Discovery of AMG 232, a potent, selective, and orally bioavailable MDM2-p53 inhibitor in clinical development

Article

pubs.acs.org/jmc

Discovery of AMG 232, a Potent, Selective, and Orally Bioavailable

MDM2−p53 Inhibitor in Clinical Development

Daqing Sun,*^,†Zhihong Li,^†Yosup Rew,^†Michael Gribble,^†Michael D. Bartberger,^∥Hilary P. Beck,^†

Jude Canon,^⊥Ada Chen,^†Xiaoqi Chen,^†David Chow,^†Jeﬀrey Deignan,^†Jason Duquette,^†

John Eksterowicz,^†Benjamin Fisher,^†Brian M. Fox,^†Jiasheng Fu,^†Ana Z. Gonzalez,^†

Felix Gonzalez-Lopez De Turiso,^†Jonathan B. Houze,^†Xin Huang,^#Min Jiang,^§Lixia Jin,^§Frank Kayser,^†

Jiwen (Jim) Liu,^†Mei-Chu Lo,^†Alexander M. Long,^#Brian Lucas,^†Lawrence R. McGee,^†Joel McIntosh,^†

Jeﬀ Mihalic,^†Jonathan D. Oliner,^⊥Tao Osgood,^⊥Matthew L. Peterson,^∇Philip Roveto,^†Anne Y. Saiki,^⊥

Paul Shaﬀer,^#Maria Toteva,^‡Yingcai Wang,^†Yu Chung Wang,^⊥Sarah Wortman,^‡Peter Yakowec,^#

Xuelei Yan,^†Qiuping Ye,^§Dongyin Yu,^⊥Ming Yu,^†Xiaoning Zhao,^†Jing Zhou,^†Jiang Zhu,^†

Steven H. Olson,^†and Julio C. Medina^†

‡

§

^†Departments of Therapeutic Discovery, Pharmaceutics, and Pharmacokinetics and Drug Metabolism, Amgen Inc., 1120 Veterans

Boulevard, South San Francisco, California, 94080, United States

⊥

^∥Departments of Therapeutic Discovery, and Oncology Research, Amgen Inc., One Amgen Center Drive, Thousand Oaks,

California 91320, United States

^#Departments of Therapeutic Discovery, and ^∇Pharmaceutics, Amgen Inc., 360 Binney Street, Cambridge, Massachusetts 02142,

United States

S

* Supporting Information

ABSTRACT: We recently reported the discovery of AM-8553

(1), a potent and selective piperidinone inhibitor of the

MDM2−p53 interaction. Continued research investigation of

the N-alkyl substituent of this series, focused in particular on a

previously underutilized interaction in a shallow cleft on the

MDM2 surface, led to the discovery of a one-carbon tethered

sulfone which gave rise to substantial improvements in

biochemical and cellular potency. Further investigation

produced AMG 232 (2), which is currently being evaluated

in human clinical trials for the treatment of cancer. Compound

2 is an extremely potent MDM2 inhibitor (SPR K_D= 0.045

nM, SJSA-1 EdU IC₅₀= 9.1 nM), with remarkable

pharmacokinetic properties and in vivo antitumor activity in the SJSA-1 osteosarcoma xenograft model (ED₅₀= 9.1 mg/kg).

inhibitors have advanced into clinical trials for the treatment of

cancer (Figure 1).¹²

INTRODUCTION

■

The tumor suppressor protein p53 plays a pivotal role in growth

suppression and apoptosis of cancer cells,^1,2and p53 pathway

inactivation may be required for advanced tumor formation.³

Roughly half of human cancers have wild-type p53, whose

functions are regulated by human murine double minute 2

(MDM2).⁴⁻⁶MDM2 directly binds to and blocks the N-terminal

transcriptional activation domain of p53, promotes export of p53

from nucleus to the cytoplasm, and induces degradation of p53

via ubiquitination through its E3 ligase activity.^1,2As a result, the

disruption of the MDM2−p53 interaction has been regarded as

an attractive strategy for the activation of the p53 pathway.⁷In

fact, studies with small molecule inhibitors of the MDM2−p53

interaction have yielded signiﬁcant reductions in tumor growth

both in vitro and in vivo.⁸⁻¹¹Recently, several of these MDM2

RESULTS AND DISCUSSION

■

Previously, we reported the discovery of AM-8553 (1; Figure

2)¹³as a potent and selective piperidinone inhibitor of the

MDM2−p53 interaction. Compound 1 inhibited the MDM2−

p53 interaction in a biochemical HTRF assay¹⁴with an IC₅₀of

1.1 nM. Its activity on the human SJSA-1 tumor cell line ranged

from an IC₅₀of 0.072 μM in an EdU cell proliferation assay¹⁴to

an IC₅₀of 0.76 μM in a p21 induction assay¹⁴in the presence of

10% human serum. Furthermore, this compound demonstrated

tumor growth inhibition in the SJSA-1 xenograft model.

Received: November 13, 2013

A

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a

Figure 1. Chemical structures and potencies of RG7112, RG7388, SAR299155, and 2. K_Din the surface plasmon resonance (SPR) spectroscopy

binding assay.¹⁴^bIC₅₀in the EdU proliferation assay (SJSA-1, 10% human serum).¹⁴

a

Figure 2. Chemical structure and potency of 1. K_Din the surface

plasmon resonance (SPR) spectroscopy binding assay.¹⁴^bIC₅₀in the

EdU proliferation assay (SJSA-1, 10% human serum).¹⁴

The cocrystal structure of 1 with human MDM2 protein is

shown in Figure 3.¹³The potency of 1 was improved with a focus

on the three critical hydrophobic binding pockets and an

interaction with H96. The C5 aryl group ﬁlls the Leu26_(p53)

pocket and engages in a face-to-face π-stacking interaction with

H96, while the C6 aryl group occupies the Trp23_(p53)binding

cavity. The hydroxyl group is exposed to solvent and the ethyl

group directed into the Phe19_(p53)pocket by the conformational

Figure 3. Cocrystal structure of 1 bound to human MDM2 (17−111) at

constraint induced by the α-substitution. In addition, the

2.0 Å resolution and depicting proposed electrostatic interactions with

conformation of the piperidinone ring is stabilized by

H96 and G58 “shelf” region (circled). White labels indicate the positions

incorporation of a methyl group at the C3 position. Finally, the

normally occupied by key p53 residues. MDM2 residues H96, G58,

M62, N59, and F55 are labeled in yellow. Cocrystallized water molecules

are shown in red. PDB code: 4ERF.

carboxylate of 1 interacts with the adjacent, presumably, charged

imidazole of the H96 side chain of MDM2.

Analysis of our own in-house MDM2 cocrystal structures

along with those in the literature suggested a region of the

MDM2 protein which, while not accessed to a signiﬁcant extent

by inhibitors published to date, might serve as a potential source

for improved binding aﬃnity. The right side of Figure 3 depicts

this shallow, predominantly hydrophobic cleft adjacent to the

Phe19 binding pocket and to the right of the N-alkyl group of 1. It

is bordered on the edges by F55, N59, and M62. In this paper, we

designate this region the “glycine shelf”, as G58 serves as the

approximate center and entry point to this extended, relatively

ﬂat part of the protein surface. While hydrophobic, this section of

the binding site in the cocrystal structure is shallow and largely

solvent exposed. We felt that additional research investigation of

the N-alkyl group in 1, especially in this shelf region, might oﬀer

an opportunity to increase potency while maintaining favorable

selectivity and low intrinsic clearance in human hepatocytes. In

this article, we report the continued investigational eﬀorts which

led to the discovery of AMG 232 (2, Figure 1), a highly potent

MDM2 inhibitor (K_D= 0.045 nM, EdU IC₅₀= 9.1 nM), with

remarkable pharmacokinetic properties and in vivo eﬃcacy in the

SJSA-1 osteosarcoma xenograft model (ED₅₀= 9.1 mg/kg).

The investigation began with the replacement of the secondary

alcohol in 1 with various alcohol or nitrile groups (Table 1). The

tertiary alcohols 3 and 4 were potent in the biochemical assay in

the absence of human serum but suﬀered a larger potency

decrease than 1 upon addition of human serum. Compound 5,

containing a two-carbon spacer between the tertiary alcohol

moiety and the carbon adjacent to the ring nitrogen, exhibited a

similar potency as 1 in both the biochemical and the cell-based

assays. Changing from the tertiary alcohol to a secondary alcohol

(6 and 7)¹⁵slightly improved the biochemical potency but

decreased the cellular potency. Similarly, replacing the secondary

B

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Table 1. Initial Modiﬁcation of the N-Alkyl Group

a

b

Potency data are reported as the mean and standard deviation of at least two runs. HS = human serum.

alcohol with a cyanomethyl or 2-cyano-2-methylpropanyl group

resulted in a potency loss in the EdU assay (8 and 9 vs 1).

To improve the binding aﬃnity to the MDM2 protein, we

incorporated a sulfonamide moiety to search for additional

interactions with the glycine shelf. As shown in Table 2,

sulfonamide 10 had similar biochemical potency but weaker

cellular potency in the presence of human serum as compared to

1. Increasing the S-alkyl group size from methyl to t-butyl or

cyclopropyl results in a substantial improvement in cellular

potency (11 and 12 vs 10). N-Methylation of sulfonamide 11

was well tolerated (13), while N-methylation of 12 resulted in 2-

fold improvement in the EdU cellular assay (IC₅₀for 14 of 16 nM

vs IC₅₀for 12 of 37 nM). Furthermore, expanding the ethyl

group (Phe19 pocket) attached to the carbon adjacent to the ring

nitrogen to a cyclopropyl or an isopropyl group resulted in the

sulfonamides 15 and 16, which were 2−3-fold more potent than

14 in the EdU assay (IC₅₀for 15 of 5.3 nM and IC₅₀for 16 of 6.5

nM vs IC₅₀for 14 of 16 nM). Unfortunately, the majority of these

sulfonamides proved to be less stable than 1 in human

hepatocytes¹⁶and exhibited moderate to high clearance in rat

(Table 2).¹⁷Reversing the sulfonamide orientation provided

compounds 17, 18, and 19, which maintained potency but

yielded no improvement in metabolic stability.¹⁸

Encouraged by the increased potency imparted by the

sulfonamide functionality, we decided to explore the less polar

and potentially more stable sulfone moiety to maintain the

C

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Table 2. Sulfonamide and Reverse Sulfonamide Piperidinone Derivatives

a

b

Potency data are reported as the mean and standard deviation of at least two runs. HS = human serum.

D

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Table 3. Sulfone Piperidinone Derivatives

a

b

Potency data are reported as the mean and standard deviation of at least two runs. HS = human serum.

favorable interaction with the glycine shelf and improve the

stability in human hepatocytes and high clearance in rat. As

revealed in Table 3, compound 20, containing a one-carbon

tethered sulfone, was noticeably more potent in both the

biochemical assay and the cell-based assay than 1 and 21 which

contains a two-carbon spacer. With the one-carbon tethered

E

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Table 4. Combination of Alkyl Groups at R¹and R²

biochemical potency

cellular potency (SJSA-1)

Rat PK (iv, 0.5 mg/kg)

CL(L/h/kg)/t_1/2(h)

a

Compd

R¹

R²

HTRF IC₅₀(nM)

EdU (10%HS) IC₅₀(nM)

hHep CL_int(μL/min/10⁶cells)

1

1.1 0.5

0.10 0.04

0.10 0.03

0.1 0.03

0.6 0.02

0.5 0.1

72 14

3.0 0.8

8.1 0.04

4.3 1.2

7.1 4.3

7.5 2.3

1.6 0.8

4.8 0.6

6.0 4.1

14 1.8

9.1 2.8

8.8 5.2

2.5 1.5

3.4 0.8

9.5 2.5

110 15

3.4 0.9

4.0 2.5

3.0

11

1.2/3.5

0.47/3.4

n/d

25

31

32

33

34

35

36

24

37

2

t-Bu

i-Pr

Me

Et

CF₃CH₂

Me

26

n/d

7.9

4.6

16

0.62/3.9

0.78/7.5

0.19/18

1.1/3.8

n/d

i-Pr

t-Bu

c-Pr

c-Bu

Et

0.10 0.01

0.1 0.02

0.20 0.03

0.2 0.05

0.6 0.4

12

11

0.85/6.7

2.1/3.8

0.66/2.4

0.78/8.0

0.96/8.7

1.4/3.7

4.5/0.35

n/d

Me

4.1

6.3

20

i-Pr

t-Bu

c-Pr

c-Bu

Et

38

39

40

20

41

42

43

0.4 0.3

0.20 0.03

0.1 0.01

0.30 0.o8

0.9 0.1

20

9.2

n/d

11

Me

i-Pr

c-Pr

0.30 0.08

0.10 0.04

2.7/9.0

0.94/7.7

Me

6.1

a

b

Potency data are reported as the mean and standard deviation of at least two runs. HS = human serum.

sulfone as the key feature required for potent inhibition,

numerous sulfone derivatives were synthesized to deﬁne the

optimum alkyl group in the sulfone side chain. In general,

sulfones were more potent than sulfonamides as illustrated in

Table 3. Many of the sulfone derivatives 22−30 were typically

10−20-fold more potent than 1 in both the p21 induction and

the EdU proliferation assays. The t-butyl sulfone 25 was the most

potent compound among the piperidinone sulfone derivatives,

with an IC₅₀of 3.0 nM in the EdU assay.

On the basis of the observations exempliﬁed in Tables 2 and 3,

we explored combinations of alkyl groups at R¹and R²

(compounds 31−43, Table 4) in order to improve pharmaco-

kinetic properties while maintaining favorable potency. Im-

portantly, many of those compounds not only maintain favorable

cellular potency but also display low intrinsic clearance in human

hepatocytes and in vivo clearance in rat. To discover a molecule

suitable for preclinical development, the highly potent sulfone

inhibitors in Table 4 were proﬁled in the CYP inhibition, time

dependent inhibition (TDI) of CYP 3A4, and PXR induction

assays. Compounds 2, 25, and 43 were found to have minimal

liabilities in these assays. However, both compound 25 and 43

had moderate to high clearance and poor oral exposure in the

cynomolgus monkey. Gratifyingly, compound 2 exhibited

excellent pharmacokinetic properties in cynomolgus monkey

(CL = 0.51 L/h/kg, %F = 51), allowing it to be selected for

further development.

the proliferation of HCT116 p53^wtand p53^−/−tumor cells in

vitro¹⁹as previously reported for compound 1.¹³HCT116 p53^wt

and p53^−/−tumor cells were incubated with 2 for 16 h, and the

percentage of cell-growth inhibition was measured in a BrdU

proliferation assay. The BrdU selectivity data demonstrated that

2 has remarkable p53-dependent cellular inhibition (Figure 4). It

displayed substantial growth inhibition of wild-type p53 cells

(IC₅₀= 10 nM) and no growth inhibition of p53-deﬁcient cells

(IC₅₀> 25 μM).

In the pharamacodynamic assay²⁰with MDM2-ampliﬁed

SJSA-1 osteosarcoma tumor cells, compound 2 demonstrated

The dissociation constant (K_D) of 2 was measured as 0.045

nM in a surface plasmon resonance (SPR) spectroscopy binding

assay.¹⁴Compound 2 was an extremely potent MDM2 inhibitor

in all assays, especially in the EdU proliferation assay (IC₅₀= 9.1

nM). The selectivity of 2 was evaluated by examining its eﬀect on

Figure 4. Cell activity of 2 is p53 dependent. In HCT116 p53^wtand

p53^−/−cells, the percentage of BrdU positive cells was measured 16 h

post compound treatment by ﬂow cytometry. DMSO control was

designated as 0% inhibition.

F

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signiﬁcant time and concentration dependent p21 induction

relative to the vehicle control, consistent with observations

previously reported for compound 1.¹³A 30-fold induction of

p21 mRNA was observed 4 h after the last dose after QD

adminstration for 4 days at 50 mg/kg (Figure 5). This data

indicated that compound 2 achieves an on-target activation of the

p53 pathway.

crystallography to a resolution of 1.7 Å (Figure 7).²²25 occupies

the three critical binding pockets as was seen with the earlier

Figure 5. Pharmacodynamic study: treatment with 2 caused time and

concentration dependent induction of p21 mRNA in SJSA-1 tumor

xenografts. *p < 0.05. Female athymic nude mice were implanted

subcutaneously with 5 × 10⁶SJSA-1 cells. When tumors reached ∼175

mm³, 50 mg/kg of 2 or vehicle was administered orally once daily (QD)

for 4 days. Mice were sacriﬁced on day 4 at 1, 2, 4, 8, and 24 h postdose

(n = 5/group). Tumors were immediately removed and snap-frozen.

p21 mRNA levels were measured by quantitative RT-PCR. Tumors

treated with vehicle served as a negative control and indicated the

baseline p21 mRNA level. Data are represented as mean p21 fold

induction over vehicle and error bars represent SEM of data from ﬁve

mice. Concentrations in plasma were analyzed by LC/MS/MS.

