diphenyl ether renin inhibitors: Filling the S1 pocket of renin via the S3 pocket

Bioorganic & Medicinal Chemistry Letters 21 (2011) 4836–4843

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Biphenyl/diphenyl ether renin inhibitors: Filling the S1 pocket of renin via the S3 pocket Jing Yuan a,⇑, Robert D. Simpson a, Wei Zhao a, Colin M. Tice a, Zhenrong Xu a, Salvacion Cacatian a, Lanqi Jia a, Patrick T. Flaherty a, Joan Guo a, Alexey Ishchenko a, Zhongren Wu a, Brian M. McKeever a, Boyd B. Scott a, Yuri Bukhtiyarov a, Jennifer Berbaum a, Reshma Panemangalore a, Ross Bentley b, Christopher P. Doe b, Richard K. Harrison a, Gerard M. McGeehan a, Suresh B. Singh a, Lawrence W. Dillard a, John J. Baldwin a, David A. Claremon a a b

Vitae Pharmaceuticals, 502 West Office Center Drive, Fort Washington, PA 19034, USA GlaxoSmithKline, 709 Swedeland Road, King of Prussia, PA 19406, USA

a r t i c l e

i n f o

Article history: Received 18 April 2011 Revised 8 June 2011 Accepted 10 June 2011 Available online 17 June 2011 Keywords: Renin inhibitors Hypertension Biphenyl Diphenyl ether Structure-based design X-ray crystallography

a b s t r a c t Structure-based design led to the discovery of a novel class of renin inhibitors in which an unprecedented phenyl ring filling the S1 site is attached to the phenyl ring filling the S3 pocket. Optimization for several parameters including potency in the presence of human plasma, selectivity against CYP3A4 inhibition and improved rat oral bioavailability led to the identification of 8d which demonstrated antihypertensive efficacy in a transgenic rat model of human hypertension. Ó 2011 Elsevier Ltd. All rights reserved.

The renin angiotensin aldosterone system (RAAS) is a central mechanism by which mammalian blood pressure is controlled.1 The aspartyl protease renin performs the first and rate-limiting step in the RAAS, cleavage of the decapeptide angiotensin I from the N-terminus of the glycoprotein angiotensinogen (Fig. 1). Although renin has long been viewed as a desirable target for antihypertensives, identification of orally bioavailable, low molecular weight inhibitors proved challenging. Efforts by many research groups during the 1980s focused on modified peptides, leading to the identification of potent compounds such as remikiren (1).2,3 However, none of these compounds was ultimately deemed suitable for full development.4 Finally in 2007, Novartis brought the first renin inhibitor, aliskiren (2), to market.5,6 Comparison of the structural features of remikiren (1, Fig. 1) to the amino acid residues surrounding the cleavage site of the natural substrate angiotensinogen illustrates the correspondence between groups occupying the S1, S2 and S3 pockets7 in the respective structures (PDB code: 3D91). Publication of the first X-ray structure of human renin (PDB code: 1RNE)8 revealed that the S1 and S3 pockets form a contiguous superpocket, suggesting ⇑ Corresponding author. E-mail address: [email protected] (J. Yuan). 0960-894X/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.bmcl.2011.06.043

that it might be possible to attach the group filling S3 to the group occupying S1. Early attempts to reduce this idea to practice led to compounds with substantially reduced potency.9,10 However, extensive structure-based design work at Ciba-Geigy, including deletion of the peptide backbone, led to the identification of 2.11–13 The X-ray crystal structure of 2 bound to renin (PDB code: 2V0Z) confirmed that the methoxyphenyl group occupies S3 and that the two i-Pr groups occupy S1 and S10 . We recently described the discovery of potent, orally bioavailable alkyl amine renin inhibitor 3 (Fig. 1).14 This compound connects the residues filling the S1 and S3 pockets through a bridging piperidine linker. Inspection of the X-ray structure of 3 bound to renin (PDB code: 3GW5) suggested an alternate inhibitor design strategy where S1 could be filled by direct attachment of a suitable group to the ortho-position of the phenyl ring occupying S3. In this Letter, we describe the successful realization of this design strategy to afford potent, orally bioavailable renin inhibitors. Application of Contour™, a proprietary computational structure-based drug design program, initially identified the phenoxy group represented in general structure 4 as a candidate to fill the S1 site, when combined with an appropriate group at R1 (Table 1). Introduction of the phenoxy group while retaining the cyclohexylmethyl group at R1 (4a) substantially reduced potency

