Discovery of Benzoazepinequinoline (BAQ) Derivatives as Novel

Subscriber access provided by UNIV OF NEW ENGLAND ARMIDALE

Article

Discovery of Benzoazepinequinoline (BAQ) Derivatives as Novel, Potent, Orally Bioavailable Respiratory Syncytial Virus Fusion Inhibitors Xiufang Zheng, Chungen Liang, Lisha Wang, Baoxia Wang, Yongfu Liu, Song Feng, Jim Zhen Wu, Lu Gao, Lichun Feng, Li Chen, Tao Guo, Hong C Shen, and Hongying Yun J. Med. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jmedchem.8b01394 • Publication Date (Web): 19 Oct 2018 Downloaded from http://pubs.acs.org on October 19, 2018

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Journal of Medicinal Chemistry

Discovery of Benzoazepinequinoline (BAQ) Derivatives as Novel, Potent, Orally Bioavailable Respiratory Syncytial Virus Fusion Inhibitors Xiufang Zheng†, Chungen Liang†, Lisha Wang†, Baoxia Wang†, Yongfu Liu†, Song Feng†, Jim Zhen Wu†, Lu Gao†, Lichun Feng†, Li Chen†, Tao Guo‡, Hong C. Shen† and Hongying Yun†, * †Roche

Pharma Research and Early Development，Roche Innovation Center Shanghai, Bldg 5, 720 Cailun

Road, Shanghai, 201203 China ‡International

Discovery Service Unit, Research Service Division, WuXi AppTec (Shanghai) Co., Ltd.

Lane 31, Yiwei Road, Waigaoqiao, Shanghai, 200131 China KEYWORDS: Respiratory Syncytial Virus (RSV), Antiviral, Fusion inhibitors, Benzoazepine, Quinoline, Oxetane ABSTRACT: A novel benzoazepinequnoline (BAQ) series was discovered as RSV fusion inhibitors. BAQ series was originated from compound 2, a hit from similarity-based virtual screening. In SAR exploration, benzoazepine allowed modifications in the head moiety. Benzylic sulfonyl on benzoazepine and 6-Me on quinolone were crucial for good anti-RSV activity. Although the basic amine in the head portion was crucial for anti-RSV activity, the attenuated basicity was required to reduce Vss. Introducing oxetane to the head portion led to discovery of compound 1, which demonstrated single-digit nM anti-RSV activity against different RSV strains, reasonable oral exposure in plasma and 78-fold higher exposure in lung. Compound 1 also displayed > 2 log viral reduction in a female BALB/c mice RSV model by BID oral dosing at 12.5 mg/kg. A single resistant mutant at L138F in fusion protein proved compound 1 to be a RSV fusion inhibitor.

ACS Paragon Plus Environment

Journal of Medicinal Chemistry 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Introduction As a negative-sense single-stranded RNA virus, human respiratory syncytial virus (RSV) belongs to the family of Paramyxoviridae.1 The most common cause of human RSV infection is acute upper and lower respiratory tract infection in infants and young children. Almost all children by age of three can be infected by RSV at least once.2 RSV infection is normally mild and associated with upper respiratory tract symptoms in normal adults and elder children. However, populations associated with high-risk factors, such as premature birth, congenital heart disease, chronic pulmonary diseases, and immunocompromised conditions, are more prone to mortality and morbidity due to lower respiratory tract infections. Severe RSV infection often leads to bronchiolitis and pneumonia with an increased chance of morbidity or mortality in young children and immunocompromised adults.2,3 Furthermore, the sequela of severe RSV infection at young age is recurrent wheezing or asthma,4 and RSV-related mortality rate becomes higher in high-risk populations.5,6 Although numerous attempts in inactivated subunits and live-attenuated vaccination are under development, there is no available vaccine for RSV infection. Palivizumab, a humanized monoclonal antibody against RSV fusion protein, was approved for prophylaxis in high-risk infants in 1998. However, palivizumab showed no efficacy in the treatment of established RSV infection.7 Ribavirin, an aerosol formulation comprising ribavirin, is the only approved antiviral therapy for RSV infection. Due to limited efficacy and side effects, it is rarely used in clinic.8 As such, safe and effective therapy for RSV infection is a high unmet medical need.


Page 2 of 42

Page 3 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


OH HO

HO

N

N H 2N

N

N H

O

N

N

N N H

N

JNJ-2408068

Cl

O H N O S O

N N

N N N N N

N N

OH

BMS-433771

N N N N N

O O S HN

N

GS-5806

OH

N

N NH 2

OH

OH

O N

N H

TMC-353121

N N N

N

N

Cl

OH

MDT-637

O

F

N

F

N

NH 2

N

F

N

JNJ-53718678

7

Figure 1. Selected RSV fusion inhibitors under development. RSV Fusion (F) protein is a surface glycoprotein on the viral envelope. The F protein plays a major role for the virus entry into host cells together with the G surface glycoprotein.9 Depletion of the RSV G protein can only reduce the efficiency of entry process but cannot terminate the entry process. Moreover, the RSV F protein also promotes syncytia formation between infected cell and adjacent cells. Inhibition of viral entry and spread by targeting RSV F protein has emerged as a promising treatment for RSV infected patients. Small molecule RSV F protein inhibitors have great potential to decrease the duration and severity of respiratory symptoms, and the subsequent risk of prolonged hospitalization and complications. In the past, several structurally different and potent RSV F protein inhibitors were reported.10-13 A number of RSV F inhibitors had successfully progressed to preclinical stage, but only a few had entered clinical development.14 JNJ-2408068 was a picomolar RSV F inhibitor (EC50: 0.16 nM) with low cytotoxicity against several lab strains. However its long tissue retention time in the tested species hindered its further development.11 Another picomolar RSV F inhibitor, TMC-353121, exhibited anti-RSV efficacy in inhibiting RSV replication prophylactically and therapeutically.12 BMS-433771 was the first RSV F inhibitor to demonstrate oral bioavailability,13 but was terminated for further development likely due to business reason. VP-14637, a nanomolar RSV F inhibitor, was formulated and used as an inhaled dry



Page 4 of 42

powder product called MDT-637 in Phase I trials. This molecule demonstrated good tolerance in healthy human subjects.15 Most recently, GS-5806 achieved proof-of-concept in human RSV challenge studies and showed a significant viral load reduction (4.2 log10) and a significant disease severity reduction.16 In a Phase IIa trial with the RSV-A Memphis 37b virus inoculated healthy adults, daily oral dosing JNJ53718678, a picomolar RSV F inhibitor, reduced viral load and clinical severity, which established clinical proof of concept of RSV treatment by using RSV F inhibitor. 17 In our endeavors to develop highly potent RSV F inhibitors, we used a similarity-based virtual screening approach and generated several hits as chemistry starting points. In our similarity-based virtual screening, MOS (2D similarity) and ROCS (3D similarity) were used in parallel to screen the Roche Smart library. JNJ-2408068, TMC-353121and BMS-433771 were chosen as reference compounds for the virtual screening. In our previous publication, we reported compound 7 as a validated hit as RSV F inhibitor by this approach.18 In this article, we describe the discovery of benzoazepinequnoline (BAQ) series derived from compound 2, another hit from the similarity-based virtual screening approach (Figure 2). h

HN OH N

HN

NH 2

N

2

Ring expansion

R

2

1

R

R

N

N X

BAQ

Figure 2. BAQ series originated from ring expansion of compound 2. Results and Discussion As aforementioned, BAQ series was originated from compound 2. We were delighted to see that the reasonable anti-RSV activity of compound 2 in CPE assay (EC50=0.22 M). Further exploration disclosed that ring size reduction to 5-membered ring 3 resulted in a total loss of anti-RSV activity and strong cytotoxicity. The anti-RSV activity remained the same with ring expansion to a 7-membered ring (4, EC50=0.22 M).18


Page 5 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Table 1. SAR results of R2 of the scaffold derived from compound 2a HN OH

NH 2

2

R

N

Compd

2

3

4 a EC : 50

R2 N

N

N

EC50 (µM)a

CC50 (µM)

TIb

0.22

21

95

>100

2.9

0

0.22

21

95

the concentration of compound that reduced 50% cytopathic effect of RSV Long strain infected HEp-2 cells.

CC50: the concentration of compound that manifests cytotoxicity towards 50% of uninfected HEp-2 cells.

b

Therapeutic index (TI) is the ratio of CC50/EC50.

To explain the correlation of ring size to anti-RSV activity, we did the conformation analysis (Figure 3). Compound 3 (a 5-membered ring analogue) adopted almost a planar conformation. Compound 2 (a 6-membered ring analogue) adopted a conformation with nearly 40-degree dihedral angle between quinoline and tetrahydroisoquinoline. Finally, compound 4 (a 7-membered ring analogue) adopted a conformation with almost 90-degree dihedral angle between quinoline and benzoazepine. These observations gave us a hint that the active RSV inhibitors may prefer a non-planar scaffold with a dihedral angle between two ring systems up to 90 degrees.



Page 6 of 42

Figure 3. Conformation of 2, 3 and 4 at the lowest energy. Compound 2 in orange, compound 3 in pink and compound 4 in green.

An exploration on the head portion (Rh) of compound 2 revealed a flat SAR (Table 2). All the modifications on Rh resulted in a significant loss of antiviral activity. Furthermore, derivatives with basic terminals (2b, 2d and 2f) demonstrated better antiviral activity than the non-basic terminal derivatives (2a, 2c and 2g). The chain length between the two amino groups of Rh also impacted on activity. For example, two-carbon linker derivatives (2d and 2f) provided better anti-RSV activity than a three- and a four-carbon linker derivatives (2b and 2e).

Table 2. SAR exploration of the head portion (Rh) of the scaffold derived from compound 2a h

HN

R

N

Compd

2

N

Rh

NH 2 OH

EC50 (µM)a

CC50 (µM)

TIb

0.22

21

95


Page 7 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


2a

OH

22

66

3

N H2

7.8

8.4

1

> 100

89

0

2.3

15

7

> 100

7.4

0

2.3

> 100

43

15

80

5

OH

2b

2c

OH N H2

2d

N H2

2e H N

2f

NH 2 NH

H N

2g

NH 2 O

a EC : 50



b


Further profiling of compound 2 revealed a very low permeability (0  10-6 cm/s) in the PAMPA assay, which indicated a very low cellular permeability potential. Thus compound 2 was not suitable for oral dosing. Considering the flat SAR of the derivatives of compound 2 on Rh and poor permeability of compound 2, we turned our attention to compound 4 (BAQ). Compound 4 showed identical activity and TI to compound 2. However, different from compound 2, compound 4 allowed further modification on Rh without sacrificing potency. For example, compound 5, a close analogue to 2d, showed the identical good activity to compound 4. Moreover, the PAMPA of compound 5 was 0.99, which was categorized as medium to high cell permeability.



Page 8 of 42

Table 3. Head portion (Rh) exploration on the scaffold derived from compound 4a h

HN

R

N

N

Compd

Rh

TIb

0.22

21

95

0

0.20

23

115

0.99

OH

N H2

5 a EC : 50

CC50 (µM)

NH 2

4

PAMPA Peff

EC50 (µM)a

(10-6 cm/s)


CC50: the concentration of compound that manifests cytotoxicity towards 50% of uninfected HEp-2 cells. b Therapeutic index (TI) is the ratio of CC50/EC50.

Subsequently, the rat single-dose pharmacokinetics (SDPK) of compound 5 showed reasonable oral exposure with moderate oral bioavailability (Table 4). Both in vitro rat liver microsomal clearance and in vivo clearance were medium to high, which needed to be further optimized. The high volume of distribution (Vss) was most likely due to the lipophilic basic nature of compound 5.

