Synthesis and molecular modeling of novel tetrahydro-β-carboline derivatives with phosphodiesterase 5 inhibitory and anticancer properties

J. Med. Chem. 2011, 54, 495–509 495

DOI: 10.1021/jm100842v

Synthesis and Molecular Modeling of Novel Tetrahydro-β-carboline Derivatives with

Phosphodiesterase 5 Inhibitory and Anticancer Properties

Heba A. Mohamed,^†,Nancy M. R. Girgis,^†,Rainer Wilcken,^‡Matthias R. Bauer,^‡Heather N. Tinsley,^§Bernard D. Gary,^§

Gary A. Piazza,^§Frank M. Boeckler,^‡and Ashraf H. Abadi*^,†

^†Department of Pharmaceutical Chemistry, Faculty of Pharmacy and Biotechnology, German University in Cairo, Cairo 11835, Egypt,

^‡Laboratory for Molecular Design and Pharmaceutical Biophysics, Department of Pharmaceutical and Medicinal Chemistry,

Institute of Pharmacy, Eberhard-Karls-University, Tu€bingen, Auf der Morgenstelle 8, 72076 Tu€bingen, Germany, and

^§Division of Drug Discovery, Department of Biochemistry and Molecular Biology, Southern Research Institute,

The University of Alabama at Birmingham, 2000 Ninth Avenue South, Birmingham, Alabama 35205, United States.

The first and second author contributed equally to this work

Received July 7, 2010

New derivatives based upon the tetrahydro-β-carboline-hydantoin and tetrahydro-β-carboline-piper-

azinedione scaffolds were synthesized. All compounds were evaluated for their ability to inhibit PDE5 in

vitro, and numerous compounds with IC₅₀values in the low nanomolar range were identified including

compounds derived from ^L-tryptophan. Compounds with high potency versus PDE5 were then evaluated

for inhibitory activity against other PDEs to assess isozyme selectivity. Compound 5R,11aS-5-(3,4-

dichlorophenyl)-2-ethyl-5,6,11,11a-tetrahydro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)dione

14 showed a selectivity index of >200 for cGMP hydrolysis by PDE5 versus PDE11. Meanwhile,

6R,12aR-6-(2,4-dichlorophenyl)-2-ethyl-2,3,6,7,12,12a-hexahydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-

1,4dione 45 demonstrated strong potency for inhibition of PDE11 with an IC₅₀value of 11 nM,

representing the most potent PDE11 inhibitor thus far reported. Docking experiments differentiated

between active and inactive analogues and revealing the conformational, steric, and lipophilic necessities for

potent PDE5 inhibition. Many derivatives, including potent PDE5 inhibitors, were able to inhibit the

growth of the MDA-MB-231 breast tumor cell line with low micromolar potency.

Introduction

PDE5 has received considerable attention over the years for

having been successfully developed as a therapeutic target for

erectile dysfunction. PE5 inhibitors may also have benefits for

pulmonary hypertension, cardioprotection, and enhancing

cognitive function.^3-5

Cyclic adenosine monophosphate (cAMP^a) and cyclic gua-

nosine monophosphate (cGMP) are important second mes-

sengers, responsible for transducing various extracellular

signals to the intracellular milieu. The intracellular levels of

cyclic nucleotides are controlled by the activities of cyclases,

which catalyze the formation of cyclic nucleotides from their

nucleotide triphosphate precursors and phosphodiesterases

(PDEs), which hydrolyze the cyclic nucleotide bond to termi-

nate the signal.¹On the basis of sequence homology and

regulatory properties, the PDEs can be classified into 11

distinct isozyme families. Often, there is more than one gene

product within each family and each gene generates multiple

splice variants.²PDE5 is a cGMP specific PDE that is expressed

in vascular smooth muscle and lung platelets, although recent

reports demonstrate that it is also overexpressed in various

carcinomas including breast, colon, lung, and bladder cancers.

Recent studies have demonstrated that PDE5 may serve as

a novel target for the cancer chemopreventive drug, sulindac.⁶

Moreover, certain tadalafil related analogues have shown tumor

cell growth inhibitory properties versus colorectal HT-29

cancer cell line that appear to be associated with their ability

to inhibit PDE5.⁷Other reports have revealed anticancer

activities to various tetrahydro-β-carbolines through different

mechanisms, including inhibition of topoisomerase II,⁸cyclin

dependent kinases,⁹and mitotic kinesin Eg5.¹⁰These obser-

vations suggest thattadalafilcan serveas a good starting point

for the discovery of novel anticancer compounds. However,

traditional PDE5 inhibitors are unable to induce cGMP

signaling in tumor cells and thus lack anticancer activity.

Three PDE5 inhibitors, tadalafil (Figure 1), sildenafil, and

vardenafil, have received Food and Drug Administration

(FDA) approval for the treatment of erectile dysfunction and

pulmonary hypertension. Although these drugs share a com-

mon mechanism of action, they have different pharmaco-

kinetic profiles, PDE isozyme selectivities, and interaction

mechanism with PDE5.^11-13For example, tadalafil has a long

half-life of 17.5 h and can also inhibit PDE11. The interaction

of tadalafil with PDE5 involves hydrogen bonding with the

conserved Gln817, which is essential for nucleotide recognition

*To whom correspondence should be addressed. Phone:

þ20227590716. Fax: þ20227581041. E-mail: ashraf.abadi@guc.edu.eg.

^aAbbreviations: cAMP, cyclic adenosine monophosphate; cGMP,

cyclic guanosine monophosphate; PDEs, phosphodiesterases; PDE5,

phosphodiesterase5; PDE3B, phosphodiesterase3B; PDE4, phospho-

diesterase4; FDA, Food and Drug Administration; t-butyl, tertiary-

butyl; CH₂Cl₂, methylene chloride; NOESY, nuclear Overhauser effect

spectroscopy; R_f, retardation factor; μM, micromolar; PDB, Protein

Data Bank; TMS, tetramethylsilane; TLC, thin layer chromatography;

IMAP, immobilized metal affinity for phosphochemicals; FP, fluores-

cence polarization; TAMRA-cGMP, tetramethylrhodamine-cGMP.

r

2010 American Chemical Society

Published on Web 12/28/2010

pubs.acs.org/jmc

496 Journal of Medicinal Chemistry, 2011, Vol. 54, No. 2

Mohamed et al.

and selectivity, and π-π stacking with Phe820. Unlike silde-

nafil and vardenafil, the rigid chemical structure of tadalafil,

which has only one rotatable bond, contributes to its binding

affinity because entropy is lost upon protein binding. Addi-

tionally, tadalafil has no direct or water mediated interaction

with the metal ions that are present in the binding pocket

of PDE5.^14-16

Chemically, tadalafil (Figure 1) is a tetrahydro-β-carboline-

piperazinedione with two chiral carbons that are both of the R

configuration, a methyl substituent on the piperazinedione

nitrogen, and a pendant 1,3-benzodioxol substituent attached

to C-6.

To expand the scope of structure-activity relationships

with this class of compounds, herein we report the discovery

of novel tadalafil analogues with the tetrahydro-β-carboline-

imidazoline and the tetrahydro-β-carboline-piperazinedione

scaffolds; the chiral carbons are of the R,R, R,S, S,R, or S,S

configuration, the terminal ring nitrogen substituent is methyl,

ethyl, butyl, or t-butyl, and the pendant aryl is 2,4-dichloro-

phenyl, 3,4-dichlorophenyl, or 2,6-dichlorophenyl.

To evaluate the biochemical and biological relevance of

these derivatives, they were assayed for their ability to inhibit

the recombinant human PDE5 enzyme and to inhibit tumor cell

growth usingthehumanMDA-MB-231breast tumorcell line.

acid esters.¹⁷The respective ester was subjected to Pictet-

Spengler reaction with 2,4-dichlorobenzaldehyde, 3,4-dichloro-

benzadehyde, and 2,6-dichlorobenzaldehyde. The reaction of

the ester with 2,4-dichlorobenzaldehyde and 3,4-dichloroben-

zadehyde gave both the cis and trans isomers as expected.

Because the cis and trans isomers are diastereomers, they were

separated by column chromatography using CH₂Cl₂as an

eluent. The reaction of the ester with the 2,6-dichlorobenzal-

dehyde yielded exclusively the cis isomer. Previously reported

data indicated that the trans isomer is the thermodynamically

favored isomer while the cis-isomer is kinetically favored.^18-21

A bulky tryptophan ester substituent, particularly benzyl,

favors the formation of the corresponding trans isomer.

Conversely, an allyl ester favors the formation of the cis

isomer upon reaction with an aldehyde.^18-21We found that

an aromatic aldehyde with bulky substituents at positions 2

and 6 of the benzene ring favors the formation of the cis

isomer upon reaction with tryptophan methyl ester, probably

due to restricted rotation. The Pictet-Spengler reaction was

carried out at room temperature under nonstereospecific

conditions. As expected, the kinetics were in favor of the cis

isomer.^18-21Thus, the formation of diastereomers 3, 5, and 7

exceeded the formation of diastereomers 4, 6, and 8, respec-

tively, with % de=11%, 27%, and 25%, respectively, mean-

while, for the isomers 1 and 9, there was no preferentiality

relative to 2 and 10.

Chemistry

Reaction of the respective pure cis- or trans-isomers of 1,3-

disubstituted tetrahydro-β-carboline (1-10) with commer-

cially available ethyl- and t-butyl isocyanate produced the

corresponding cis- and trans-hydantoins (11-30).

