substantially weaker inhibitor of CYP3A4 than
2, while 5 did not differ significantly from 2. As predicted, 4 was less
potent than 3. Analogue 6 was also prepared because the (tetrahy-
dropyran-4-yl)methyl group has the advantage of not introducing
an additional chiral center. Interestingly, although 6 was less potent
against the enzyme in buffer than 2 or 3, it experiences only a 2ꢁ
loss in potency when assayed in the presence of human plasma and
its CYP3A4 IC50 value was >30 μM. Pharmacokinetic parameters of
3 were determined in rat and cynomolgus monkey (Table 2). The
1
2
3
4
5
6
7
8
9
0.40
0.48
0.56
5.5
0.33
6.2
2.2
0.73
0.47
0.53
0.65
0.82
0.48
4.5
1.06
13.1
69
4600
28000
0.42
2.7
0.15
1.7
7100
>30000
2910
0.27
9.4
1.1
0.30
>30000
a Assay protocols are provided in the Supporting Information. b Assay
c
results are the average of at least two replicates. Inhibition of 0.3 nM of
purified recombinant human renin in buffer was measured. d Inhibition of 36
pM of purified recombinant human renin in buffer was measured. e IC50 in
f
the presence of human plasma. Inhibition of CYP3A4 in human liver
microsomes was measured.
corresponding ether oxygen in epimeric (S)-(tetrahydropyran-3-yl)-
methyl compound 4 was unable to form this interaction and was
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LETTER
Table 2. PK Parametersa
IV dose
oral dose
(mg/kg)
oral AUC(0ꢀt)
IV CL
compd no.
species
ratb
(mg/kg)
(ng h/mL)
IV t1/2 (h)
(mL/min kg)
Vss (L/kg)
F (%)
3
3
1
2
3
7
8
9
1
2
3
9
1
9
2.0
1.6
1.5
0.8
1.6
2.0
1.0
1.0
1.0
1.0
1.0
0.5
10.0
8.0
8.2
4.0
8.2
10.0
2.0
1.0
2.0
2.0
1.0
1.0
21
301
167
269
226
477
154
130
172
404
149
484
39.6
16.2
18.3
51.0
15.9
7.3
55
36
62
19
1.2
9.4
7.5
29.5
5.5
37
rat
rat
48
39
rat
47
101
16
rat
30
rat
128
2.5
38
monkeyb
monkey
monkey
monkey
dog
14.7
23.3
14.0
12.3
16.3
9.9
1.5
9.8
5.0
4.7
4.7
4.0
1.4
9.9
5.3
18
11.7
9.5
29.6
11.0
12.9
11
dog
41
a Compounds were administered as fumarate salts. b PK parameters for 1 in rat, marmoset, and dog have been reported previously. Concentrations of 1
were determined by a bioassay. See ref 18.
Scheme 1. Preparation of 3a
a Teoc = 2-(trimethylsilyl)ethoxycarbonyl; CDI = carbonyl diimidazole.
oral bioavailability of 3 in both species was lower than that of 2 but
superior to that of 1.18 Further synthesis was directed toward the
identification of a compound that combined the potency and
minimal CYP3A4 inhibition of 3 with improved oral bioavailability.
In the course of earlier modeling and SAR studies around 2, it
had been shown that the cyclohexylmethyl susbstituent that fills
the S1 pocket could also be attached to the carbon atom R to the
basic amine, albeit with a modest loss in potency. For example,
primary amine 7had an IC50 of 2.2 nM against human renin in buffer.
Intriguingly, 7 had significantly greater oral bioavailability in rat than
the related regioisomeric compound 820, in which the cyclohexyl-
methyl moiety is attached to the carbon atom β to the basic amine.