Table 4. The DMPK profile of compound 5a RLMb Compd

(mL/min/kg)

AUC0-INF

AUC0-INF

(iv 2 mg/kg)

(po 10 mg/kg)

F (%) in vivo CL (mL/min/kg)

5

23 (medium)

(h*ng/mL)

(h*ng/mL)

324

510


89 (high)

Vss (L/kg)

106

32

Page 9 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


a The

SDPK study in male Wistar rats was carried out according to the standard procedures described in the supporting

information. Plasma clearance (CL), volume of distribution at steady state (Vss), area under the curve (AUC), oral bioavailability (F), rat liver microsomal clearance (RLM).

In the resistant mutation selection study, a single mutant at D331N in RSV F protein was identified in compound 5 resistant virus at passage 7. The anti-RSV activity was dropped from 0.20 M to 2.3 M in resistant virus. It proved that compound 5 was targeting on RSV fusion protein.

Encouraged by the good potency, reasonable oral absorption, and modeling understanding, we selected compound 5 as a new chemistry starting point for further optimization. Chemistry modification was initiated at R1 on quinoline. Selected data are summarized in Table 5. A Cl-walk on quinolone revealed that 6-position was critical for antiviral potency. SAR exploration at the 6-position disclosed that the 6-Me analogue (5f) demonstrated the best potency and the largest TI although it still showed strong cytotoxicity like the other R1 derivatives. The cytotoxicity cannot be addressed by optimizing R1 only, but was eventually addressed by the combined optimization on both R1 and X, which was described in Table 6.

Table 5. SAR exploration of R1 of the quinolone scaffold derived from compound 5a NH 2

HN 1

6

R

N

N

Compd

R1

EC50 (µM)a

CC50 (µM)

TIb

5

H

0.20

23

115

5a

5-Cl

6.6

23

3

5b

6-Cl

0.12

>3.2

>27



Page 10 of 42

5c

7-Cl

>3.2

7.5

2

5d

8-Cl

1.1

25

23

5e

6-OMe

0.32

4.8

15

5f

6-Me

0.073

9.4

129

5g

6-F

1.5

8.0

5

5h

6-CF3

1.1

6.7

6

5i

6-SO2Me

5.6

13

2

a EC : 50



b


As described in Table 4, compound 5 showed medium to high in vitro and in vivo clearance. Based on the metabolic identification (Met ID) study in mouse and rat liver microsomes (Figure 4), the benzoazepine moiety was identified as a metabolic “hot spot” generating 90% of metabolites in rat liver microsomes. In particular, the benzylic position (X in Figure 2) and benzene ring of benzoazepine were highly likely to be accountable for metabolism. In the head moiety, amide and acid were generated by oxidation of the carbon adjacent to the terminal amine. All these Met ID results cast lights on how to further optimize compound 5.

Figure 4. Met ID of compound 5 in rat and mouse liver microsomes.


Page 11 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Our optimization strategy focused on reducing the electronic density of benzene (Table 6). Two approaches were explored. The first was to introduce fluorine to benzene to block a metabolic “hot spot”, which unfortunately failed (6f and 6g). The other approach was to replace the benzyl carbon (X in Figure 2) with heteroatoms, which was proven to be very successful. In particular, introducing the electron withdrawing group SO2 led to discovery of compounds 6a and 6h (R1 = H or Me and X = SO2), which led to significantly improved metabolic stability. The liver microsomal clearance in tested species were low. In addition, compounds 6a and 6h also demonstrated superior anti-RSV activity and good TI compared to other modifications. Compound 6h demonstrated the best anti-RSV activity (0.004 M) and TI (16,500), as shown in Table 6. Table 6. X optimization impacts on both potency and XLM

1

NH 2

HN

R

2

R

N

3'

N X

HLM/RLM/MLM Compd

R1

R2

X

EC50 (µM)a

CC50 (µM)

TIb (mL/min/kg)c

5

H

H

CH2

0.20

23

115

10/23/19

6a

H

H

SO2

0.011

94

8,545

0/3/0

6b

H

H

O

0.23

>10

>43

14/20/25

6c

Me

H

CF2

0.062

6.6

106

NA

6d

Me

H

NH

0.23

12

52

5/43/19

6e

Me

H

0.31

55

177

0/21/11

HN

O

N H

6f

Me

3’-F

CH2

0.46

>3.2

>7

8/33/31

6g

Me

2’-F

CH2

0.10

>3.2

>32

6/160/24



6h a EC : 50

Me

H

SO2

0.004

66

Page 12 of 42

16,500

5/8/1


CC50: the concentration of compound that manifests cytotoxicity towards 50% of uninfected HEp-2 cells. b Therapeutic index (TI) is the ratio of CC50/EC50. cScaled intrinsic clearance of compounds in human, rat and mouse liver microsomes (HLM, RLM or MLM).

In the next round of optimization, the head portion (Rh) was re-explored with the optimal 6-Me and X as SO2 (Table 7). We observed a much broader tolerance of substitution on Rh. The length of alkyl chain between two amino groups was identified to be crucial for antiviral activity. Compound 6h and compound 7a were optimal for excellent anti-RSV activity (EC50=0.004 M and 0.001 M, respectively). Methyl substitution on either terminal amine (7d and 7e) or alkyl chain (7f to 7i) reduced the ant-RSV activity. Replacing the terminal amine by heterocycles (from 7j to 7m) significantly reduced the anti-RSV activity. Oxetanylation of terminal amine (7s) led to much reduced anti-RSV activity. Acetylation of the terminal amine (7p) totally removed the anti-RSV activity. Clearly introducing electron withdrawing group at the terminal amine would most likely reduce anti-RSV activity. In addition, replacing terminal amine with hydroxyl group (7q) resulted in a 10-fold decrease in anti-RSV activity compared to compound 6h. It was noteworthy that oxetane analogues (7r and 1) both demonstrated excellent anti-RSV activity at single-digit nM level (EC50=0.005 M and 0.002 M, respectively). Compound 1 also demonstrated the highest therapeutic index (TI >50,000) throughout the whole chemical series in the CPE assay. Table 7. SAR exploration at the Rh position with fixed 6-Me and X = SO2a h

HN N

R

N S O O


Page 13 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Compd

Rh N H2

6h 7a

N H2 N H2

7b 7c

N H2

EC50 (µM)a

CC50 (µM)

TIb

0.004

66

21,500

0.001

21

21,000

0.007

92

13,142

0.027

21

778

7d

H N

0.014

29

2,071

7e

N

0.037

89

2,405

7f

NH 2

0.010

25

2,500

0.014

86

6,143

0.025

25

1,000

0.012

23

1,917

0.15

40

267

0.54

>100

>185

N H2

7g

7h

NH 2

N H2

7i

7j

7k

NH N

O N



7l

N

H N

7m

Page 14 of 42

0.28

22

79

0.071

23

324

0.039

>100

>2,564

0.026

22

846

3.1

66

21

0.041

>100

>2,439

0.005

86

17,200

0.031

>100

3,226

0.002

>100

>50,000

O

7n

N H2

N H2

7o

O H N

7p

O

OH

7q O

7r

7s

1 a EC : 50

N H2

H N O NH 2 O



Since compound 1 was identified with the best potency and therapeutic index, it was further profiled against different RSV strains in CPE assay. To our delight, compound 1 demonstrated consistent and high anti-RSV activity against different RSV strains in CPE and plaque reduction assays. Regarding selectivity, compound 1 showed no activity to influenza H1N1 in CPE assay (Table 8), which indicated that it did not target on a broad-antiviral host target.


Page 15 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Table 8. Anti-RSV activity and selectivity of compound 1a RSV Long

RSV A2

RSV B

Plaque reduction

Influenza H1N1

EC50 (µM)

EC50 (µM)

EC50 (µM)

EC90 (µM)

EC50 (µM)

0.002

0.004

0.002

0.003

100

Compd 1 a

EC50: the concentration of compound that reduced 50% cytopathic effect of RSV Long, A2 and B strains, plaque

reduction in long strain and influenza H1N1 strain infected HEp-2 cells.

ADME and in vivo efficacy assessment of selected BAQs. The physicochemical properties and ADME of selected BAQs were listed in Table 9. The mlLogD had correlation with pKb of the head amine except for compound 7q. Compound 7q, the one without ionizable head amine showed high lipophilicity (mlLogD), good permeability but low aqueous solubility. On the contrary, all the other terminal amino analogues demonstrated good solubility. In term of pKb, it was known that oxetane as a carbonyl surrogate can reduce pKb of adjacent amine.20 Introducing oxetane next to the terminal amine in compound 1 led to the reduction of the pKb more than 2 log units compared to any other analogues containing a terminal amino group. The impact of oxetane on the pKb was quickly diminished with the elongated distance to terminal amine such as compound 7r. Compounds 1 and 7r demonstrated high clearance in liver microsomal clearance assay. Except for compound 6h, all the tested analogues showed good permeability in PAMPA assay. In addition, the plasma protein binding of tested compounds were medium to high.

Table 9: Physicochemical and ADME properties of compounds 1, 5, 6h, 7q and 7r. Compd

pKbb

Lysac

head amine

(µg/mL)

HLM/RLM/MLM

PAMPAe

PPBf

(10−6 cm/s)

(human/mouse)

mlLogDa d (mL

min−1 kg−1)

1

2.20

8.0

482

17/95/204

0.73

2.4/11

5

0.73

10.4

>460

10/23/19

0.99

NA



a

Page 16 of 42

6h

0.88

10.4

>466

5/8/1

0.12

7.8/8

7q

2.57

none

9

36/81/120

1.53

NA

7r

1.12

9.9

>480

12/37/23

0.59

3/10

Machine learning LogD. b Predicted by MoKa v2.0.

c

Lyophilisation solubility assay (LYSA) (µg/mL). d Scaled

intrinsic clearance of compounds in human, rat and mouse liver microsomes (HLM, RLM or MLM).

e

Parallel

Artificial Membrane Permeability Assay (10-6 cm/s). e Plasma protein binding assay (% unbound fraction).

In the SDPK study in male Wistar rats (Table 10), compounds 1, 5, 7q and 7r all demonstrated reasonable oral exposure. Compound 6h did not show any oral exposure, which could be due to its low cellular permeability. The most important finding was that the Vss of compound 1 was as low as that of compound 7q, which did not contain a basic head group. It is well known that basic lipophilic molecules typically lead to a high volume of distribution, which may cause unexpected accumulation of molecule in undesired tissues. Hence the project team made the decision to choose compound 1 for further characterization. Table 10. Rat SDPK profiles of selected BAQ compoundsa Compd

1

5

6h

7q

7r

t1/2 (iv, h)

1.7

12

4.5

1.0

4.1

Vss (iv, L/kg)

4.1

73

17

2.5

20

51

106

97

33

53

686

324

346

717

466

558

510

0

1640

1267

CL (iv) (mL min−1 kg−1) Plasma AUC(0-∞) (iv, µg/L*hr) AUC(0-∞) (po, µg/L*hr)


Page 17 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


F (%) a

16

32

0

59

29

The SDPK study in male Wistar rats was carried out according to the standard procedures. (iv: 2 mg/kg; po: 10

mg/kg). Major parameters, including plasma clearance (CL), volume of distribution at steady state (Vss), T1/2 (i.v.), area under the cure (AUC), and oral bioavailability (F) are reported.