Treatment of (1-10) with chloroacetyl chloride provided

the corresponding chloroacetyl derivative(31-40). The β-car-

boline-piperazinedione derivatives with the respective N-alkyl

substituents were obtained by reaction of the respective

chloroethanone derivative with different primary amines,

The syntheses of the 1,3-disubstituted tetrahydro-β-carbo-

lines (Scheme 1), the tetrahydro-β-carboline-hydantoins

(Scheme 2), and the tetrahydro-β-carboline-piperazinediones

(Scheme 3) are shown. Both ^D- and L-tryptophan methyl ester

were synthesized by a general synthetic procedure for amino

Scheme 2^a

^aConditions: (a) ethyl or t-butylisocyanate/2-butanone/reflux; R =

-C₂H₅or -C-(CH₃)₃, X,Y = 2,4-dichloro, 3,4-dichloro, or 2,6-di-

chloro. For exact individual assignment, see Table 2. Stereocenters are

labeled.

Figure 1. Chemical structure of tadalafil with the points of mod-

ifications encircled.

Scheme 1. D or L-Tryptophan, D- or L-Tryptophan Methyl Ester, and 1,3-Disubstituted Tetrahydro-β-arboline^a

^aConditions: (a) CH₃COCl/methanol/reflux/6 h, basify and extract with CH₂Cl₂; (b) 2,4-dichlorobenzaldehyde, 3,4-dichlorobenzadehyde, or 2,6-

dichlorobenzaldehyde/TDA/4 days/RT. For exact indivividual assignment, see Table 1. Stereocenters are labeled.

Article

Journal of Medicinal Chemistry, 2011, Vol. 54, No. 2 497

Scheme 3

Conditions: (a) Cl-CO-CH₂-Cl/chloroform/NaHCO₃/RT/1 h; (b) methyl, ethyl, butyl or t-butyl amine/methanol/reflux/16hrs. For exact individual

assignment, see Tables 3 and 4. Stereocenters are labeled

Table 1. Chemical Structure, Growth Inhibition of MDA-MB-231 Breast Tumor Cell Line, and PDE5 Inhibitory Activity of 1,3-Disubstituted-

tetrahydro-β-carbolines (1-10)

% growth

inhibition

at 10 μM^a

growth inhibition

IC₅₀(μM) and

CI at p < 0.05^b

% PDE 5

inhibition at

10 μM^a

PDE 5 inhibition

IC₅₀(μM) and CI

at p < 0.05^b

compd

stereochemistry

X,Y

1

2

1R,3R

1S,3R

1S,3S

1R,3S

1R,3R

1S,3R

1S,3S

1R,3S

1R,3R

1S,3S

2,4-dichloro

3,4-dichloro

2,6-dichloro

29

78

50

28

43

100

62

48

25

52

ND

6.73 (2.21-12.5)

ND

65

40

55

82

78

50

18

24

25

15

7.2 (5.51-9.01)

ND

3

ND

0.57 (0.55-0.77)

1.61 (1.30-1.98)

ND

4

ND

5

6

4.62 (4.09-5.23)

6.98 (5.93-8.22)

ND

7

8

9

10

^aCalculated from duplicate values. ^bCalculated from at least 8 concentrations, each with quadruple replicates, p < 0.05.

including methylamine, ethylamine, butylamine, and t-butyl-

amine in refluxing methanol (41-68).

Mass spectrometry of all derivatives showed molecular ion

peaks at M^þ, M^þþ 2, and M^þþ 4 due to the isotopic nature

and number of chlorine atoms in the molecule. Most of the

1,3-disubstituted-tetrahydro-β-carboline derivatives deriva-

tives (1-10) showed the base peaks as molecular ion peaks,

indicating their stable nature.

The infrared spectra of (1-10) showed bands at a stretching

frequency around 3400 cm^-1for the N-H stretching and a

band around 1730 cm^-1for the ester carbonyl stretching. On

the other hand, the β-carboline-hydantoin derivatives (11-30)

showed two carbonyl stretching bands because the hydantoin

carbonyls are not equivalent as one of the carbonyls is flanked

between two nitrogen atoms and the other between carbon

and nitrogen.

The stretching vibrations of the two carbonyls of the

piperazinedione derivatives (41-68) were generally at lower

wave numbers relative to their hydantoin analogues. This may

be due to the higher ring strain of the five-membered

ring relative to the six-membered ring, producing higher

double bond characteristics of the carbonyls in the hydantoin

series.

The optical rotations of derivatives (5-8) showed that they

are optically active, with those derived from ^D-tryptophan

dextrorotatory and those derived from ^L-tryptophan levoro-

tatory. The optical rotation angle of a 0.1% ethanolic solution of

derivative 5 relative to 7 and compound 6 relative to 8 showed

equivalent values but opposite signs, indicating their enantio-

meric nature.

The assignment of cis/trans-stereochemistry for the tetra-

hydro-β-carbolines (1-10) was based on a detailed study of

¹³C NMR spectroscopic data well established in previous

literature.^22,23The ¹³C NMR signals for C-1 and C-3 are

more shielded in the trans-isomer compared to cis-isomer with

Δδ 3-4 ppm, which may be due to 1,3-diaxial spatial inter-

actions present in the trans-isomer. The tetrahydropyridine

ring exists in half-chair conformation where the ester at C-3 is

equatorially located. No analogous difference was detected

with C-3 in compounds (1-10). Moreover, a 500 MHz 2D

NOESY experiment that was carried out for cis-1 and trans-2

showed a clear correlation between the C-1 and C3-hydrogens

as expected for a cis-relationship with a coupling constant of

4.1 Hz (Supporting Information).

Moreover, a correlation exists between the R_fvalue on TLC

and the stereochemistry of the 1,3-disubstituted tetrahydro-β-

carboline (1-10) and their hydantoin analogues (11-30) where

the cis-isomer issystematicallylesspolar than thetrans-isomer

for the tetrahydro-β-carboline. However, in the hydantoin

series, the polarity is reversed, thus the cis-isomer becomes

more polar than the trans-isomer. For example, the R_fvalues

of the 1R,3R, 1S,3R, 1S,3S, and 1R,3S isomers of the (1-4)

and theircorrespondingN-ethylhydantoinanalogues(11-14)

using CH₂Cl₂as the mobile phase were 0.19, 0.12, 0.19, and

0.12 and 0.16, 0.28, 0.16, and 0.28, respectively.

The ¹H NMR signals for the proton at C-1 of derivatives

1-10 appeared at δ 5.2-6.2. Cyclization to the corresponding

hydantoin (11-30) or piperazinedione derivatives (41-68)

causes a large downfield shift to δ 5.8-7.0; this is mostly due

to the electron withdrawing effect of the carbonyl in the

neighboring ring.

Results and Discussion

All of the final compounds and intermediates synthesized

were screened for in vitro tumor cell growth inhibition activity

using the human MDA-MB-231 breast tumor cell line and for

498 Journal of Medicinal Chemistry, 2011, Vol. 54, No. 2

Mohamed et al.

Table 2. Chemical Structure, Growth Inhibition of MDA-MB-231 Breast Tumor Cell Line, and PDE5 Inhibitory Activity of Tetrahydro-β-carboline-

hydantoins (11-30)

% growth

inhibition at

10 μM^a

growth inhibition

IC₅₀(μM) and CI

at p < 0.05^b

% PDE 5

inhibition at

10 μM^a

PDE 5 inhibition

IC₅₀μM and CI at

p < 0.05^b

compd

stereochemistry

X,Y

R

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

30

5R,11aR

5S,11aR

5S,11aS

5R,11aS

5R,11aR

5S,11aR

5S,11aS

5R,11aS

5R,11aR

5S,11aS

5R,11aR

5S,11aR

5S,11aS

5R,11aS

5R,11aR

5S,11aR

5S,11aS

5R,11aS

5R,11aR

5S,11aS

2,4-dichloro

3,4-dichloro

2,6-dichloro

2,4-dichloro

3,4-dichloro

2,6-dichloro

-C₂H₅

39

54

29

68

50

60

78

18

55

52

68

65

22

69

99

58

70

60

38

15

ND

95

90

85

95

78

60

90

18

37

80

73

75

95

82

65

80

88

52

25

0.010 (0.005-0.024)

0.090 (0.08-0.11)

0.85 (0.67-1.07)

0.007 (0.004-0.009)

0.15 (0.10-0.24)

2.86 (2.40-3.40)

8.75 (6.90-10.64)

0.30 (0.23-0.40)

ND

5.37 (4.71-6.11)

ND

4.78 (1.03-12.12)

ND

-C₂H₅

ND

-C-(CH₃)₃

2.26 (1.52-3.33)

0.64 (0.50-0.80)

ND

0.59 (0.41-0.85)

1.48 (1.19-1.84)

4.14 (3.27-5.23)

0.070 (0.06-0.08)

1.87 (1.53-2.28)

6.38 (5.01-8.12)

2.31 (1.79-2.97)

0.012 (0.005-0.024)

ND

1.26 (1.04-1.53)

3.25 (1.46-4.0)

ND

1.43 (0.94-3.79)

ND

^aCalculated from duplicate values. ^bCalculated from at least 8 concentrations, each with quadruple replicates, p < 0.05.

Table 3. Chemical Structure, Growth Inhibition of MDA-MB-231 Breast Tumor Cell Line, and PDE5 Inhibitory Activity of 1,2,3-Trisubstituted-

tetrahydro-β-carbolines (31-40)

% growth

inhibition at

10 μM^a

growth inhibition

IC₅₀(μM) and CI

at p < 0.05^b

% PDE 5

inhibition at

10 μM^a

PDE 5 inhibition

IC₅₀μM and CI

at p < 0.05^b

compd

stereochemistry

X,Y

31

32

33

34

35

36

37

38

39

40

1R,3R

1S,3R

1S,3S

1R,3S

1R,3R

1S,3R

1S,3S

1R,3S

1R,3R

1S,3S

2,4-dichloro

3,4-dichloro

2,6-dichloro

98

55

98

40

50

43

55

15

30

0.56 (0.52-0.61)

0.07 (0.05-0.11)

ND

66

25

99

38

40

30

20

40

27

67

2.30 (1.71-3.08)

ND

0.38 (0.25-0.44)

ND

1.24 (1.05-1.48)

ND

0.73 (0.64-0.83)

^aCalculated from duplicate values. ^bCalculated from at least 8 concentrations, each with quadruple replicates, p < 0.05.

inhibition of recombinant human PDE5 at a single concentra-

tion of 10 μM. For compounds showing >60% inhibition, the

IC₅₀was determined by testing a range of eight concentrations

with quadruple replicates per concentration, tadalafil used as

a positive control. The results are shown in Tables 1-5.