Figure 7. Cocrystal structure of 25 bound to human MDM2 (17−111)

at 1.7 Å resolution. White labels indicate positions normally occupied by

key p53 residues. H96 and G58 are labeled in yellow. Cocrystallized

water molecules are shown in red. PDB code: 4OAS.

structures with 1 and other piperidinone compounds.¹³The C5

aryl group ﬁlls the Leu26 _(p53)pocket and engages in a face-to-

face π-stacking interaction with H96, while the C6 aryl group

occupies the Trp23

directed into the Phe19

constraint induced by the α-substitution. Additionally, the

carboxylate of 25 interacts with the imidazole on the H96 side

chain of MDM2.

binding cavity. The ethyl group is

(p53)

In the SJSA-1 osteosarcoma xenograft model, treatment with 2

signiﬁcantly inhibited tumor growth at each dose compared to

the vehicle with an ED₅₀of 9.1 mg/kg (Figure 6). Unbound EC₅₀

associated with ED₅₀was calculated as 2.8 nM.²¹The 60 mg/kg

QD dose of 2 caused complete tumor regression in 10 of 12 mice.

While cocrystallization attempts of 2 with human MDM2

protein unfortunately proved unsuccessful, the cocrystal

structure of 25, an analogue of 2, was obtained by X-ray

pocket by the conformational

(p53)

The lactam side chain plays an important role in the enhanced

potency of 25, as illustrated in the bound structure. The sulfone

moiety properly projects the t-butyl group to the glycine shelf

region to maximize the hydrophobic contact while the ethyl

moiety binds to the Phe19_(p53)pocket. In addition, the sulfone

function is situated ca. 3.5 Å from the α-carbon of G58,

suggesting a CH···O type interaction with this residue.²³

To conﬁrm the absolute and relative stereochemical assign-

ments of 2, the single crystal structure was obtained, and the unit

cell containing 2 and ethanol is shown in Figure 8 with a

resolution of 0.84 Å.²⁴Superimposition of 2 onto the cocrystal

structure of 25 (Figure 9) illustrates that the binding

conformation of 25 is essentially identical to the conformation

of 2 crystallized in the absence of protein. The sulfone side chain

is bound in the extended low energy conformation. This

observation conﬁrms and validates our previously reported

eﬀorts to stabilize the conformation of both the core and

substituents in this piperidinone class of inhibitors,¹³predispos-

ing the inhibitor for optimal MDM2 binding with minimal

conformational cost.

Figure 6. Treatment with 2 inhibited the growth of SJSA-1 tumors in

vivo. * p < 0.05. SJSA-1 cells (5 × 10⁶) were implanted subcutaneously

into female athymic nude mice. Treatment with vehicle or 2 at 7.5, 15,

30, or 60 mg/kg QD by oral gavage began on day 11 when tumors had

reached ∼200 mm³(n = 12/group). Tumor sizes were measured twice

per week. Data represent mean tumor volumes, and the error bars

represent SEM of data from 12 mice.

CHEMISTRY

■

The synthesis of alcohol and nitrile derivatives (3−9) is

described in Schemes 1−3. Aldehyde intermediate 44 was

obtained via an asymmetric synthetic route reported previ-

G

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a

Scheme 1

Figure 8. Single crystal structure of 2 with ethanol highlighting an

intermolecular hydrogen bond between the two components.

a

Reagents and conditions: (a) CH₃MgBr, 25 °C, 43−100%; (b)

(COCl)₂, TEA, DMSO, −60 °C, 100%; (c) NaIO₄, RuCl₃, CH₃CN/

CCl₄/H₂O, 22−78%; (d) KHMDS, Ph₃P⁺CH₂OCH₃Cl⁻; 3 N aq HCl,

90%; (e) Dess−Martin periodinane, 79−93%; (f) NaBH₄, 91%.

Sulfonamides 10−16 were synthesized as shown in Scheme 4.

Reductive amination of aldehyde 44 with PMBNH₂, followed by

removal of the PMB group aﬀorded primary amine 54.

Sulfonylation of the amine in 54 with MsCl and oxidative

cleavage of the alkene gave methyl sulfonamide 10. Alternatively,

Mitsunobu reaction of primary alcohols 55−57 with various

sulfonamides (R¹SO₂NHR²) in the presence of CMBP²⁶

provided compounds 11−16. The synthesis of primary alcohol

55 was previously reported.²⁵Alcohols 56−57 and 61−64 were

prepared according to the procedures described for the synthesis

of 55, replacing (S)-(+)-2-amino-1-butanol with the appropriate

amino alcohols.²⁷

Scheme 5 describes the synthesis of the reverse sulfonamides

17−19. Protection of the primary alcohol 56, followed by

conversion of the alkene to the methyl ester, gave 58. Mitsunobu

reaction of 58 with benzyl mercaptan in the presence of CMBP

provided benzyl sulﬁde 59. Transformation of benzyl sulﬁde 59

into a sulfonyl chloride in the presence of iodosobenzene and

HCl²⁸was followed by the reaction of this sulfonyl chloride with

the various amines to aﬀord compounds 17−19.

Figure 9. Single crystal structure of 2 aligned with cocrystal structure of

25 with MDM2 (PDB: 4OAS). 2 is shown in yellow and 25 is shown in

blue.

ously.²⁵Grignard reaction of 44 with CH₃MgBr, followed by a

Swern oxidation, aﬀorded ketone 45. Grignard reaction of 45

with CH₃MgBr and oxidative cleavage of the alkene in 46 with

catalytic ruthenium tetraoxide provided acid 3. In a similar

manner, conversion of 44 to the homologated aldehyde 47 via

vinyl ether formation was followed by sequential transformations

to yield alcohols 4, 6, and 7 (Scheme 1).

Similarly, the two-carbon homologated tertiary alcohol 5 was

synthesized from the aldehyde 47 via methyl ketone 49 (Scheme

2). Conversion of 47 into the primary amide 50, followed by

dehydration under (CF₃CO)₂O and TEA conditions, provided

nitrile 8 (Scheme 2).

The synthesis of α,α-dimethyl cyano compound 9 started with

oxidation of the alkene of 51¹³to a 1,2-diol. Protection of the diol

as its acetonide, followed by conversion of the ester to the

aldehyde, gave 52. Finally, compound 9 was obtained by a

Horner−Wadsworth−Emmons reaction of 52 with diethyl

cyanomethylphosphonate, followed by reduction of the alkene,

methylation of the cyanide, deprotection of the acetonide group,

and oxidation of the diol (Scheme 3).

Sulfone derivatives (2, 20, and 22−43), having a one carbon

spacer between the sulfone moiety and the methine adjacent to

the ring nitrogen, were synthesized either via a Mitsunobu

reaction of primary alcohols 55−57 and 61−64 with various

thiols in the presence of CMBP (Scheme 6) or via a nucleophilic

ring-opening reaction of the bicyclic iminium ether²⁵derived

H

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a

Scheme 2

Scheme 4

a

Reagents and conditions: (a) KHMDS, Ph₃P⁺CH₂OCH₃Cl⁻; 3 N aq

HCl, 76%; (b) MeMgBr, 25 °C, 98−100%; (c) Dess−Martin

periodinane, 97%; (d) NaIO₄, RuCl₃, CH₃CN/CCl₄/H₂O, 55−75%;

(e) NaH₂PO₄, NaClO₂, 27%; (f) isobutyl chloroformate, N-

methylmorpholine, NH₄OH, 98%; (g) (CF₃CO)₂O, TEA, 56%.

a

Reagents and conditions: (a) PMBNH₂, NaBH(OAc)₃, DCE, 100%;

(b) ceric ammonium nitrate, H₂O/CH₃CN, 53%; (c) MsCl, pyridine,

47%; (d) NaIO₄, RuCl₃, CH₃CN/CCl₄/H₂O, 32−62%; (e)

R¹SO₂NHR², CMBP, toluene, 50−90%.

a

Scheme 3

a

Scheme 5

a

Reagents and conditions: (a) OsO₄, NMO, 66%; (b) 2,2-

dimethoxypropane, TsOH monohydrate, 91%; (c) LiBH₄, 100%; (d)

Dess−Martin periodinane, 85%; (e) diethyl cyanomethylphosphonate,

DMPU, NaH, 96%; (f) H₂, Pd/C, 58%; (g) CH₃I, LiHMDS, 63%; (h)

3 N aq HCl, 100%; (i) NaIO₄, RuCl₃, CH₃CN/CCl₄/H₂O, 65%.

a

Reagents and conditions: (a) TBDPSCl, imidazole, DMF, 92%; (b)

NaIO₄, RuCl₃, CH₃CN/CCl₄/H₂O, 2 M (trimethylsilyl)diazomethane

in Et₂O, 1 M TBAF in THF, 53%; (c) PhCH₂SH, CMBP, toluene,

81%; (d) iodosobenzene, HCl, R₁R₂NH, 54−79%; (e) LiOH, 56−

71%.

from the primary alcohols 55−57 and 61−63, with either

thiolates or sulﬁnic acid sodium salts (Scheme 7).

Finally, compound 21, a sulfone derivative containing a two-

carbon spacer, was prepared from a Horner−Wadsworth−

Emmons reaction of 52 with diethyl methylsulfonylmethyl-

phosphonate, followed by hydrogenation and removal of the

acetonide group (Scheme 8).

CONCLUSION

■

In summary, research investigation of 1 led to the discovery of

sulfone analogues that feature substantial improvements in

biochemical and cellular potency. The enhanced potency of the

sulfone-derived inhibitors can be rationalized by the favorable

I

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a

Scheme 6

Scheme 7

a

Reagents and conditions: (a) Tf₂O, 2,6-dimethylpyridine, −40 to 0

°C, 100%; (b) method A, R¹SNa, DMF, 82%; method B, R¹S(O)ONa,

AcCN, 37−80%; method C, R¹SH, base, DMF, 27−84%; (c) NaIO₄,

RuCl₃, CH₃CN/CCl₄/H₂O, 37−68%.

a

Scheme 8

a

Reagents and conditions: (a) TMSCH₂SH, CMBP, toluene, 40−

50%; (b) 1 M TBAF in THF, 45−57%; (c) NaIO₄, RuCl₃, CH₃CN/

CCl₄/H₂O, 30−62%; (d) R¹SH, CMBP, toluene, 32−91%.

hydrophobic interaction between the alkyl sulfone and the

glycine shelf, which was observed in the cocrystal structure of 25

with MDM2. Further investigation eﬀorts produced 2, a highly

potent MDM2 inhibitor (SPR K_D= 0.045 nM, EdU IC₅₀= 9.1

nM), with remarkable pharmacokinetic properties and in vivo

antitumor activity in the SJSA-1 osteosarcoma xenograft model

(ED₅₀= 9.1 mg/kg). Compound 2 is being evaluated in human

clinical trials for the treatment of cancer.¹²

EXPERIMENTAL SECTION

■

General Chemistry. All reactions were conducted under an inert gas

atmosphere (nitrogen or argon) using a Teﬂon-coated magnetic stir bar

at the temperature indicated. Commercial reagents and anhydrous

solvents were used without further puriﬁcation. Removal of solvents was

conducted by using a rotary evaporator, and residual solvent was

removed from nonvolatile compounds using a vacuum manifold

maintained at approximately 1 Torr. All yields reported are isolated

yields. Preparative reversed-phase high pressure liquid chromatography

(RP-HPLC) was performed using an Agilent 1100 series HPLC and

Phenomenex Gemini C18 column (5 μm, 100 mm × 30 mm i.d.),

eluting with a binary solvent system A and B using a gradient elusion (A,

H₂O with 0.1% triﬂuoroacetic acid (TFA); B, CH₃CN with 0.1% TFA)

with UV detection at 220 nm. All bioassayed compounds were puriﬁed

to ≥95% purity as determined by an Agilent 1100 series HPLC with UV

detection at 220 nm using the following method: Zorbax SB-C8 column

(3.5 μm, 150 mm × 4.6 mm i.d.); mobile phase, A = H₂O with 0.1%

TFA, B = CH₃CN with 0.1% TFA; gradient, 5−95% B (0.0−15.0 min);

ﬂow rate, 1.5 mL/min. Low-resolution mass spectral (MS) data were

a

Reagents and conditions: (a) n-BuLi, diethyl methylsulfonylmethyl-

phosphonate, 70%; (b) H₂, Pd/C, 83%; (c) TFA, THF, 64%; (d)

NaIO₄, RuCl₃, CH₃CN/CCl₄/H₂O, 55%.

determined on an Agilent 1100 series LCMS with UV detection at 254

nm and a low resolution electrospray mode (ESI). High-resolution mass

spectra (HRMS) were obtained on an Agilent 6510 Q-TOF MS with a

Agilent 1200 LC on the front end. ¹H NMR spectra were obtained on a

Bruker Avance III 500 (500 MHz) or Bruker Avance II 400 (400 MHz)

spectrometer. Chemical shifts (δ) are reported in parts per million

(ppm) relative to residual undeuterated solvent as an internal reference.

The following abbreviations were used to explain the multiplicities: s =

J

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single; d = doublet, t = triplet, q = quartet, dd = doublet of doublets, dt =

doublet of triplets, m = multiplet, br = broad.

After the mixture was stirred at 0 °C for 30 min, a solution of 44 (564

mg, 1.27 mmol) in THF (10 mL) was added at 0 °C dropwise. After

being stirred at 25 °C for 12 h, the reaction was quenched (satd NH₄Cl),

extracted (2 × EtOAc), and washed (brine). The combined organic

layers were dried (Na₂SO₄) and concentrated under reduced pressure.

The residue was puriﬁed by ﬂash column chromatography (SiO₂, 15%

and 20% EtOAc/hexanes) to provide a crude vinyl ether.

To a solution of the crude product above in acetonitrile (7.8 mL) was

added 3 N hydrochloric acid (4.4 mL, 13.0 mmol), and the resulting

solution was stirred at 25 °C for 1.5 h. The reaction was extracted (2 ×

EtOAc) and washed (brine). The combined organic layers were dried

(Na₂SO₄) and concentrated under reduced pressure to provide 47 (2.09

g, 90%) as a pale-yellow ﬁlm. ¹H NMR (400 MHz, CDCl₃) δ ppm 9.82

(s, 1 H), 7.24 (d, J = 8.4 Hz, 2 H), 7.08−7.18 (m, 2 H), 7.05 (d, J = 7.8

Hz, 2 H), 6.99 (s, 1 H), 6.80 (d, J = 7.2 Hz, 1 H), 5.76−5.95 (m, 1 H),

5.09−5.24 (m, 2 H), 4.74 (d, J = 10.6 Hz, 1 H), 3.46−3.63 (m, 1 H),

3.06−3.24 (m, 2 H), 2.45−2.69 (m, 3 H), 1.97 (d, J = 19.6 Hz, 1 H),

1.81−1.94 (m, 2 H), 1.33−1.47 (m, 1 H), 1.19 (s, 3 H), 0.46 (t, J = 7.5

Hz, 3 H). MS (ESI) 458.0 [M + H]⁺.

(3S,5R,6S)-3-Allyl-5-(3-chlorophenyl)-6-(4-chlorophenyl)-1-

((3S)-5-hydroxyhexan-3-yl)-3-methylpiperidin-2-one (48). To a

solution of 47 (540 mg, 1.18 mmol) in THF (12 mL) was added methyl

magnesium bromide (1.4 M in toluene, 2.52 mL, 3.53 mmol) at 0 °C.

The reaction was allowed to warm to 25 °C and stirred for 3 h. The

reaction was quenched (satd NH₄Cl), extracted (2 × EtOAc), and

washed (brine). The combined organic layers were dried (Na₂SO₄) and

concentrated under reduced pressure to provide 48 (558 mg, 100%) as a

mixture of diastereomers.

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-

((3S,5R)-5-hydroxyhexan-3-yl)-3-methyl-2-oxopiperidin-3-yl)-

acetic Acid (6) and 2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlor-

ophenyl)-1-((3S,5S)-5-hydroxyhexan-3-yl)-3-methyl-2-oxopi-

peridin-3-yl)acetic Acid (7). 2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-

(4-chlorophenyl)-3-methyl-2-oxo-1-((S)-5-oxohexan-3-yl)piperidin-3-

yl)acetic acid was prepared as a white foam from 48 according to a

similar procedure described for the synthesis of 3. ¹H NMR (400 MHz,

CDCl₃) δ ppm 7.23−7.27 (m, 2 H), 7.06−7.17 (m, 4 H), 7.00 (t, J = 1.8

Hz, 1 H), 6.81 (s, 1 H), 4.92 (d, J = 10.8 Hz, 1 H), 3.54−3.63 (dd, J =

12.0, 12.0 Hz,1 H), 3.07 (m, 3 H), 2.67 (d, J = 15.8 Hz, 1 H), 2.50−2.60

(dd, J = 4.0, 12.0 Hz, 1 H), 2.19 (s, 3 H), 2.12 (t, J = 12.0 Hz, 1 H), 1.97−

2.07 (m, 1 H), 1.88−1.96 (m, 1 H), 1.39 (s, 3 H), 1.21−1.32 (m, 1 H),

0.37 (t, J = 7.5 Hz, 3 H). MS (ESI) 490.0 [M + H]⁺, 488.0 [M − H]⁻.

To a solution of the keto acid above (24.0 mg, 0.049 mmol) in ether

(0.4 mL) and MeOH (0.1 mL) was added sodium borohydride (9.26

mg, 0.245 mmol) at 0 °C. The reaction was allowed to warm to 25 °C.