Since the in vivo anti-RSV model was run in mice, the additional SDPK in IRC mice was run with compound 1 at different doses (Table 11). The Vss in mouse SDPK was very close to the one measured in rat SDPK. The oral exposure was over dose proportional, which could be due to the saturation of metabolic enzymes at high doses. In the medium dose (200 mg/kg), the lung exposure was also analyzed. It was very interesting to see the exposure of lung was 78-fold higher than plasma. It is known that basic drugs are bound to certain binding sites containing phosphatidylserine.21 Phosphatidylserine is highly expressed in the surface of epithelium cells in lung. Compound 1 could accumulate in lung as a counter ion to phosphatidylserine, which is quite acidic. This is perhaps the most convincing explanation of high exposure of compound 1 in lung. To our RSV fusion inhibitor project, high exposure in lung was an advantage. As we previously mentioned, an overly high Vss might lead to unexpected accumulation and potential toxicity risk for basic molecule. We utilized oxetane to control pKb of head amine and minimized the potential risk of toxicity. Table 11. Mouse SDPK profiles of compound 1a Dose (mg/kg) t1/2 (po, h)

50

200

450

1.3

2.3

2.9

Vss (iv, L/kg)

4.7

CL (iv) (mL min−1 kg−1)

222

AUC(0-∞) (iv, µg/L*hr)

150

Plasma AUC(0-∞) (po, µg/L*hr)

1,352

9,884

44,532

F (%)

36

66

132



Lung

AUC(0-∞) (po, µg/L*hr)

AUC(0-∞), lung/AUC(0-∞), plasma (po) a The

NA

769,641

NA

NA

78

NA

SDPK study in male ICR mice was carried out according to the standard procedures. (iv: 2 mg/kg; po: 50, 150,

450 mg/kg). Major parameters, including plasma clearance (CL), volume of distribution at steady state (Vss), T1/2 (i.v.), area under the cure (AUC), and oral bioavailability (F) are reported.

Finally, compound 1 was tested in a mouse RSV model (Figure 5). This compound demonstrated 1 log unit of viral titer reduction at 12.5 mg/kg by oral dosing twice a day (BID). The anti-RSV efficacy was dose dependent. The RSV titer was below detectable limit at 50 mg/kg and 150 mg/kg, po, BID. Typically, an antiviral drug needs to have enough exposure to maintain antiviral activity, and one criterion is that the plasma Cmin is higher than IC90. In our case, the oral T1/2 of 50 mg/kg was only 1.25 h, the total plasma drug concentration was below IC90 after 8 hours. It could be assumed that at 12.5 mg/kg, po, BID dosing, the in vivo efficacy was not likely to be observed. However, considering the high lung exposure of compound 1, we speculated that the observed good anti-RSV efficacy at such a low dose was a result of the high lung exposure and longer lung T1/2 of compound 1. Unfortunately, we were unable to test the drug concentration in lung in RSV mouse model due to the limited amount of samples. We plan to provide more evidences on the relationship of pKb and lung distribution in our succeeding manuscript.

Log (p.f.u./g lung)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 18 of 42

4 3.5 3 2.5 2 1.5 1 0.5 0

Baseline

Vehicle

150 mg/kg

50 mg/kg

25 mg/kg

Figure 5. In vivo Reduction of RSV Titer in Mouse Lung with compound 1 (po, BID)a


12.5 mg/kg

Page 19 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


a

Female BALB/c mice were used for in vivo efficacy assay. Animals were anesthetized and to the

animals were intranasally administrated RSV long strain (5 105 plaque-forming units [PFU]). After oral administration of compound 1 for 4 days, the mice were euthanized with CO2, and lungs were harvested and analyzed for viral titer.

There were more examples carrying oxetane with diverse modifications on 6-position and SO2 replacement in our previous publication.19 Among those modification, compound 1 remained the top molecule considering anti-RSV activity, TI, DMPK and toxicity profiles. In the resistant mutation selection study, a single mutant at L138F in RSV F protein was identified in compound 1 resistant virus at the passage 7. The anti-RSV activity of compound 1 was dropped from 2 nM to 2.56 M, which proved that the target of compound 1 was indeed RSV fusion protein. The single resistant mutation study provided us the opportunity to seek the potential binding site of compound 1 in RSV F protein. Actually, L138 located quite close to the binding pocket of RSV pre-fusion protein with JNJ-2408068 reported by McLellan, et al.22 We docked compound 1 into the same threefoldsymmetric cavity in profusion RSV F-protein. We observed that the 6-Me entered into a small hydrophobic pocket and picked more hydrophobic interaction with protein, such as Met 396. The oxygen atoms of sulfone formed strong hydrophobic interaction with Phe 140. The model suggested that the oxetane moiety did not form any interaction with the protein, and it mainly controlled the conformation. All these 3 factors possibly accounted for 100-fold potency increase compared to original hit, compound 2. The docking model also demonstrated that the Leu 138 was very close to the binding pocket, and L138F mutation may change the conformation of the binding pocket. This appeared to elucidate the huge drop of the antiviral activity of compound 1 to this resistant virus.



a)

b)

Figure 6. Docking model of compound 1 binding to a three-fold symmetric cavity in prefusion RSV F glycoprotein (the docking procedure was described in the supporting information). a) Side view of the compound 1 in the binding pocket of prefusion FSV F glycoprotein (shown in white). Leu138 was shown as a stick with color in orange. Compound 1 was shown as ball-and-stick, with carbon atoms colored in green, nitrogen atoms in blue, and oxygen atoms in red; b) 2D ligand-interaction diagram was generated in MOE. The interactions of compound 1 with RSV F main-chain and side-chain atoms were shown as blue and green dotted lines, respectively. And the arrowhead pointed to the acceptor.

Chemistry As exemplified by the synthesis of compound 1, BAQ deriatives described herein were mostly prepared according to the procedure shown in Scheme 1. Starting with 2-sulfanylbenzoic acid 8, esterification with methanol gives methyl 2-sulfanylbenzoate 9, which was followed by annulation with 2-chloro ethylamine to afford 3,4-dihydro-1,4-benzothiazepin-5(2H)-one 10. Reduction of compound 10 with LiAlH4 provided 2,3,4,5-tetrahydro-1,4-benzothiazepine 11, which was converted to 13 via acetylation and oxidation. The subsequent deacylation afforded the desired building block 14, which underwent a coupling reaction with 2,4-dichloro-6-methyl-quinoline to generate the key intermediate 15. Another coupling with 3(aminomethyl)-N,N-dibenzyloxetan-3-amine yielded compound 16, which was finally de-benzylated to afford 1 after HPLC purification. The structure of compound 1 could be successfully assigned by


Page 20 of 42

Page 21 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


spectroscopic analysis on the basis of 1 H, 13C, H-H COSY, H-C HSQC, and H-C HMBC NMR experiments. This efficient synthetic route was successfully applied to the synthesis of the BAQ analogues described in this paper. Scheme 1a O

O

a

HO

O

HS

HS 8

d

e

11

14

O

h

N

NH 2

HN O

i

N

S O O

g

S O O

Bn N Bn

HN

Cl N

H N

S O O 13

12

15

S

f

N

S

N

c

O

N

H N

O

S 10

9

O

aReagents

H N

b

N

N S O O

16

S O O 1

and conditions: (a) H2SO4, MeOH, reflux, 18 h, 91% (crude); (b) 2-chloroethanamine hydrochloride,

NaH, DMF, rt, overnight, 47%; (c) LiAlH4, THF, reflux, 18 h, 90%; (d) Ac2O, Et3N, DCM, 0 oC to rt, 1 h; (e) mCPBA, DCM, 0 oC to rt, 1 h; (f) NaOH, EtOH/H2O, reflux, overnight, 77%; (g) 2,4-dichloro-6-methyl-quinoline, 1-butanol, MW., 160 oC, 2 h; (h) 3-(aminomethyl)-N,N-dibenzyloxetan-3-amine, PdCl2(dppf), dppf, t-BuONa, dioxane, MW., 120 oC, 1 h, 65%; (i) Pd(OH)2, H2, MeOH, rt, 16 h, 17%.

Conclusions In summary, we have discovered a novel series of BAQ derivatives as RSV fusion inhibitors derived from compound 2, a similarity-based virtual screening hit. In the SAR exploration of the BAQ series, the combination of sulfonyl group at the benzylic position of benzoazepine and 6-Me on quinolone led to singledigit nM anti-RSV compounds (1, 6h, 7a, 7b and 7r). Sulfonyl group replacement at the



benzylic position of benzoazepine also significantly improved the metabolic stability (compound 6a vs. compound 5). Although the basic terminal amine on the head region was crucial for anti-RSV activity, it was discovered that reduced basicity was important for reducing Vss. Oxetane was introduced adjacent to the terminal NH2, which led to discovery of compound 1 with much reduced Vss and intact anti-RSV activity. Compound 1 was the most balanced molecule in the BAQ series. It demonstrated good anti-RSV activity against different RSV strains in CPE assays (IC50: 2 nM in long strain, 4 nM in A2 strain, and 2 nM in B strain), and reasonable oral exposure in plasma and 78-fold higher exposure in lung. It also demonstrated good anti-RSV activity (> 2 log units) in a female BALB/c mice model by BID oral dosing at 12.5 mg/kg. In the resistant mutation selection study, a single mutant at L138F in RSV F protein was identified in compound 1 resistant virus, which confirmed compound 1 as a RSV fusion inhibitor. Based on its favorable in vitro and in vivo properties, compound 1 was considered as a promising clinical candidate for RSV treatment. EXPERIMENTAL SECTION Synthetic Chemistry General Comments. Intermediates and final compounds were purified by flash chromatography using one of the following instruments: i) Biotage SP1 system and the Quad 12/25 Cartridge module. ii) ISCO combi-flash chromatography instrument. Silica gel Brand and pore size: i) KPSIL 60 Å, particle size: 40-60 µM; ii) CAS registry NO: Silica Gel: 63231-67-4, particle size: 47-60 micron silica gel; iii) ZCX from Qingdao Haiyang Chemical Co., Ltd, pore: 200-300 or 300-400. Intermediates and final compounds were purified by preparative HPLC on reversed phase column using X BridgeTM Perp C18 (5 µm, OBDTM 30 × 100 mm) column or SunFireTM Perp C18 (5 µm, OBDTM 30 × 100 mm) column. LC/MS spectra were obtained using a MicroMass Plateform LC (WatersTM alliance 2795-ZQ2000). Standard LC/MS conditions were as follows (running time 6 minutes): Acidic condition: A: 0.1% formic acid in H2O; B: 0.1% formic acid in acetonitrile; Basic condition: A: 0.01% NH3·H2O in H2O; B: acetonitrile; Neutral condition: A: H2O; B: acetonitrile. Mass spectra (MS): generally only ions which indicate the parent mass are reported, and unless otherwise stated the mass ion quoted is the positive mass ion (M+H)+.


Page 22 of 42

Page 23 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


The microwave assisted reactions were carried out in a Biotage Initiator Sixty. NMR Spectra were obtained using Bruker Avance 400MHz. All reactions involving air-sensitive reagents were performed under an argon atmosphere. Reagents were used as received from commercial suppliers without further purification unless otherwise noted. All of the final compounds had purities greater than 95% based upon HPLC, LC/MS, and 1H NMR analyses. All of the reported yields are for isolated products and are not optimized. General Synthetic Procedures for the Synthesis of BAQ Series with N-[(3-Aminooxetan-3-yl)methyl]2-(1,1-dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4-amine (1) Exemplified. To a cooled solution of concentrated sulfuric acid (72 g) in methanol (1.5 L) at 0 oC, was added 2sulfanylbenzoic acid (8, 300 g, 1.95 mol) in portions under argon atmosphere. After being refluxed with stirring for 18 hours, the reaction mixture was concentrated in vacuo. The residue was diluted with water (800 mL), basified with a saturated aqueous solution of sodium bicarbonate to about pH 7, and extracted with dichloromethane (600 mL  3). The combined organic layers were washed with brine (800 mL), dried over sodium sulfate, filtered and concentrated in vacuo to afford methyl 2-sulfanylbenzoate (9, 300 g, yield 91%) as a light yellow oil, which was used for the next step without further purification. To a cooled solution of methyl 2-sulfanylbenzoate (9, 200 g, 1.19 mol) in tetrahydrofuran and N,Ndimethylformamide (2 L, V/V = 1/1) was added 2-chloroethanamine hydrochloride (138 g, 1.19 mol) at 0 oC

followed by sodium hydride (143 g, 3.57 mol, 60% in mineral oil) in portions. After being stirred at