For the 1,3-disubstituted tetrahydro-β-carbolines (1-10),

most of the derivatives were inactive against PDE5 at 10 μM,

with only 3 compounds (1, 4, and 5) showing activity below

the screening dose. Two of the active compounds have a 2,4-

dichlorophenyl substituent while one has a 3,4-dichlorophenyl

substituent. Conversely, all derivatives with the 2,6-dichlor-

ophenyl substituent (9 and 10) were inactive. This indicates

that PDE5 inhibitory activity is dependent on the positional

isomerization state of the compounds.

For the hydantoins (11-30), only derivatives with 2,4-

dichloro or 3,4-dichloro substitution at C-5 were able to inhibit

PDE5. Again, all derivatives with the 2,6-dichloro substitution

were inactive, confirming the importance of positional isomers

for PDE5 inhibitory activity. The most active compounds

were either 5R,11aS or 5R,11aR configuration. Additionally,

all compounds with the S configuration at the C-5 were

inactive or less active than their R congeners, indicating that

the stereochemical configuration of the carbon derived from

the aldehyde and not the stereochemical aspects of the carbon

derived from the tryptophan amino acid is an important

determinant of PDE5 inhibitory activity. This observation

expands the chemical space from which potent PDE5 inhibi-

tors may be discovered and presents an important economic

advantage, as the natural ^L-tryptophan is much more

widely available and cheaper than the non-natural ^D-isomer.

Additionally, the compounds derived from ^L-tryptophan

were more active than those derived from ^D-tryptophan

(e.g., 14 > 11, 24 > 21 and 28 > 25), further supporting

these conclusions.

The nature of the alkyl side chain group on the hydantoin

ring does not significantly influence the potency. It seems that

a wide range of substituents on the hydantoin nitrogen appear

to be tolerated without affecting PDE5 inhibitory activity.

Most of the 1,2,3-trisubstituted tetrahydro-β-carboline de-

rivatives 31-40 were inactive as PDE5 inhibitors. However,

three compounds (31, 33, and 40) did show moderate activity

with IC₅₀values of 0.38-2.3 μM. Compounds 33 and 40 were

Article

Journal of Medicinal Chemistry, 2011, Vol. 54, No. 2 499

Table 4. Chemical Structure, Growth Inhibition of MDA-MB-231 Breast Tumor Cell Line, and PDE5 Inhibitory Activity of Tetrahydro-β-carboline-

piperazinedione Derivatives (41-68)

% growth

inhibition at

10 μM^a

growth inhibition

IC₅₀(μM)^band

CI at p < 0.05

% PDE 5

inhibition at

10 μM^a

PDE 5 inhibition

IC₅₀μM^band CI

at p < 0.05

compd

stereochemistry

X,Y

R

41

42

43

44

45

46

47

48

49

50

51

52

53

54

55

56

57

58

59

60

61

62

63

64

65

66

67

68

6R,12aR

6S,12aR

6S,12aS

6R,12aS

6R,12aR

6S,12aR

6S,12aS

6R,12aS

6R,12aR

6S,12aR

6S,12aS

6R,12aS

6R12aR

6S,12aS

6R,12aR

6S,12aR

6S,12aS

6R,12aS

6R,12aR

6S,12aR

6S,12aS

6R,12aS

6R,12aR

6S,12aR

6S,12aS

6R,12aS

6R,12aR

6S,12aS

2,4-dichloro

3,4-dichloro

2,6-dichloro

2,4-dichloro

3,4-dichloro

2,6-dichloro

-CH₃

-C₂H₅

-C₄H₉

10

65

70

23

30

66

52

20

30

44

35

50

31

24

45

65

82

42

15

70

71

30

36

40

58

50

16

20

ND

3.26 (2.52-4.20)

7.10 (4.70-9.33)

ND

95

90

82

95

91

90

85

95

80

45

67

78

40

28

95

69

78

89

94

70

78

70

35

30

35

85

21

0.050 (0.03-0.068)

1.20 (0.82-2.34)

0.85 (0.70-1.03)

0.050 (0.041-0.067)

0.002 (0.0006-0.008)

0.26 (0.14-0.48)

0.99 (0.87-1.13)

0.010 (0.006-0.022)

1.34 (0.60-2.87)

ND

1.54 (1.21-1.95)

ND

3.30 (1.11-4.60)

2.04 (1.86-2.22)

ND

0.27 (0.19-0.37)

1.11 (0.56-2.21)

4.46 (3.70-5.36)

0.29 (0.23-0.35)

0.19 (0.09-0.43)

1.26 (0.70-2.35)

1.35 (1.08-1.69)

0.26 (0.20-0.32)

ND

1.20 (0.66-2.18)

3.28 (2.75-3.86)

ND

-C₄H₉

-C-(CH₃)₃

ND

2.92 (1.61-5.29)

2.59 (1.48-4.56)

ND

1.27 (1.11-1.45)

ND

^aCalculated from duplicate values. ^bCalculated from at least 8 concentrations, each with quadruple replicates, p < 0.05.

Table 5. PDE Isozyme Selectivity of the Most Active Tetrahydro-β-carboline-hydantoins and Piperazinedione Derivatives

PDE3B

inhibition IC₅₀

PDE4B

inhibition IC₅₀

PDE5A inhibition

PDE3B

PDE11A inhibition

IC₅₀(μM,cGMP)^binhibition IC₅₀(μM,cGMP)^band (μM,cAMP)^band IC₅₀(μM, cAMP)^band IC₅₀, (μM, (cGMP)^bselectivity

compd

11

and CI at p < 0.05

(μM,cAMP)^b

CI at p < 0.05

and CI at p < 0.05

index^a

0.010 (0.007-0.013)

0.007 (0.005-0.011)

0.002 (0.0009-0.006)

>50

34 (24.8-47.1)

3.60 (2.80-4.60) 13.00 (10.9-15.6)

>50

2.00 (1.4-2.90)

0.31 (0.25-0.38)

1.50 (1.28-1.80)

0.011 (0.008-0.015)

0.05 (0.044-0.055)

31.0

208.3

5.50

14

45

0.104 (0.09-0.14)

0.30 (0.23-0.38)

tadalafil 0.003 (0.0007-0.005)

16.60

^aIC₅₀of PDE11A (cGMP)/IC₅₀of PDE5A (cGMP). ^bCalculated from at least 8 concentrations, each with quadruple replicates, p < 0.05.

of the S,S configuration and compound 40 was the only

derivative with the pendant 2,6-dichlorophenyl that showed

PDE5 inhibitory activity. This demonstrates that these deri-

vatives may be following a different trend, which could be due

to their being alkyl halides and, therefore, having an alternate

means of interacting with PDE5.

We then expanded the hydantoin ring to a six-membered

piperazinedione system. Again, PDE5 inhibitory activity was

limited to derivatives with pendant 2,4-dichlorophenyl or 3,4-

dichlorophenyl, whereas those derivatives with the 2,6-di-

chlorophenyl substituent were inactive, confirming that the

potency for PDE5 inhibition is related to the nature of the

positional isomers. Furthermore, these observations indicate

that those derivatives with the 2,6-dichlorophenyl moiety

may be sterically locked in a conformation that prevents

binding to PDE5.

derived from the amino acid. In the piperazinediones, we intro-

duced four substituents on the nitrogen of the piperazinedione

ring with 2,4-dichlorophenyl, viz methyl, ethyl, butyl, and

tert-butyl. The order of PDE inhibitory activity was ethyl >

methyl > tert-butyl > butyl as shown by compounds 41 and

44 versus 45 and 48 versus 59 and 62 versus 55 and 58,

respectively. This indicates that there is an optimum size for

the N substitution and steric parameters are of importance in

this regard.

Compounds 11, 14, and 45 were the most potent of these

derivatives and showed comparable potency to tadalafil for

PDE5 inhibition. Therefore, we determined their isozyme

selectivity by comparing their potency for PDE5 inhibition

to their potency for inhibition of PDE3, PDE4, and PDE11.

As summarized in Table 5, all three compounds as well as the

reference compound tadalafil lacked the ability to inhibit

either cAMP or cGMP hydrolysis by PDE3B at concentra-

tions up to 50 μM. Interestingly, compounds 11 and 14

showed mild to moderate inhibition of the cAMP selective

PDE4B despite tadalafil having little effect on the activity of

The 6R,12aR or 6R,12aS rather than the 6S,12aR or

6S,12aS were more potent for PDE5 inhibition, confirming

that PDE5 inhibition is predominantly determined by the

stereochemical state of C-6 rather than the chiral carbon

500 Journal of Medicinal Chemistry, 2011, Vol. 54, No. 2

Mohamed et al.

˚

Figure 3. Docked pose of compound 11 (orange) in the cavity of

1UDU superimposed on tadalafil, white, featuring van der Waals

spheres on the chlorine atoms illustrating the steric requirements

of 11.