J. Yuan et al. / Bioorg. Med. Chem. Lett. 21 (2011) 4836–4843

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Figure 1. The N-terminal sequence of angiotensinogen is shown highlighting the amino acids in the P3–P10 positions. The P1, P2 and P3 groups in 1 (remikiren), the P10 , P1 and P3 groups in 2 (aliskiren) and the P1 and P3 groups in 3 are indicated.

Table 1 SAR of diphenyl ethers 4

OH

MeO

N

H O

H N 1 2 O R R

NHMe

4

Compd c

3 4a 4b 4c 4d 4e

R1

R2

IC50a,b,c (nM)

— c-hexCH2 i-Bu t-BuCH2 Me Me

— H H H H Me

0.5 596 8.8 420 126 2900

PRAa,b,d (nM) 13

CYP 3A4e (lM) 1

313

1.1

1340

1.9

a

See Ref. 14 for assay protocols. IC50 values are the average of at least two replicates. c Inhibition of 0.3 nM of purified recombinant human renin in buffer was measured. d IC50 in the presence of human plasma. e Inhibition of CYP3A4 in human liver microsomes was measured. b

compared to 3, as expected. However, truncation of R1 to i-Bu (4b), the P1 sidechain in the natural substrate, greatly increased potency. The closely related t-BuCH2 group (4c) was much less potent. Further reducing the size of R1 to Me (4d) reduced potency compared to 4b. The geminal dimethyl substitution was also unfavorable (4e).

The identification of potent compound 4b provided the first validation of our design. As a next step, we surveyed replacement of the substituted ethylene diamine chain in 4 with various cyclic moieties incorporating basic amines to give compounds of general structure 5 (Table 2. R3 = aminocycloalkyl, azacycloalkyl, etc.). Presentation of the basic amine as part of a cyclic structure was intended to retain the key coulombic interactions with the catalytic aspartates and provide entropic benefit by removing torsional degrees of freedom present in the earlier compound 4. From this effort, we identified (3S,1R)-3aminocyclopentane-1-carboxylic acid derivative 5a with modest potency (Table 2). By contrast, the (3R,1R) and (3R,1S)-isomers 5b and 5c were substantially less potent. Modeling of 5a in the renin binding site suggested that additional hydrophobic interactions with the protein could be attained by introduction of a small lipophilic substituent at the ortho-position of the distal ring of the diphenyl ether which occupies S1. 2-Methylphenoxy compound 5d proved to be 10 more potent than 5a. Further examination of our model indicated that a hydroxyl group at the 4-position of the cyclopentane ring syn to the 3-amino group would enjoy a favorable interaction with the Asp32 carboxylate. Thus, 5e exhibited 7 greater potency than 5d, while the epimer 5f was only slightly more potent than 5d. Unfortunately, the more potent analogs of general structure 5 also inhibited CYP3A4 with IC50 values 61 lM. Rat pharmacokinetic parameters were determined for 5e; the compound showed good oral bioavailability but rapid clearance (Table 3). The rat liver microsome half life (RLM t1/2) of 5e was 18 min, suggesting that oxidative metabolism might well be the mechanism of clearance. Introduction of an ortho-fluorine on the proximal phenyl ring was expected to lock the diphenyl ether into

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Table 2 SAR of 3-aminocyclopentane carboxamides 5

OH

MeO X

6 5

Compd

R3

O 3

4

R3

N

H

X

Y

IC50a,b,c (nM)

O Y

PRAa,b,d (nM)

CYP 3A4e (lM)

5a

NH 2

H

H

195

5b

NH 2

H

H

1437

5c

NH 2

H

H

>5000

5d

NH 2

H

2-Me

19

306

0.4

H

2-Me

2.7

32

0.5

H

2-Me

11

233

1.0

3-F

2-Me

1.2

19

0.4

RLM t1/2 (min)