room temperature overnight, the reaction mixture was poured into ice-water and extracted with ethyl acetate (900 mL  4). The organic layers were combined, washed with brine (900 mL  3), dried over sodium sulfate and concentrated in vacuo. The residue was stirred in a mixture of ethyl acetate and petroleum ether (300 mL, V/V = 1/1) for 1 hour. The solid was collected by filtration and dried in vacuo to afford 3,4dihydro-1,4-benzothiazepin-5(2H)-one (10, 100 g, yield 47%). To a bottle containing a cooled suspension of lithium aluminum hydride (44 g, 1.17 mol) in dry tetrahydrofuran (1.5 L) was added 3,4-dihydro-1,4-benzothiazepin-5(2H)-one (10, 150 g, 0.84 mol) in



portions at 0 oC. After being refluxed for 18 hours, the reaction mixture was cooled to 0 oC, followed by addition of water (25 mL) dropwise. The reaction mixture was then filtered through a pad of celite and washed with dichloromethane. The filtrate was dried over sodium sulfate and evaporated in vacuo to afford 2,3,4,5-tetrahydro-1,4-benzothiazepine (11, 125 g, yield 90%), which was used for the next step without further purification. To a solution of 2,3,4,5-tetrahydro-1,4-benzothiazepine (11, 5 g, 30.3 mmol) in dry dichloromethane (100 mL) was added triethylamine (5.06 mL, 36.3 mmol) at room temperature, followed by the dropwise addition of acetic anhydride (3.43 mL, 36.3 mmol) at 0 oC under nitrogen. The resulting solution was stirred for 1 hour whilst allowing the temperature to rise slowly to room temperature. The mixture was washed with brine (50 mL × 2), dried over sodium sulfate, filtered and concentrated in vacuo to afford 1-(2,3-dihydro1,4-benzothiazepin-4(5H)-yl)ethanone (12, 6.28 g, quantitative yield) as a yellow oil, which was used for next step without further purification. To a cooled solution of 1-(2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)ethanone (12, 6.27 g, 30.2 mmol) in dichloromethane (100 mL) was added a suspension of 3-chloroperoxybenzoic acid (20.9 g, 90.8 mmol, 75% purity) in dichloromethane (50 mL) at 10 oC. After the addition, the resulting mixture was stirred for 1 hour whilst allowing the temperature to rise slowly to room temperature. The mixture was washed with a saturated aqueous solution of sodium carbonate (100 mL  2), a saturated aqueous solution of sodium sulfite (100 mL  2) and brine (100 mL) in sequence. The organic layer was dried over sodium sulfate, filtered and concentrated in vacuo. The residue was stirred in diethyl ether (50 mL) and the solid was collected by filtration and dried in vacuo to afford 1-(1,1-dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)ethanone (13, 6 g, yield 83%) as a white powder. To a solution of 1-(1,1-dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)ethanone (13, 6 g, 25 mmol) in ethanol (25 mL) was added sodium hydroxide (5 g, 125 mmol) and water (28 mL). The mixture was refluxed overnight and then concentrated in vacuo. The residue was extracted by ethyl acetate (60 mL  4).


Page 24 of 42

Page 25 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


The combined organic layers were extracted by hydrochloric acid (80 mL, 3 N). The acidic aqueous layer was washed with ethyl acetate (60 mL  2), then basified with a saturated aqueous solution of sodium bicarbonate to pH > 7, and extracted with ethyl acetate (60 mL  4). The combined organic layers were dried over sodium sulfate, filtered and concentrated in vacuo to afford 2,3,4,5-tetrahydro-1,4benzothiazepine1,1-dioxide (14, 3.8 g, yield 77%), MS obsd. (ESI+) [(M+H)+] 198. 1H NMR (400 MHz, DMSO-d6) δ ppm 7.89 (dd, J = 1.2, 7.6 Hz, 1 H), 7.56 (t, J = 7.6 Hz, 1 H), 7.47 (t, J = 7.6 Hz, 1 H), 7.42 (d, J = 7.6 Hz, 1 H), 4.04 (s, 2 H), 3.32 - 3.30 (m, 2 H), 3.30 - 3.25 (m, 2 H), 2.64 (s, 1 H). To a solution of 2,4-dichloro-6-methyl-quinoline (2.12 g, 10 mmol) in 20 mL of 1-butanol was added 2,3,4,5-tetrahydro-1,4-benzothiazepine1,1-dioxide (14, 2.16 g, 11 mmol). The mixture was stirred at 160 ºC under microwave irradiation for 2 hours. After being cooled to room temperature, the mixture was diluted with dichloromethane, washed with brine, dried over anhydrous sodium sulfate. The solvent was evaporated under reduced pressure, and the residue was purified by chromatography to give 8-(4-chloro-6methyl-quinolin-2-yl)-6,7,8,9-tetrahydro-5-thia-8-aza-benzocycloheptene 5,5-dioxide (15, 3.7 g, yield 99%), which was used for next step without further purification. MS obsd. (ESI+) [(M+H)+] 373. A mixture of 4-(4-chloro-6-methylquinolin-2-yl)-8-methoxy-2,3,4,5-tetrahydro-1,4-benzothiazepine 1,1dioxide

(15,

100

mg,

0.27

mmol),

sodium

tert-butoxide

bis(diphenylphosphino)ferrocene-palladium(II)dichloride

(22

(52 mg,

mg, 0.027

0.54

mmol), mmol),

1,1'1,1'-

bis(diphenylphosphino)ferrocene (15 mg, 0.027 mmol) and 3-(aminomethyl)-N,N-dibenzyloxetan-3-amine (76 mg, 0.27 mmol) in 1,4-dioxane (4 mL) was stirred at 120 ºC under microwave irradiation for 1 hour. The resulting mixture was concentrated in vacuo. The residue was purified by preparative HPLC to afford N-[[3-(dibenzylamino)oxetan-3-yl]methyl]-2-(1,1-dioxo-3,5-dihydro-2H-1lambda6,4-benzothiazepin-4yl)-6-methyl-quinolin-4-amine (16, 108 mg, yield 65%) as a white powder. MS obsd. (ESI+) [(M+H)+] 619. A

mixture

of

N-[[3-(dibenzylamino)oxetan-3-yl]methyl]-2-(1,1-dioxo-3,5-dihydro-2H-1lambda6,4-

benzothiazepin-4-yl)-6-methyl-quinolin-4-amine (108 mg, 0.17 mmol), 10% palladium hydroxide on active carbon (14 mg) in methanol (6 mL) was stirred for 16 hours at room temperature under hydrogen



atmosphere (1 bar). The resulting mixture was concentrated in vacuo. The residue was purified by preparative

HPLC

to

afford

N-[(3-aminooxetan-3-yl)methyl]-2-(1,1-dioxido-2,3-dihydro-1,4-

benzothiazepin-4(5H)-yl)-6-methylquinolin-4-amine (1, 13 mg, yield 17%). m.p. 229.3  229.7 oC. MS obsd. (ESI+) [(M+H)+] 439. HRMS calcd [(M+H)+] 439.17984, measured [(M+H)+] 439.17988. 1H NMR (400 MHz, DMSO-d6) δ ppm 7.96 (d, J = 7.07 Hz, 1 H), 7.88 (dd, J = 1.26, 7.83 Hz, 1 H), 7.69 (s, 1H), 7.62 (dt, J = 1.14, 7.52 Hz, 1 H), 7.47 (t, J = 7.67 Hz, 1 H), 7.34 (d, J = 8.34 Hz, 1 H), 7.25 (dd, J = 1.52, 8.59 Hz, 1 H), 6.45 (br t, J = 5.56 Hz, 1 H), 6.20 (s, 1 H), 5.09 (br s, 2 H), 4.45 (d, J = 6.06 Hz, 2 H), 4.32 - 4.42 (m, 2 H), 3.63 (br t, J = 4.55 Hz, 2 H), 3.55 (d, J = 5.31 Hz, 2 H), 2.53 - 2.81 (m, 2 H), 2.36 (s, 3 H). 13C

NMR (101 MHz, DMSO-d6) δ 155.6, 151.6, 146.5, 141.1, 137.7, 133.5, 133.3, 131.2, 129.9, 128.2,

127.5, 127.0, 120.5, 115.5, 85.5, 82.2, 57.0, 55.3, 51.9, 49.7, 45.0, 41.0, 40.9, 40.8, 21.4. For the following BAQ series, except described specifically, all the analogues are prepared in analogy to 1 from the commercially available building blocks. 1-Amino-3-[[2-(3,4-dihydro-1H-isoquinolin-2-yl)-4-quinolyl]amino]propan-2-ol (2). MS obsd. (ESI+) [(M+H)+] 349. 1H NMR (400 MHz, CD3OD) δ ppm 8.21 (d, J = 8.08 Hz, 1 H), 7.88 (d, J = 8.34 Hz, 1 H), 7.78 (t, J = 7.71 Hz, 1 H), 7.50 (t, J = 7.71 Hz, 1 H), 7.34 - 7.41 (m, 1 H), 7.28 - 7.34 (m, 3 H), 6.23 (s, 1 H), 4.88 - 4.93 (m, 2 H), 4.29 (br d, J = 6.06 Hz, 1 H), 3.96 (t, J = 5.81 Hz, 2 H), 3.60 - 3.78 (m, 2 H), 3.23 - 3.31 (m, 1 H), 3.16 (t, J = 5.81 Hz, 2H), 3.04 (br dd, J = 9.98, 12.25 Hz, 1 H). 3-[[2-(3,4-Dihydro-1H-isoquinolin-2-yl)-4-quinolyl]amino]propane-1,2-diol (2a). MS obsd. (ESI+) [(M+H)+] 350. 1H NMR (400 MHz, CDCl3) δ ppm 7.75 (br d, J = 8.31 Hz, 1 H), 7.60 (br d, J = 8.19 Hz, 1 H), 7.37 (br t, J = 7.70 Hz, 1 H), 7.27 - 7.31 (m, 1 H), 7.18 - 7.25 (m, 3 H), 7.11 (t, J = 7.57 Hz, 1 H), 5.69 (br s, 1 H), 4.77 (s, 2 H), 4.09 - 4.18 (m, 1 H), 3.84 - 3.95 (m, 3 H), 3.78 (dd, J = 5.26, 11.37 Hz, 1 H), 3.24 - 3.51 (m, 2 H), 3.01 (t, J = 5.75 Hz, 2 H). N'-[2-(3,4-Dihydro-1H-isoquinolin-2-yl)-4-quinolyl]propane-1,3-diamine (2b). MS obsd. (ESI+) [(M+H)+] 333. 1H NMR (400 MHz, CD3OD) δ ppm 8.22 (d, J = 8.38 Hz, 1 H), 7.88 (d, J = 7.82 Hz, 1 H),


Page 26 of 42

Page 27 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


7.77 (t, J = 7.76 Hz, 1 H), 7.49 (t, J = 7.44 Hz, 1 H), 7.29 - 7.40 (m, 4 H), 6.11 (s, 1 H), 4.90 - 4.94 (m, 2 H), 3.96 (t, J = 5.93 Hz, 2 H), 3.71 (t, J = 7.03 Hz, 2 H), 3.10 - 3.31 (m, 4 H), 2.22 (quin, J = 7.37 Hz, 2 H). 1-[[2-(3,4-Dihydro-1H-isoquinolin-2-yl)-4-quinolyl]amino]propan-2-ol

(2c).

MS

obsd.

(ESI+)

[(M+H)+] 334. 1H NMR (400 MHz, CD3OD) δ ppm 7.97 - 8.13 (m, 1 H), 7.60 - 7.85 (m, 2 H), 7.45 (ddd, J = 1.10, 7.11, 8.32 Hz, 1 H), 7.18 - 7.33 (m, 4 H), 6.12 (s, 1 H), 4.80 - 4.86 (m, 2 H), 4.15 (dt, J = 4.74, 6.67 Hz, 1 H), 3.89 (t, J = 5.95 Hz, 2 H), 3.41 - 3.55 (m, 2 H), 3.10 (t, J = 5.95 Hz, 2 H), 1.30 (d, J = 6.39 Hz, 3 H). N'-[2-(3,4-Dihydro-1H-isoquinolin-2-yl)-4-quinolyl]ethane-1,2-diamine

(2d).