Figure 2. Side view of the crystal structure 1UDU with 2.83 A

resolution (yellow) containing a tadalafil molecule (green) in the

active side. Another PDE5 structure from the PDB with a higher

˚

resolution of 2.30 A that was cocrystallized with sildenafil, 2H42

(light blue), is superposed onto 1UDU to account for unresolved

amino acids in the flexible loop region of 1UDU.

this PDE isoform. The effects of the compounds on cAMP

and cGMP hydrolysis by PDE11A showed that the piperazi-

nedione derivative 45 and tadalafil were the most efficacious

and potent for PDE11 inhibition. Furthermore, compound 45

represents the most potent PDE11 inhibitor reported to date

withanIC₅₀value of11nM. Thehydantoinderivatives11and

14 were more selective for inhibition of cGMP hydrolysis by

PDE5 compared to inhibition of cGMP hydrolysis by PDE11A

with selectivity indices of 31 and 208, respectively. Conversely,

tadalafil possessed a selectivity index of just 16.6 for PDE5

versus PDE11, suggesting thatcompounds11and14possess a

lower potential for side effects due to their reduced effect upon

PDE11. Additionally, these studies demonstrate that the

PDE5/PDE11 selectivity of these compounds is primarily

influenced by the size and interactions of the tetracycle terminal

ring and not the pendant aryl.

We performed in silico docking studies in order to elucidate

the structural basis for the binding of the compounds.

Tadalafil has been cocrystallized with human PDE5 (PDB

1UDU) at a low resolution of 2.8 A, while sildenafil has been

cocrystallized with human PDE5 (PDB 2H42) at a better

˚

resolution of 2.3 A. Analysis of the crystal structures showed

Figure 4. Docked pose of compound 11 (orange) in the cavity of

1UDU (yellow). The flexible loop from 2H42 containing Ile665 is

superimposed for comparison (light blue).

ꢀ

˚

with the central scaffold of tadalafil. The 4-chloro substituent

is buried deeply within the central hydrophobic cavity. The

geometry of the hydrogen bond with Gln817 as well as the π-

stacking interaction with Phe820, as observed in tadalafil, is

also conserved.The interaction with Gln817 is believed to

restrict the rotational freedom of the side chain of Gln817

which is critical for cGMP binding.²⁴

Compound 12 differs from 11 only in one stereocenter. The

inversion of the 5R center to 5S induces a 180° flip of the

central scaffold. This has consequences both on the proposed

binding mode (i.e., the docking pose possessing the optimal

consensus score) and on the measured affinity. The hydrogen

bond between Gln817 and the NH-group of the indole moiety

present in both tadalafil and all potent novel compounds of R

configuration is lost. Additionally, the added carbonyl group

clashes with the carbonyl of Gln817 (red spheres), leading to

significantly reduced binding affinity. However, the π-π stack-

ing interaction with Phe820 is conserved as shown in Figure 5.

The biological data revealed the 2,6-dichloro analogues to

be substantially impaired PDE5 inhibitors. The docked pose

that some residues in close proximity to the binding site are

not resolved in 1UDU. Superimposing the two crystal struc-

tures highlights the difference between 1UDU and 2H42:

following Tyr664, which is a part of the flexible loop, Ile665

is present in 2H42 but not in 1UDU. From this structural

comparison, it seems reasonable to assume that this isoleucine

residue could form substantial hydrophobic interactions with

the N-alkyl substituent of tadalafil or its analogues as shown

in Figure 2.

Compound 11, which was found to inhibit PDE5 with low

nanomolar affinity, docked into the PDE5 active site in

1UDU in a very similar manner to that of tadalafil as shown

in Figure 3. However, the effects of the hydrophobic side

chains at the five-membered ring can not be reproduced due

to the missing hydrophobic loop region containing Ile665

(Figure 4). Compound 11 has the same stereoconfiguration as

tadalafil at the carbon where the dichlorobenzene ring is

attached, leading to a rather close match of the docked pose

Article

Journal of Medicinal Chemistry, 2011, Vol. 54, No. 2 501

Figure 6. Effects of 2,6-dichloro substitution at the aromatic ring

system exemplified by the docked pose of 53 (orange) superimposed

on tadalafil, white.

Figure 5. Docked pose of compound 12, orange, superimposed on

tadalafil, white.

active among all of the compounds presented here for growth

inhibition with a submicromolar IC₅₀value. Again, none of the

active compounds were of the 2,6-dichlorophenyl derivatives.

In addition to PDE5 inhibitors, this series of compounds were

alkylating agents and this may contribute to their activity.

For the piperazinedione derivatives 41-68, seven com-

pounds showed growth inhibitory activity, with IC₅₀values

in the range of 1.2-7.1 μM. As with PDE5 inhibition, none of

the active growth inhibitors were of 2,6-dichlorphenyl sub-

stituent, confirming that such an isomer is deleterious to both

PDE5 and growth inhibitory activity. Additionally, all active

compounds were of the S configuration at the C-6 position,

indicating stereochemical preferentiality for this series of

compounds. The active derivatives had different substituents

on the terminal piperazinedione N, demonstrating that a wide

variety of substituents can be tolerated at this residue. Finally,

all of the active derivatives demonstrated appreciable potency

for PDE5 inhibition. Together, these data suggest that mod-

ifications to the terminal ring of the tetracycle in tetrahydro-β-

carboline-hydantoin and tetrahydro-β-carboline-piperazine-

dione skeletons improves the PDE5 selectivity and tumor cell

growth inhibitory properties of this group of compounds.

of compound 53 demonstrates that the steric requirements of

the compound (chloro substituents shown as green spheres)

exceed the size of the binding pocket (shown as a transparent

surface). These steric hindrances likely reduce binding affinity.

Additionally, the missing 4-chloro substituent in 53 leaves the

deep hydrophobic cavity unaddressed, as shown in Figure 6.

Because of previous reports that PDE5 may serve as a novel

targetfor the discovery of new anticancer drugs, the tumorcell

growth inhibitory activity of these derivatives was measured

versus the human MDA-MB-231 breast tumor cell line, a cell

line that has been shown to rely on PDE5 for growth and

survival.⁶Among the 1,3-disubstituted tetrahydro-β-carbo-

lines 1-10, only compounds 2, 6, and 7 showed IC₅₀values

of <10 μM. The three compounds share the 1S chiral carbon,

and they all possess 2,4-dichlorophenyl or 3,4-dichlorophenyl

pendant aryls. This stereochemistry preference is opposite to

that observed with PDE5 inhibition, whereas the pendant aryl

positional isomer preference is the same.

Only seven compounds of the hydantoin derivatives 11-30

showed growth inhibitory activity, with IC₅₀values of 0.64-

5.37 μM. Again, all the active compounds had either 2,4-

dichlorophenyl or 3,4-dichlorophenyl substitution but none

of them are 2,6-dichlorophenyl derivatives. This supports the

dependence of the growth inhibitory activity on the substitu-

tion patterns at the aromatic ring. Conversely, the compounds

were of variable stereochemistry at the chiral carbons, indi-

cating insensitivity to chirality issues within this series of

compounds. Interestingly, all growth inhibitory compounds

in this series were also PDE5 inhibitors, although potency

for PDE5 inhibition did not correlate with growth inhibition

and may reflect additional factors when comparing activities

within cells compared with effect observed in isolated

enzymes.

Conclusions

The present structure-activity relationship study revealed

the following first-time essential findings for interaction be-

tween tetrahydro-β-carboline-hydantoins and tetrahydro-β-

carboline-piperazinedione derivatives and PDE5 including:

the interaction between the alkyl side chain on the N of the

terminal ring and Ile665 and hydrophobic interaction of the

pendant aryl with Met816, this being in addition to the known

H-bonding of the indole NH with Gln817 and π-π stacking

between the indole ring and the phenyl of Phe820. The

stereochemistry of the chiral carbon derived from the alde-

hyde, not the one derived from the amino acid, is the most

crucial feature for PDE5 inhibition; this expands the chemical

space from which novel PDE5 inhibitors might be explored.

The size and lipophilicity of the aryl attached to the stereo-

carbon derived from the aldehyde modulates activity as well.

The cross interaction with PDE11 can be modulated by the

nature of the terminal ring; this may evoke new studies to

improve PDE5/PDE11 selectivity or to bend toward one of

For the chloroethanone derivatives 31-40, only three com-

pounds showed appreciable growth inhibitory effect, namely

31, 32, and 34. Compound 32 differs from 31 by one stereo-

center, and its growth inhibitory potency is almost 10 times

that of 31. This indicates that the potency of these compounds

for growth inhibition is related to the stereochemistry of the

C-1 derived from the aldehyde. Compound 32 was the most

502 Journal of Medicinal Chemistry, 2011, Vol. 54, No. 2

Mohamed et al.

them. Evaluation of the new 68 derivatives in a cancer cell line

screening showed that the antiproliferative activity is not

necessarily parallel to the PDE5 inhibition; however, the latter

may be among the contributing factors to the antiproliferative

activity and thus it is important to strike the right balance

between selectivity/nonselectivity of cGMP-PDEs inhibition.

The new emerging indications for PDE5 inhibitors confirm

the need to the discovery of more inhibitors of PDE5 for novel

indications.

Ar), 7.35 (d, 1H, J=7.0 Hz, Ar), 7.26-7.15 (m, 4H, Ar þ NH),

7.04-6.93 (m, 2H, Ar), 5.82 (s, 1H, CHPh), 4.03-4.00 (m, 1H,

CHCOOCH₃), 3.83 (s, 3H, OCH₃), 3.25 (dd, 1H, J=4.3/7.0 Hz,

CH_aH_b), 3.23-3.04 (m, 1H, CH_aH_b). ¹³C NMR: δ 172.82, 136.16,

135.01, 134.11, 132.10, 131.23, 130.02, 128.06, 126.81, 123.40,

122.26, 119.83, 118.28, 110.97, 109.33, 56.55 (C1), 54.53, 52.40

(C3), 25.28. MS: m/z 378 (M^þþ 4), m/z 376 (M^þþ 2), m/z 374

(M^þ), m/z 217 (100%). Elemental analysis: calculated for (C₁₉H₁₆-

Cl₂N₂O₂) C, H, N.