OH 5e

18

NH 2 OH 5f

NH 2 OH 5g

31

NH 2 a b c d e

See Ref. 14 for assay protocols. IC50 values are the average of at least two replicates. Inhibition of 0.3 nM of purified recombinant human renin in buffer was measured. IC50 in the presence of human plasma. Inhibition of CYP3A4 in human liver microsomes was measured.

a favorable conformation and reduce oxidative metabolism; a modest increase in potency and RLM t1/2 of 31 min were observed (5g). In peptidomimetic renin inhibitors, analogs with aryl rings at P1 are much less potent than compounds with aliphatic groups.15 Furthermore, the majority of the previously described classes of potent, non-peptidic renin inhibitors that bind to the closed form of renin fill S1 with an aliphatic group;16 however, workers at Sanofi-Aventis have recently described inhibitors which deploy a phenyl ring in S1.17 Modeling suggested that replacement of the diphenyl ether with a biphenyl system would be tolerated while, potentially, offering superior intrinsic metabolic stability. Unsubstituted biphenyl analog 6a demonstrated moderate overall potency (Table 4). Modeling indicated that small lipophilic substituents could be accomodated at the 3-position of the distal

phenyl ring of the biphenyl ring system. Thus, 6c (Y = 3-Me) was more potent than both the unsubstituted biphenyl 6a and the 2and 4-methyl isomers 6b and 6d. Another improvement in potency was obtained with 3-chloro compound 6e. Modeling indicated that the chlorine interacted favorably with Val30 and Val120 in the S1 pocket. Unfortunately, 6e suffered a greater loss in potency in the presence of plasma than 6c. However, both 6c and 6e were more stable in RLM than 5e. Introduction of small lipophilic substituents at the 3-position of the proximal phenyl ring was predicted to increase potency by locking the biphenyl system into a favorable conformation for binding to renin; this was borne out with fluoro analog 6f and chloro analog 6i. Introduction of fluorine at the 4-position (6g) and particularly the 5-position (6h) reduced potency. As seen with 6e, chloro analogs 6i–k, while potent when assayed in buffer, suffered >20 losses in potency in presence of plasma.

Table 3 Rat pharmacokinetic parametersa,b

a b c

Compd

Oral Cmax (ng/mL)

Oral tmax (h)

Oral AUC(0–t) ng h/mL

Oral t1/2 (h)

iv CL (mL/min.kg)

Vss (L/kg)

F (%)

3 5e 6i 7bc 8a 8b 8d

73 185 60 5.6 45 71 111

3.3 4.3 3.3 0.2 2.7 2.3 3.3

472 728 264 23 186 295 338

nd 3.8 4.1 50 15.3 4.9 3.1

33 66 39 50 80 58 53

nd 12 9 39 25 19 83

13 34 8 2 14 13 19

All compounds were dosed as fumarate salts at 2 mg/kg iv. All compounds were dosed as fumarate salts at 10 mg/kg po except 7b. Oral dose 7 mg/kg.

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J. Yuan et al. / Bioorg. Med. Chem. Lett. 21 (2011) 4836–4843 Table 4 SAR of biphenyls 6

OH OH

MeO

N

H 6

X

5

a b c d e

NH2 O Y

4

3

Compd

X

Y

IC50a,b,c (nM)

PRAa,b,d (nM)

CYP 3A4e (lM)

6a 6b 6c 6d 6e 6f 6g 6h 6i 6j 6k

H H H H H 3-F 4-F 5-F 3-Cl 3-F 3-Cl

H 2-Me 3-Me 4-Me 3-Cl 3-Me 3-Me 3-Me 3-Me 3-Cl 3-Cl

37 23 10 20 2.1 1.4 7.1 4700 1.6 1.9 3.7

420 126 137 405 79 12 50

4.0 0.5 1.6 0.9 0.2 0.8

35 47 131

0.2 0.5 2.0

RLM t1/2 (min)

67 73

102

See Ref. 14 for assay protocols. IC50 values are the average of at least two replicates. Inhibition of 0.3 nM of purified recombinant human renin in buffer was measured. IC50 in the presence of human plasma. Inhibition of CYP3A4 in human liver microsomes was measured.