MS

obsd.

(ESI+)

[(M+H)+] 319. 1H NMR (400 MHz, CD3OD) δ ppm 8.04 - 8.28 (m, 1 H), 7.84 - 7.99 (m, 1 H), 7.79 (dt, J = 1.16, 7.73 Hz, 1 H), 7.51 (ddd, J = 1.16, 7.09, 8.25 Hz, 1 H), 7.29 - 7.37 (m, 4 H), 6.16 (s, 1 H), 4.90 4.93 (m, 2 H), 3.84 - 4.03 (m, 4 H), 3.35 - 3.43 (m, 2 H), 3.04 - 3.31 (m, 2 H). N'-[2-(3,4-Dihydro-1H-isoquinolin-2-yl)-4-quinolyl]butane-1,4-diamine

(2e).

MS

obsd.

(ESI+)

[(M+H)+] 347. 1H NMR (400 MHz, CD3OD) δ ppm 8.22 (d, J = 7.91 Hz, 1 H), 7.87 (d, J = 8.19 Hz, 1 H), 7.77 (t, J = 7.78 Hz, 1 H), 7.42 - 7.56 (m, 1 H), 7.29 - 7.39 (m, 4 H), 6.07 (s, 1 H), 4.90 - 4.92 (m, 2 H), 3.95 (t, J = 5.93 Hz, 2 H), 3.63 (t, J = 6.60 Hz, 2 H), 3.12 - 3.31 (m, 2 H), 3.06 (br t, J = 7.27 Hz, 2 H), 1.83 - 1.99 (m, 4 H). 1-[2-[[2-(3,4-Dihydro-1H-isoquinolin-2-yl)-4-quinolyl]amino]ethyl]guanidine

(2f).

The

synthetic

procedure was described in the Supporting Information. MS obsd. (ESI+) [(M+H)+] 361. 1H NMR (400 MHz, CD3OD) δ ppm 8.12 (d, J = 8.31 Hz, 1 H), 7.82 (d, J = 8.31 Hz, 1 H), 7.72 (t, J = 7.64 Hz, 1 H), 7.44 (t, J = 7.58 Hz, 1 H), 7.25 - 7.34 (m, 4 H), 6.09 (s, 1 H), 4.86 (s, 2 H), 3.92 (t, J = 5.81 Hz, 2 H), 3.75 (br t, J = 5.69 Hz, 2 H), 3.62 (br t, J = 5.62 Hz, 2 H), 3.11 (t, J = 5.75 Hz, 2 H).

2-[[2-(3,4-Dihydro-1H-isoquinolin-2-yl)-4-quinolyl]amino]ethylurea (2g). The synthetic procedure was described in the Supporting Information. MS obsd. (ESI+) [(M+H)+] 362. 1H NMR (400 MHz, CD3OD) δ



Page 28 of 42

ppm 8.00 (d, J = 8.19 Hz, 1 H), 7.76 (d, J = 8.31 Hz, 1 H), 7.67 (t, J = 7.70 Hz, 1 H), 7.19 - 7.41 (m, 5 H), 6.16 (s, 1 H), 4.86 (s, 2 H), 3.92 (t, J = 5.87 Hz, 2 H), 3.35 - 3.58 (m, 4 H), 3.09 (t, J = 5.81 Hz, 2 H).

1-Amino-3-[(2-isoindolin-2-yl-4-quinolyl)amino]propan-2-ol (3). MS obsd. (ESI+) [(M+H)+] 335. 1H NMR (400 MHz, CD3OD) δ ppm 7.86 - 8.06 (m, 1 H), 7.68 - 7.77 (m, 1 H), 7.58 (s, 1 H), 7.38 - 7.46 (m, 2 H), 7.31 - 7.38 (m, 2 H), 7.23 - 7.29 (m, 1 H), 5.90 (s, 1 H), 4.89 - 4.97 (m, 4 H), 3.98 - 4.18 (m, 1 H), 3.43 - 3.58 (m, 2 H), 3.01 (dd, J = 3.73, 13.02 Hz, 1 H), 2.85 (dd, J = 7.95, 12.96 Hz, 1 H). 1-Amino-3-[2-(1,3,4,5-tetrahydro-benzo[c]azepin-2-yl)-quinolin-4-ylamino]-propan-2-ol

(4).

MS

obsd. (ESI+) [(M+H)+] 363. 1H NMR (400 MHz, CD3OD) δ ppm 8.05 - 8.03 (d, J = 7.6 Hz, 1 H), 7.78 7.76 (d, J = 7.6 Hz, 1 H), 7.72 - 7.68 (m, 1 H), 7.52 - 7.50 (m, 1 H), 7.44 - 7.40 (m, 1 H), 7.23 - 7.19 (m, 3 H), 6.12 (s, 1 H), 4.97 (s, 2 H), 4.16 - 4.11 (m, 3 H), 3.56 - 3.55 (d, J = 6 Hz, 2 H), 3.20 - 3.16 (dd, J = 2.8, 12.8 Hz,1 H), 3.11 - 3.08 (m, 2 H), 2.99 - 2.94 (m, 1 H), 1.99 - 1.98 (m, 2 H). N-[2-(1,3,4,5-Tetrahydro-2H-2-benzazepin-2-yl)quinolin-4-yl]ethane-1,2-diamine

(5).

MS

obsd.

(ESI+) [(M+H)+] 333. HRMS calcd [(M+H)+] 333.20737, measured [(M+H)+] 333.20677. 1H NMR (400 MHz, CD3OD) δ ppm 7.91 - 7.89 (m, 1 H), 7.62 - 7.60 (m, 1 H), 7.52 - 7.45 (m, 2 H), 7.21 - 7.12 (m, 3 H), 5.96 (s, 1 H), 4.10 (s, 2 H), 3.51 - 3.47 (t, J = 6.0 Hz, 1 H), 3.04 - 2.98 (m, 5 H), 2.84 (s, 2 H), 1.94 - 1.91 (m, 2 H). N-[5-Chloro-2-(1,3,4,5-tetrahydro-2H-2-benzazepin-2-yl)quinolin-4-yl]ethane-1,2-diamine (5a). MS obsd. (ESI+) [(M+H)+] 367. 1H NMR (400 MHz, CD3OD) δ ppm 7.46 - 7.42 (m, 2 H), 7.27 - 7.23 (m, 1 H), 7.16 - 7.11 (m, 3 H), 7.02 (66, J = 7.6, 1.2 Hz, 1 H), 5.97 (s, 1 H), 4.79 (s, 2 H), 4.13 (br s, 2 H), 3.38 - 3.21 (m, 2 H), 3.04 - 3.02 (m, 2 H), 2.96 (t, J = 6.0 Hz, 2 H), 1.91 - 1.86 (m, 2 H). N-[6-Chloro-2-(1,3,4,5-tetrahydro-2H-2-benzazepin-2-yl)quinolin-4-yl]ethane-1,2-diamine (5b). MS obsd. (ESI+) [(M+H)+] 367. 1H NMR (400 MHz, CD3OD) δ ppm 7.87 (d, J = 2.4 Hz, 1 H), 7.47 - 7.42 (m,


Page 29 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


2 H), 7.34 (dd, J = 9.0, 2.2 Hz, 1 H), 7.15 - 7.09 (m, 3 H), 5.99 (s, 1 H), 4.76 (s, 2 H), 4.11 (br s, 2 H), 3.38 - 3.32 (m, 2 H), 3.00 - 2.93 (m, 4 H), 1.87 (t, J = 5.2 Hz, 2 H). N-[7-Chloro-2-(1,3,4,5-tetrahydro-2-benzazepin-2-yl)-4-quinolyl]ethane-1,2-diamine (5c). MS obsd. (ESI+) [(M+H)+] 367. 1H NMR (400 MHz, CD3OD) δ ppm 7.71 (d, J = 8.84 Hz, 1 H), 7.48 (d, J = 2.02 Hz, 1 H), 7.38 (d, J = 7.07 Hz, 1 H), 7.09 - 7.13 (m, 1 H), 7.02 - 7.07 (m, 2 H), 6.97 (dd, J = 2.02, 8.84 Hz, 1 H), 5.88 (s, 1 H), 4.66 (s, 2 H), 4.04 (br s, 2 H), 3.37 (s, 1 H), 3.29 - 3.31 (m, 1 H), 2.85 - 2.94 (m, 4 H), 1.80 (br s, 2 H). N-[8-Chloro-2-(1,3,4,5-tetrahydro-2H-2-benzazepin-2-yl)quinolin-4-yl]ethane-1,2-diamine (5d). MS obsd. (ESI+) [(M+H)+] 367. 1H NMR (400 MHz, CD3OD) δ ppm 7.24 (d, J = 8.0 Hz, 1 H), 7.60 (d, J = 6.8 Hz, 1 H), 7.55 (d, J = 7.6 Hz, 1 H), 7.13 - 7.09 (m, 3 H), 6.98 (t, J = 8.0 Hz, 1 H), 6.03 (s, 1 H), 4.89 (s, 2 H), 4.20 (br s, 2 H), 3.46 (t, J = 6.0 Hz, 2 H), 3.07 - 3.02 (m, 4 H), 1.92 (t, J = 5.2 Hz, 2 H). N-[6-Methoxy-2-(1,3,4,5-tetrahydro-2H-2-benzazepin-2-yl)quinolin-4-yl]ethane-1,2-diamine

(5e).

MS obsd. (ESI+) [(M+H)+] 363. 1H NMR (400 MHz, CD3OD) δ ppm 7.60 (d, J = 9.2 Hz, 1 H), 7.50 (d, J = 6.4 Hz, 1 H), 7.42 (d, J = 2.4 Hz, 1 H), 7.25 - 7.18 (m, 4 H), 6.01 (s, 1 H), 4.93 (s, 2 H), 4.13 (br s, 2 H), 3.89 (s, 3 H), 3.53 (t, J = 6.4 Hz, 2 H), 3.10 - 3.01 (m, 4 H), 1.96 (s, 2 H). N-[6-Methyl-2-(1,3,4,5-tetrahydro-2H-2-benzazepin-2-yl)quinolin-4-yl]ethane-1,2 diamine (5f). MS obsd. (ESI+) [(M+H)+] 347. 1H NMR (400 MHz, DMSO-d6) δ ppm 7.68 (s, 1 H), 7.55 (d, J = 6.8 Hz, 1 H), 7.29 (d, J = 8.4 Hz, 1 H), 7.21 (d, J = 6.8 Hz, 1 H), 7.13 - 7.05 (m, 3 H), 6.67 (br s, 1 H), 6.00 (s, 1 H), 4.79 (s, 2 H), 4.11 (br s, 2 H), 3.52 (s, 2 H), 3.05 - 2.99 (m, 4 H), 2.35 (s, 3 H), 1.77 (s, 2 H). N-[6-Fluoro-2-(1,3,4,5-tetrahydro-2H-2-benzazepin-2-yl)quinolin-4-yl]ethane-1,2-diamine (5g). MS obsd. (ESI+) [(M+H)+] 351. 1H NMR (400 MHz, CD3OD) δ ppm 7.60 - 7.52 (m, 2 H), 7.47 (d, J = 6.8 Hz, 1 H), 7.26 - 7.21 (m, 1 H), 7.17 - 7.12 (m, 3 H), 6.04 (s, 1 H), 4.82 (s, 2 H), 4.15 (br s, 2 H), 3.45 (t, J = 6.4 Hz, 2 H), 3.06 - 2.99 (m, 4 H), 1.91 (t, J = 5.4 Hz, 2 H).