(1S,3R)Methyl-1-(2,4-Dichlorophenyl)-2,3,4,9-tetrahydro-1H-

β-carboline-3-carboxylate (2). Yield 34%; yellow powder; mp

130-133 °C. R_f= 0.12 (CH₂Cl₂). IR (cm^-1): 3388 (-NH-),

1735(-CO-).¹HNMR:δ7.64 (s, 1H, NH), 7.56 (d, 1H, J=7.0 Hz,

Ar), 7.47 (s, 1H, NH), 7.00 (d, 1H, J = 7.0 Hz, Ar), 7.27-7.12

(m, 5H, Ar), 5.97 (s, 1H, CHPh), 3.92-3.88 (m, 1H, CHCOO-

CH₃), 3.74 (s, 3H, OCH₃), 3.30 (dd, 1H, J=4.6/7.6 Hz, CH_aH_b),

3.17-3.15 (m, 1H, CH_aH_b). ¹³C NMR: δ 173.09, 137.15, 136.29,

134.73, 134.51, 131.16, 130.83, 129.78, 127.27, 126.63, 122.48,

119.82, 118.38, 111.08, 109.43, 52.40(C1), 52.27, 51.15(C3), 24.39.

MS: m/z 378 (M^þþ 4), m/z 376 (M^þþ 2), m/z 374 (M^þ, 100%).

Elemental analysis: calculated for (C₁₉H₁₆Cl₂N₂O₂) C, H, N.

(1S,3S)Methyl-1-(2,4-Dichlorophenyl)-2,3,4,9-tetrahydro-1H-

β-carboline-3-carboxylate (3). Yield 45%; white powder; mp

90-92 °C; R_f=0.19 (CH₂Cl₂). IR (cm^-1): 3433 (-NH-), 1728

(-CO-). ¹H NMR: δ 7.90 (s, 1H, NH), 7.54-7.43 (m, 2H, Ar),

7.38 (d, 1H, J=7.2 Hz, Ar), 7.26-7.10 (m, 5H, Ar þ NH), 5.8 (s,

1H, CHPh), 3.90 -3.87 (m, 1H, CHCOOCH₃), 3.82 (s, 3H,

OCH₃), 3.25 (dd, 1H, J=4.3/7.6 Hz, CHaH_b), 3.01-2.90 (m,

1H, CH_aHb). ¹³C NMR: δ 172.90, 136.15, 135.10, 134.65, 133.15,

132.12, 131.45. 130.0, 129.38, 128.04, 126.85, 122.01, 119.8, 115.92,

110.96, 109.37, 56.57 (C1), 51.36 (C3), 25.38. MS: m/z 378

(M^þþ 4), m/z 376 (M^þþ 2), m/z 374 (M^þ, 100%). Elemental

analysis: calculated for (C₁₉H₁₆Cl₂N₂O₂) C, H, N.

Experimental Section

Chemistry. Mass spectra were measured using a Jeol MSta-

tion, double-focusing magnetic sector (MS 700). Melting points

were determined on a Buchi melting point apparatus and are

uncorrected. IR spectra were recorded on Perkin-Elmer Paragon

1

1000 using KBr-KCl plates. H NMR and ¹³C NMR spectra

were recorded on Jeol Eclipse, 400 and 500 MHz device using

CDCl₃as a solvent and TMS as a standard, chemical shifts (δ)

were reported in parts per million (ppm) downfield from TMS,

and J is expressed in Hz. The following abbreviations are used

for multiplicity of NMR signals: s=singlet, d=doublet, q =

quartet, m = multiplet, dd = doublet of doublet, br = broad.

Elemental analyses were performed by the Microanalytical

Unit, Faculty of Science, Cairo University; found values were

within (0.4% of the theoretical ones unless otherwise indicated.

Column chromatography was performed using silica-gel 70-

230 mesh. Reaction progress was monitored by TLC, performed

on precoated silica gel plates (ALUGRAM SIL G/UV254), and

detection of the components was made by short UV light. All

reactions were carried out under nitrogen or argon. All chemi-

cals and solvents were obtained from Sigma-Aldrich and were

used without further purification. All the described compounds

had >95% purity as determined by GC/MS and the areas under

the peaks.

(1R,3S)Methyl-1-(2,4-Dichlorophenyl)-2,3,4,9-tetrahydro-1H-

β-carboline-3-carboxylate (4). Yield 30%; white powder; mp

128-130 °C; R_f= 0.12 (CH₂Cl₂). IR (cm^-1): 3396 (-NH-),

1739 (-CO-). ¹H NMR: δ 7.80 (s, 1H, NH), 7.49 (s, 1H, NH),

7.46 (d, 1H, J=7.2 Hz, Ar), 7.19-7.11 (m, 5H, Ar), 6.94 (d, 1H,

J=7.0 Hz, Ar), 5.30 (s, 1H, CHPh), 3.74 (s, 3H, OCH₃), 3.68-

3.62 (m, 1H, CHCOOCH₃), 3.30 (dd, 1H, J=4.6/7.6 Hz, CHaH_b),

3.16-3.14 (m, 1H, CH_aHb). ¹³C NMR: δ 173.17, 136.27, 134.70,

134.50, 131.12. 131.0, 129.77, 127.27, 126.65, 122.46, 120.01,

119.82, 118.39, 111.06, 109.46, 52.38 (C1), 51.28 (C3), 51.14.

MS: m/z 378 (M^þþ 4), m/z 376 (M^þþ 2), m/z 374 (M^þ, 100%).

Elemental analysis: calculated for (C₁₉H₁₆Cl₂N₂O₂) C, H, N.

(1R,3R)Methyl-1-(3,4-dichlorophenyl)-2,3,4,9-tetrahydro-1H-

β-carboline-3-carboxylate (5). Yield 35%; yellow powder; mp

90-92 °C; R_f= 0.15 (CH₂Cl₂). IR (cm^-1): 3396 (-NH-), 1734

(-CO-). ¹H NMR: δ 9.0 (s, 1H, NH), 7.51-7.44 (m, 5H, Ar þ

NH), 7.25 (d, 1H, J=7.2 Hz, Ar), 7.20-7.10 (m, 2H, Ar), 5.23 (s,

1H, CHPh), 3.96 (dd, J=4.0/7.6 Hz, 1H, CH_aH_b,), 3.82 (s, 3H,

OCH₃), 3.05-3.01 (m, 1H, CHCOOCH₃), 3.00-2.95 (m, 1H,

CH_aH_b). ¹³C NMR: δ 172.88, 140.98, 136.25, 133.30, 133.12,

132.74, 130.92, 130.58, 127.94, 126.90, 122.35, 119.89, 118.37,

111.03, 109.34, 57.73 (C1), 56.64, 52.42 (C3), 25.43. MS:m/z 378

(M^þþ 4), m/z 376 (M^þþ 2), m/z 374 (M^þ, 100%). Elemental

Procedure for Preparation of ^D-andL-Tryptophan Methyl Ester.¹³

A 250 mL round-bottomed flask containing methanol (100 mL)

was cooled in an ice-water bath. Acetyl chloride are (8 mL) was

then added dropwise to the flask over a period of 8 min using a

graduated dropping funnel while the flask was still in the ice bath.

The solution was stirred for 5 min before adding D or L-tryptophan

(15 g, 68.77 mmol) in one portion. The solution was then heated

to reflux and left for 5 h. It was then left to cool to room

temperature, and the solvent was removed under reduced pressure

to give tryptophan methyl ester hydrochloride. The free base was

obtained by adding dilute NH₄OH and extracting the base using

CH₂Cl₂(5 ꢀ 50 mL). The organic layer was dried over anhy-

drous Na₂SO₄and put under reduced pressure to give a yellow-

ish solid. This solid was then used without further purification.

General Procedures for the Preparation of Methyl 1-(Aryl)-

2,3,4,9-tetrahydro-1H-β-carboline-3-carboxylate. The appropri-

ate tryptophan methyl ester (15.52 g, 60.93 mmol) was added to

the appropriate dichlorobenzaldehyde (11.72 g, 66.95 mmol)

and dissolved in CH₂Cl₂(100 mL). The solution was cooled to

0 °C in an ice bath. Trifluoroacetic acid (3 mL) was then added

to the solution dropwise while still in the ice bath, and the

mixture was left to stir for 4 days at room temperature and under

N₂atmosphere. The reaction mixture was made basic by adding

dilute NH₄OH solution and then extracted with CH₂Cl₂(3 ꢀ

100 mL). The organic layer was washed with water and brine

and dried over anhydrous Na₂SO₄. It was then filtered and

evaporated under reduced pressure.

analysis: calculated for (C₁₉H₁₆Cl₂N₂O₂), C, H, N. [R]²⁰

þ58.35 (c=0.1, EtOH)

=

D

(1S,3R)Methyl-1-(3,4-dichlorophenyl)-2,3,4,9-tetrahydro-1H-

β-carboline-3-carboxylate (6). Yield 20%; yellow powder; mp

140-143 °C; R_f=0.10 (CH₂Cl₂). IR (cm^-1):3345 (-NH-), 1710

1

(-CO-). HNMR: δ 7.55 (d, 1H, J=7.2 Hz,, Ar), 7.50-7.02

(m, 3H, Ar), 7.20-7.10 (m, 5H, Ar), 5.49 (s, 1H, CHPh),

3.98-3.95 (m, 1H, CHCOOCH₃), 3.73 (s, 3H, OCH₃), 3.37 (d,

1H, J=8.0 Hz, CH_aH_b), 3.29 (dd, 1H, J=4.6/8.0 Hz, CH_aH_b,).

¹³C NMR: δ 172.87, 148.02, 136.27, 135.02, 132.95, 132.55, 130.72,

130.57, 128.02, 126.69, 122.51, 119.85, 118.45, 111.09, 108.57,

53.94 (C1), 52.59 (C3), 52.40, 24.17. MS: m/z 378 (M^þþ 4), m/z

376 (M^þþ 2), m/z 374 (M^þ, 100%). Elemental analysis:

The residue was purified and the isomers were separated by

column chromatography on silica gel eluting with CH₂Cl₂to

give the respective cis-isomer first followed by the trans-isomer.