Many analogs of general structure 6 posessed submicromolar IC50 values against CYP3A4. An X-ray structure of unsubstituted biphenyl 6a complexed to renin (PDB code: 3Q3T) confirmed the modeled binding pose (Fig. 2). Strong electrostatic interactions of the protonated cyclopentylamine with the Asp32 and Asp215 side chain carboxylates provide a key, anchoring interaction. In addition, the hydroxyl substituent on the cyclopentane donates a hydrogen bond to the Asp32 carboxylate. The tertiary alcohol moiety hydrogen bonds with Ser219, as observed in the crystal structure of 3 bound to renin (PDB code: 3GW5). The ether oxygen at the terminus of the methoxybutyl chain occupying the S3 subpocket forms a hydrogen bond with the backbone NH of Tyr14, as seen for the analogous groups in 2 (PDB code: 2V0Z) and 3 (PDB code: 3GW5). The S3– S1 superpocket filled by the biphenyl moiety, has the phenyl ring in the S3 pocket interacting with the positive edge of Phe117 and

the second phenyl ring in the S1 pocket interacting with Tyr75. Modeling based on 3Q3T predicted that the m-positions of both the phenyl rings have ample room to allow small substituents to occupy hydrophobic spaces in the S1 and S3 pockets and provide improved affinity as observed for 6c, 6e–g and 6i–k. In rat, 6i demonstrated reduced iv clearance, consistent with its longer RLM t1/2, but reduced orally bioavailability, compared to 5e (Table 3). In addition, the IC50 value for 6i in the presence of human plasma (PRA) was unsatisfactory due to projected inferior efficacy in vivo. Our subsequent efforts were directed to identify analogs with improved PRA, reduced CYP3A4 inhibition and increased oral bioavailability. In earlier work with related compounds, our general approach to improving PRA and reducing CYP inhibition was to introduce additional polar functionality into the molecule.18 Specifically, in these earlier compounds, removal of the hydroxyl group and

Figure 2. X-ray structure of 6a bound to renin (PDB code: 3Q3T). The molecular surface corresponding to certain amino acid residues of the binding site was not rendered to provide an unobstructed view of the inhibitor and its interactions.

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replacement of the 4-methoxybutyl chain occupying S3sp with an acylated 2-aminoethoxy chain greatly improved PRA inhibition. By contrast, in this series, the same modification decreased potency both in buffer and in the PRA (7a vs 6f). On the other hand, retention of the hydroxyl and replacement of the 4-methoxybutyl chain with a 3-(methoxycarbonylamino)propyl chain afforded 7b with improved PRA (Table 5). Replacement of fluorine with chlorine (7c) was well tolerated. 3-(Acetylamino)propyl analogs 7d and 7e also had good activity in the PRA and offered a modest reduction in CYP3A4 inhibition. Compound 7b was advanced to rat PK. Despite its long RLM t1/2, 7b was subject to rapid clearance in vivo and had low oral bioavailability (Table 3). Replacement of the piperidine ring in 6 with morpholine offered an alternative strategy to increase overall polarity. Effecting this change in 6f gave 8a, with a 3 reduction in renin potency but a 30 reduction in CYP3A4 inhibition (Table 6). Changing fluorine

to chlorine (8b) and methyl to ethyl (8c) improved renin potency (Table 6). Combining, these afforded 8d with an excellent in vitro profile. Interestingly, replacing the ethyl group in 8d with isopropyl (8e) had little effect on renin potency but led to a substantial increase in CYP3A4 inhibition. Oral bioavailability was demonstrated for 8a, 8b and 8d, but clearance remained high for all three compounds (Table 3). Compounds 8a–f also demonstrated excellent selectivity for renin over three other aspartyl proteases: bsecretase, cathepsin D and cathepsin E (<10% inhibition at 10 lM). The antihypertensive efficacy of 8d was demonstrated in a double transgenic rat (dTGR) model of human hypertension, in which the animals express both human renin and human angiotensinogen (Fig. 3).19,20 Oral administration of 10 mg/kg of 8d led to a statistically significant reduction in mean arterial blood pressure (MABP) sustained for more than 12 h. At the nadir, MABP was reduced by >20 mm Hg.