Page 30 of 42

N-[2-(1,3,4,5-Tetrahydro-2H-2-benzazepin-2-yl)-6-(trifluoromethyl)quinolin-4-yl]ethane-1,2diamine (5h). MS obsd. (ESI+) [(M+H)+] 401. 1H NMR (400 MHz, DMSO-d6) δ ppm 8.36 (s, 1 H), 7.57 7.45 (m, 3 H), 7.12 (br s, 3 H), 6.99 (s, 1 H), 6.03 (s, 1 H), 4.81 (s, 2 H), 4.15 (br s, 2 H), 3.29 (br s, 2 H), 3.01 (s, 2 H), 2.82 (s, 2 H), 1.77 (s, 2 H). N-[6-(Methylsulfonyl)-2-(1,3,4,5-tetrahydro-2H-2-benzazepin-2-yl)quinolin-4-yl]ethane-1,2-diamine (5i). MS obsd. (ESI+) [(M+H)+] 411. 1H NMR (400 MHz, DMSO-d6) δ ppm 8.518 (s, 0.5 H), 8.514 (s, 0.5 H), 7.76 - 7.73 (m, 1 H), 7.61 (d, J = 6.4 Hz, 0.5 H), 7.52 (d, J = 6.8 Hz, 0.5 H), 7.47 - 7.34 (m, 1 H), 7.13 - 7.07 (m, 3 H), 6.23 (s, 0.5 H), 6.04 (s, 0.5 H). 4.83 (d, J = 6.4 Hz, 2 H), 4.34 (br s, 2 H), 3.15 - 3.28 (m, 6 H), 3.00 (s, 2 H), 2.81 (t, J = 6.4 Hz, 1 H), 1.76 (br s, 2 H). N-[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)quinolin-4-yl]ethane-1,2-diamine

(6a).

MS obsd. (ESI+) [(M+H)+] 383. 1H NMR (400 MHz, CD3OD) δ ppm 7.87 (m, 1 H), 7.81 (d, J = 7.83 Hz, 2 H), 7.55 (m, 1 H), 7.47 (m, 1 H), 7.33 (m, 2 H), 7.04 (m, 1 H), 5.94 (m,1 H), 5.12 (s, 2 H), 3.57 (t, J = 4.6 Hz, 2 H), 3.43 (t, J = 6.3 Hz, 2 H), 3.33 (m, 2 H), 2.97 (t, J = 6.4 Hz, 2 H). N-[2-(2,3-Dihydro-1,4-benzoxazepin-4(5H)-yl)quinolin-4-yl]ethane-1,2-diamine (6b). Compound 6b was prepared in analogy to 1 by using 2,3,4,5-tetrahydro-1,4-benzoxazepine hydrochloride (6b-c, see Supporting Information for its synthesis) instead of 2,3,4,5-tetrahydro-1,4-benzothiazepine1,1-dioxide 14. MS obsd. (ESI+) [(M+H)+] 335. 1H NMR (400 MHz, CD3OD) δ ppm 8.61 (s, 1 H), 7.90 (d, J = 7.6 Hz, 1 H), 7.60 - 7.51 (m, 3 H), 7.23 (br s, 2 H), 7.09 - 6.98 (m, 2 H), 6.05 (s, 1 H), 4.89 (s, 2 H), 4.27 (s, 2 H), 4.25 (s, 2 H), 3.61 (s, 2 H), 3.15 (s, 2 H). N-[2-(5,5-Difluoro-1,3,4,5-tetrahydro-2H-2-benzazepin-2-yl)-6-methylquinolin-4-yl]ethane-1,2diamine (6c). Compound 6c was prepared in analogy to 1 by using 5,5-difluoro-2,3,4,5-tetrahydro-1Hbenzazepine (6c-c, see Supporting Information for its synthesis) instead of 2,3,4,5-tetrahydro-1,4benzothiazepine1,1-dioxide 14. MS obsd. (ESI+) [(M+H)+] 383. 1H NMR (400 MHz, CD3OD) δ ppm 7.89 (s, 1 H), 7.72 - 7.70 (d, J = 6.8 Hz, 1 H), 7.70 - 7.67 (d, J = 6.8 Hz, 2 H), 7.65 - 7.42 (m, 3 H), 6.00 (s, 1


Page 31 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


H), 5.02 (s, 2 H), 4.23 - 4.20 (t, 2 H), 3.88 - 3.78 (t, 2 H), 3.58 - 3.55 (t, 2 H), 2.71 - 2.61 (m, 2 H), 2.46 (s, 3 H). N-[6-Methyl-2-(1,2,3,5-tetrahydro-4H-1,4-benzodiazepin-4-yl)quinolin-4-yl]ethane-1,2-diamine (6d). MS obsd. (ESI+) [(M+H)+] 348. 1H NMR (400 MHz, CD3OD) δ ppm 7.90 (s, 1 H), 7.67 (d, J = 8.4 Hz, 1 H), 7.58 (d, J = 8.4 Hz, 1 H), 7.42 (d, J = 7.2 Hz, 1 H), 7.12 (t, J = 8.0 Hz, 1 H), 6.88 (t, J = 7.2 Hz, 1 H), 6.82 (d, J = 8.0 Hz, 1 H), 5.95 (s, 1 H), 4.87 (s, 2 H), 4.00 (t, J = 4.8 Hz, 2 H), 3.72 (t, J = 6.0 Hz, 2 H), 3.46 (t, J = 4.8 Hz, 2 H), 3.18 (t, J = 6.0 Hz, 2 H), 2.49 (s, 3 H). 4-{4-[(2-Aminoethyl)amino]-6-methylquinolin-2-yl}-1,3,4,5-tetrahydro-2H-1,4-benzodiazepin-2-one (6e). MS obsd. (ESI+) [(M+H)+] 362. 1H NMR (400 MHz, CD3OD) δ ppm 7.64 (s, 1 H), 7.44 - 7.40 (m, 2 H), 7.28 (dd, J = 1.6, 8.4 Hz, 1 H), 7.18 (dd, J = 1.6, 8.0 Hz, 1 H), 7.09 (dd, J = 0.8, 8.0 Hz, 1 H), 6.98 (dd, J = 0.8, 8.0 Hz, 1 H), 6.00 (s, 1 H), 4.87 (s, 2 H), 4.72 (s, 2 H), 3.40 (t, J = 6.4 Hz, 2 H), 2.95 (t, J = 6.4 Hz, 2 H), 2.40 (s, 3 H). N-[2-(7-Fluoro-1,3,4,5-tetrahydro-2H-2-benzazepin-2-yl)-6-methylquinolin-4-yl]ethane-1,2-diamine (6f). MS obsd. (ESI+) [(M+H)+] 365. 1H NMR (400 MHz, CD3OD) δ ppm 7.82 (s, 1 H), 7.58 (d, J = 8.4 Hz, 1 H), 7.45 (m, 1 H), 7.17 (m, 1 H), 6.94 (m, 1 H), 6.84 (m, 1 H), 6.03 (d, J = 4.0 Hz, 1 H), 4.87 (s, 2 H), 3.95 (m, 4 H), 3.56 (t, J = 6.4 Hz, 2 H), 3.10 (m, 4 H), 2.48 (s, 3 H). N-[2-(8-Fluoro-1,3,4,5-tetrahydro-2H-2-benzazepin-2-yl)-6-methylquinolin-4-yl]ethane-1,2-diamine (6g). MS obsd. (ESI+) [(M+H)+] 365. 1H NMR (400 MHz, CD3OD) δ ppm 7.68 (s, 1 H), 7.48 (d, J = 8.4 Hz, 1 H), 7.28 (m, 2 H), 7.12 (m, 1 H), 6.83 (m, 1 H), 5.93 (s, 1 H), 4.80 (s, 2 H), 4.09 (s, 2 H), 3.46 (t, J = 6.0 Hz, 2 H), 3.00 (m, 4 H), 2.40 (s, 3 H), 1.90 (m, 2 H). N-[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4-yl]ethane-1,2diamine (6h). MS obsd. (ESI+) [(M+H)+] 397. 1H NMR (400 MHz, CD3OD) δ ppm 7.96 (d, J = 7.58 Hz, 1 H), 7.86 (br d, J = 7.33 Hz, 1 H), 7.67 (s, 1 H), 7.62 (t, J = 7.08 Hz, 1 H), 7.39 - 7.46 (m, 2 H), 7.28 (d, J



= 8.69 Hz, 1 H), 6.06 (s, 1 H), 5.15 (s, 2 H), 4.53 (br s, 2 H), 3.49 - 3.66 (m, 4 H), 3.06 - 3.17 (m, 2 H), 2.41 (s, 3 H). N-[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4-yl]propane-1,3diamine (7a). MS obsd. (ESI+) [(M+H)+] 411. 1H NMR (400 MHz, CD3OD) δ ppm 7.98 (dd, J = 1.01, 7.83 Hz, 1 H), 7.83 (d, J = 7.07 Hz, 1 H), 7.56 - 7.64 (m, 2 H), 7.39 - 7.47 (m, 2 H), 7.27 (dd, J = 1.77, 8.59 Hz, 1 H), 6.00 (s, 1 H), 5.14 (s, 2 H), 4.40 - 4.67 (m, 2 H), 3.58 (t, J = 4.93 Hz, 2 H), 3.41 (t, J = 6.82 Hz, 2 H), 2.84 (t, J = 6.95 Hz, 2 H), 2.41 (s, 3 H), 1.91 (quin, J = 6.88 Hz, 2 H). N-[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4-yl]butane-1,4diamine (7b). MS obsd. (ESI+) [(M+H)+] 425. 1H NMR (400 MHz, CD3OD) δ ppm 8.01 - 7.95 (m, 1 H), 7.92 -7.86 (m, 1 H), 7.83 - 7.76 (m, 1 H), 7.69 - 7.61 (m, 1 H), 7.61 - 7.54 (m, 1 H), 7.50 - 7.42 (m, 1 H), 7.39 - 7.31 (m, 1 H), 5.96 (s, 1 H), 5.20 (br s, 2 H), 4.54 (br s, 2 H), 3.63 (br s, 2 H), 3.47 (br s, 2 H), 3.37 (s, 2 H), 2.42 (s, 3 H), 2.27 - 2.24 (m, 3 H), 1.86 (d, J = 3.28 Hz, 3 H). N-[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4-yl]pentane-1,5diamine (7c). MS obsd. (ESI+) [(M+H)+] 439. 1H NMR (400 MHz, CD3OD) δ ppm 7.91 - 8.07 (m, 1 H), 7.78 - 7.89 (m, 1 H), 7.74 (s, 1 H), 7.62 (q, J = 7.33 Hz, 1 H), 7.41 - 7.55 (m, 2 H), 7.29 - 7.36 (m, 1 H), 6.00 (s, 1 H), 5.16 (s, 2 H), 4.37 - 4.71 (m, 2 H), 3.52 - 3.70 (m, 2 H), 3.39 - 3.44 (m, 2 H), 3.28 - 3.37 (m, 2 H), 2.24 (s, 3 H), 1.71 - 1.84 (m, 4 H), 1.54 - 1.64 (m, 2 H). N-[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4-yl]-N'methylethane-1,2-diamine (7d). MS obsd. (ESI+) [(M+H)+] 411. 1H NMR (400 MHz, CD3OD) δ ppm 8.03 - 8.0 (m, 3 H), 7.87 - 7.84 (d, J = 8.8 Hz, 1 H), 7.71 - 7.70 (d, J = 1.2 Hz, 1 H), 7.57 - 7.53 (m, 2 H), 6.07 (s, 1 H), 5.40 (s, 2 H), 4.56 (s, 2 H), 3.96 - 3.93 (dd, J = 6.0, 6.4 Hz, 2 H), 3.75 - 3.73 (q, J = 4.4 Hz, 2 H), 3.43 - 3.40 (q, J = 6 Hz, 2 H), 2.80 (s, 3 H), 2.46 (s, 3 H).