After being stirred at 25 °C for 1 h, the reaction was quenched (10%

citric acid) and extracted (2 × EtOAc). The combined organic layers

were washed (brine), dried (Na₂SO₄), and concentrated under reduced

pressure. The residue was puriﬁed by RP-HPLC (30−70% A/B,

gradient elution) to provide 6 as the more polar isomer. ¹H NMR (400

MHz, CDCl₃) δ ppm 7.22−7.27 (m, 2 H), 7.07−7.19 (m, 2 H), 6.95−

7.06 (m, 3 H), 6.71 (dt, J = 7.6, 1.6 Hz, 1 H), 4.57 (d, J = 10.2 Hz, 1 H),

3.77−3.89 (m, 1 H), 2.99−3.15 (m, 2 H), 2.69 (d, J = 14.9 Hz, 1 H), 2.23

(t, J = 13.6 Hz, 1 H), 1.81−1.99 (m, 2 H), 1.52−1.64 (m, 1 H), 1.47 (s, 3

H), 1.39 (d, J = 2.0 Hz, 1 H), 1.28 (m, 1 H), 1.19 (d, J = 6.3 Hz, 3 H),

0.60 (t, J = 7.4 Hz, 3 H). HRMS (ESI) m/z 492.1709 [M + H]⁺

(C₂₆H₃₁Cl₂NO₄requires 492.1703). Further elution provided 7 as the

less polar isomer. ¹H NMR (400 MHz, CDCl₃) δ ppm 7.04−7.27 (m, 5

H), 6.91−7.04 (m, 2 H), 6.70 (dt, J = 7.7, 1.5 Hz, 1 H), 4.47 (d, J = 10.4

Hz, 1 H), 3.75 (ddd, J = 12.4, 6.1, 0.7 Hz, 1 H), 3.15−3.27 (m, 1 H), 2.96

(d, J = 14.5 Hz, 1 H), 2.76 (d, J = 14.7 Hz, 1 H), 2.18 (t, J = 13.8 Hz, 1 H),

2.02−2.09 (m, 1 H), 1.59−1.71 (m, 2 H), 1.50 (s, 3 H), 1.26 (dd, J =

16.8, 6.8 Hz, 1 H), 1.08−1.21 (m, 1 H), 0.99−1.07 (m, 3 H), 0.89 (s, br,

3 H). HRMS (ESI) m/z 492.1709 [M + H]⁺(C₂₆H₃₁Cl₂NO₄requires

492.1703).

(3S,5R,6S)-3-Allyl-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-

methyl-1-((S)-2-oxopentan-3-yl)piperidin-2-one (45). To a sol-

ution of 44 (258 mg, 0.581 mmol) in THF (5.8 mL) was added

methylmagnesium bromide (1.4 M in toluene/tetrahydrofuran (75/25),

1.24 mL, 1.74 mmol) at 0 °C. The reaction was allowed to warm to 25

°C. After being stirred at 25 °C for 3 h, the reaction was quenched (satd

NH₄Cl), extracted (3 × EtOAc), and washed (2 × water and 1 × brine).

The combined organic layers were dried (Na₂SO₄) and concentrated

under reduced pressure. The residue was puriﬁed by ﬂash column

chromatography (SiO₂, 30%, 40%, and 50% EtOAc/hexanes) to provide

(3S,5R,6S)-3-allyl-5-(3-chlorophenyl)-6-(4-chlorophenyl)-1-((3S)-2-

hydroxypentan-3-yl)-3-methylpiperidin-2-one (200 mg, 0.434 mmol,

75%) as a mixture of two diastereomers. MS (ESI) 460.0 [M + H]⁺.

To a solution of oxalyl dichloride (78.0 μL, 0.869 mmol) in DCM

(1.5 mL) at −60 °C was added a solution of DMSO (93.0 μL, 1.30

mmol) in DCM (1.5 mL) under nitrogen. After being stirred for 2 min, a

solution of the alcohol above (200 mg, 0.434 mmol) in DCM (1.5 mL)

was added and the resulting solution was stirred for 15 min at −60 °C.

Triethylamine (305 μL, 2.17 mmol) was added to the reaction solution.

After being stirred at 25 °C for 20 min, the reaction was quenched

(water), extracted (2 × EtOAc), washed (2 × brine), and dried

(Na₂SO₄). The organic solution was concentrated under reduced

pressure and the residue was puriﬁed by ﬂash column chromatography

(SiO₂, 20% and 30% EtOAc/hexanes) to provide 45 (200 mg, 0.436

mmol, 100%) as a colorless foam. ¹H NMR (400 MHz, CDCl₃) δ ppm

7.23 (d, J = 8.8 Hz, 2 H), 7.07−7.17 (m, 2 H), 6.91−7.07 (m, 3 H), 6.78

(dt, J = 7.4, 1.6 Hz, 1 H), 5.80−5.94 (m, 1 H), 5.16−5.24 (m, 2 H), 4.52

(d, J = 10.8 Hz, 1 H), 3.32 (ddd, J = 13.6, 10.7, 3.1 Hz, 1 H), 3.11 (dd, J =

7.0, 5.7 Hz, 1 H), 2.62 (d, J = 7.4 Hz, 2 H), 2.20 (t, J = 13.7 Hz, 1 H), 2.15

(s, 3 H), 2.05−2.13 (m, 1 H), 1.98 (dd, J = 13.8, 3.2 Hz, 1 H), 1.79 (ddd,

J = 14.0, 7.9, 5.7 Hz, 1 H), 1.27 (s, 3 H), 0.64 (t, J = 7.5 Hz, 3 H). MS

(ESI) 458.0 [M + H]⁺.

(3S,5R,6S)-3-Allyl-5-(3-chlorophenyl)-6-(4-chlorophenyl)-1-

((S)-2-hydroxy-2-methylpentan-3-yl)-3-methylpiperidin-2-one

(46). A solution of methylmagnesium bromide (1.4 M in toluene, 0.623

mL, 0.873 mmol) was added to a solution of 45 (0.100 g, 0.218 mmol) in

THF (4 mL) was added at 0 °C. The reaction was allowed to warm to 25

°C. After being stirred at 25 °C for 4 h, an additional methylmagnesium

bromide (0.156 mL, 0.218 mmol) was added and the reaction was

stirred for another 2 h. The reaction was quenched (satd NH₄Cl), and

extracted (EtOAc). The combined organic layers were washed (brine),

dried (MgSO₄), and ﬁltered, and the ﬁltrate was concentrated under

reduced pressure. The residue was puriﬁed by ﬂash column

chromatography (SiO₂, 40% EtOAc/hexanes) to provide 46 (44.0

mg, 43%) as a light-yellow oil. MS (ESI) 474.2 [M + H]⁺.

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

2-hydroxy-2-methylpentan-3-yl)-3-methyl-2-oxopiperidin-3-

yl)acetic acid (3). To a rapidly stirring solution of 46 (0.044 g, 0.093

mmol) in H₂O/CCl₄/MeCN (0.9 mL:0.45 mL:0.45 mL) was added

sodium periodate (0.079 g, 0.371 mmol), followed by ruthenium(III)

chloride hydrate (2.09 mg, 9.27 μmol). After being stirred vigorously for

36 h, the reaction was acidiﬁed (10% citric acid) and diluted (DCM).

The insoluble material was ﬁltered through Celite to remove. The ﬁltrate

was extracted (2 × DCM) and the combined organic layers were washed

(brine), dried (MgSO₄), and concentrated under reduced pressure. The

residue was puriﬁed by ﬂash column chromatography (SiO₂, 40%

1

EtOAc/hexanes) to give 3 (10.0 mg, 22%) as colorless oil. H NMR

(400 MHz, CDCl₃) δ ppm 7.21−7.38 (m, 2 H), 7.01−7.21 (m, 4 H),

6.95 (m, 1 H), 6.73 (dt, J = 1.52, 7.53 Hz, 1 H), 4.40 (d, J = 10.56 Hz, 1

H), 3.24−3.37 (m, 1 H), 2.78−2.95 (m, 2 H), 2.44−2.55 (m, 1 H),

2.10−2.31 (m, 3 H), 1.68 (m, 1 H), 1.50 (s, 3 H), 1.35 (s, 3 H), 1.10 (s, 3

H), 0.37 (t, J = 7.63 Hz, 3 H). HRMS (ESI) m/z 492.1708 [M + H]⁺

(C₂₆H₃₁Cl₂NO₄requires 492.1703).

(S)-3-((3S,5R,6S)-3-Allyl-5-(3-chlorophenyl)-6-(4-chlorophen-

yl)-3-methyl-2-oxopiperidin-1-yl)pentanal (47). To a solution of

dried (methoxymethyl)triphenylphosphonium chloride (1.96 g, 5.71

mmol) in THF (10 mL) was added KHMDS (0.5 M in toluene, 10.2

mL, 5.08 mmol) at −78 °C. The color of the solution turned blood red.

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

5-hydroxy-5-methylhexan-3-yl)-3-methyl-2-oxopiperidin-3-

yl)acetic Acid (4). To a solution of 48 (100 mg, 0.211 mmol) and water

(5.70 μL, 0.316 mmol) in DCM (2.3 mL) was added Dess−Martin

periodinane (134 mg, 0.316 mmol) at 25 °C. After being stirred at 25 °C

K

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for 40 min, the reaction was quenched (1 M Na₂S₂O_3,2 mL), extracted

(2 × DCM), and washed (2 × satd NaHCO₃and brine). The combined

organic layers were dried (Na₂SO₄) and concentrated under reduced

pressure. The residue was puriﬁed by ﬂash column chromatography

(SiO₂, 25% and 30% EtOAc/hexanes) to aﬀord (3S,5R,6S)-3-allyl-5-(3-

chlorophenyl)-6-(4-chlorophenyl)-3-methyl-1-((S)-5-oxohexan-3-yl)-

piperidin-2-one (93.0 mg, 93%) as a colorless ﬁlm. ¹H NMR (400 MHz,

CDCl₃) δ 7.23 (d, J = 8.6 Hz, 2 H), 7.03−7.17 (m, 4 H), 6.93−7.01 (m, 1

H), 6.84 (s, 1 H), 5.78−5.94 (m, 1 H), 5.18 (d, J = 3.9 Hz, 1 H), 5.15 (s, 1

H), 4.80 (d, J = 10.8 Hz, 1 H), 3.55−3.68 (m, 1 H), 2.98−3.20 (m, 2 H),

2.58 (s, 3 H), 2.18 (s, 3 H), 1.93−2.05 (m, 1 H), 1.83−1.92 (m, 2 H),

1.28−1.37 (m, 1 H), 1.18 (s, 3 H), 0.40 (t, J = 7.5 Hz, 3 H). MS (ESI)

472.1 [M + H]⁺.

Compound 4 was prepared as a white foam from ((3S,5R,6S)-3-allyl-

5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-methyl-1-((S)-5-oxohexan-

3-yl)piperidin-2-one according to a similar procedure described for the

synthesis of 3. ¹H NMR (400 MHz, CDCl₃) δ ppm 7.25 (d, J = 7.8 Hz, 2

H), 6.98−7.18 (m, 4 H), 6.95 (t, J = 1.8 Hz, 1 H), 6.70 (d, J = 7.6 Hz, 1

H), 4.90−5.38 (m, 2 H), 4.67−4.81 (m, 1 H), 3.51 (s, 1 H), 2.98−3.13

(m, 2 H), 2.70 (d, J = 15.1 Hz, 1 H), 2.19 (t, J = 13.8 Hz, 1 H), 1.93 (d, J =

13.3 Hz, 2 H), 1.48 (s, 4 H), 1.16−1.28 (m, 7 H), 0.53 (s, br, 3 H).

HRMS (ESI) m/z 506.1862 [M + H]⁺(C₂₇H₃₃Cl₂NO₄requires

506.1859).

7.08 (m, 2 H), 6.93 (s, 1 H), 6.84 (d, J = 8.0 Hz, 1 H), 5.88 (m, 1 H),

5.15−5.25 (m, 2 H), 4.65 (d, J = 8.0 Hz, 1 H), 3.45 (dd, J = 8.0, 12.0 Hz,

1 H), 3.19 (m, 1 H), 2.94 (m, 1 H), 2.63 (m, 2 H), 2.35 (dd, J = 4.0, 12.0

Hz, 1 H), 2.22 (t, J = 12 Hz, 1 H), 1.95−2.15 (m, 1 H), 1.95 (dd, J = 4.0,

12 Hz, 1 H), 1.55 (m, 1 H), 1.32 (s, 3 H), 0.50 (t, J = 8.0 Hz, 3 H).

Compound 8 was prepared from the nitrile above according to a

similar procedure described for the synthesis of 3. ¹H NMR (400 MHz,

CDCl₃) δ ppm 7.20−7.28 (m, 2 H), 7.00−7.18 (m, 4 H), 6.95 (s, 1 H),

6.82 (d, J = 8.0 Hz, 1 H), 4.65 (d, J = 8.0 Hz, 1 H), 3.42 (dd, J = 8.0, 12.0

Hz, 1 H), 3.20 (m, 1 H), 2.90−3.05 (m, 3 H), 2.42 (dd, J = 4.0, 12.0 Hz, 1

H), 2.32 (t, J = 12 Hz, 1 H), 2.00−2.15 (m, 1 H), 1.95 (dd, J = 4.0, 12 Hz,

1 H), 1.52 (m, 1 H), 1.52 (s, 3 H), 0.48 (t, J = 8.0 Hz, 3 H). HRMS (ESI)

m/z 473.1403 [M + H]⁺(C₂₅H₂₆Cl₂N₂O₃requires 473.1393).

(2S)-2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-3-

((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-3-methyl-2-oxopi-

peridin-1-yl)butanal (52). To a solution of 51 (1.06 g, 2.23 mmol)

and 4-methylmorpholine 4-oxide (393 mg, 3.35 mmol) in DCM (15.8

mL) was added osmium(VIII) oxide polymer-bound, 1% DVB (56.8

mg, 2.23 μmol). After being vigorously stirred at 25 °C for 48 h,

additional osmium(VIII) oxide polymer-bound, 1% DVB (56.8 mg, 2.23

μmol) was added and the resulting solution was vigorously stirred at 25

°C for another 48 h. The resin was ﬁltered and washed (DCM). The

combined organic layers were washed (brine), dried (Na₂SO₄), and

concentrated under reduced pressure. The residue was puriﬁed by ﬂash

column chromatography (SiO₂, 53% and 63% EtOAc/hexanes) to

provide the diol (746 mg, 66%). MS (ESI) 508.0 [M + H]⁺.

To a solution of the product above (749 mg, 1.47 mmol) in DCM

(8.2 mL) was added p-toluenesulfonic acid monohydrate (14.0 mg,

0.074 mmol) and 2,2-dimethoxypropane (8.15 mL, 66.3 mmol). After

being stirred at 25 °C for 3 h, the reaction mixture was concentrated

under reduced pressure and the residue was dissolved (EtOAc and satd

NaHCO₃) and extracted (3 × EtOAc). The combined organic layers

were washed (brine), dried (Na₂SO₄), and concentrated under reduced

pressure. The residue was puriﬁed by ﬂash column chromatography

(SiO₂, 27% and 37% EtOAc/hexanes) to provide the protected diol as a

colorless ﬁlm (732 mg, 91%). MS (ESI) 548.0 [M + H]⁺.

To a solution of the crude product above (734 mg, 1.34 mmol) in

ether (12 mL) was added lithium borohydride (58.3 mg, 2.68 mmol) at

0 °C. After being stirred at 0 °C for 30 min, the reaction was quenched

(ice cold 10% citric acid), extracted (2 × EtOAc), and washed (brine).

The combined organic layers were dried (Na₂SO₄) and concentrated

under reduced pressure to provide (3R,5R,6S)-5-(3-chlorophenyl)-6-

(4-chlorophenyl)-3-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1-((S)-

1-hydroxybutan-2-yl)-3-methylpiperidin-2-one as a colorless ﬁlm (693

mg, 100%). MS (ESI) 518.0 [M + H]⁺. The crude product was used in

the next step without further puriﬁcation.

Compound 52 was prepared from (3R,5R,6S)-5-(3-chlorophenyl)-6-

(4-chlorophenyl)-3-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1-((S)-

1-hydroxybutan-2-yl)-3-methylpiperidin-2-one according to a similar

procedure described for the synthesis of 49. MS (ESI) 518.0 [M + H]⁺.

(4S)-4-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-3-

((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-3-methyl-2-oxopi-

peridin-1-yl)hexanenitrile (53). To a solution of diethyl cyanome-

thylphosphonate (126 μL, 0.799 mmol) and DMPU (480 μL, 3.99

mmol) in THF (1.3 mL) was added 60% sodium hydride in mineral oil

(24.0 mg, 0.599 mmol) at 0 °C. The mixture was stirred for 30 min and

then treated with a solution of 52 (207 mg, 0.399 mmol) in THF (1.33

mL). After being stirred for 4 h, the reaction was quenched (water),

extracted (2 × EtOAc), and washed (brine). The combined organic

layers were dried (Na₂SO₄) and concentrated under reduced pressure.

The residue was puriﬁed by ﬂash column chromatography (SiO₂, 30−

40% EtOAc/hexanes, gradient elution) to provide the conjugated nitrile

(218 mg, 96%) as a colorless liquid. MS (ESI) 541.2 [M + H]⁺.

To a solution of the product above (208 mg, 0.384 mmol) in EtOH

(13 mL) was added 10% palladium on activated carbon (40.9 mg, 0.038

mmol). The reaction mixture was stirred under a hydrogen balloon.