Table 5 SAR of polar side chain analogs 7

OH R5

R4

N H

NH2 O

Y

X

a b c d e

Compd

X

Y

R4

R5

IC50a,b,c (nM)

PRAa,b,d (nM)

CYP 3A4e (lM)

7a 7b 7c 7d 7e

F F Cl F Cl

Me Me Me Me Me

H OH OH OH OH

MeOCONHCH2CH2O– MeOCONH(CH2)3– MeOCONH(CH2)3– MeCONH(CH2)3– MeCONH(CH2)3–

5.2 1.6 1.1 1.6 2.1

52 4.4 2.8 1.7 4.4

0.5 3.8 1.1 5.3 3.5

RLM t1/2 (min) 567 460 2033

See Ref. 14 for assay protocols. IC50 values are the average of at least two replicates. Inhibition of 0.3 nM of purified recombinant human renin in buffer was measured. IC50 in the presence of human plasma. Inhibition of CYP3A4 in human liver microsomes was measured.

Table 6 SAR of morpholine analogs 8

OH MeO

O OH

N

H

NH2 O

Y

X

a b c d e

Compd

X

Y

IC50a,b,c (nM)

PRAa,b,d (nM)

CYP 3A4e (lM)

RLM t1/2 (min)

8a 8b 8c 8d 8e 8f

F Cl F Cl Cl Cl

Me Me Et Et i-Pr MeS

3.8 1.8 2.5 1.1 0.7 2.3

17 11 8.1 4.0 5.2 26

25 18 17 10 0.4 2

99

See Ref. 14 for assay protocols. IC50 values are the average of at least two replicates. Inhibition of 0.3 nM of purified recombinant human renin in buffer was measured. IC50 in the presence of human plasma. Inhibition of CYP3A4 in human liver microsomes was measured.

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Vehicle (n=6)

Change from Baseline(mmHg)

30

10mg/kg po (n=6)

20 10 0 -10 -20 -30 -40 0

6

12

18

24

Time (hrs) Figure 3. Effect of 8d on mean arterial blood pressure in dTGR.

The syntheses of piperidines 12 and 16, which comprise the left hand sides of the target compounds 5, 6 and 7, are shown in Scheme 1.21 (R)-Boc-nipecotic acid 9 was converted to the corresponding Weinreb amide and treated with various substituted diphenyl ether (10a) or biphenyl (10b) lithiums to afford ketone 11. Addition of (4-methoxybutyl)magnesium chloride to ketone 11 afforded the desired Boc-protected alcohol as the major product (75–80%). To avoid elimination of the tertiary alcohol, the Boc group was removed under mild aqueous conditions to afford key intermediate 12. Similarly, addition of Grignard reagent 14, which was generated from protected amine 13, to ketone 11b afforded, after aqueous work up, aminoalcohol 15 as the major product. The amino group in 15 was treated with acetic anhydride or methyl chloroformate, followed by removal of the Boc group under mild conditions, to afford key piperidine intermediates 16. The stereochemistry of the newly formed carbinol center in both 12 and 16 was initially assigned based on the results obtained in analogous reactions reported previously.14 These assignments were subsequently confirmed by the X-ray structure of derived compound 6a (PDB code: 3Q3T). The synthesis of morpholine intermediate 27 is shown in Scheme 2. Epoxide 17 was opened by amine 18 under basic