Page 32 of 42

Page 33 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


N'-[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4-yl]-N,Ndimethylethane-1,2-diamine (7e). MS obsd. (ESI+) [(M+H)+] 425. 1H NMR (400 MHz, CD3OD) δ ppm 8.05 - 8.01 (m, 3 H), 7.88 - 7.86 (d, J = 8.8 Hz, 1 H), 7.70 - 7.68 (d, J = 1.2 Hz, 1 H), 7.56 - 7.52 (m, 2 H), 6.05 (s, 1 H), 5.41 (s, 2 H), 4.56 (s, 2 H), 3.96 - 3.93 (dd, J = 6.0, 6.4 Hz, 2 H), 3.75 - 3.73 (q, J = 4.4 Hz, 2 H), 3.43 - 3.40 (q, J = 6.0 Hz, 2 H), 3.01 (s, 6 H), 2.45 (s, 3 H). N~2~-[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4-yl]propane-1,2diamine (7f). MS obsd. (ESI+) [(M+H)+] 411. 1H NMR (400 MHz, CD3OD) δ ppm 7.97 (dd, J = 7.83, 1.01 Hz, 1 H), 7.82 (d, J = 7.33 Hz, 1 H), 7.69 - 7.56 (m, 2 H), 7.46 - 7.38 (m, 2 H), 7.28 (dd, J = 8.46, 1.64 Hz, 1 H), 6.05 (s, 1 H), 5.13 (s, 2 H), 4.48 (br s, 2 H), 3.56 (m, 2 H), 3.42 - 3.49 (m, 3 H), 2.41 (s, 3 H), 1.38 (d, J = 5.81 Hz, 3 H). N~1~-[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4-yl]propane-1,2diamine (7g). MS obsd. (ESI+) [(M+H)+] 411. 1H NMR (400 MHz, CD3OD) δ ppm 7.91 (dd, J = 7.83, 1.01 Hz, 1 H), 7.83 (d, J = 7.33 Hz, 1 H), 7.69 (s, 1 H), 7.58 (d, J = 1.26 Hz, 1 H), 7.44 (d, J = 8.59 Hz, 1 H), 7.38 - 7.31 (m, 1 H), 7.28 (dd, J = 8.59, 1.77 Hz, 1 H), 6.04 (s, 1 H), 5.13 (br s, 2 H), 4.48 (br s, 2 H), 3.56 (t, J = 4.67 Hz, 2 H), 3.52 - 3.41 (m, 3 H), 2.41 (s, 3 H), 1.38 (d, J = 6.06 Hz, 3 H). N~2~-[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4-yl]-2methylpropane-1,2-diamine (7h). MS obsd. (ESI+) [(M+H)+] 425. 1H NMR (400 MHz, CD3OD) δ ppm 7.98 (dd, J = 7.83, 1.26 Hz, 1 H), 7.86 (d, J = 6.82 Hz, 1 H), 7.70 (s, 1 H), 7.61 (td, J = 7.45, 1.26 Hz, 1 H), 7.47 - 7.42 (m, 2 H), 7.29 (dd, J = 8.59, 1.77 Hz, 1 H), 6.09 (s, 1 H), 5.16 (s, 2 H), 4.53 (br s, 2 H), 3.58 (t, J = 4.80 Hz, 2 H), 3.27 (s, 2 H), 2.43 (s, 3 H), 1.27 (s, 6 H). N~1~-[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4-yl]-2methylpropane-1,2-diamine (7i). MS obsd. (ESI+) [(M+H)+] 425. 1H NMR (400 MHz, CD3OD) δ ppm 7.97 (d, J = 7.83 Hz, 1 H), 7.85 (d, J = 7.58 Hz, 1 H), 7.70 (s, 1 H), 7.60 (t, J = 7.33 Hz, 1 H), 7.48 - 7.38



(m, 2 H), 7.29 - 7.27 (m, 1 H), 6.09 (s, 1 H), 5.15 (br s, 2 H), 4.52 (br s, 2 H), 3.57 (br s, 2 H), 3.26 (s, 2 H), 2.43 (s, 3 H), 1.34 - 1.20 (m, 6 H). 2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methyl-N-[2-(piperazin-1yl)ethyl]quinolin-4-amine (7j). MS obsd. (ESI+) [(M+H)+] 466. 1H NMR (400 MHz, DMSO-d6) δ ppm 7.14 - 7.18 (m, 1 H), 7.04 (d, J = 7.58 Hz, 1 H), 6.74 - 6.82 (m, 2 H), 6.58 - 6.65 (m, 2 H), 6.47 (dd, J = 1.52, 8.59 Hz, 1 H), 5.22 (s, 1 H), 4.33 (s, 2 H), 3.72 (br s, 2 H), 2.77 (br t, J = 4.67 Hz, 2 H), 2.67 (t, J = 6.57 Hz, 2 H), 2.55 - 2.57 (m, 2 H), 2.13 (t, J = 4.80 Hz, 4 H), 1.94 (t, J = 6.57 Hz, 2 H), 1.79 (br s, 2 H), 1.60 (s, 3 H). 2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methyl-N-[2-(morpholin-4yl)ethyl]quinolin-4-amine (7k). MS obsd. (ESI+) [(M+H)+] 467. 1H NMR (400 MHz, CDCl3) δ ppm 8.05 (dd, J = 1.01, 7.83 Hz, 1 H), 7.66 (d, J = 7.33 Hz, 1 H), 7.51 (dt, J = 1.14, 7.52 Hz, 2 H), 7.38 (t, J = 7.66 Hz, 1 H), 7.33 (d, J = 8.63 Hz, 1 H), 7.28 (br s, 1 H), 5.88 (s, 1 H), 5.62 (br s, 1 H), 5.13 (s, 2 H), 4.6 (br s, 2 H), 3.74 - 3.81 (m, 4 H), 3.58 (br s, 2 H), 3.31 (q, J = 5.31 Hz, 2 H), 2.82 (t, J = 5.94 Hz, 2 H), 2.50 - 2.61 (m, 4 H), 2.46 (s, 3 H). 2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methyl-N-[2-(piperidin-1yl)ethyl]quinolin-4-amine (7l). MS obsd. (ESI+) [(M+H)+] 465. 1H NMR (400 MHz, CDCl3) δ ppm 8.04 (d, J = 7.6 Hz, 1 H), 7.67 (d, J = 7.6 Hz, 1 H), 7.51 (m, 2 H), 7.38 (t, J = 7.6 Hz, 1 H), 7.30 (m, 2 H), 5.88 (s, 1 H), 5.74 (m, 1 H), 5.12 (s, 2 H), 4.6 (br s, 2 H), 3.58 (s, 2 H), 3.28 (m, 2 H), 2.75 (t, J = 8.4 Hz, 2 H), 2.46 (m, 7 H), 1.62 (m, 4 H), 1.52 (m, 2 H). 2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methyl-N-(piperidin-2ylmethyl)quinolin-4-amine (7m). MS obsd. (ESI+) [(M+H)+] 451. 1H NMR (400 MHz, CD3OD) δ ppm 8.07 (d, J = 7.6 Hz, 1 H), 7.93 (s, 1 H), 7.85 (d, J = 7.6 Hz, 1 H), 7.73 - 7.70 (m, 2 H), 7.62 - 7.58 (m, 2 H), 6.02 (s, 1 H), 5.35 (s, 2 H), 4.51 (s, 2 H), 3.75 - 3.71 (m, 4 H), 3.51 - 3.40 (m, 2 H), 2.98 - 2.92 (m, 1 H), 2.46 (s, 3 H), 2.11 - 2.05 (m, 1 H), 1.96 - 1.90 (m, 2 H), 1.78 - 1.56 (m, 3 H).


Page 34 of 42

Page 35 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


N-[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4-yl]glycinamide (7n). The synthetic procedure was described in the Supporting Information. MS obsd. (ESI+) [(M+H)+] 411. 1H

NMR (400 MHz, CD3OD) δ ppm 8.51 (s, 1 H), 8.12 (s, 1 H), 8.07 (d, J = 7.6 Hz, 1 H), 8.00 (d, J = 7.6

Hz, 1 H), 7.87 (d, J = 8.8 Hz, 1 H), 7.67 - 7.12 (m, 2 H), 7.57 (t, J = 8.0 Hz, 1 H), 5.31 (s, 2 H), 4.66 (br s, 2 H), 4.25 (s, 2 H), 3.78 (s, 2 H), 2.52 (s, 3 H). N~2~-[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4-yl]glycinamide (7o). MS obsd. (ESI+) [(M+H)+] 411. 1H NMR (400 MHz, DMSO-d6) δ ppm 7.87 (d, J = 7.83 Hz, 1 H), 7.82 (d, J = 7.33 Hz, 1 H), 7.74 - 7.64 (m, 2 H), 7.52 - 7.41 (m, 2 H), 7.36 - 7.30 (m, 1 H), 7.30 - 7.19 (m, 2 H), 7.10 (t, J = 5.56 Hz, 1 H), 5.87 (s, 1 H), 5.00 (br s, 2 H), 4.40 (br s, 2 H), 3.86 (d, J = 5.81 Hz, 2 H), 3.59 (br s, 2 H), 3.32 (s, 1 H), 2.36 (s, 3 H). N-(2-{[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4yl]amino}ethyl)acetamide (7p). MS obsd. (ESI+) [(M+H)+] 439. 1H NMR (400 MHz, CD3OD) δ ppm 8.16 (s, 1 H), 7.95 (d, J = 1.8 Hz, 1 H), 7.86 (d, J = 1.9 Hz, 1 H), 7.62 (t, J = 3.6 Hz, 2 H), 7.46 (t, J = 3.8 Hz, 1 H), 7.31 (d, J = 2.1 Hz, 1 H), 7.22 (d, J = 2.1 Hz, 1 H), 6.84 (s, 1 H), 6.20 (s, 1 H), 5.10 (s, 2 H), 4.43 (br s, 2 H), 3.62 (s, 2 H), 3.32 (m, 4 H), 2.34 (s, 3 H), 1.89 (s, 3 H). 2-{[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4-yl]amino}ethanol (7q). MS obsd. (ESI+) [(M+H)+] 398. HRMS calcd [(M+H)+] 398.15329, measured [(M+H)+] 398.15313. 1H

NMR (400 MHz, CD3OD) δ ppm 8.08 (d, J = 7.6 Hz, 1 H), 7.91 (s, 1 H), 7.85 (d, J = 7.6 Hz, 1 H), 7.74

- 7.67 (m, 2 H), 7.59 - 7.56 (m, 2 H), 6.09 (s, 1 H), 5.29 (s, 2 H), 4.51 (s, 2 H), 3.83 (t, J = 5.6 Hz, 2 H), 3.74 - 3.72 (m, 2 H), 3.62 (t, J = 5.6 Hz, 2 H), 2.47 (s, 3 H). N-[3-(Aminomethyl)oxetan-3-yl]-2-(1,1-dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6methylquinolin-4-amine (7r). MS obsd. (ESI+) [(M+H)+] 439. HRMS calcd [(M+H)+] 439.17984, measured [(M+H)+] 439.17975. 1H NMR (400 MHz, CD3OD) δ ppm 7.98 - 7.96 (m, 1 H), 7.58 - 7.56 (d, J