After being stirred at 25 °C for 1.5 h, the catalyst was ﬁltered using a

short plug of silica gel and washed (EtOAc). The combined organic

solutions were concentrated under reduced pressure. The residue was

puriﬁed by ﬂash column chromatography (SiO₂, 35% and 40% EtOAc/

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

6-hydroxy-6-methylheptan-3-yl)-3-methyl-2-oxopiperidin-3-

yl)acetic Acid (5). Compound 5 was prepared as a white solid from 47

1

according to a similar procedure described for the synthesis of 4. H

NMR (400 MHz, CDCl₃) δ ppm 7.25 (d, J = 8.4 Hz, 2 H), 7.16 (dd, J =

1.9, 1.1 Hz, 1 H), 7.11 (d, J = 7.6 Hz, 1 H), 6.94 (t, J = 1.8 Hz, 3 H), 6.70

(d, J = 7.6 Hz, 1 H), 5.01−5.25 (m, 2 H), 4.37 (d, J = 10.4 Hz, 1 H), 3.06

(d, J = 15.3 Hz, 2 H), 2.93−3.03 (m, 1 H), 2.71 (d, J = 15.3 Hz, 1 H),

2.20 (s, 1 H), 2.02 (s, 1 H), 1.78−1.97 (m, 2 H), 1.37−1.56 (m, 6 H),

1.22 (d, J = 5.5 Hz, 6 H), 0.55 (t, J = 7.5 Hz, 3 H). HRMS (ESI) m/z

520.2024 [M + H]⁺(C₂₈H₃₅Cl₂NO₄requires 520.2016).

(S)-3-((3S,5R,6S)-3-Allyl-5-(3-chlorophenyl)-6-(4-chlorophen-

yl)-3-methyl-2-oxopiperidin-1-yl)pentanamide (50). To a sol-

ution of 47 (147 mg, 0.321 mmol) in 2-methylpropan-2-ol/2-methyl-2-

butene (3: 1, 4.4 mL) was added a solution of sodium chlorite (290 mg,

3.21 mmol) and dihydrogen sodium phosphate (17.3 μL, 0.289 mmol)

in water (1.6 mL). After being stirred at rt for 20 min, the reaction was

quenched (satd NH4Cl), diluted (EtOAc), washed (brine), dried

(MgSO₄), and concentrated to give (S)-3-((3S,5R,6S)-3-allyl-5-(3-

chlorophenyl)-6-(4-chlorophenyl)-3-methyl-2-oxopiperidin-1-yl)-

1

pentanoic acid (41.0 mg, 27%). H NMR (400 MHz, CDCl₃) δ ppm

7.25 (m, 2 H), 7.00−7.20 (m, 4 H), 6.95 (s, 1 H), 6.75 (d, J = 8.0 Hz, 1

H), 5.85 (m, 1 H), 5.15−5.25 (m, 2 H), 4.65 (d, J = 8.0 Hz, 1 H), 3.35

(m, 1 H), 3.10−3.25 (m, 2 H), 2.63 (m, 2 H), 2.50 (dd, J = 4.0, 12.0 Hz, 1

H), 1.85−2.10 (m, 3 H), 1.45 (m, 1 H), 1.25 (s, 3 H), 0.50 (t, J = 8.0 Hz,

3 H).

To a solution of the acid above (40.0 mg, 0.084 mmol) and 1-

methylmorpholine (13.0 μL, 0.118 mmol) in THF (0.5 mL) was added

isobutyl chlorocarbonate (13.1 μL, 0.101 mmol) followed by aqueous

ammonium hydroxide (28%, 11.2 mL, 0.169 mmol) at 0 °C. After being

stirred at 0 °C for 30 min, the reaction was quenched (satd NH₄Cl),

diluted (EtOAc), washed (brine), dried (MgSO₄), and concentrated.

The residue was puriﬁed by ﬂash column chromatography (SiO₂, 30−

50% EtOAc/hexanes, gradient elution) to give 50 (39.0 mg, 98%). MS

(ESI) 473.2 [M + H]⁺.

(S)-3-((3S,5R,6S)-3-Allyl-5-(3-chlorophenyl)-6-(4-chlorophen-

yl)-3-methyl-2-oxopiperidin-1-yl)pentanenitrile (8). A solution of

50 (39.0 mg, 0.082 mmol) and triethylamine (57.3 μL, 0.412 mmol) in

THF (1.2 mL) was treated with triﬂuoroacetic anhydride (28.8 μL,

0.206 mmol) at 0 °C. After being stirred at 0 °C for 1.5 h, the reaction

was warmed to 25 °C and stirred for another 1 h. The reaction was

quenched (satd NH₄Cl), extracted (EtOAc), washed (brine), dried

(MgSO₄), and concentrated. The residue was puriﬁed by ﬂash column

chromatography (SiO₂, 20% EtOAc/hexanes) to provide (S)-3-

((3S,5R,6S)-3-allyl-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-methyl-

2-oxopiperidin-1-yl)pentanenitrile (21.0 mg, 56%). ¹H NMR (400

MHz, CDCl₃) δ ppm 7.20−7.28 (m, 2 H), 7.10−7.20 (m, 2 H), 6.95−

L

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hexanes) to provide 53 (120 mg, 58%) as a colorless ﬁlm. MS (ESI)

543.1 [M + H]⁺.

(Na₂SO₄), and concentrated under reduced pressure to give the free

amine. To a solution of the amine above in DMF (0.34 mL) was added

methanesulfonyl chloride (53.0 μL, 0.680 mmol) and pyridine (66.0 μL,

0.820 mmol) successively at 0 °C. After being stirred at 25 °C for 12 h,

the reaction was acidiﬁed (10% citric acid) and extracted (2 × EtOAc)

and washed (brine). The combined organic layers were dried (Na₂SO₄)

and concentrated under reduced pressure. The residue was puriﬁed by

RP-HPLC (45−80% A/B, gradient elution) to provide N-((S)-2-

((3S,5R,6S)-3-allyl-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-methyl-

2-oxopiperidin-1-yl)butyl)methanesulfonamide (33.7 mg, 47%) as a

white powder. ¹H NMR (400 MHz, CDCl₃) δ ppm 7.23 (d, J = 8.4 Hz, 2

H), 7.10−7.19 (m, 2 H), 7.02 (d, J = 7.0 Hz, 2 H), 6.93−6.98 (m, 1 H),

6.85 (dd, J = 6.7, 1.7 Hz, 1 H), 5.73−5.99 (m, 1 H), 5.12−5.25 (m, 2 H),

4.84−4.97 (m, 1 H), 4.71 (d, J = 10.6 Hz, 1 H), 3.53−3.68 (m, 1 H),

3.05−3.24 (m, 3 H), 2.98 (s, 3 H), 2.62 (d, J = 7.2 Hz, 2 H), 2.09−2.24

(m, 1 H), 1.90 (dd, J = 13.7, 3.1 Hz, 2 H), 1.44−1.62 (m, 1 H), 1.27 (s, 3

H), 0.58 (t, J = 7.5 Hz, 3 H). MS (ESI) 523.0 [M + H]⁺.

Compound 10 was prepared from N-((S)-2-((3S,5R,6S)-3-allyl-5-(3-

chlorophenyl)-6-(4-chlorophenyl)-3-methyl-2-oxopiperidin-1-yl)-

butyl)methanesulfonamide according to a similar procedure describled

for the synthesis of 3. ¹H NMR (400 MHz, CDCl₃) δ ppm 7.25 (d, J =

8.2 Hz, 2 H), 7.10−7.18 (m, 2 H), 7.00−7.10 (m, 2 H), 6.97 (s, 1 H),

6.83 (d, J = 7.2 Hz, 1 H), 4.87−4.97 (m, 1 H), 4.74 (d, J = 10.4 Hz, 1 H),

3.44−3.65 (m, 1 H), 3.10−3.33 (m, 2 H), 3.02−3.09 (m, 1 H), 2.99 (s, 3

H), 2.96 (m, 1 H), 2.77 (d, J = 12.0 Hz, 1 H), 2.36 (d, J = 12.0 Hz, 1 H),

1.94−2.05 (m, 1 H), 1.77−1.92 (m, 1 H), 1.52−1.59 (m, 1 H), 1.50 (s, 3

H), 0.58 (t, J = 7.3 Hz, 3 H). HRMS (ESI) m/z 541.1333 [M + H]⁺

(C₂₅H₃₀Cl₂N₂O₅S requires 541.1325).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-(1,1-dimethylethylsulfonamido)butan-2-yl)-3-methyl-2-oxo-

piperidin-3-yl)acetic Acid (11). 55 (203 mg, 0.454 mmol) and 2-

methylpropane-2-sulfonamide (130 mg, 0.948 mmol) were dissolved in

anhydrous toluene (4.5 mL). Cyanomethylenetributylphosphorane

(451 mg, 1.68 mmol) was transferred to the reaction vessel via syringe.

The reaction mixture was stirred 40 °C in a preheated oil bath, and an

additional 2-methylpropane-2-sulfonamide (130 mg, 0.948 mmol) was

added. After being stirred at 40 °C for 48 h, the mixture was partitioned

between ethyl acetate and satd ammonium chloride. The aqueous phase

was extracted (2 × EtOAc), washed (brine), dried (Na₂SO₄), and

ﬁltered, and the ﬁltrate was concentrated in vacuo. The residue was

puriﬁed by ﬂash column chromatography (SiO₂, 0−30% EtOAc/

hexanes, gradient elution) to give N-((S)-2-((3S,5R,6S)-3-allyl-5-(3-

chlorophenyl)-6-(4-chlorophenyl)-3-methyl-2-oxopiperidin-1-yl)-

butyl)-2-methylpropane-2-sulfonamide (201 mg, 78%) as an oﬀ-white

solid. MS (ESI) m/z = 565 [M + H]⁺.

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

5-cyano-5-methylhexan-3-yl)-3-methyl-2-oxopiperidin-3-yl)-

acetic Acid (9). To a solution of 53 (120 mg, 0.221 mmol) in THF (1.1

mL) was added lithium diisopropylamide (2 M in heptane/THF/

ethylbenzene, 552 μL, 1.10 mmol) at −78 °C. After being stirred for 5

min at −78 °C, iodomethane (94.0 μL, 1.51 mmol) was added and the

resulting solution was stirred at −78 °C for 30 min. The reaction was

allowed to warm to 25 °C and stirred for 12 h. The reaction was

quenched (satd NH₄Cl) and extracted (2 × EtOAc), and the combined

organic layers were washed (brine), dried (Na₂SO₄), and concentrated

under reduced pressure. The residue was puriﬁed by ﬂash column

chromatography (SiO₂, 30% and 40% EtOAc/hexanes) to provide a

mixture of dimethylated product and monomethylated product, which

was resubjected to the methylation conditions described below.

To a solution of the crude product above in THF (1.1 mL) was added

lithium diisopropylamide (2 M in heptane/THF/ethylbenzene, 552 μL,

1.10 mmol) at −78 °C. After being stirred for 5 min at −78 °C,

iodomethane (94 μL, 1.51 mmol) was added and the resulting solution

was stirred at −78 °C for 30 min. The reaction was allowed to warm to

25 °C and stirred for 12 h. The reaction was quenched (satd NH₄Cl)

and extracted (2 × EtOAc), and the combined organic layers were

washed (brine), dried (Na₂SO₄), and concentrated under reduced

pressure. The residue was puriﬁed by RP-HPLC (60−90% A/B,

gradient elution) to aﬀord the tertiary nitrile (77.0 mg, 63%). MS (ESI)

557.2 and 571.2 [M + H]⁺.

To a solution of the product above (77.0 mg, 0.135 mmol) in THF

(2.7 mL) was added 3 N hydrochloric acid in water (1.35 μL, 4.04

mmol) at 25 °C. After being stirred at 25 °C for 4 h, the reaction was

diluted (brine) and extracted (2 × EtOAc). The combined organic

layers were washed (brine), dried (Na₂SO₄), and concentrated under

reduced pressure to provide the corresponding diol (71.6 mg, 100%) as

a colorless foam. MS (ESI) 531.2 [M + H]⁺.

Compound 9 was prepared as a white solid from (4S)-4-((3R,5R,6S)-

5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-(2,3-dihydroxypropyl)-3-

methyl-2-oxopiperidin-1-yl)-2,2-dimethylhexanenitrile according to a

similar procedure described for the synthesis of 3. ¹H NMR (400 MHz,

CDCl₃) δ ppm 7.27 (m, 2 H), 7.00−7.20 (m, 4 H), 6.97 (t, J = 1.7 Hz, 1

H), 6.82 (dt, J = 7.4, 1.4 Hz, 1 H), 4.82 (d, J = 10.6 Hz, 1 H), 3.12 (m, 1

H), 3.01 (d, J = 15.1 Hz, 1 H), 2.75 (m, 3 H), 2.54 (t, J = 12.0 Hz, 1 H),

2.03−2.12 (m, 1 H), 2.01 (s, 1 H), 1.90 (dd, J = 14.0, 2.6 Hz, 1 H), 1.52

(s, 3 H), 1.43 (s, 3 H), 1.35−1.41 (m, 1 H), 1.31 (s, 3 H), 1.23 (d, J =

13.9 Hz, 1 H), 0.33 (t, J = 7.3 Hz, 3 H). HRMS (ESI) m/z 515.1869 [M

+ H]⁺(C₂₈H₃₂Cl₂NO₃requires 515.1863).

(S)-2-((3S,5R,6S)-3-Allyl-5-(3-chlorophenyl)-6-(4-chlorophen-

yl)-3-methyl-2-oxopiperidin-1-yl)butan-1-ammonium 2,2,2-tri-

ﬂuoroacetate (54). To a solution of 44 (300 mg, 0.675 mmol) and (4-

methoxyphenyl)methanamine (131 μL, 1.01 mmol) in DCE (4.5 mL)

was added sodium triacetoxyborohydride (429 mg, 2.03 mmol) at 0 °C

in several portions. After being stirred at 25 °C for 18 h, the reaction was

quenched (ice-cold satd aqueous NaHCO₃) and extracted (2 × DCM).

The combined organic layers were washed (brine) and concentrated

under reduced pressure to provide the PMB-protected amine (382 mg,

100%), which was used in the next step without further puriﬁcation. MS

(ESI) 565.2 [M + H]⁺.

To a solution of the crude amine above (370 mg, 0.654 mmol) in

acetonitrile (8 mL) and water (1.6 mL) was added ceric ammonium

nitrate (2.87 g, 5.23 mmol) at 25 °C. After being stirred at rt for 2 days,

the reaction was quenched (brine), extracted (3 × EtOAc), and washed

(brine). The combined organic layers were dried (Na₂SO₄) and

concentrated under reduced pressure. The residue was puriﬁed by RP-

HPLC (35−70% A/B, gradient elution) to provide 54 (158 mg, 53%) as

a pale-yellow powder. MS (ESI) 445.1 [M + H]⁺.

Compound 11 was prepared from N-((S)-2-((3S,5R,6S)-3-allyl-5-(3-

chlorophenyl)-6-(4-chlorophenyl)-3-methyl-2-oxopiperidin-1-yl)-

butyl)-2-methylpropane-2-sulfonamide according to a similar procedure

1

described for the synthesis of 3. H NMR (500 MHz, CD₃OD-d₄) δ

6.79−7.45 (m, 8 H), 4.94−5.05 (m, 1 H), 3.86−4.05 (m, 1 H), 3.32−

3.40 (m, 1 H), 2.97−3.07 (m, 1 H), 2.88−2.97 (m, 1 H), 2.72−2.83 (m,

1 H), 2.64 (d, J = 13.45 Hz, 1 H), 2.33−2.49 (m, 1 H), 1.96−2.10 (m, 1

H), 1.75−1.89 (m, 1 H), 1.52−1.66 (m, 1 H), 1.45 (s, 3 H), 1.30−1.42

(m, 9 H), 0.46 (t, J = 7.58 Hz, 3 H). HRMS (ESI) m/z 583.1802 [M +

H]⁺(C₂₈H₃₆Cl₂NO₅S requires 583.1795).

Compounds 12−16 were prepared from 55, 56, and 57, respectively,

according to a similar procedure described for the synthesis of 11,

substituting methanesulfonylchloride for the appropriate sulfonyl

chloride.