Modeling indicated that 9, in which an (R)-(tetrahydropyran-3-yl)-
methyl group is attached in the (S) configuration R to the basic
amine, suitably positioned the pyran oxygen to accept a hydrogen
bond from Thr 77. Compound 9proved to have comparable potency
to 1and 3against renin both in buffer and in the PRA assay (Table 1)
and had an IC50 value >30 μM against CYP3A4. Encouragingly, the
oral bioavailabilities of 9 in rat and monkey were 37% and 18%,
respectively, surpassing both 1and 3(Table 2). Furthermore, the oral
bioavailability of 9 in dog was greater than that of 1. The free fraction
of 9 in plasma was determined in four species, including human,
and ranged from 22% in dog to 29% in monkey. These results are
consistent with the observed 3ꢁ loss in potency when enzyme inhibi-
tion was measured in the absence and presence of plasma.
Compound 9 was selected for further in vitro and in vivo
evaluation. At a 10 μM concentration, 9 caused <10% inhibition of
three other human aspartyl proteases: β-secretase, cathepsin D, and
cathepsin E. In addition, on the basis of its PRA potency, 9 demon-
strated >1000ꢁ selectivity for renin, over a panel of >150 receptors,
ion channels, and enzymes.23,24
Compound 9 was nonmutagenic in the Ames assay, when tested
in either the presence or absence of S9-mix, and was not genotoxic in
the mouse lymphoma L5178Y TK( test system.
The efficacy of 9 at 10 mg/kg po was compared to 1 at the
same dose in double transgenic rats (dTGR)25 engineered to express
human renin and angiotensinogen (Figure 2). These animals have
severe hypertension dependent on human renin. Compound 9 pro-
vided a greater reduction in mean arterial blood pressure (MAP) at
t = 24 h and a longer duration of action.
In a single ascending dose study in normal volunteers, dose
proportionate increases in plasma drug levels of 9 were observed with
doses from 40 mg to 1000 mg.26
X-ray crystal structures were obtained of both 3 (PDB code:
3Q5H, Figure 3A) and 9 (PDB code: 3Q4B Figure 3B) bound to
renin. The general binding modes and interactions observed in both
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LETTER
Figure 2. Mean arterial pressure (MAP) responses to 1 and 9 in dTGR.
alkyl amine chemotype (PDB codes: 3KM4, 3GW5).19,20 Thus, in
both cases, the protonated secondary amine is positioned between
the catalytic aspartates, Asp32 and Asp215. The urea and carbamate
NH's both donate hydrogen bonds to the backbone carbonyl oxygen
of Gly217. These favorable hydrogen bonds, and an intraligand
hydrogen bond between the carbamate NH and the ether oxygen of
the group occupying the S3sp subsite, serve to mitigate the repulsion
between the carbonyl of Gly217 and the ether oxygen of the ligands.
The tetrahydropyran oxygens of both 3 and 9 form hydrogen bonds
to Thr77, validating our design.
’ CONCLUSION
Structure based optimization of a series of nonpeptidic alkyl
amine renin inhibitors allowed the rational incorporation of addi-
tional polar functionality and a change in the attachment point of the
group occupying the S1 pocket. The additional polar functionality
maintained potency against renin, while improving the selectivity of
the compounds, and led to the identification of clinical candidate 9.
This compound demonstrated >1000ꢁ selectivity over related and
unrelated off-targets, >15% oral bioavailability in three species, and
oral efficacy in a dTGR model of hypertension.
’ ASSOCIATED CONTENT
S
Supporting Information. Representative synthetic
b
schemes and procedures; NMR, MS, and HPLC data on new
compounds; assay protocols; and crystallography procedures.
This material is available free of charge via the Internet at
http://pubs.acs.org.
Figure 3. Crystal structures of 3 and 9 bound to renin: (A) Compound
3 bound to renin, PDB code: 3Q5H. (B) Compound 9 bound to renin,
PDB code: 3Q4B. Selected hydrogen bonds are depicted as dashed yellow
lines. The molecular surface corresponding to certain amino acid residues
from the flap region of the binding site, including Thr77, was not rendered
to provide an unobstructed view of the inhibitor and its interactions.
’ AUTHOR INFORMATION
Corresponding Author
*Phone: 215-461-2042. Fax: 215-461-2006. E-mail: ctice@
vitaerx.com.
complexes were as expected on the basis of molecular modeling and
the previously published X-ray crystal structures of members of the
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LETTER
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