(1R,3R)Methyl-1-(2,4-dichlorophenyl)-2,3,4,9-tetrahydro-1H-

β-carboline-3-carboxylate (1). Yield 35%; yellow powder; mp

88-92 °C; R_f=0.19 (CH₂Cl₂). IR (cm^-1): 3406 (-NH-), 1734

(-CO-). ¹H NMR: δ 7.48 (s, 1H, NH), 7.51 (d, 1H, J=7.0 Hz,

Article

Journal of Medicinal Chemistry, 2011, Vol. 54, No. 2 503

calculated for (C₁₉H₁₆Cl₂N₂O₂) C, H, N. [R]²⁰_D=þ69.5 (c =

0.1, EtOH).

4.40 (dd, 1H, J=4.3/7.0 Hz, CHC(O)N), 3.58-3.55 (m, 3H,

NCH₂þ CH_aH_b), 3.08-3.05 (m, 1H, CH_aH_b), 1.23 (t, 3H, J =

7.2 Hz, CH₃). ¹³C NMR: δ 171.29, 154.39, 136.67, 135.06, 134.68,

133.98, 132.21, 129.44, 129.37, 128.16, 125.95, 123.14, 120.37,

118.50, 111.31, 107.61, 57.93 (C1), 51.46 (C3), 33.76, 22.46,

13.46. MS: m/z 417 (M^þþ 4), m/z 415 (M^þþ 2), m/z 413 (M^þ),

m/z 252 (100%). Elemental analysis: calculated for (C₂₁H₁₇Cl₂-

N₃O₂) C, H, N.

(1S,3S)Methyl-1-(3,4-dichlorophenyl)-2,3,4,9-tetrahydro-1H-

β-carboline-3-carboxylate (7). Yield 38%; yellow powder; mp

88-91 °C; R_f=0.15 (CH₂Cl₂). IR (cm^-1): 3395 (-NH-), 1734

(-CO-). ¹H NMR: δ 7.51-7.44 (m, 5H, Ar þ NH), 7.25 (d, 1H,

J=7.2 Hz, Ar), 7.20-7.10 (m, 2H, Ar), 5.24 (s, 1H, CHPh), 3.95

(dd, 1H, J=4.0/8.0 Hz, CHCOOCH₃), 3.82 (s, 3H, OCH₃), 3.25

(dd, 1H, J=4.6/7.6 Hz, CH_aH_b), 3.05-2.95 (m, 1H, CH_aH_b).

¹³C NMR: δ 172.78, 140.81, 136.27, 133.16, 133.12, 132.78,

130.919, 130.62, 127.99, 127.51, 126.88, 122.37, 119.90, 118.37,

111.04, 109.32, 57.70 (C1), 52.42 (C3), 25.36. MS: m/z 378

(M^þþ 4), m/z 376 (M^þþ 2), m/z 374 (M^þ, 100%). Elemental

analysis: calculated for (C₁₉H₁₆Cl₂N₂O₂) C, H, N. [R]²⁰_D=-58.35

(c=0.1, EtOH)

(1R,3S)Methyl-1-(3,4-dichlorophenyl)-2,3,4,9-tetrahydro-1H-

β-carboline-3-carboxylate (8). Yield 25%; yellow powder; mp

139-142 °C; R_f= 0.10 (CH₂Cl₂). IR (cm^-1): 3386 (-NH-),

1731 (-CO-). ¹H NMR: δ 7.55 (d, 2H, J=7.0 Hz, Ar þ NH),

7.40-7.35 (m, 3H, Ar þ NH), 7.27 (d, 2H, J = 7.0 Hz, Ar),

7.21-7.10 (m, 2H, Ar), 5.45 (s, 1H, CHPh), 3.97-3.93 (m, 1H,

CHCOOCH₃), 3.73 (s, 3H, OCH₃), 3.31-2.26 (m, 1H, CH_aH_b),

3.18 (dd, 1H, J=4.6/8.0 Hz, CH_aH_b,). ¹³C NMR: δ 173.13, 139.96,

136.04, 135.80, 132.57, 130.45, 130.22, 129.07, 128.84, 126.98,

124.06, 121.91, 119.77, 118.01, 110.99, 108.63, 53.20 (C1), 52.36

(C3), 25.72. MS: m/z 378 (M^þþ 4), m/z 376 (M^þþ 2), m/z 374

(M^þ, 100%). Elemental analysis: calculated for (C₁₉H₁₆Cl₂-

N₂O₂) C, H, N. [R]²⁰_D=-69.5 (c=0.1, EtOH)

(1R,3R)Methyl-1-(2,6-dichlorophenyl)-2,3,4,9-tetrahydro-1H-

β-carboline-3 carboxylate (9). Yield 55%; red powder; mp 102-

105 °C; R_f=0.13 (100% CH₂Cl₂). IR (cm^-1): 3389 (-NH-),

1729 (-CO-). ¹H NMR: δ 7.58-7.52 (m, 3H, Ar þ NH), 7.28-

7.27 (m, 3H, Ar þ NH), 7.26-7.25 (m, 3H, Ar), 6.29 (s, 1H,

CHPh), 4.14-4.06 (m, 1H, CHCOOCH₃), 3.85 (s, 3H, OCH₃),

3.32 (dd, 1H, J=4.0/8.0 Hz, CH_aH_b), 3.03-2.99 (m, 1H, CH_aH_b).

¹³C NMR: δ 172.25, 138.07, 136.20, 135.88, 134.37, 130.47, 129.40,

127.84, 126.60, 121.82, 119.66, 118.07, 112.06, 111.02, 56.83

(C1), 54.07, 52.21 (C3), 37.04, 27.15. MS: m/z 378 (M^þþ 4), m/z

376 (M^þþ 2), m/z 374 (M^þ), m/z 217 (100%). Elemental analysis:

calculated for (C₁₉H₁₆Cl₂N₂O₂) C, H, N.

(1S,3S)Methyl-1-(2,6-dichlorophenyl)-2,3,4,9-tetrahydro-1H-

β-carboline-3-carboxylate (10). Yield 57%; red powder; mp 101-

103 °C; R_f=0.13 (100% CH₂Cl₂). IR (cm^-1): 3386 (-NH-), 1730

(-CO-). ¹H NMR: δ 7.51-7.50 (m, 3H, Ar þ NH), 7.43 (d, 1H,

J=7.2 Hz, Ar), 7.26-7.25 (m, 3H, Ar þ NH), 7.13-7.11 (m, 2H,

Ar), 6.21(s, 1H, CHPh), 4.07-4.04 (m, 1H, CHCOOCH₃), 3.83(s,

3H, OCH₃), 3.25 (dd, 1H, J=4.0/8.0 Hz, CH_aH_b), 2.99-2.96 (m,

1H, CH_aH_b). ¹³C NMR: δ 173.13, 136.96, 135.80, 134.3, 132.57,

130.46, 129.08, 128.84, 126.98, 121.91, 119.77, 118.01, 110.99,

108.64, 57.21 (C1), 54.21, 52.36 (C3), 36.88, 25.72. MS: m/z 378

(M^þþ 4), m/z 376 (M^þþ 2), m/z 374 (M^þ, 100%). Elemental

analysis: calculated for (C₁₉H₁₆Cl₂N₂O₂) C, H, N.

(5S,11aR) 5-(2,4-Dichlorophenyl)-2-ethyl-5,6,11,11a-tetrahy-

dro-1H-imidazo [1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione (12).

Yield 57%; yellow powder; mp 126-129 °C; R_f=0.28 (CH₂Cl₂).

1

IR (cm^-1): 3312 (-NH-), 1767, 1710 (-CO-). H NMR: δ

7.87 (s, 1H, NH), 7.53 (d, 1H, J=7.0 Hz, Ar), 7.31-7.12 (m, 6H,

Ar), 6.68 (s, 1H, CHPh), 4.51 (dd, 1H, J=4.3/7.6 Hz, CHC-

(O)N), 3.63-3.61 (m, 3H, NCH₂þ CH_aH_b), 2.92-2.89 (m, 1H,

CH_aH_b), 1.24 (t, 3H, J=7.2 Hz, CH₃). ¹³C NMR: δ 172.51,

155.24, 136.56, 135.80, 135.28, 133.57, 130.29, 130.17, 129.37,

128.10, 125.92, 123.11, 120.34, 118.46, 111.31, 107.61, 54.71

(C1), 49.13 (C3), 33.76, 22.46, 13.46. MS: m/z 417 (M^þþ 4), m/z

415 (M^þþ 2), m/z 413 (M^þ), m/z 252 (100%). Elemental

analysis: calculated for (C₂₁H₁₇Cl₂N₃O₂) C, H, N.

(5S,11aS) 5-(2,4-Dichlorophenyl)-2-ethyl-5,6,11,11a-tetrahy-

dro-1H-imidazo [1⁰,5⁰:1,6] pyrido[3,4-b]indole-1,3(2H)-dione (13).

Yield 68%; yellow powder; mp 226-228 °C; R_f=0.16 (CH₂Cl₂).

1

IR (cm-1): 3331 (-NH-), 1706 (-CO-), 1624 (-CO-). H

NMR: δ 7.90 (s, 1H, NH), 7.57 (d, 2H, J=7.8 Hz, Ar), 7.25-

7.18 (m, 5H, Ar), 6.41 (s, 1H, CHPh), 4.38 (dd, 1H, J=4.6/7.8 Hz,

CHC(O)N), 3.56-3.49 (m, 3H, NCH₂þ CH_aH_b), 3.07-3.04

(m, 1H, CH_aH_b), 1.20 (t, 3H, J=7.2 Hz, CH₃). ¹³C NMR: δ

171.27, 154.33, 136.65, 135.04, 134.62, 133.98, 132.17, 129.37,

129.35, 128.10, 125.30, 123.08, 120.31, 118.45, 111.30, 107.54,

57.89 (C1), 51.41 (C3), 33.72, 22.42, 13.42. MS: m/z 418 (M^þþ

4), m/z 416 (M^þþ 2), m/z 414 (M^þ, 100%). Elemental analysis:

calculated for (C₂₁H₁₇Cl₂N₃O₂) C, H, N.