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-(cyclopropanesulfonamido)butan-2-yl)-3-methyl-2-oxopi-

1

peridin-3-yl)acetic Acid (12). H NMR (400 MHz, CDCl₃) δ ppm

7.23 (m, 2 H), 6.90−7.18 (m, 5 H), 6.78−6.90 (m, 1 H), 4.76 (d, J =

10.56 Hz, 1 H), 3.61 (m, 1 H), 3.01−3.28 (m, 3 H), 2.92 (d, J = 14.09

Hz, 1 H), 2.77 (d, J = 14.28 Hz, 1 H), 2.28−2.49 (m, 2 H), 1.96−2.07

(m, 1 H), 1.77−1.92 (m, 1 H), 1.39−1.64 (m, 4 H), 1.08−1.23 (m, 2 H),

0.92−1.08 (m, 2 H), 0.54 (t, J = 7.53 Hz, 3 H).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-(N,2-dimethylpropan-2-ylsulfonamido)butan-2-yl)-3-methyl-

2-oxopiperidin-3-yl)acetic Acid (13). ¹H NMR (500 MHz,

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-3-

methyl-1-((S)-1-(methylsulfonamido)butan-2-yl)-2-oxopiperi-

din-3-yl)acetic Acid (10). Lithium hydroxide (2 N aqueous solution,

0.340 mL, 0.680 mmol) was added to a solution of 54 (74.0 mg, 0.140

mmol) in DCM at 0 °C, and the resulting solution was stirred for 5 min

at 0 °C. The solution was extracted (2 × DCM), washed (brine), dried

M

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CD₃OD-d₄) δ 7.10−7.40 (m, 5 H), 6.96−7.09 (m, 3 H), 4.79 (d, J =

10.76 Hz, 1 H), 4.40 (s, br, 1 H), 3.32−3.35 (m, 1 H), 2.88−3.01 (m, 4

H), 2.71−2.88 (m, 2 H), 2.63 (d, J = 13.20 Hz, 1 H), 2.43 (t, J = 13.69

Hz, 1 H), 1.94−2.06 (m, 1 H), 1.78−1.92 (m, 1 H), 1.58−1.71 (m, 1 H),

1.32−1.48 (m, 12 H), 0.51 (t, J = 7.21 Hz, 3 H). HRMS (ESI) m/z

597.1955 [M + H]⁺(C₂₉H₃₈Cl₂N₂O₅S requires 597.1951).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-3-

methyl-1-((S)-1-(N-methylcyclopropanesulfonamido)butan-2-

yl)-2-oxopiperidin-3-yl)acetic Acid (14). ¹H NMR (400 MHz,

CDCl₃) δ ppm 7.24 (s, br, 2 H), 7.10−7.17 (m, 2 H), 6.80−7.09 (m, 4

H), 4.79 (d, J = 10.8 Hz, 1 H), 4.13−4.30 (m, 1 H), 2.99−3.12 (m, 2 H),

2.89 (s, 3 H), 2.64−2.81 (m, 2 H), 2.45 (t, J = 13.8 Hz, 1 H), 2.33 (m, 1

H), 1.88 (dd, J = 13.9, 2.7 Hz, 2 H), 1.54−1.66 (m, 1 H), 1.50−1.54 (m,

3 H), 1.22 (d, J = 4.5 Hz, 2 H), 1.02 (dd, J = 8.0, 3.9 Hz, 2 H), 0.51 (t, J =

7.4 Hz, 3 H). HRMS (ESI) m/z 581.1643 [M + H]⁺(C₂₈H₃₄Cl₂N₂O₅S

requires 581.1638).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-cyclopropyl-2-(N-methylcyclopropanesulfonamido)ethyl)-3-

methyl-2-oxopiperidin-3-yl)acetic Acid (15). ¹H NMR (400 MHz,

CD₃OD-d₄) δ ppm 7.31 (s, br, 2H), 7.11 (m, 3H), 7.02−7.05 (m, 3H),

4.82 (d, J = 8 Hz, 1H), 4.40 (s, br, 1H), 3.42 (m, 1H), 3.07 (m, 1H), 2.99

(d, J = 12 Hz, 1H), 2.94 (s, 3H), 2.70 (d, J = 12 Hz, 1H), 2.57 (s, br, 1H),

2.35 (t, J = 8 Hz, 1H), 2.20 (s, br, 1H), 2.08 (m, 1H), 1.73 (s, br, 1H),

1.44 (s, 3H), 1.06 (d, br, 4H), 0.40 (s, br, 1H), 0.29 (s, br, 1H), −0.28 (s,

br, 1H), −0.71 (s, br, 1H.; HRMS (ESI) m/z 593.1642 [M + H]⁺

(C₂₉H₃₄Cl₂N₂O₅S requires 593.1638).

6.95 (s, 1 H), 6.61−6.67 (m, 1 H), 4.68−4.73 (m, 1 H), 4.37−4.47 (m, 1

H), 3.69 (s, 3 H), 3.47−3.56 (m, 1 H), 3.17−3.28 (m, 1 H), 2.91 (m, 1

H), 2.85 (m, 1 H), 2.33−2.46 (m, 1 H), 2.05−2.23 (m, 2 H), 1.47 (s, 3

H), 1.14 (s, 9 H), 0.15−0.28 (m, 2 H), −0.46 to −0.33 (m, 1 H), −0.89

to −0.69 (m, 1 H). MS (ESI) 728.2 [M + H]⁺.

To a solution of the methyl ester above (578 mg, 0.793 mmol) in

THF (3.2 mL) was added TBAF (1 M in THF, 2.40 mL, 2.38 mmol) at 0

°C, and the reaction mixture was allowed to warm to 25 °C for 6 h. The

reaction was quenched (satd NH₄Cl), extracted (2 × EtOAc), and

washed (brine). The combined organic layers were dried (Na₂SO₄) and

concentrated under the reduced pressure. The residue was puriﬁed by

ﬂash column chromatography (SiO₂, 10% and 50% EtOAc/hexanes) to

provide 58 (388 mg, 100%) as a colorless foam. ¹H NMR (400 MHz,

CDCl₃) δ ppm 6.99−7.26 (m, 7 H), 6.77 (d, J = 7.4 Hz, 1 H), 4.87 (d, J =

10.0 Hz, 1 H), 3.70 (s, 3 H), 3.58 (dd, J = 11.0, 4.5 Hz, 1 H), 3.32−3.47

(m, 1 H), 3.14 (t, J = 10.5 Hz, 2 H), 2.75−2.93 (m, 2 H), 2.06−2.23 (m,

2 H), 1.40 (s, 3 H), 0.77−0.94 (m, 1 H), 0.61 (dd, J = 4.6, 3.8 Hz, 2 H),

0.16−0.33 (m, 1 H), 0.00−0.13 (m, 1 H). MS (ESI) 490.0 [M + H]⁺.

Methyl 2-((3R,5R,6S)-1-((S)-2-(Benzylthio)-1-cyclopropyleth-

yl)-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-methyl-2-oxopi-

peridin-3-yl)acetate (59). A mixture of 58 (0.213 g, 0.434 mmol),

phenylmethanethiol (0.102 mL, 0.869 mmol), and 2-

(tributylphosphoranylidene)acetonitrile (0.233 mL, 0.869 mmol) was

heated at 100 °C for 2 h. The reaction mixture was cooled to 25 °C, and

diluted (EtOAc), washed (satd NH₄Cl and brine), dried (Na₂SO₄), and

concentrated. The residue was puriﬁed by ﬂash column chromatography

(SiO₂, 0−50% EtOAc/hexanes, gradient elution) to provide 59 (210

mg, 81%). ¹H NMR (400 MHz, CDCl₃) δ ppm 7.00−7.60 (m, 12 H),

6.95 (d, J = 8.0 Hz, 1 H), 4.82 (d, J = 10.6 Hz, 1 H), 3.92 (s, 2 H), 3.85 (s,

3 H), 3.50 (m, 1 H), 3.35 (m, 1 H), 3.08 (d, J = 12 Hz, 1 H), 3.00 (d, J =

12 Hz, 1 H), 2.62 (m, 2 H), 2.35 (dd, J = 4.0, 12.0 Hz, 1 H), 2.20 (dd, J =

4.0, 12.0 Hz, 1 H), 1.65 (s, 3 H), 1.65 (m, 1 H), 0.75 (m, 1 H), 0.45 (m, 1

H), 0.05 (m, 1 H), −0.65 (m, 1 H). MS (ESI) 596.0 [M + H]⁺.

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-cyclopropyl-2-(N,N-dimethylsulfamoyl)ethyl)-3-methyl-2-ox-

opiperidin-3-yl)acetic Acid (17). To a heterogeneous mixture of 59

(0.03 g, 0.05 mmol) and iodosobenzene (0.037 g, 0.166 mmol) in ether

(30 mL) was added hydrogen chloride (2.0 M in ether, 3.60 mL, 7.20

mmol) gradually with vigorously stirring. The resulting mixture was

stirred for 30 min and concentrated under reduced pressure. The

solution of the dried residue in DCM (2 mL) was added to a mixture of

dimethylamine (0.011 g, 0.251 mmol) and N-ethyl-N-isopropylpropan-

2-amine (0.032 g, 0.251 mmol) in DCM (1 mL). The resulting mixture

was stirred at 25 °C for 1 h and was diluted (DCM), washed (water and

brine), dried (Na₂SO₄), and concentrated under reduced pressure. The

residue was puriﬁed by ﬂash column chromatography (SiO₂, 20−70%,

gradient elution) to give methyl 2-((3R,5R,6S)-5-(3-chlorophenyl)-6-

(4-chlorophenyl)-1-((S)-1-cyclopropyl-2-(N,N-dimethylsulfamoyl)-

ethyl)-3-methyl-2-oxopiperidin-3-yl)acetate (23.0 mg, 79%). ¹H NMR

(400 MHz, CDCl₃) δ ppm 7.27 (s, br, 3 H), 7.07−7.16 (m, 3 H), 6.95 (s,

1 H), 6.83−6.89 (m, 1 H), 4.83 (d, J = 10.56 Hz, 1 H), 4.19 (t, J = 12.42

Hz, 1 H), 3.07−3.19 (m, 2 H), 2.90 (s, 6 H), 2.73−2.84 (m, 2 H), 2.60

(s, br, 1H), 2.50 (t, J = 13.79 Hz, 1 H), 1.87 (dd, J = 13.79, 2.84 Hz, 2H),

1.53 (s, 3 H), 0.18−0.44 (m, 2 H), −0.28 (s, br, 1 H), −1.07 (s, br, 1 H).

MS (ESI) 567.0 [M + H]⁺.

2-((3R,5R,6S)-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-meth-

yl-1-((S)-3-methyl-1-(N-methylcyclopropanesulfonamido)-

1

butan-2-yl)-2-oxopiperidin-3-yl)acetic Acid (16). H NMR (500

MHz, CD₃OD-d₄) δ ppm 7.29 (s, br, 4 H), 7.07−7.16 (m, 2 H), 6.98−

7.06 (m, 2 H), 4.97 (d, J = 11.0 Hz, 1 H), 4.13−4.40 (m, 1 H), 3.48 (ddd,

J = 13.8, 11.0, 3.1 Hz, 1 H), 3.01 (d, J = 13.5 Hz, 1 H), 2.87−2.96 (m, 4

H), 2.60−2.71 (m, 2 H), 2.51−2.60 (m, 1 H), 2.38 (t, J = 13.7 Hz, 1 H),

2.23 (dquin, J = 9.5, 6.7 Hz, 1 H), 2.07 (dd, J = 13.7, 3.2 Hz, 1 H), 1.40 (s,

3 H), 0.95−1.22 (m, 4 H), 0.69 (d, J = 6.60 Hz, 3 H), 0.63 (d, J = 6.60

Hz, 3 H). HRMS (ESI) m/z 595.1799 [M + H]⁺(C₂₉H₃₆Cl₂N₂O₅S

requires 595.1795).

Methyl 2-((3R,5R,6S)-5-(3-chlorophenyl)-6-(4-chlorophenyl)-

1-((S)-1-cyclopropyl-2-hydroxyethyl)-3-methyl-2-oxopiperi-

din-3-yl)acetate (58). To a solution of 56 (1.15 g, 2.51 mmol) and

imidazole (0.439 g, 6.45 mmol) in DMF (6.5 mL) was added tert-

butylchlorodiphenylsilane (0.963 mL, 3.76 mmol) via syringe. The

yellow solution was stirred at 25 °C for 12 h under nitrogen. The

reaction was quenched (water) and extracted (diethyl ether). The

combined organic layers and washed (brine), dried (Na₂SO₄), ﬁltered,

and concentrated in vacuo. Flash column chromatography (SiO₂, 0−

10% EtOAc/hexanes, gradient elution) gave the crude product (1.60 g,

92%). MS (ESI) 696.2 [M + H]⁺.

2-((3R,5R,6S)-1-((S)-2-(tert-Butyldiphenylsilyloxy)-1-cyclopropy-

lethyl)-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-methyl-2-oxopiperi-

din-3-yl)acetic acid was prepared as a pale-yellow foam from the TBDPS

protected alcohol according to a similar procedure described for the

synthesis of 3. ¹H NMR (400 MHz, CDCl₃) δ ppm 7.62 (ddd, J = 7.2,

5.9, 1.4 Hz, 4 H), 7.37−7.49 (m, 8 H), 7.12−7.26 (m, 3 H), 7.05 (s, 1 H),

6.97 (s, 1 H), 6.60−6.69 (m, 1 H), 4.67−4.78 (m, 1 H), 4.26−4.40 (m, 1

H), 3.43−3.58 (m, 1 H), 3.12−3.23 (m, 1 H), 2.96−3.12 (m, 1 H), 2.72

(s, 1 H), 2.40−2.56 (m, 1 H), 2.24−2.35 (m, 1 H), 1.95−2.07 (m, 1 H),

1.52 (s, 3 H), 1.39−1.48 (m, 1 H), 1.15 (s, 9 H), 0.21 to −0.34 (m, 2 H),

−0.4 to −0.31 (m, 1 H), −0.95−0.76 (m, 1 H). MS (ESI) 714.3 [M +

H]⁺.

To a solution of the sulfonamide above (0.023 g, 0.04 mmol) in

THF/H₂O (3: 1, 3.0 mL) was added lithium hydroxide (0.079 mL,

0.079 mmol). The reaction mixture was stirred at 25 °C for 4 h. The

reaction was quenched (satd NH₄Cl), extracted (2 × EtOAc), and

washed (brine). The residue was puriﬁed by RP-HPLC (10−90% A/B,

gradient elution) to provide 17 (16.0 mg, 71%). ¹H NMR (400 MHz,

CDCl₃) δ ppm 7.10−7.19 (m, 2 H), 7.02−7.08 (m, 4 H), 6.88 (m, 1 H),

6.78 (m, 1 H), 4.77 (m, 1 H), 4.10 (m, 1 H), 3.02−3.10 (m, 2 H), 2.79 (s,

6 H), 2.64−2.70 (m, 2 H), 2.32−2.55 (m, 2 H), 1.70−1.81 (m, 2 H),

1.45 (s, 3 H), 0.30 (s, br, 1 H), 0.18 (s, br, 1 H), −0.32 (s, br, 1 H), −1.18

(s, br, 1 H). HRMS (ESI) m/z 567.1487 [M + H]⁺(C₂₇H₃₂Cl₂N₂O₅S

requires 567.1482).

To a solution of the acid above (1.40 g, 1.96 mmol) in MeOH (3.1

mL) and benzene (13 mL) was added 2.0 M (trimethylsilyl)-

diazomethane in hexanes (1.96 mL, 3.92 mmol) at 0 °C dropwise.

After being stirred at 0 °C for 1 h, the reaction mixture was concentrated

under reduced pressure. The residue was puriﬁed by ﬂash column

chromatography (SiO₂, 10% and 20% EtOAc/hexanes) to provide

methyl 2-((3R,5R,6S)-1-((S)-2-(tert-butyldiphenylsilyloxy)-1-cyclopro-

pylethyl)-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-methyl-2-oxopi-

1

Compounds 18 and 19 were prepared from 59 according to a similar

procedure described for the synthesis of 17, substituting dimethylamine

for pyrrolidine, azetidine, respectively.

peridin-3-yl)acetate (992 mg, 70%) as a pale-yellow foam. H NMR

(400 MHz, CDCl₃) δ ppm 7.64 (ddd, J = 11.2, 8.0, 1.4 Hz, 4 H), 7.34−

7.49 (m, 8 H), 7.15−7.21 (m, 2 H), 7.10−7.13 (m, 1 H), 7.02 (s, 1 H),

N

dx.doi.org/10.1021/jm401753e | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1.37 (s, 3 H), 0.67 (d, J = 6.6 Hz, 3 H), 0.50 (d, J = 6.9 Hz, 3 H); MS

(ESI) 540.2 [M + H]⁺.

Compounds 26, 28−33, 35, 38, and 40, were prepared from 55−57,

and 61−64, respectively, according to a similar procedure described for

the synthesis of 20.

1-cyclopropyl-2-(pyrrolidin-1-ylsulfonyl)ethyl)-3-methyl-2-ox-

1

opiperidin-3-yl)acetic Acid (18). H NMR (400 MHz, CDCl₃) δ

ppm 7.21−7.37 (m, 3 H), 7.07−7.19 (m, 3 H), 6.98 (s, 1 H), 6.89 (d, J =

6.65 Hz, 1 H), 4.84 (d, J = 10.56 Hz, 1 H), 4.00 (m, 4 H), 2.30−3.305

(m, 10 H), 1.81 (s, br, 1 H), 1.52 (s, 3 H), 0.17−0.42 (m, 2 H), −0.28 (s,

br, 1 H), −1.08 (s, br, 1 H). HRMS (ESI) m/z 579.1482 [M + H]⁺

(C₂₈H₃₂Cl₂N₂O₅S requires 579.1482).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-((cyclopropylmethyl)sulfonyl)butan-2-yl)-3-methyl-2-oxopi-

1

peridin-3-yl)acetic Acid (26). H NMR (400 MHz, CDCl₃) δ ppm

7.23−7.27 (m, 2 H), 7.00−7.22 (m, 4 H), 6.94−6.98 (m, 1 H), 6.85 (dt,

J = 7.1, 1.5 Hz, 1 H), 4.95 (d, J = 10.8 Hz, 1 H), 4.17 (t, J = 12.1 Hz, 1 H),

3.35 (t, J = 9.7 Hz, 1 H), 3.13 (ddd, J = 13.6, 10.9, 2.7 Hz, 1 H), 2.83−

3.07 (m, 4 H), 2.77 (d, J = 14.9 Hz, 1 H), 2.39 (t, J = 13.8 Hz, 1 H),

2.07−2.22 (m, 1 H), 1.91 (dd, J = 13.9, 2.7 Hz, 1 H), 1.48 (s, 3 H), 1.40−

1.47 (m, 1 H), 1.12−1.25 (m, 1 H), 0.75−0.85 (m, 2 H), 0.37−0.48 (m,

5 H). HRMS (ESI) m/z 566.1538 [M + H]⁺(C₂₈H₃₃Cl₂NO₅S requires

566.1529).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-(cyclopentylsulfonyl)butan-2-yl)-3-methyl-2-oxopiperidin-3-

yl)acetic Acid (28). ¹H NMR (400 MHz, CDCl₃) δ ppm 7.25 (m, 2 H),

7.12 (m, 4 H), 6.96 (s, 1 H), 6.85 (d, J = 8.0 Hz, 1 H), 4.97 (d, J = 8.0 Hz,

1 H), 4.10 (t, J = 12 Hz, 1 H), 3.36 (m, 2 H), 3.12 (m, 1 H), 3.00 (d, J =

16 Hz, 1 H), 2.76 (d, J = 16.0 Hz, 2 H), 2.40 (t, J = 12 Hz, 1 H), 2.09 (m,

5 H), 1.80−1.95 (m, 3 H), 1.70 (m, 2 H), 1.49 (s, 3 H), 1.46 (m, 1 H),

0.40 (t, J = 8.0 Hz, 3 H). HRMS (ESI) m/z 580.1692 [M + H]⁺

(C₂₉H₃₅Cl₂NO₅S requires 580.1686).