= 6.8 Hz, 1 H), 7.44 - 7.40 (m, 2 H), 7.32 - 7.30 (t, 1 H), 7.19 (m, 2 H), 5.99 (s, 1 H), 5.54 (s, 2 H), 4.98 4.97 (d, J = 6.0, 4 H), 4.64 - 4.63 (m, 2 H), 3.51 (s, 2 H), 3.30 (s, 2 H), 2.34 (s, 3 H). N-[2-(1,1-Dioxido-2,3-dihydro-1,4-benzothiazepin-4(5H)-yl)-6-methylquinolin-4-yl]-N'-(oxetan-3yl)ethane-1,2-diamine (7s). MS obsd. (ESI+) [(M+H)+] 453, 1H NMR (400 MHz, CD3OD) δ ppm 8.10 (dd, J = 1.01, 7.83 Hz, 1 H), 7.88 (br d, J = 6.57 Hz, 2 H), 7.51 - 7.75 (m, 4 H), 6.03 (s, 1 H), 5.30 (s, 2 H), 4.76 - 4.87 (m, 2 H), 4.42 - 4.65 (m, 4 H), 3.99 - 4.22 (m, 1 H), 3.66 - 3.90 (m, 2 H), 3.48 - 3.66 (m, 2 H), 2.94 (br s, 2 H), 2.48 (s, 3 H). CPE assay. CPE assay was performed to assess the protective effects of compounds on cell viability. Plates (96-well) were seeded with 6000 Hep-2 cells per well. Cells were infected the next day with RSV at MOI 0.02 to produce an approximately 90% cytopathic effect after 5 days. Cells were incubated during this period in the presence or absence of serial dilutions of compounds. The viability of cells was assessed after 5 days by CCK-8 (Dojindo Molecular Technologies, Inc.). Results were expressed as 50% effective concentrations (EC50) and 50% cell cytotoxicity (CC50) values. Plaque reduction assays were carried out by infecting Hep-2 cell monolayers with 0.5 ml of 200 PFU/ml of RSV Long strain per well of a 12-well plate with or without the presence of serial diluted compounds. After 2 hours, cells were overlaid with DMEM/F12 containing 4% FBS and 0.55% agarose and compounds. Plates were incubated for 3 days and cells were then fixed with 4% paraformaldehyde for 6 hours. The agarose plugs were removed, and viral plaques were visualized by immunostaining. Cells were blocked with 1× TBS buffer with 1% BSA-0.5% Triton X-100. Plates were then incubated in the presence of a mouse anti-RSV monoclonal antibody (NCLRSV3; Novocastra) at 1:300 dilution followed by a rabbit anti-mouse horseradish peroxidase-labeled secondary antibody. The plaque staining was developed with 4-chloro-1-naphthol in the presence of hydrogen peroxide, and plaques were counted. In vivo efficacy study. Six-week-old female BALB/c mice were purchased from Jackson Laboratories and fed a standard diet and water ad libitum. Animals were housed in specific-pathogen-free conditions. The


Page 36 of 42

Page 37 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Animal Care Committee of Roche Innovation center China approved the protocol. Animals were anesthetized intraperitoneally with ketamine/xylazine before any intranasal administration. RSV long strain (5105 plaque-forming units [PFU] for all animal experiments), drugs, and controls were given in 100 µL volumes. Animals were euthanized with CO2, and lungs were harvested. For histopathologic analysis, the left lower lobe of the lung was removed and inflated with 10% formalin. Specimens were fixed, paraffin embedded, stained, and analyzed. ASSOCIATED CONTENTS SUPPORTING INFORMATION Detailed experimental procedures for the synthesis of analogues 2f, 2g and 7n, intermediates, 6b-c and 6c-c. 1H NMR,

13C

NMR, DEPT-135, H-H COSY, HSQC and HMBC Spectrum of Compound 1,

pharmacokinetic studies and molecular modeling. Molecular formula strings of all the compounds in this article. These materials are available free of charge via the Internet at http://pubs.acs.org. AUTHOR INFORMATION Corresponding authors *Hongying Yun (primary contact and paper writing): Phone: +86 21 28946727. Fax: +86 21 50790293. Email: [email protected]. NOTES The authors declare no competing financial interest. ABBREVIATION RSV, respiratory syncytial virus; RSV F, respiratory fusion; BAQ, benzoazepinequinoline; PAMPA, parallel artificial membrane permeability assay; LYSA, lyophilisation solubility assay; ADME, absorption, distribution, metabolism, and excretion; mlLogD, machine learning LogD; MW, microwave irradiation; PPB, plasma protein binding; SDPK, single-dose pharmacokinetics; PE, petroleum ether; EA, ethyl acetate; HPLC, high performance liquid chromatography; IPA, isopropanol; PD, pharmacodynamics; PK,



pharmacokinetic; prep-HPLC, preparative high performance liquid chromatography; SD, standard deviation. ACKNOWLEDGEMENTS We are grateful to Qingshan Gao, Ying Ji, Wei Li, Wenzhi Chen, Yang Lu, Yongguo Li, Liqin Chen, Peilan Ding, Wei Zhang, Hongxia Qiu, Yi Zhang, Yuxia Zhang, Sheng Zhong, Ki Sun, Rong Zhao, Shuang Ren, Jian Xin and Rong Zhao for purification of the final compounds and the analytical assistance. We thank the process group, including Jin She and Yi Ren, for their assistance with large scale synthesis campaigns. We also would like to thank Kunlun Xiang for CPE, plaque reduction and resistant mutation selection assays. In addition, we thank Wuxi and ChemPartner chemists for working together with us in some building block synthesis. REFERENCE 1. Borchers, A. T.; Chang, C.; Gershwin, M. E.; Gershwin, L. J. Respiratory syncytial virus-a comprehensive review. Clin. Rev. Allergy Immunol. 2013, 45, 331−379. 2. Piedimonte, G.; Perez, M. K. Alternative mechanisms for respiratory syncytial virus (RSV) infection and persistence: could RSV be transmitted through the placenta and persist into developing fetal lungs? Curr. Opin. Pharmacol. 2014, 16, 82−88. 3. Wright, M.; Piedimonte, G. Respiratory syncytial virus prevention and therapy: past, present, and future. Pediatr. Pulmonol. 2011, 46, 324−347. 4. Sigurs, N. Epidemiologic and clinical evidence of a respiratory syncytial virus reactive airways disease link. Am. J. Crit. Care Med. 2001, 163 (no 3), S2−6. 5. Falsey, A. R.; Hennessey, P. A.; Formica, M. A.; Cox, C.; Walsh, E. E. Respiratory syncytial virus infection in elderly and high-risk adults. N. Engl. J. Med. 2005, 352 (17), 1749−1759. 6. Boeckh, M.; Englund, J.; Li, Y.; Miller, C.; Cross, A.; Fernandez, H.; Kuypers, J.; Kim, H.; Gnann, J.; Whitley, R. Randomized controlled multicenter trial of aerosolized ribavirin for respiratory syncytial


Page 38 of 42

Page 39 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


virus upper respiratory tract infection in hematopoietic cell transplant recipients. Clin. Infect. Dis. 2007, 44, 245−249. 7. The Impact-RSV Study Group. Palivizumab, a humanized respiratory syncytial virus monoclonal antibody, reduces hospitalization from respiratory syncytial virus infection in high-risk infants. Pediatrics 1998, 102, 531−537. 8. Ventre, K.; Randolph, A. G. Ribavirin for respiratory syncytial virus infection of the lower respiratory tract in infants and young children. Cochrane Database Syst. Rev. 2010, DOI: 10.1002/14651858, CD000181. 9. Zhao, X.; Singh, M.; Malashkevich, V. N.; Kim, P. S. Structural characterization of the human respiratory syncytial virus fusion protein core. PNAS. 2000, 97, 14172−14177. 10. Ding, W. D.; Mitsner, B.; Krishnamurthy, G.; Aulabaugh, A.; Hess, C. D.; Zaccardi, J.; Cutler, M.; Feld, B.; Gazumyan, A.; Raifeld, Y.; Antonia Nikitenko, A.; Lang, S. A.; Gluzman, Y.; O’Hara, B.; Ellestad, G. A. Novel and specific respiratory syncytial virus inhibitors that target virus fusion. J. Med. Chem. 1998, 41, 2671−2675. 11. Andries, K.; Moeremans, M.; Gevers, T.; Willebrords, R.; Sommen, C.; Lacrampe, J.; Janssens, F.; Wyde, P. R. Substituted benzimidazoles with nanomolar activity against respiratory syncytial virus. Antiviral Res. 2003, 60, 209−219. 12. Bonfanti, J. F.; Meyer, C.; Doublet, F.; Fortin, J.; Muller, P.; Queguiner, L.; Gevers, T.; Janssens, P.; Szel, H.; Willebrords, R.; Timmerman, P.; Wuyts, K.; van Remoortere, P.; Janssens, F.; Wigerinck, P.; Andries, K. Selection of a respiratory syncytial virus fusion inhibitor clinical candidate. 2. Discovery of a morpholinopropylaminobenzimidazole derivative (TMC353121). J. Med. Chem. 2008, 51, 875−896. 13. Yu, K. L.; Sin, N.; Civiello, R. L.; Wang, X. A.; Combrink, K. D.; Gulgeze, H. B.; et al. Respiratory syncytial virus fusion inhibitors. Part 4: Optimization for oral bioavailability. Bioorg. Med. Chem. Lett. 2007, 17, 895−901.



14. Sun, Z.; Pan, Y.; Jiang, S.; Lu, L. Respiratory syncytial virus entry inhibitors targeting the F protein. Viruses 2013, 5, 211−225. 15. Douglas, J. L.; Panis, M. L.; Ho, E.; Lin, K. Y.; Krawczyk, S. H.; Grant, D. M.; Cai, R.; Swaminathan, S.; Cihlar, T. Inhibition of respiratory syncytial virus fusion by the small molecule VP-14637 via specific interactions with F protein. J. Virol. 2003, 77, 5054−5064. 16. Mackman, R. L.; Sangi, M.; Sperandio, D.; Parrish, J. P.; Eisenberg, E.; Perron, M.; et al. Discovery of an oral respiratory syncytial virus (RSV) fusion inhibitor (GS-5806) and clinical proof of concept in a human RSV challenge study. J. Med. Chem. 2015, 58, 1630−1643. 17. Stevens, M.; Rusch, S.; Huntjens, D.; Lounis, N.; Mari, N. K.; Remmerie, B.; Roymans, D.; Koul A.; Verloes, R.; DeVincenzo, J.; Kim, Y.; Harrison, L.; Meals, E. A.; DeVincenzo, J.; DeVincenzo, J.; Boyers, A.; Fok-Seang, J., Antiviral activity of oral JNJ-53718678 in healthy adult volunteers challenged with respiratory syncytial virus: a placebo-controlled study. J. Infect. Dis. 2018, 218(5), 748−756. 18. Zheng, X.; Wang, L.; Wang, B.; Miao, K.; Xiang, K.; Feng, S.; Gao, L.; Shen, H. C.; and Yun, H.; Discovery of piperazinylquinoline derivatives as novel respiratory syncytial virus fusion inhibitors. ACS Med. Chem. Lett. 2015, 7, 558−562. 19. Liang, C.; Yun, H.; Wang, L.; Gao, L.; Feng, S.; Wong, J. C.; Wu, J. Z.; Liu, Y.; Feng, L.; Chen, L.; Huang, M.; Guo, T.; Wu, X.; Zheng, X. Compounds for the Treatment and Prophylaxis of Respiratory Syncytial Virus Disease. WO2013/020993A1 Feb 14, 2013. 20. Burkhard, J. A.; Wuitschik, G.; Rogers-Evans, M.; Muller, K.; Carreira E. M. Oxetanes as versatile elements in drug discovery and synthesis. Angew. Chem. Int. Ed. 2010, 49, 9052. 21. Hanada, K.; Akimoto, S.; Mitsui, K.; Mihara, K.; Ogata, H. Enantioselective tissue distribution of the basic drugs disopyramide, flecainide and verapamil in rats: role of plasma protein and tissue phosphatidylserine binding. Pharmaceutical Research 1998, 15, 1250–1256. 22. Battles, M. B.; Langedijk, J. P.; Furmanova-Hollenstein, P.; Chaiwatpongsakorn, S.; Costello, H. M.; Kwanten, L.; Vranckx, L.; Vink, P.; Jaensch, S.; Jonckers, T. H. M.; Koul, A.; Arnoult, E.; Peeples, M.


Page 40 of 42

Page 41 of 42 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


E.; Roymans, D.; McLellan, J. S. Molecular mechanism of respiratory syncytial virus fusion inhibitors. Nat. Chem. Biol. 2016, 12, 87–93.



Table of Contents graphic HN

N H2

N H2

HN

OH N

N

O N

N

S O

4 RSV Long IC50/CC50: 0.223/ 20.8 μM HTS hit

1

O

RSV Long IC50/CC50: 0.002/ >100 μM BAQ with good in vivo efficacy


Page 42 of 42

Discovery of Benzoazepinequinoline (BAQ) Derivatives as Novel

Recommend Documents