(5R,11aS) 5-(2,4-Dichlorophenyl)-2-ethyl-5,6,11,11a-tetrahy-

dro-1H-imidazo [1⁰,5⁰:1,6] pyrido[3,4-b]indole-1,3(2H)-dione (14).

Yield 45%; yellow powder; mp 125-128 °C; R_f=0.28 (CH₂Cl₂).

1

IR (cm^-1): 3331 (-NH-), 1704 (-CO-), 1623 (-CO-). H

NMR: δ 7.96 (s, 1H, NH), 7.51 (d, 2H, 8.0 J=Hz, Ar), 7.26-

7.12 (m, 5H, Ar), 6.21 (s, 1H, CHPh), 4.51-4.48 (m, 1H,

CHC(O)N), 3.63-3.60 (m, 3H, NCH₂þ CH_aH_b), 2.92-2.88

(m, 1H, CH_aH_b), 1.24 (t, 3H, J=7.2 Hz, CH₃). ¹³C NMR: δ

172.50, 155.20, 136.55, 135.78, 135.23, 133.57, 130.22, 130.17,

130.13, 128.03, 125.89, 123.06, 120.28, 118.42, 111.27, 107.08,

54.64 (C1), 49.09 (C3), 33.91, 23.33, 13.46. MS: m/z 418 (M^þþ

4), m/z 416 (M^þþ 2), m/z 414 (M^þ), m/z 378 (100%). Elemental

analysis: calculated for C₂₁H₁₇Cl₂N₃O₂C, H, N.

(5R,11aR) 5-(3,4-Dichlorophenyl)-2-ethyl-5,6,11,11a-tetrahy-

dro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione (15).

Yield 52%; white powder; mp 229-232 °C; R_f= 0.18 (CH₂Cl₂).

1

IR (cm^-1): 3332 (-NH-), 1706 (-CO-), 1623 (-CO-). H

NMR: δ 7.88 (s, 1H, NH), 7.58 (d, 1H, J=7.0 Hz, Ar), 7.38 (s,

1H, Ar), 7.26-7.12 (m, 5H, Ar), 5.77 (s, 1H, CHPh), 4.35 (dd,

1H, J=4.6/9.7 Hz, CHC(O)N), 3.57-3.50 (m, 3H, NCH₂þ

CH_aH_b), 3.08-3.06 (m, 1H, CH_aH_b), 1.15 (t, 3H, J=7.2 Hz, CH₃).

¹³C NMR: δ 171.19, 154.79, 139.06, 136.86, 132.99, 132.73,

132.19, 130.88, 129.72, 127.13, 126.04, 123.16, 120.38, 118.61,

111.32, 107.52, 57.90 (C1), 55.60, 53.45 (C3), 33.74, 13.46. MS:

m/z 418 (M^þþ 4), m/z 416 (M^þþ 2), m/z 414 (M^þ, 100%).

Elemental analysis: calculated for (C₂₁H₁₇Cl₂N₃O₂) C, H, N.

(5S,11aR) 5-(3,4-Dichlorophenyl)-2-ethyl-5,6,11,11a-tetrahy-

dro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione (16).

Yield 49%; yellow powder; mp 100-102 °C; R_f=0.36 (CH₂-

Cl₂). IR (cm^-1): 3389 (-NH-), 1704 (-CO-), 1624 (-CO-).

¹H NMR: δ 7.94 (s, 1H, NH), 7.57 (d, 1H, J = 8.0 Hz, Ar),

7.44-7.41 (m, 2H, Ar), 7.30 (d, 1H, J=8.0 Hz, Ar), 7.26-7.21

(m, 4H, Ar), 6.26 (s, 1H, CHPh), 4.28-4.23 (m, 1H, CHC(O)N),

3.93 (q, 2H, J=7.2 Hz, NCH₂), 3.61-3.58 (m, 1H, CH_aH_b),

General Procedures for the Preparation of 5-(Aryl)-2-alkyl-

5,6,11,11a-tetrahydro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-

1,3(2H)-dione. An appropriate amount of 1,3-disubstituted tetra-

hydro-β-carbolines 1-10 (0.38 g, 1 mmol) was dissolved in methyl

ethyl ketone (10 mL) and stirred. Excess amount of ethyl or tert-

butyl isocyanate (1.6 mmol) was added, and the mixture was then

stirred and left to reflux under N₂atmosphere for 16 h. The solvent

was evaporated under reduced pressure, and the residue was

purified by column chromatography on silica gel eluting with

CH₂Cl₂. The solution containing the desired product from the

column was evaporated under reduced pressure to give the pure

powder of the desired product.

(5R,11aR) 5-(2,4-Dichlorophenyl)-2-ethyl-5,6,11,11a-tetrahy-

dro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione (11).

Yield 58%; yellow powder; mp 223-226 °C; R_f=0.16 (100%

1

CH₂Cl₂). IR (cm^-1): 3321 (-NH-), 1764, 1705 (-CO-). H

NMR: δ 7.79 (s, 1H, NH), 7.57 (d, 1H, J=7.0 Hz, Ar), 7.50-

7.49 (m, 1H, Ar), 7.30-7.12 (m, 5H, Ar), 6.45 (s, 1H, CHPh),

2.92-2.89 (m, 1H, CH_aH_b), 1.14 (t, 3H, J=7.2 Hz, CH₃). ¹³

NMR: δ 172.28, 154.84, 139.21, 136.68, 133.42, 133.25, 131.15,

C

504 Journal of Medicinal Chemistry, 2011, Vol. 54, No. 2

Mohamed et al.

130.00, 129.12, 127.62, 126.20, 125.98, 123.28, 120.40, 118.58,

111.35, 108.62, 53.11 (C1), 50.93 (C3), 33.88, 13.51. MS: m/z 418

(M^þþ 4), m/z 416 (M^þþ 2), m/z 414 (M^þ, 100%). Elemental

analysis: calculated for (C₂₁H₁₇Cl₂N₃O₂) C, H, N.

111.26, 107.40, 58.26, 57.26 (C1), 51.33 (C3), 29.70, 28.74, 22.53.

MS: m/z 445 (M^þþ 4), m/z 443 (M^þþ 2), m/z 441 (M^þ), m/z 252

(100%). Elemental analysis: calculated (C₂₃H₂₁Cl₂N₃O₂) C, H, N.

(5S,11aR) 5-(2,4-Dichlorophenyl)-2-tert-butyl-5,6,11,11a-tet-

rahydro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione

(22). Yield 47%; yellowish powder; mp 205-208 °C; R_f=0.60

(5S,11aS) 5-(3,4-Dichlorophenyl)-2-ethyl-5,6,11,11a-tetrahy-

dro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione (17).

Yield 67%; yellow powder; mp 230-234 °C; R_f=0.17 (CH₂Cl₂).

IR (cm^-1): 3363 (-NH-), 1708 (-CO-), 1699(-CO-). ¹H

NMR: δ 7.74 (s, 1H, NH), 7.58 (dd, 1H, J=2.0/8.0 Hz, Ar), 7.38

(d, 1H, 8.0 Hz, Ar), 7.26-7.19 (m, 5H, Ar), 5.75 (s, 1H, CHPh),

4.36 (dd, 1H, J=4.3/6.8 Hz, CHC(O)N), 3.55-3.50 (m, 3H,

NCH₂þ CH_aH_b), 3.09-3.02 (m, 1H, CH_aH_b), 1.21 (t, 3H, J=

7.2 Hz, CH₃). ¹³C NMR: δ 171.14, 154.78, 139.00, 136.82, 133.02,

132.75, 132.16, 130.89, 129.67, 127.10, 126.03, 123.16, 120.39,

118.61, 111.32, 107.52, 57.90(C1), 55.60(C3), 33.74, 22.379,

13.46. MS: m/z 418 (M^þþ 4), m/z 416 (M^þþ 2), m/z 414

(M^þ, 100%). Elemental analysis: calculated for (C₂₁H₁₇Cl₂-

N₃O₂) C, H, N.

1

(CH₂Cl₂). IR (cm^-1): 3312 (-NH-), 1764, 1698 (-CO-). H

NMR: δ 7.86 (s, 1H, NH), 7.53-7.50 (m, 2H, Ar), 7.28-7.15 (m,

5H, Ar), 6.64 (s, 1H, CHPh), 4.39 (dd, 1H, J = 4.3/7.0 Hz,

CHC(O)N), 3.51 (dd, 1H, J=4.3/7.0 Hz, CH_aH_b), 2.88-2.85

(m, 1H, CH_aH_b), 1.61 (s, 9H, C(CH₃)₃). ¹³C NMR: δ 173.67,

156.09, 136.13, 135.08, 133.35, 131.36, 130.40, 130.20, 130.07,

128.10, 125.97, 122.97, 120.24, 118.44, 111.21, 106.96, 58.27,

54.14 (C1), 48.94 (C3), 28.68, 23.61. MS: m/z 445 (M^þþ 4), m/z

443 (M^þþ 2), m/z 441 (M^þ), m/z 57.1 (100%). Elemental analysis:

calculated (C₂₃H₂₁Cl₂N₃O₂) C, H, N.

(5S,11aS) 5-(2,4-Dichlorophenyl)-2-tert-butyl-5,6,11,11a-tet-

rahydro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione

(23). Yield 58%; white powder; mp 288-290 °C; R_f=0.42 (CH₂-

Cl₂). IR (cm^-1): 3309 (-NH-), 1702 (-CO-), 1624 (-CO-).