2-((3R,5R,6S)-1-((S)-2-(Azetidin-1-ylsulfonyl)-1-cyclopropy-

lethyl)-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-methyl-2-oxo-

piperidin-3-yl)acetic Acid (19). ¹H NMR (400 MHz, CDCl₃) δ ppm

7.24−7.28 (m, 3 H), 7.07−7.17 (m, 3 H), 6.97 (m, 1 H), 6.81−6.88 (m,

1 H), 4.82 (d, J = 10.56 Hz, 1 H), 3.45−3.65 (m, 4 H), 3.10−3.20 (m, 2

H), 2.80−2.95 (m, 2 H), 2.45−2.80 (m, 2 H), 1.80−2.10 (m, 6 H), 1.80

(s, br, 1 H), 1.51 (s, 3 H), 0.18−0.44 (m, 2 H), 0.28 (s, br, 1 H), −1.07 (s,

br, 1 H). HRMS (ESI) m/z 593.1644 [M + H]⁺(C₂₉H₃₄Cl₂N₂O₅S

requires 593.1638).

(3S,5R,6S)-3-Allyl-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-

methyl-1-((S)-1-(((trimethylsilyl)methyl)thio)butan-2-yl)-

piperidin-2-one (60). To a solution of 55 (125 mg, 0.280 mmol) in

toluene (1 mL) was added cyanomethylenetributylphosphorane (300

μL, 1.12 mmol) and (trimethylsilyl)methanethiol (150 μL 1.12 mol) at

25 °C. The mixture was heated to 110 °C for 2 h. The reaction was

cooled down to rt, quenched (satd NH₄Cl), extracted (2 × EtOAc), and

washed (brine). The combined organic layers were dried over Na₂SO₄

and ﬁltered, and the ﬁltrate was concentrated under reduced pressure.

The residue was puriﬁed by ﬂash column chromatography (SiO₂, 10, 20,

40% EtOAc/hexanes) to provide 60 (65.0 mg, 42%) as a white foam. ¹H

NMR (400 MHz, CDCl₃) δ ppm 7.20−7.25 (m, 2 H), 7.05−7.20 (m, 2

H), 6.90−7.05 (m, 3 H), 6.75 (d, J = 8.0 Hz, 1 H), 5.88 (m, 1 H), 5.14−

5.20 (m, 2 H), 4.67 (d, J = 8.0 Hz, 1 H), 3.40 (dd, J = 8.0, 12.0 Hz, 1 H),

3.11 (m, 1 H), 2.83 (m, 1 H), 2.60 (m, 2 H), 2.48 (dd, J = 4.0, 12 Hz, 1

H), 2.13 (t, J = 12.0 Hz, 1 H), 2.05 (m, 1 H), 1.89 (dd, J = 4.0, 12 Hz, 1

H), 1.82 (d, J = 12.0 Hz, 1 H), 1.73 (d, J = 12.0 Hz, 1 H), 1.57 (m, 1 H),

1.27 (s, 3 H), 0.50 (t, J = 8.0 Hz, 3 H), 0.16 (s, 9 H).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-3-

methyl-1-((S)-1-(oxetan-3-ylsulfonyl)butan-2-yl)-2-oxopiperi-

1

din-3-yl)acetic Acid (29). H NMR (400 MHz, CD₃OD-d₄) δ ppm

7.28 (s, br, 3 H), 7.07−7.22 (m, 3 H), 7.00−7.07 (m, 1 H), 6.97 (dt, J =

6.65, 1.76 Hz, 1 H), 4.94−5.01 (m, 2 H), 4.87−4.93 (m, 4 H), 4.55−4.76

(m, 1 H), 3.94−4.20 (m, 1 H), 3.41 (ddd, J = 13.60, 10.96, 2.84 Hz, 1 H),

2.98−3.07 (m, 1 H), 2.95 (d, J = 13.69 Hz, 1 H), 2.61 (d, J = 13.69 Hz, 1

H), 2.29 (t, J = 13.69 Hz, 1 H), 2.02−2.10 (m, 2 H), 1.46−1.63 (m, 1 H),

1.34−1.43 (m, 3 H), 0.39 (t, J = 7.63 Hz, 3 H). HRMS (ESI) m/z

568.1326 [M + H]⁺(C₂₇H₃₁Cl₂NO₆S requires 568.1322).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-3-

methyl-2-oxo-1-((S)-1-(phenylsulfonyl)butan-2-yl)piperidin-3-

yl)acetic Acid (30). ¹H NMR (400 MHz, CDCl₃) δ ppm 7.93 (d, J = 8

Hz, 2 H), 7.71 (m, 1 H), 7.61 (m, 2 H), 7.27(m, 2 H), 7.14 (m, 4 H),

6.99 (s, 1 H), 6.88 (d, J = 8.0 Hz, 1 H), 5.04 (d, J = 8.0 Hz, 1 H), 4.24 (dd,

J = 16, 12 Hz, 1 H), 3.41 (m, 1 H), 3.15 (m, 1 H), 3.05 (d, J = 16 Hz, 1

H), 2.88 (dd, J = 16 Hz, 1 H), 2.79 (d, J = 16 Hz, 1 H), 2.50 (t, J = 16 Hz,

1 H), 2.08 (m, 1 H), 1.94 (dd, J = 12, 4 Hz, 1 H), 1.58 (s, 3 H), 1.46 (m, 1

H), 0.37 (t, J = 8 Hz, 3 H). HRMS (ESI) m/z 588.1378 [M + H]⁺

(C₃₀H₃₁Cl₂NO₅S requires 588.1373).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-3-

methyl-1-((S)-1-(methylsulfonyl)butan-2-yl)-2-oxopiperidin-3-

yl)acetic Acid (20). 60 (60.0 mg, 0.118 mmol) was dissolved in a

solution of tetrabutylammonium ﬂuoride (1 M in THF, 3.5 mL, 3.50

mmol) at 25 °C. After the resulting solution was stirred at 25 °C for 16 h,

the solvents were removed under reduced pressure. The residue was

puriﬁed by ﬂash column chromatography (SiO₂, 10, 20, 40% EtOAc/

hexanes) to provide (3S,5R,6S)-3-allyl-5-(3-chlorophenyl)-6-(4-chlor-

ophenyl)-3-methyl-1-((S)-1-(methylthio)butan-2-yl)piperidin-2-one

(32.0 mg, 57%). ¹H NMR (400 MHz, CDCl₃) δ ppm 7.14−7.21 (m, 2

H), 7.05−7.15 (m, 2 H), 6.95−7.05 (m, 3 H), 6.76 (d, J = 8.0 Hz, 1 H),

5.85 (m, 1 H), 5.15−5.20 (m, 2 H), 4.62 (d, J = 8.0 Hz, 1 H), 3.35 (dd, J

= 8.0, 12.0 Hz, 1 H), 3.12 (m, 1 H), 2.85 (m, 1 H), 2.60 (m, 2 H), 2.48

(dd, J = 4.0, 12 Hz, 1 H), 2.13 (s, 3 H), 1.95−2.20 (m, 2 H), 1.89 (dd, J =

4.0, 12 Hz, 1 H), 1.56 (m, 1 H), 1.28 (s, 3 H), 0.52 (t, J = 8.0 Hz, 3 H).

Compound 20 was prepared from (3S,5R,6S)-3-allyl-5-(3-chlor-

ophenyl)-6-(4-chlorophenyl)-3-methyl-1-((S)-1-(methylthio)butan-2-

yl)piperidin-2-one according to a similar procedure described for the

synthesis of 3. ¹H NMR (400 MHz, CDCl₃) δ ppm 7.27 (m, 2 H), 7.12

(m, 4 H), 6.95 (s, 1 H), 6.84 (d, J = 8.0 Hz, 1 H), 4.90 (d, J = 8.0 Hz, 1

H), 4.24 (m, 1 H), 3.37 (m, 1 H), 3.13 (m, 1 H), 2.99 (s, 3 H), 2.97 (m, 1

H), 2.90 (d, J = 12 Hz, 1 H), 2.76 (d, J = 16 Hz, 1 H), 2.37 (t, J = 16.0 Hz,

1 H), 2.14 (m, 1 H), 1.90 (d, J = 12 Hz, 1 H), 1.48 (s, 3 H), 1.45 (m, 1

H), 0.42 (t, J = 8.0 Hz, 3 H). HRMS (ESI) m/z 526.1217 [M + H]⁺

(C₂₅H₂₉Cl₂NO₅S requires 526.1216).

2-((3R,5R,6S)-1-((S)-1-(tert-Butylsulfonyl)-4,4,4-triﬂuorobu-

tan-2-yl)-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-methyl-2-

1

oxopiperidin-3-yl)acetic Acid (31). NMR (400 MHz, CDCl₃) δ

ppm 7.27 (s, 2 H), 7.03−7.23 (m, 4 H), 6.97 (s, 1 H), 6.87 (dt, J = 7.0,

1.6 Hz, 1 H), 5.07 (d, J = 10.8 Hz, 1 H), 4.11 (dd, J = 13.2, 11.2 Hz, 1 H),

3.86 (t, J = 10.3 Hz, 1 H), 3.09−3.27 (m, 2 H), 2.98 (dd, J = 13.2, 2.1 Hz,

1 H), 2.78−2.90 (m, 2 H), 2.41 (t, J = 13.8 Hz, 1 H), 2.11−2.28 (m, 1

H), 1.95 (dd, J = 13.9, 2.7 Hz, 1 H), 1.49 (s, 3 H), 1.43 (s, 9 H). HRMS

(ESI) m/z 622.1401 [M + H]⁺(C₂₈H₃₂Cl₂F₃NO₅S requires 622.1403).

2-((3R,5R,6S)-1-((S)-1-(tert-Butylsulfonyl)propan-2-yl)-5-(3-

chlorophenyl)-6-(4-chlorophenyl)-3-methyl-2-oxopiperidin-3-

1

yl)acetic Acid (32). H NMR (500 MHz, CDCl₃) δ ppm 7.21−7.28

(m, 2 H), 7.08−7.15 (m, 4 H), 6.94−6.99 (m, 1 H), 6.83 (d, J = 7.1 Hz, 1

H), 4.95 (d, J = 10.8 Hz, 1 H), 4.24 (dd, J = 13.2, 11.3 Hz, 1 H), 3.70 (1

H, m), 3.00−3.10 (m, 2 H), 2.72 (d, J = 15.2 Hz, 1 H), 2.66 (dd, J = 2.45,

12.5, 1 H), 2.41 (t, J = 13.7 Hz, 1 H), 1.88 (d, J = 13.7 Hz, 1 H), 1.49 (s, 3

H), 1.43 (s, 9 H), 1.27 (d, J = 6.85 Hz, 3 H). HRMS (ESI) m/z 554.1539

[M + H]⁺(C₂₇H₃₃Cl₂NO₅S requires 554.1529).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-3-

methyl-1-((S)-3-methyl-1-(methylsulfonyl)butan-2-yl)-2-oxopi-

peridin-3-yl)acetic Acid (42). Compound 42 was prepared from 57

2-((3R,5R,6S)-1-((S)-1-(tert-Butylsulfonyl)-3-methylbutan-2-

1

according to a similar procedure described for the synthesis of 20. H

yl)-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-methyl-2-oxopi-

1

NMR (500 MHz, CD₃OD-d₄) δ ppm 7.29 (s, br, 4 H), 7.08−7.16 (m, 2

H), 7.03−7.08 (m, 1 H), 6.98 (dt, J = 7.2, 1.4 Hz, 1 H), 5.09 (d, J = 11

Hz, 1 H), 4.07 (dd, J = 13.9, 10.5 Hz, 1 H), 3.56 (ddd, J = 13.8, 10.9, 2.9

Hz, 1 H), 3.32−3.35 (m, 1 H), 3.20−3.29 (m, 1 H), 3.09 (s, br, 3 H),

2.99 (d, J = 13.5 Hz, 1 H), 2.62 (d, J = 13.5 Hz, 1 H), 2.29 (t, J = 13.6 Hz,

1 H), 2.18 (dq, J = 14.2, 6.9 Hz, 1 H), 2.05 (dd, J = 13.8, 3.1 Hz, 1 H),

peridin-3-yl)acetic Acid (33). H NMR (500 MHz, CD₃OD-d₄) δ

ppm 7.18−8.01 (m, 3 H), 7.04−7.17 (m, 3 H), 6.98−7.03 (m, 1 H), 5.15

(d, J = 11.3 Hz, 1 H), 3.96 (dd, J = 13.8, 10.4 Hz, 1 H), 3.57 (ddd, J =

13.6, 11.1, 2.9 Hz, 1 H), 3.25−3.29 (m, 1 H), 3.10 (dd, J = 13.7, 1.71 Hz,

1 H), 2.99 (d, J = 13.7 Hz, 1 H), 2.61 (d, J = 13.5 Hz, 1 H), 2.30 (t, J =

13.7 Hz, 1 H), 2.11−2.25 (m, 1 H), 2.01−2.10 (m, 1 H), 1.45 (s, 9 H),

O

dx.doi.org/10.1021/jm401753e | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

1.37 (s, 3 H), 0.65 (d, J = 6.6 Hz, 3 H), 0.51 (d, J = 6.9 Hz, 3 H). HRMS

(ESI) m/z 582.1845 [M + H]⁺(C₂₉H₃₇Cl₂NO₅S requires 582.1842).

2-((3R,5R,6S)-1-((S)-2-(tert-Butylsulfonyl)-1-cyclopropyleth-

yl)-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-methyl-2-oxopi-

synthesis of 3. ¹H NMR (400 MHz, CDCl₃) δ ppm 7.24−7.26 (m, 2 H),

7.01−7.20 (m, 4 H), 6.93−6.98 (m, 1 H), 6.85 (d, J = 7.0 Hz, 1 H), 4.94

(d, J = 10.6 Hz, 1 H), 4.15 (t, J = 12.1 Hz, 1 H), 3.24−3.37 (m, 1 H),

2.92−3.18 (m, 4 H), 2.71−2.82 (m, 2 H), 2.38 (t, J = 13.8 Hz, 1 H),

2.06−2.21 (m, 1 H), 1.92 (dd, J = 13.7, 2.7 Hz, 1 H), 1.48 (s, 3 H), 1.42−

1.46 (m, 1 H) 1.44 (t, J = 7.5 Hz, 3 H), 0.41 (t, J = 7.5 Hz, 3 H). HRMS

(ESI) m/z 540.1378 [M + H]⁺(C₂₆H₃₁Cl₂NO5S requires 540.1373).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-(cyclopropylsulfonyl)butan-2-yl)-3-methyl-2-oxopiperidin-3-

yl)acetic Acid (23). Method B: To a solution of (3S,5S,6R,8S)-8-allyl-

6-(3-chlorophenyl)-5-(4-chlorophenyl)-3-ethyl-8-methyl-2,3,5,6,7,8-

hexahydrooxazolo[3,2-a]pyridin-4-ium triﬂuoromethanesulfonate

(75.0 mg, 0.130 mmol) in acetonitrile (1.3 mL) was added

cyclopropanesulﬁnic acid sodium salt (50.0 mg, 0.390 mmol) at 25

°C. After being stirred at 90 °C for 1 day, the reaction was quenched

(satd NH₄Cl) and extracted (2 × EtOAc), and the combined organic

layers were washed (2 × brine), dried (Na₂SO₄), and ﬁltered, and the

ﬁltrate was concentrated under reduced pressure. The residue was

puriﬁed by ﬂash column chromatography (SiO₂, 35% and 45% EtOAc/

hexanes) to provide (3S,5R,6S)-3-allyl-5-(3-chlorophenyl)-6-(4-chlor-

ophenyl)-1-((S)-1-(cyclopropylsulfonyl)butan-2-yl)-3-methylpiperi-

din-2-one (55.3 mg, 80%) as a colorless liquid.