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conditions to give intermediate 19, which was further cyclized and protected to afford morpholine benzyl ether 21.22 The benzyl group of compound 21 was removed by hydrogenation to afford alcohol 22, which was oxidized to acid 24 using trichloroisocyanuric acid (23) and TEMPO in excellent yield. It was determined experimentally that Grignard additions to morpholine ketones such as 26 occur under chelation control. This dictated that the order of addition of the methoxybutyl group and the biphenyl moiety be reversed from that used in the synthesis of piperidine intermediates 12 and 16. Thus, acid 24 was converted to Weinreb amide 15 and treated with (4-methoxybutyl)magnesium chloride to afford ketone 26. A variety of substituted biphenyl lithiums (10b) was added to ketone 16 to give, after removal of the Boc group under mild conditions, the desired morpholine intermediate 27 as the major product which was utilized in the synthesis of analogs 8. The assembly of ureas 4 is shown in Scheme 3. Commercially available Boc-amino acids 28 were converted to the corresponding methyl amides 29, which were reduced with Red-Al to afford secondary amines 30 in reasonable yields. After protecting the secondary amine with Teoc, the Boc group of compounds 31 was selectively removed using p-toluenesulfonic acid under vaccum at 60 °C to give primary amines 32. Amines 32 were then activated with p-nitrophenyl chloroformate to form their p-nitrophenylcarbamates, which were reacted with piperidine 12a (X, Y = H) to afford the urea. Finally, the Teoc group was removed with Et4N+F in acetonitrile to give the desired analogs 4. The formation of target compounds 5–8 is shown in Scheme 4. Commercially available stereoisomers of Boc-protected amino esters 34 were hydrolyzed to the carboxylic acids 33b under mild conditions. The Boc-protected amino acids 33 were then coupled with piperidines 12 or 16 or morpholines 27 under standard amide formation conditions, followed by Boc removal, to afford 5–8.23 Scheme 5 shows the synthesis of target compound 7a. Ketone 11b was reduced by NaBH4 to alcohol 35 as a mixture of diastereoisomers, which was alkylated to give ester 36. Ester 36 was reduced to alcohol 37, which was elaborated to primary amine 38 in three steps. The diastereomers of 38 were separated by preparative HPLC. Methyl carbamate formation and removal of the Boc group gave piperidine 39. Coupling the (1S,3R,4S)-isomer of 33b with 39, followed by deprotection afforded 7a. In conclusion, we have described the structure based design of a topologically novel class of renin inhibitors in which an unprecedented phenyl ring filling the S1 site is attached to the phenyl ring filling S3. The predicted binding mode was confirmed by X-ray

Scheme 1. Synthesis of piperidine intermediates. Reagents and conditions: (a) MeNHOMeHCl, EDC; i-Pr2NEt, CH2Cl2, rt, 16 h; (b) 10a, THF, 20 to 10 °C, 30 min to 1 h; or 10b, THF, 78 °C to rt, 16 h; (c) for 11a: MeO(CH2)4MgCl, THF, 10 °C to 78 °C to rt, 16 h; for 11b, MeO(CH2)4MgCl, THF, 78 °C to rt, 16 h; (d) 1:1 2 M aq HCl/MeCN, rt, 16 h; (e) Mg, THF, reflux, 2.5 h; (f) 11b, THF, 78 °C to rt, 16 h; (g) for 16a: MeOCOCl, DMAP, Et3N, CH2Cl2; for 16b: Ac2O, DMAP, Et3N, CH2Cl2.

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Scheme 2. Synthesis of morpholine intermediates. Reagents and conditions: (a) NaOH, MeOH/H2O, 40 °C, 2 h; (b) NaOH, toluene, H2O, 65 °C, 16 h; (c), (Boc)2O, K2CO3, 3:1 acetone/H2O, 0 °C to rt, 30 min; (d) Pd–C, H2, MeOH, rt; (e) TEMPO, NaBr, NaHCO3, acetone; (f) MeNHOMe, HBTU, HOBt, DIEA, DMF; (g) MeO(CH2)4MgCl, THF, 20 °C, 10 min; (h) 10b, THF, toluene, 20 °C, 10 min; (i) 1:1 2 M aq HCl/MeCN, rt, 16 h.