1

peridin-3-yl)acetic Acid (35). H NMR (400 MHz, CDCl₃) δ ppm

7.20−7.26 (m, 3 H), 7.03−7.17 (m, 3 H), 6.93−6.97 (m, 1 H), 6.82−

6.92 (m, 1 H), 4.95 (d, J = 10.6 Hz, 1 H), 4.30 (t, J = 12.0 Hz, 1 H), 3.14

(ddd, J = 13.6, 10.8, 2.6 Hz, 1 H), 3.07 (d, J = 15.1 Hz, 1 H), 2.92 (d, J =

11.9 Hz, 1 H), 2.79 (d, J = 15.1 Hz, 1 H), 2.72 (t, J = 9.6 Hz, 1 H), 2.45 (t,

J = 13.8 Hz, 1 H), 1.88 (dd, J = 13.6, 2.4 Hz, 2 H), 1.50 (s, 3 H), 1.44 (s,

9H), 0.30−0.44 (m, 1 H), 0.17−0.30 (m, 1 H), −0.37 to −0.26 (m, 1

H), −1.15 to −1.05 (m, 1 H). HRMS (ESI) m/z 580.1692 [M + H]⁺

(C₂₉H₃₅Cl₂NO₅S requires 580.1686).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-(isopropylsulfonyl)-3,3-dimethylbutan-2-yl)-3-methyl-2-oxo-

1

piperidin-3-yl)acetic Acid (38). H NMR (500 MHz, DMSO-d₆) δ

ppm 12.36 (s, 1H), 7.81 (br s, 1H), 7.48 (s, br, 1H), 7.25 (s, br, 1H), 7.22

(t, J = 7.8 Hz, 1H), 7.15 (ddd, J = 7.8. 2.0, 0.7 Hz, 1H), 7.04 (t, J = 1.7 Hz,

1H), 7.00 (d, br, J = 7.8 Hz, 1H), 5.05 (d, J = 11.2 Hz, 1H), 3.90 (dd, J =

13.7, 11.0, 2.6 Hz, 1H), 3.70 (ddd, J = 13.7, 11.0, 2.6 Hz, 1H), 3.45 (m,

2H), 3.12 (dd, J = 13.4, 2.0, 1H), 2.97 (d, J = 13.9 Hz, 1H), 2.14 (t, J =

13.2 Hz, 1H), 2.03 (dd, J = 13.4 Hz, 1H), 1.32 (d, J = 6.8 Hz, 3H), 1.31

(d, J = 6.6 Hz, 3H), 1.23 (s, 3H), 0.62 (s, 9H). HRMS (ESI) m/z

582.1845 [M + H]⁺(C₂₉H₃₇Cl₂NO₅S requires 582.1842).

Compound 23 was prepared from (3S,5R,6S)-3-allyl-5-(3-chlor-

ophenyl)-6-(4-chlorophenyl)-1-((S)-1-(cyclopropylsulfonyl)butan-2-

yl)-3-methylpiperidin-2-one according to a similar procedure described

for the synthesis of 3. ¹H NMR (400 MHz, CDCl₃) δ ppm 7.22−7.26

(m, 2 H), 6.99 −7.19 (m, 4 H), 6.91−6.97 (m, 1 H), 6.76−6.89 (m, 1

H), 4.91 (d, J = 10.8 Hz, 1 H), 4.21 (dd, J = 13.5, 11.3 Hz, 1 H), 3.31 (t, J

= 10.3 Hz, 1 H), 3.12 (ddd, J = 13.6, 10.9, 2.6 Hz, 1 H), 2.88−3.02 (m, 2

H), 2.76 (d, J = 14.9 Hz, 1 H), 2.31−2.49 (m, 2 H), 2.06−2.24 (m, 1 H),

1.90 (dd, J = 13.7, 2.7 Hz, 1 H), 1.40−1.56 (m, 1 H), 1.47 (s, 3 H), 1.23−

1.36 (m, 2 H), 1.01−1.16 (m, 2 H), 0.42 (t, J = 7.5 Hz, 3 H). HRMS

(ESI) m/z 552.1377 [M + H]⁺(C₂₇H₃₁Cl₂NO₅S requires 552.1373).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-(isopropylsulfonyl)butan-2-yl)-3-methyl-2-oxopiperidin-3-

yl)acetic Acid (24). Method C: To a solution of (3S,5S,6R,8S)-8-allyl-

6-(3-chlorophenyl)-5-(4-chlorophenyl)-3-ethyl-8-methyl-2,3,5,6,7,8-

hexahydrooxazolo[3,2-a]pyridin-4-ium triﬂuoromethanesulfonate

(61.0 mg, 0.105 mmol) in DMF (0.5 mL) was added cesium carbonate

(206 mg, 0.633 mmol) and 2-propanethiol (59.2 μL, 0.633 mmol). After

being stirred at 25 °C for 1.5 h, the recation was quenched (satd

NH₄Cl), extracted (2 × EtOAc), and washed (2 × brine). The

combined organic layers were dried (Na₂SO₄) and concentrated under

reduced pressure. The residue was puriﬁed by ﬂash column

chromatography (SiO₂, 7% and 15% EtOAc/hexanes) to provide

(3S,5R,6S)-3-allyl-5-(3-chlorophenyl)-6-(4-chlorophenyl)-1-((S)-1-

(isopropylthio)butan-2-yl)-3-methylpiperidin-2-one (26.5 mg, 50%) as

a colorless liquid. ¹H NMR (400 MHz, CDCl₃) δ ppm 7.22 (d, J = 8.6

Hz, 2 H), 7.07−7.18 (m, 2 H), 6.86−7.05 (m, 3 H), 6.77 (dt, J = 7.4, 1.4

Hz, 1 H), 5.69−5.96 (m, 1 H), 5.06−5.23 (m, 2 H), 4.66 (d, J = 10.6 Hz,

1 H), 3.42 (dd, J = 12.6, 10.9 Hz, 1 H), 3.12 (ddd, J = 13.6, 10.6, 3.1 Hz, 1

H), 2.90 (dt, J = 13.4, 6.7 Hz, 1 H), 2.68−2.80 (m, 1 H), 2.52−2.67 (m, 3

H), 2.10−2.19 (m, 1 H), 1.99−2.09 (m, 1 H), 1.88 (dd, J = 13.5, 3.1 Hz,

1 H), 1.56−1.61 (m, 1 H), 1.31 (d, J = 6.6 Hz, 3 H), 1.30 (d, J = 6.6 Hz, 3

H), 1.26−1.29 (m, 3 H), 0.51 (t, J = 7.5 Hz, 3 H). MS (ESI) 504.2 [M +

H]⁺.

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-cyclobutyl-2-(isopropylsulfonyl)ethyl)-3-methyl-2-oxopiper-

idin-3-yl)acetic Acid (40). ¹H NMR (400 MHz, CDCl₃) δ ppm 7.24−

7.30 (m, 2 H), 7.08−7.15 (m, 4 H), 6.98 (m, 1 H), 6.87 (d, J = 7.0 Hz, 1

H), 5.02 (d, J = 10.0 Hz, 1 H), 3.99 (t, J = 12.1 Hz, 1 H), 3.49 (m, 1 H),

2.95−3.18 (m, 6 H), 2.65−2.80 (m, 2 H), 2.42 (t, J = 12.0 Hz, 1 H),

1.80−1.95 (m, 3 H), 1.58 (m, 1 H), 1.46 (s, 3 H), 1.44 (d, J = 5 Hz, 6 H),

0.81 (m, 1 H). HRMS (ESI) m/z 580.1691 [M + H]⁺(C₂₉H₃₅Cl₂NO₅S

requires 580.1686).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-(ethylsulfonyl)butan-2-yl)-3-methyl-2-oxopiperidin-3-yl)-

acetic Acid (22). To a solution of 55 (300 mg, 0.652 mmol) in DCM (6

mL) was added triethylamine (270 μL, 1.96 mmol) and methane-

sulfonic anhydride (170 mg, 0.977 mmol) successively at 0 °C. After

being stirred at 25 °C for 2 h, the reaction was quenched with 10%

aqueous citric acid and extracted (DCM), and the combined extracts

were washed (water), dried (Na₂SO₄), and ﬁltered. The ﬁltrate was

concentrated to give (3S,5S,6R,8S)-8-allyl-6-(3-chlorophenyl)-5-(4-

chlorophenyl)-3-ethyl-8-methyl-2,3,5,6,7,8-hexahydrooxazolo[3,2-a]-

pyridin-4-ium triﬂuoromethanesulfonate. ¹H NMR (500 MHz, DMSO-

d₆) δ ppm 7.95 (s, br, 1 H), 7.34−7.60 (m, 2 H), 7.18−7.34 (m, 4 H),

7.13 (dt, J = 7.5, 1.3 Hz, 1 H), 5.88 (m, 1 H), 5.37 (dd, J = 16.8, 1.6 Hz, 1

H), 5.28 (dd, J = 10.0, 2.0 Hz, 1 H), 5.16 (d, J = 10.8 Hz, 1 H), 5.06 (t, J =

9.8 Hz, 1 H), 4.78 (dd, J = 9.5, 7.1 Hz, 1 H), 4.45 (m, J = 2.7 Hz, 1 H),

3.88−3.98 (m, 1 H), 2.66−2.85 (m, 2 H), 2.33 (t, J = 13.4 Hz, 1 H), 1.99

(dd, J = 13.7, 3.4 Hz, 1 H), 1.32 (s, 3 H), 0.94 (m, 1 H), 0.59 (t, J = 7.2

Hz, 1 H), 0.41−0.53 (m, 1 H). MS (ESI) 428.2 [M + H]⁺.

Method A: To a solution of the triﬂuoromethanesulfonate above

(86.0 mg, 0.150 mmol) in DMF (0.7 mL) was added sodium

ethanethiolate (38.0 mg, 0.450 mmol). After being stirred at 25 °C

for 1.5 h, the reaction was quenched (satd NH₄Cl), extracted (2 ×

EtOAc), and washed (2 × brine). The combined organic layers were

dried (Na₂SO₄) and ﬁltered, and the ﬁltrate was concentrated under

reduced pressure. The residue was puriﬁed by ﬂash column

chromatography (SiO₂, 10% and 20% EtOAc/hexanes) to provide

(3S,5R,6S)-3-allyl-5-(3-chlorophen(yl)-6-(4-chlorophenyl)-1-((S)-1-

(ethylthio)butan-2-yl)-3-methylpiperidin-2-one (60.0 mg, 82%) as a

colorless liquid. ¹H NMR (400 MHz, CDCl₃) δ ppm 7.22 (d, J = 8.4 Hz,

2 H), 7.08−7.18 (m, 2 H), 6.90−7.03 (m, 3 H), 6.72−6.81 (m, 1 H),

5.76−5.98 (m, 1 H), 5.18 (d, J = 4.5 Hz, 1 H), 5.15 (s, 1 H), 4.64 (d, J =

10.6 Hz, 1 H), 3.32−3.45 (m, 1 H), 3.13 (m, 1 H), 2.73−2.90 (m, 1 H),

2.49−2.68 (m, 5 H), 2.14 (t, J = 13.7 Hz, 1 H), 1.98−2.10 (m, 1 H), 1.89

(dd, J = 13.5, 3.1 Hz, 1 H), 1.57 (dd, J = 6.9, 2.6 Hz, 1 H), 1.24−1.33 (m,

6 H), 0.52 (t, J = 7.5 Hz, 3 H). MS (ESI) 490.2 [M + H]⁺.

Compound 24 was prepared from (3S,5R,6S)-3-allyl-5-(3-chlor-

ophenyl)-6-(4-chlorophenyl)-1-((S)-1-(isopropylthio)butan-2-yl)-3-

methylpiperidin-2-one according to a similar procedure described for

the synthesis of 3. ¹H NMR (400 MHz, CDCl₃) δ ppm 7.22−7.26 (s, br,

2 H), 7.02−7.21 (m, 4 H), 6.92−6.94 (s, 1 H), 6.85 (d, J = 7.0 Hz, 1 H),

4.97 (d, J = 10.8 Hz, 1 H), 4.10 (t, J = 11.7 Hz, 1 H), 3.27−3.42 (m, 1 H),

2.98−3.16 (m, 3 H), 2.70−2.78 (m, 2 H), 2.40 (t, J = 13.9 Hz, 1 H),

2.08−2.26 (m, 1 H), 1.86−1.93 (m, 1 H), 1.49 (s, 3 H), 1.43 (d, J = 6.8

Hz, 3 H), 1.43 (d, J = 6.8 Hz, 3 H), 1.40−1.50 (m, 1 H) 0.41 (t, J = 7.5

Hz, 3 H). HRMS (ESI) m/z 554.1533 [M + H]⁺(C₂₇H₃₃Cl₂NO₅S

requires 554.1529).

Compound 22 was prepared from (3S,5R,6S)-3-allyl-5-(3-chlor-

ophenyl)-6-(4-chlorophenyl)-1-((S)-1-(ethylthio)butan-2-yl)-3-meth-

ylpiperidin-2-one according to a similar procedure described for the

2-((3R,5R,6S)-1-((S)-1-(tert-Butylsulfonyl)butan-2-yl)-5-(3-

chlorophenyl)-6-(4-chlorophenyl)-3-methyl-2-oxopiperidin-3-

yl)acetic Acid (25). Compound 25 was prepared from 55 according to

P

dx.doi.org/10.1021/jm401753e | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

1

a similar procedures described in method C. H NMR (500 MHz,

CDCl₃) δ ppm 7.21−7.26 (m, 2 H), 7.01−7.20 (m, 4 H), 6.92−6.94 (m,

1 H), 6.85 (d, J = 7.1 Hz, 1 H), 4.99 (d, J = 10.8 Hz, 1 H), 4.04 (dd, J =

13.2, 11.2 Hz, 1 H), 3.33 (t, J = 10.4 Hz, 1 H), 3.03−3.15 (m, 2 H), 2.80

(dd, J = 13.2, 2.0 Hz, 1 H), 2.72 (d, J = 15.4 Hz, 1 H), 2.43 (t, J = 13.8 Hz,

1 H), 2.08−2.23 (m, 1 H), 1.86 (dd, J = 13.7, 2.4 Hz, 1 H), 1.50 (s, 3 H),

1.46−1.49 (m, 1 H), 1.44 (s, 9 H), 0.41 (t, J = 7.6 Hz, 3 H). HRMS (ESI)

m/z 568.1687 [M + H]⁺(C₂₈H₃₅Cl₂NO₅S requires 568.1686).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-(cyclobutylsulfonyl)butan-2-yl)-3-methyl-2-oxopiperidin-3-

yl)acetic Acid (27). To a suspension of magnesium (306 mg, 12.6

mmol) in ether (7.4 mL) was added a solution of bromocyclobutane

(1.00 g, 7.41 mmol) in ether (7.4 mL) in several small portions at 25 °C.

After the initial exotherm had ceased, the mixture was further heated to

reﬂux for 1 h. The suspension was added in small portions to an ice-cold

solution of sulfuryl dichloride (3.00 g, 22.2 mmol) in DCM (12 mL).

The suspension was warmed to rt, and the volatiles were removed under

reduced pressure. The residue was dried under a vacuum and then

extracted with hexane (80 mL). The hexane suspension was ﬁltered, and

the ﬁlter cake was washed with hexanes. The combined ﬁltrates were

dried (Na₂SO₄) and concentrated under reduced pressure to give crude

cyclobutanesulfonyl chloride. The crude cyclobutanesulfonyl chloride

(1.15 g, 7.44 mmol) was added to a suspension of sodium sulﬁte (2.16 g,

17.1 mmol) in water (9.7 mL) and sodium carbonate (1.42 g, 13.4

mmol). The resulting suspension was heated to reﬂux for 1 h and then

cooled and lyophilized to remove water. Ethanol (50 mL) was added to

the residue, and the resulting mixture was heated under reﬂux for 2 h.

The mixture was ﬁltered and the ﬁltrate was concentrated under the

reduced pressure to give the crude cyclobutanesulﬁnic acid.

Compound 27 was prepared from 55 according to a similar

procedure described in method B, substituting cyclopropanesulﬁnic

acid for cyclobutanesulﬁnic acid. ¹H NMR (500 MHz, CDCl₃) δ ppm

7.21−7.27 (m, 2 H), 7.00−7.21 (m, 4 H), 6.93−6.97 (s, 1 H), 6.86 (d, J

= 7.1 Hz, 1 H), 4.97 (d, J = 10.8 Hz, 1 H), 3.98 (t, J = 12.2 Hz, 1 H), 3.76

(quin, J = 8.3 Hz, 1 H), 3.31 (t, J = 9.9 Hz, 1 H), 3.12 (ddd, J = 13.6, 10.9,

2.7 Hz, 1 H), 2.99 (d, J = 14.9 Hz, 1 H), 2.75 (d, J = 14.9 Hz, 1 H), 2.50−

2.69 (m, 3 H), 2.27−2.46 (m, 3 H), 2.04−2.19 (m, 3 H), 1.91 (dd, J =

13.7, 2.7 Hz, 1 H), 1.48−1.52 (m, 3 H), 1.41−1.47 (m, 1 H), 0.40 (t, J =

7.6 Hz, 3 H). MS (ESI) 566.2 [M + H]⁺.

2-((3R,5R,6S)-1-((S)-1-(tert-Butylsulfonyl)-3,3-dimethylbutan-

2-yl)-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-methyl-2-oxopi-

peridin-3-yl)acetic Acid (34). Compound 34 was prepared from 62

according to a similar procedure described in method C. ¹H NMR (500

MHz, CDCl₃) δ ppm 7.48 (s, br, 1 H), 7.38 (s, br, 1 H), 7.12−7.23 (m, 2

H), 7.05−7.11 (m, 2 H), 7.01 (1 H, s), 6.93−6.97 (m, 1 H), 5.23 (d, J =

11.2 Hz, 1 H), 3.99 (dd, J = 13.2, 11.5 Hz, 1 H), 3.70 (m, 1 H), 3.45 (m, 1

H), 3.00 (d, J = 15.2 Hz, 1 H), 2.77−2.84 (m, 2 H), 2.47 (t, J = 13.8 Hz, 1

H), 1.90 (dd, J = 13.9, 2.9 Hz, 1 H), 1.48 (s, 9 H), 1.45 (s, 3 H), 0.67−

0.73 (m, 9 H). HRMS (ESI) m/z 596.2003 [M + H]⁺(C₃₀H₃₉Cl₂NO₅S

requires 596.1999).

H), 1.26 (d, J = 6.8 Hz, 3 H). HRMS (ESI) m/z 540.1374 [M + H]⁺

(C₂₆H₃₁Cl₂NO₅S requires 540.1373).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-(isopropylsulfonyl)-3-methylbutan-2-yl)-3-methyl-2-oxopi-

peridin-3-yl)acetic Acid (2). Compound 2 was prepared from 57

according to a similar procedure described in method B. ¹H NMR (500

MHz, CD₃OD-d₄) δ ppm 7.00−7.23 (m, 7 H), 7.03 (m, 1 H), 5.13 (d, J

= 11 Hz, 1 H), 4.01 (dd, J = 13.7, 10.5 Hz, 1 H), 3.49−3.65 (m, 1 H),

3.32−3.37 (m, 1 H), 3.25−3.29 (m, 1 H), 3.11 (d, J = 13.7 Hz, 1 H), 2.99

(d, J = 13.7 Hz, 1 H), 2.61 (d, J = 13.5 Hz, 1 H), 2.31 (t, J = 13.7 Hz, 1 H),

2.18 (dq, J = 14, 6.9 Hz, 1 H), 2.05 (dd, J = 13.6, 2.8 Hz, 1 H), 1.41 (d, J =

6.9 Hz, 6 H), 1.37 (s, 3 H), 0.65 (d, J = 6.6 Hz, 3 H), −0.50 (d, J = 6.9 Hz,

3 H). MS (ESI) 568.0 [M + H]⁺.

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-cyclopropyl-2-(isopropylsulfonyl)ethyl)-3-methyl-2-oxopi-

peridin-3-yl)acetic Acid (39). Compound 39 was prepared from 56

according to similar procedure described in method C. ¹H NMR (400

MHz, CDCl₃) δ ppm 7.23−7.26 (m, 3 H), 7.04−7.20 (m, 3 H), 6.93−

6.97 (m, 1 H), 6.84−6.88 (m, 1 H), 4.93 (d, J = 10.8 Hz, 1 H), 4.34 (m, 1

H), 3.05−3.18 (3 H, m, 3 H), 2.86 (d, J = 13.3 Hz, 1 H), 2.77 (d, J = 15.3

Hz, 2 H), 2.46 (t, J = 13.8 Hz, 1 H), 1.86 (dd, J = 13.5, 2.5 Hz, 2 H), 1.51

(s, 3 H), 1.44 (d, J = 6.8 Hz, 6 H), 0.31−0.44 (m, 1 H), 0.18−2.80 (m, 1

H), −0.35−0.23 (m, 1 H), −1.15−1.02 (s, br, 1 H). HRMS (ESI) m/z

566.1536 [M + H]⁺(C₂₈H₃₃Cl₂NO₅S requires 566.1529).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-3-

methyl-1-((S)-1-(methylsulfonyl)propan-2-yl)-2-oxopiperidin-

3-yl)acetic Acid (41). Compound 41 was prepared from 61 according

1

to a similar procedure described in method C. H NMR (500 MHz,

CDCl₃) δ ppm 7.21−7.28 (m, 2 H), 7.08−7.15 (m, 4 H), 6.94−6.99 (m,

1 H), 6.83 (d, J = 7.1 Hz, 1 H), 4.95 (d, J = 10.8 Hz, 1 H), 4.44 (dd, J =

13.2, 11.3 Hz, 1 H), 4.14 (m, 1 H), 3.70 (m, 1 H), 3.00−3.10 (m, 1 H),

2.77 (dd, J = 2.45, 12.5, 1 H), 2.34 (t, J = 13.7 Hz, 1 H), 1.92 (d, J = 13.7

Hz, 1 H), 1.43 (s, 3 H), 1.27 (d, J = 6.85 Hz, 3 H). HRMS (ESI) m/z

512.1065 [M + H]⁺(C₂₄H₂₇Cl₂NO₅S requires 512.1060).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-cyclopropyl-2-(methylsulfonyl)ethyl)-3-methyl-2-oxopiperi-

din-3-yl)acetic Acid (43). Compound 43 was prepared from 56

according to a similar procedure described in method C. ¹H NMR (400

MHz, CDCl₃) δ ppm 7.27 (m, 4 H), 7.07−7.20 (m, 2 H), 6.96 (s, 1 H),

6.81−6.92 (m, 1 H), 4.87 (d, J = 10.56 Hz, 1 H), 4.44 (s, br,1 H), 3.01−

3.24 (m, 3 H), 3.00 (s, 3 H), 2.78 (m, 2 H), 2.43 (t, J = 13.79 Hz, 1 H),

1.71 1.97 (m, 2 H), 1.51 (s, 3 H), 0.33−0.49 (m, 1 H), 0.27 (s, br,1 H),

−0.26 (s, br,1 H), −1.03 (s, br, 1 H). HRMS (ESI) m/z 538.1223 [M +

H]⁺(C₂₆H₂₉Cl₂NO₅S requires 538.1216).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-3-

methyl-1-((S)-1-(methylsulfonyl)pentan-3-yl)-2-oxopiperidin-

3-yl)acetic Acid (21). To a solution of diethyl methylsulfonylmethyl-

phosphonate (48.8 mg, 0.212 mmol) in THF (1.2 mL) was added

butyllithium (1.6 M in THF, 127 μL, 0.203 mmol) at −78 °C under

nitrogen. Thirty minutes later, a solution of 52 (100 mg, 0.193 mmol) in

THF (0.8 mL) was added dropwise. After being stirred at −78 °C for 3

h, the reaction was quenched (iPrOH, EtOAc, and water). The reaction

was warmed to 25 °C, and the aqueous layer was extracted (2 × EtOAc).

The combined organics were washed (brine), dried (Na₂SO₄), ﬁltered,

and concentrated. The residue was puriﬁed by ﬂash column

chromatography (SiO₂, 0−75% EtOAc/hexanes, gradient elution)

provided (3R,5R,6S)-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-((2,2-

dimethyl-1,3-dioxolan-4-yl)methyl)-3-methyl-1-((E)-1-

(methylsulfonyl)pent-1-en-3-yl)piperidin-2-one (80.0 mg, 70%). MS

(ESI) 594.2 [M + H]⁺.

2-((3R,5R,6S)-1-((S)-2-(tert-Butylsulfonyl)-1-cyclobutylethyl)-

5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-methyl-2-oxopiperi-

din-3-yl)acetic Acid (36). Compound 36 was prepared from 63

according to a similar procedure described in method C. ¹H NMR (400

MHz, CD₃OD-d₄) δ ppm 7.29 (m, 3 H), 7.08−7.18 (m, 3 H), 7.04 (s, 1

H), 6.95−7.00 (m, 1 H), 5.06 (d, J = 10.96 Hz, 1 H), 3.87−4.00 (m, 1

H), 3.39−3.51 (m, 2 H), 2.89−3.07 (m, 3 H), 2.62 (d, J = 13.50 Hz, 1

H), 2.29 (t, J = 13.69 Hz, 1 H), 2.02 (dd, J = 13.60, 3.03 Hz, 1 H), 1.81−

1.92 (m, 1 H), 1.48−1.61 (m, 1 H), 1.44 (s, 9 H), 1.32−1.39 (m, 5 H),

1.29 (m, 1 H), 0.80−0.96 (m, 1 H). HRMS (ESI) m/z 594.1848 [M +

H]⁺(C₃₀H₃₇Cl₂NO₅S requires 594.1842).

The conjugated sulfone above (60.0 mg, 0.101 mmol) was dissolved

in ethanol (1.0 mL, 0.101 mmol) at 25 °C. Pd/C (5.37 mg, 5.05 μmol)

was added, and the reaction was allowed to stir under a hydrogen

balloon. The solvents were removed and ﬂash column chromatography

(SiO₂, 0−2% MeOH/DCM, gradient elution) provided 62 (50.0 mg,

83%).

2-((3R,5R,6S)-5-(3-Chlorophenyl)-6-(4-chlorophenyl)-1-((S)-

1-(isopropylsulfonyl)propan-2-yl)-3-methyl-2-oxopiperidin-3-

yl)acetic acid (37). Compound 37 was prepared from 61 according to

1

a similar procedure described in method C. H NMR (500 MHz,

CDCl₃) δ ppm 7.25 (m, 2 H), 7.02−7.16 (m, 4 H), 6.95 (s, 1 H), 6.83 (d,

J = 7.3 Hz, 1 H), 4.93 (d, 10.8 Hz, 1 H), 4.27 (dd, J = 13.0, 11.2 Hz, 1 H),

3.66−3.75 (m, 1 H), 3.03−3.11 (m, 2 H), 3.00 (d, J = 14.9 Hz, 1 H), 2.74

(d, J = 14.9 Hz, 1 H), 2.64 (dd, J = 13.2, 2.4 Hz, 1 H), 2.39 (t, J = 13.7 Hz,

1 H), 1.89 (dd, J = 13.8, 2.6 Hz, 1 H), 1.49 (s, 3 H), 1.42 (d, J = 6.8 Hz, 6

To a solution of 62 (75.0 mg, 0.126 mmol) in THF (1.2 mL) was

added 2,2,2-triﬂuoroacetic acid (71.7 mg, 0.629 mmol) at 25 °C. After

being stirred for 2 h, the reaction was concentrated under reduced

pressure. Flash column chromatography (SiO₂, 10% MeOH/DCM)

provided (3R,5R,6S)-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-(2,3-di-

Q

dx.doi.org/10.1021/jm401753e | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry

Article

hydroxypropyl)-3-methyl-1-(1-(methylsulfonyl)pentan-3-yl)piperidin-

2-one (45.0 mg, 64%).

Accession Codes

The coordinates of 25 with MDM2 have been deposited in the

To a solution of the diol above (45.0 mg, 0.081 mmol) and tempo

(5.57 mg, 0.036 mmol) in acetonitrile (8 mL) and pH 6.7 buﬀer (0.5

mL) at 35 °C were added solutions of sodium chlorite (23.8 mg, 0.210

mmol) in water (0.2 mL) and sodium hypochlorite (0.804 μL, 0.013

mmol) in water (0.2 mL) consecutively. After being stirred for 4 h, the

reaction was quenched (1N HCl and EtOAc). The aqueous layer was

extracted (EtOAc), and organics were washed (brine), dried (Na₂SO₄),

and concentrated. The residue was puriﬁed by ﬂash column

chromatography (SiO₂, 10% MeOH/DCM) to provide 2-

((3R,5R,6S)-5-(3-chlorophenyl)-6-(4-chlorophenyl)-3-methyl-1-((S)-

1-(methylsulfonyl)pentan-3-yl)-2-oxopiperidin-3-yl)acetic acid (7.00

PDB with accession codes 4OAS.

AUTHOR INFORMATION

Corresponding Author

*Phone: 650-244-2195. Fax: 650-837-9369. E-mail: daqings@

amgen.com.

■

Notes

The authors declare no competing ﬁnancial interest.

ACKNOWLEDGMENTS

1

■

mg, 16%). H NMR (500 MHz, CDCl₃) δ ppm 7.23−7.27 (m, 2 H),

7.14−7.18 (m, 1 H), 7.09−7.14 (m, 1 H), 7.05 (d, J = 4.89 Hz, 2 H), 6.95

(s, 1 H), 6.75 (d, J = 7.58 Hz, 1 H), 4.58 (d, J = 10.51 Hz, 1 H), 3.40 (m, 1

H), 3.00−3.20 (m, 3 H), 2.82 (d, J = 12.0 Hz, 1 H), 2.92 (s, 3 H), 2.15−

2.25 (m, 2 H), 2.05−2.10 (m, 1 H), 2.04 (m,1 H), 1.98 (s,1 H), 1.86 (m,

1 H), 1.51−1.57 (m, 1 H), 1.49 (s, 3 H), 0.61 (t, J = 7.34 Hz, 3 H).

HRMS (ESI) m/z 540.1374 [M + H]⁺(C₂₆H₃₁Cl₂NO₅S requires

540.1373).

We thank Simon Wong for the time-dependent inhibition (TDI)

determinations and Manuel Ventura and Dhanashri Bagal for the

acquisition of high resolution mass spectra.

ABBREVIATIONS USED

■

AcCN, acetonitrile; Boc, tert-butoxycarbonyl; BrdU, 5-bromo-2-

deoxyuridine; CMBP, cyanomethylenetributylphosphorane; CL,

clearance; CYP3A4, cytochrome P450 3A4; DCM, dichloro-

methane; DMF, N,N-diemethylformamide; DMSO, dimethyl-

sulfoxide; dr, diastereoselectivity ratio; EdU, 5-ethynyl-2′-

deoxyuridine; EtOAc, ethyl acetate; hPXR, human pregnane X

receptor; HTRF, homogous time-resolved ﬂuorescence;

KHMDS, potassium bis(trimethylsilyl)amide; MDM2, murine

double minute 2; MsCl, methanesulfonyl chloride; NMO, N-

methylmorpholinine N-oxide; QD, once a day dosing; qRT-

PCR, quantitative reverse transcription polymerase chain

reaction; SEM, standard error of mean; SPR, surface plasmon

resonance; TBAF, tetrabutylammonium ﬂuoride; TDI, time

dependent inhibition; TEA, triethylamine; TFA, triﬂuoroacetic

acid; THF, tetrahydrofuran

Determination of the Single Crystal Structure of 2. First, 0.75

mL of 60% EtOH/H₂O was added to 100 mg of material with stirring at

25 °C. After a couple of minutes the foam was replaced by white

crystalline material. The mixture was heated at reﬂux and then cooled to

rt slowly and allowed to stand without stirring. After a few days, large

crystals had formed. The material was collected by vacuum ﬁltration to

provide colorless needles.

A colorless rectangular rod of C₃₀H₄₁Cl₂NO₆S, approximate

dimensions 0.17 mm × 0.24 mm × 0.45 mm, was used for the X-ray

crystallographic analysis. The X-ray intensity data were measured at

100(2) K on a Bruker Kappa APEX II system equipped with a graphite

monochromator and a CuKα ﬁne-focus sealed tube (λ = 1.54178 Å)

operated at 1.2 kW power (40 kV, 30 mA). The detector was placed at a

distance of 5.0 cm from the crystal.

A total of 2464 frames were collected with a scan width of 0.5° in ω

and φ and an exposure time of 10 s/frame. The total data collection time

was 11 h. The frames were integrated with the Bruker SAINT software

package using a narrow-frame integration algorithm. The integration of

the data using a tetragonal cell yielded a total of 9052 reﬂections to a

maximum θ angle of 66.53° (0.8405 Å resolution), of which 4449 were

independent (redundancy 3.46), completeness = 92.1% (R_int= 5.21%,

R_sig= 6.70%) and 4290 (96.4%) were greater than >2σ(I) σ(F²). The

ﬁnal cell constants of a = 9.6944(4) Å, b = 9.6944(4) Å, c = 34.0641(15

)Å, α = 90°, β = 90°, γ = 90°, volume = 3201.4(2) Å³, are based upon the

reﬁnement of the XYZ-centroids of 6233 reﬂections above 20 σ(I) with

5.188° < 2θ < 133.077° . Analysis of the data showed negligible decay

during data collection. Data were corrected for absorption eﬀects using

the multiscan technique (SADABS). The ratio of minimum to

maximum apparent transmission was 0.53. The calculated minimum

and maximum transmission coeﬃcients (based on crystal size) are

0.2653 and 0.5046.

The structure was solved and reﬁned using the Bruker SHELXTL

(version 6.1) software package, using the space group P4(3), with Z = 4

for the formula unit, C₃₀H₄₁Cl₂NO₆S. The ﬁnal anisotropic full-matrix

least-squares reﬁnement on F²with 377 variables converged at R1 =

4.11%, for the observed data and wR²= 10.98% for all data. The

goodness-of-ﬁt was 1.058. The largest peak on the ﬁnal diﬀerence

electron density synthesis was 0.524 e−/Å³and the largest hole was

−0.327 e−/Å³with an RMS deviation of 0.050 e−/Å³On the basis of

the ﬁnal model, the calculated density was 1.275 g/cm³and F(000),

1304 e−.

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ASSOCIATED CONTENT

■

S

* Supporting Information

In vitro biological assays, in vivo study protocols, determination

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dx.doi.org/10.1021/jm401753e | J. Med. Chem. XXXX, XXX, XXX−XXX

Article Doi

Discovery of AMG 232, a potent, selective, and orally bioavailable MDM2-p53 inhibitor in clinical development

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