Scheme 3. Synthesis of ureas 4. Yields are shown for 4a and are representative. Reagents and conditions: (a) MeNH2HCl, HOBt, HBTU, i-Pr2NEt, DMF, 0 °C to rt, 2 h; (b) RedAl, toluene, 0 °C to rt, 16 h; (c) Teoc-OSu, K2CO3, 3:1 acetone/H2O, 0 °C to rt, 1.5 h; (d) TsOHH2O, EtOH, ether, 60–65 °C, 30 min; (e) p-NO2C6H4OCOCl, pyridine, CH2Cl2, rt, 5 min; (f) 12a (X, Y = H), i-Pr2NEt, 1:1 MeCN/CH2Cl2; (g) Et4N+F, MeCN, 50 °C, 16 h.

Scheme 4. Synthesis of amides 5–8. For definitions of X, Y, R7 and cyclopentane stereochemistry, see Tables 3–6. Reagents and conditions: (a) 1 N NaOH, 1:1 EtOH/THF; (b) HOBt, HBTU, i-Pr2NEt, DMF, rt, 5 min; (c) 1:1 2 M aq HCl/MeCN, rt, 16 h.

J. Yuan et al. / Bioorg. Med. Chem. Lett. 21 (2011) 4836–4843

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Scheme 5. Synthesis of 7a. Reagents and conditions: (a) NaBH4, CH3OH, rt, 2 h; (b) BrCH2CO2Et, NaH, THF, 0 °C to reflux, 12 h; (c) NaBH4, CH3CH2OH, rt, 16 h; (d) MsCl, Et3N, CH2Cl2, 0 °C to rt, 1 h; (e) NaN3, DMF, 80 °C, 5 h; (f) Pd–C, H2, CH3OH, rt, 16 h; (g) preparative HPLC; (h) MeOCOCl, DMAP, Et3N, CH2Cl2; (i) 1:1 2 M aq HCl/MeCN, rt, 16 h; (j) (1S,3R,4S)-33b, HOBt, HBTU, i-Pr2NEt, DMF, rt, 5 min.

crystallography. This effort culminated in the discovery of 8d, a potent, selective and orally bioavailable inhibitor of renin which effectively lowered blood pressure in an animal model of human hypertension. References and notes 1. Skeggs, L.; Kahn, J. R.; Lentz, K.; Shumway, N. P. J. Exp. Med. 1957, 106, 439. 2. Greenlee, W. J. Med. Res. Rev. 1990, 10, 173. 3. Rosenberg, S. H. In Progess in Medicinal Chemistry; Ellis, G. P., Luscombe, D. K., Eds.; Elsevier Science, 1995; Vol. 32, p 37. 4. Tice, C. M.; McGeehan, G. M.; Claremon, D. A., 7th ed. In Burger’s Medicinal Chemistry, Drug Discovery, and Development; Abraham, D. J., Rotella, D. P., Eds.; Wiley, 2010; Vol. 4, p 239. 5. Jensen, C.; Herold, P.; Brunner, H. R. Nat. Rev. Drug Disc. 2008, 7, 399. 6. Maibaum, J.; Feldman, D. L. Annu. Rep. Med. Chem. 2009, 44, 105. 7. Schechter, I.; Berger, A. Biochem. Biophys. Res. Commun. 1967, 27, 157. 8. Rahuel, J.; Priestle, J.; Gruetter, M. G. J. Struct. Biol. 1991, 107, 227. 9. Lefker, B. A.; Hada, W. A.; Wright, A. S.; Martin, W. H.; Stock, I. A.; Schulte, G. K.; Pandit, J.; Danley, D. E.; Ammirati, M. J.; Sneddon, S. F. Bioorg. Med. Chem. Lett. 1995, 5, 2623. 10. Plummer, M. S.; Shahripour, A.; Kaltenbronn, J. S.; Lunney, E. A.; Steinbaugh, B. A.; Hamby, J. M.; Hamilton, H. W.; Sawyer, T. K.; Humblet, C.; Doherty, A. M.; Taylor, M. D.; Hingorani, G.; Batley, B. L.; Rapundalo, S. T. J. Med. Chem. 1995, 38, 2893. 11. Rahuel, J.; Rasetti, V.; Maibaum, J.; Rueger, H.; Goschke, R.; Cohen, N. C.; Stutz, S.; Cumin, F.; Fuhrer, W.; Wood, J. M.; Grutter, M. G. Chem. Biol. 2000, 7, 493. 12. Webb, R. L.; Schiering, N.; Sedrani, R.; Maibaum, J. J. Med. Chem. 2010, 53, 7490. 13. Maibaum, J.; Stutz, S.; Goschke, R.; Rigollier, P.; Yamaguchi, Y.; Cumin, F.; Rahuel, J.; Baum, H. P.; Cohen, N. C.; Schnell, C. R.; Fuhrer, W.; Gruetter, M. G.; Schilling, W.; Wood, J. M. J. Med. Chem. 2007, 50, 4832.

14. Tice, C. M.; Xu, Z.; Yuan, J.; Simpson, R. D.; Cacatian, S. T.; Flaherty, P. T.; Zhao, W.; Guo, J.; Ishchenko, A.; Singh, S. B.; Wu, Z.; Scott, B. B.; Bukhtiyarov, Y.; Berbaum, J.; Mason, J.; Panemangalore, R.; Cappiello, M. G.; Mueller, D.; Harrison, R. K.; McGeehan, G. M.; Dillard, L. W.; Baldwin, J. J.; Claremon, D. A. Bioorg. Med. Chem. Lett. 2009, 19, 3541. 15. Sham, H. L.; Rempel, C. A.; Stein, H.; Cohen, J. J. Chem. Soc., Chem. Commun. 1987, 683. 16. Evolution of diverse classes of renin inhibitors through the years Tice, C. M.; Singh, S. B. In Aspartic Acid Proteases as Therapeutic Targets; Ghosh, A., Ed.; Wiley-VCH, 2010; vol. 45, p 297. 17. Scheiper, B.; Matter, H.; Steinhagen, H.; Stilz, U.; Böcskei, Z.; Fleury, V.; McCort, G. Bioorg. Med. Chem. Lett. 2010, 20, 6268. 18. Xu, Z.; Cacatian, S.; Yuan, J.; Simpson, R. D.; Jia, L.; Zhao, W.; Tice, C. M.; Flaherty, P. T.; Guo, J.; Ishchenko, A.; Singh, S. B.; Wu, Z.; McKeever, B. M.; Scott, B. B.; Bukhtiyarov, Y.; Berbaum, J.; Mason, J.; Panemangalore, R.; Cappiello, M. G.; Bentley, R.; Doe, C. P.; Harrison, R. K.; McGeehan, G. M.; Dillard, L. W.; Baldwin, J. J.; Claremon, D. A. Bioorg. Med. Chem. Lett. 2010, 20, 694. 19. Ganten, D.; Wagner, J.; Zeh, K.; Bader, M.; Michel, J. B.; Paul, M.; Zimmermann, F.; Ruf, P.; Hilgenfeldt, U.; Ganten, U. Proc. Natl. Acad. Sci. U.S.A. 1992, 89, 7806. 20. Luft, F. C.; Mervaala, E.; Muller, D. N.; Gross, V.; Schmidt, F.; Park, J. K.; Schmitz, C.; Lippoldt, A.; Breu, V.; Dechend, R.; Dragun, D.; Schneider, W.; Ganten, D.; Haller, H. Hypertension 1999, 33, 212. 21. Target compounds and key intermediates gave satisfactory 1H NMR and LC–MS data. 22. Showell, G. A.; Emms, F.; Marwood, R.; O’Connor, D.; Patel, S.; Leeson, P. D. Bioorg. Med. Chem. 1998, 6, 1. 23. Compound 8d gave the following spectral data: 1H NMR (400 MHz, CD3OD) d 7.69–7.66 (m, 1H), 7.30–7.12 (m, 4H), 6.88–6.84 (m, 2H), 6.59 (s, 2H), 4.23– 4.12 (m, 2H), 3.75–3.57 (m, 2H), 3.43–3.33 (m, 1H), 3.27–3.00 (m, 9H), 2.75– 2.54 (m, 3H), 2.23–1.79 (m, 3H), 1.68–1.12 (m, 9H), 0.86–0.75 (m, 1H); MS ESI +ve ionization, m/z 545, 547 (M+H+).