¹H NMR: δ 7.81 (s, 1H, NH), 7.53 (d, 1H, J=7.2 Hz, Ar), 7.45

(d, 1H, J=7.2 Hz, Ar), 7.25-7.16 (m, 4H, Ar), 6.99-7.01 (m,

1H, Ar), 6.42 (s, 1H, CHPh), 4.24 (q, 1H, J=7.6 Hz, CHC(O)N),

3.43 (dd, 1H, J=4.0/7.6 Hz, CH_aH_b), 2.99-2.98 (m, 1H, CH_aH_b),

1.63 (s, 9H, C(CH₃)₃). ¹³C NMR: δ 172.32, 155.32, 136.58,

135.93, 134.22, 132.93, 132.51, 131.5, 129.53, 128.71, 128.15,

125.98, 122.99, 121.7, 120.26, 118.49, 111.25, 107.36, 58.23,

57.23 (C1), 51.31 (C3), 28.72, 22.50. MS:m/z 445 (M^þþ 4), m/

z 443 (M^þþ 2), m/z 441 (M^þ, 100%).Elemental analysis:

calculated (C₂₃H₂₁Cl₂N₃O₂) C, H, N.

(5R,11aS) 5-(3,4-Dichlorophenyl)-2-ethyl-5,6,11,11a-tetrahy-

dro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione (18).

Yield 50%; yellow powder; mp 105-107 °C; R_f=0.36 (CH₂Cl₂).

1

IR (cm^-1): 3389 (-NH-), 1702 (-CO-), 1624 (-CO-). H

NMR: δ 8.10 (s, 1H, NH), 7.54 (d, 1H, J=7.4 Hz, Ar), 7.42 (d,

1H, J=7.4 Hz, Ar), 7.32-7.31 (m, 1H, Ar), 7.26-7.22 (m, 4H,

Ar), 6.26 (s, 1H, CHPh), 4.27-4.25 (m, 1H, CHC(O)N), 3.93-

3.93 (m, 3H, NCH₂þ CH_aH_b), 2.92-2.88 (m, 1H, CH_aH_b), 1.21

(t, 3H, J=7.2 Hz, CH₃). ¹³C NMR: δ 172.30, 154.84, 139.28,

136.71, 133.39, 131.12, 129.99, 129.15, 127.61, 126.03, 125.98,

123.23, 120.35, 118.56, 111.36, 108.54, 53.10 (C1), 50.92 (C3),

33.88, 22.36, 13.50. MS: m/z 418 (M^þþ 4), m/z 416 (M^þþ 2), m/

z 414 (M^þ), m/z 213 (100%). Elemental analysis: calculated for

(C₂₁H₁₇Cl₂N₃O₂) C, H, N.

(5R,11aS) 5-(2,4-Dichlorophenyl)-2-tert-butyl-5,6,11,11a-tet-

rahydro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione

(24). Yield 54%, white powder; mp 204-207 °C; R_f= 0.59

(5R,11aR) 5-(2,6-Dichlorophenyl)-2-ethyl-5,6,11,11a-tetrahy-

dro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione (19).

Yield 56%; yellow powder; mp 84-86 °C; R_f=0.21 (CH₂Cl₂).

1

(CH₂Cl₂). IR (cm^-1): 3311 (-NH-), 1698, 1622 (-CO-). H

NMR: δ 7.92 (s, 1H, NH), 7.52 (d, 1H, J=7.2 Hz, Ar), 7.47 (d,

1H, J=7.2 Hz, Ar), 7.25 (d, 1H, J=7.0 Hz, Ar), 7.25-7.13 (m,

4H, Ar), 6.71 (s, 1H, CHPh), 4.37 (q, 1H, J=7.6 Hz, CHC(O)N),

3.49 (dd, 1H, 4.0/7.6 Hz, CH_aH_b), 2.87-2.84 (m, 1H, CH_aH_b),

1.62 (s, 9H, C(CH₃)₃). ¹³C NMR: δ 73.67, 156.06, 136.51,

136.11, 135.05, 133.37, 130.37, 130.16, 130.09, 128.06, 125.96,

122.94, 120.21, 118.42, 111.21, 106.94, 58.26, 57.23, 54.09 (C1),

48.92 (C3), 28.67, 23.60. MS: m/z 445 (M^þþ 4), m/z 443 (M^þþ

2), m/z 441 (M^þ), 442 (100%). Elemental analysis: calculated for

(C₂₃H₂₁Cl₂N₃O₂) C, H, N.

1

IR (cm^-1): 3338 (-NH-), 1768, 1707 (-CO-). H NMR: δ

7.65 (s, 1H, NH), 7.68 (d, 1H, J=8.0 Hz, Ar), 7.48-7.47 (m, 1H,

Ar), 7.30-7.17 (m, 5H, Ar), 6.84 (s, 1H, CHPh), 4.43 (dd, 1H,

J=4.0/8.4 Hz, CHC(O)N), 3.96-3.92 (m, 2H, NCH₂), 3.57-

3.54 (m, 1H, CH_aH_b) 3.11-3.07 (m, 1H, CH_aH_b), 1.22 (t, 3H,

J=7.2 Hz, CH₃). ¹³C NMR: δ 171.61, 153.90, 136.48, 135.47,

134.88, 132.00, 130.27, 129.92, 129.19, 128.75, 125.88, 122.86,

120.24, 118.48, 111.14, 108.49, 57.78 (C1), 51.56 (C3), 33.63,

21.82, 13.48. MS: m/z 417 (M^þþ 4), m/z 415 (M^þþ 2), m/z 413

(M^þ), m/z 214 (100%). Elemental analysis: calculated for

(C₂₁H₁₇Cl₂N₃O₂) C, H, N.

(5R,11aR) 5-(3,4-Dichlorophenyl)-2-tert-butyl-5,6,11,11a-tet-

rahydro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione

(25). Yield 53%; yellow powder; mp 175-177 °C; R_f= 0.38

(CH₂Cl₂). IR (cm^-1): 3278 (-NH-), 1706 (-CO-), 1647

(-CO-). ¹H NMR: δ 8.88 (s, 1H, NH), 7.54-7.43 (m, 2H,

Ar), 7.26-7.17 (m, 5H, Ar), 6.42 (s, 1H, CHPh), 4.25 (dd, 1H,

J=3.6/7.6 Hz, CHC(O)N), 3.56-3.53 (m, 1H, CH_aH_b), 3.48-

3.41 (m, 1H, CH_aH_b), 1.63 (s, 9H, C(CH₃)₃). ¹³C NMR: δ 173.06,

157.86, 143.22, 136.94, 133.83, 132.57, 132.33, 131.60, 128.31,

126.21, 126.58, 125.60, 122.54, 119.89, 118.60, 111.10, 105.99,

56.61 (C1), 54.64, 52.37, 51.36(C3), 28.69, 23.37. MS:m/z 445

(M^þþ 4), m/z 443 (M^þþ 2), m/z 441 (M^þ), m/z 442 (100%).

Elemental analysis: calculated for (C₂₃H₂₁Cl₂N₃O₂) C, H, N.

(5S,11aR) 5-(3,4-Dichlorophenyl)-2-tert-butyl-5,6,11,11a-tet-

rahydro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione

(26). Yield 56%; yellow powder; mp 275-277 °C; R_f= 0.66

(CH₂Cl₂). IR (cm^-1): 3329 (-NH-), 1701 (-CO-), 1624

(5S,11aS) 5-(2,6-Dichlorophenyl)-2-ethyl-5,6,11,11a-tetrahy-

dro1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione (20).

Yield 52%; yellow powder; mp 78-80 °C; R_f=0.22 (CH₂Cl₂).

1

IR (cm^-1): 3338 (-NH-), 1768, 1707 (-CO-). H NMR: δ

7.63 (s, 1H, NH), 7.56 (d, 1H, J=8.0 Hz, Ar), 7.48-7.47 (m, 1H,

Ar), 7.30-7.14 (m, 5H, Ar), 6.84 (s, 1H, CHPh), 4.42 (dd, 1H,

J=4.0/8.6 Hz, CHC(O)N), 3.96-3.93 (m, 2H, NCH₂), 3.57-

3.54 (m, 1H, CH_aH_b) 3.10-3.07(m, 1H, CH_aH_b), 1.23 (t, 3H,

J=7.2 Hz, CH₃). ¹³C NMR: δ 171.61, 153.90, 136.48, 135.48,

134.88, 132.01, 130.27, 129.93, 129.19, 128.75, 125.87, 122.85,

120.24, 118.48, 111.15, 108.49, 57.78 (C1), 51.57 (C3), 33.63,

21.82, 13.48. MS: m/z 417 (M^þþ 4), m/z 415 (M^þþ 2), m/z 413

(M^þ), m/z 214 (100%). Elemental analysis: calculated for

(C₂₁H₁₇Cl₂N₃O₂) C, H, N.

(5R,11aR) 5-(2,4-Dichlorophenyl)-2-tert-butyl-5,6,11,11a-tet-

rahydro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione

(21). Yield 50%; yellowish powder; mp 280-282 °C; R_f=

0.41(CH₂Cl₂). IR (cm^-1): 3308 (-NH-), 1756, 1699 (-CO-).

¹H NMR: δ 7.80 (s, 1H, NH), 7.53 (d, 1H, J=8.0 Hz, Ar), 7.47 (s,

1H, Ar), 7.30-7.17 (m, 5H, Ar), 6.37 (s, 1H, CHPh), 4.24 (dd,

1H, J=4.3/7.0 Hz, CHC(O)N), 3.43 (dd, 1H, J=4.2/7.0 Hz,

1

(-CO-). H NMR: δ 7.94 (s, 1H, NH), 7.58 (d, 2H, J=8.0

Hz, Ar), 7.42-7.37 (m, 4H, Ar), 7.29-7.21 (m, 1H, Ar), 6.22 (s,

1H, CHPh), 4.13 (q, 1H, J = 7.6 Hz, CHC(O)N), 3.47-3.43 (m,

1H, CH_aH_b), 2.87-2.84 (m, 1H, CH_aH_b), 1.59 (s, 9H, C(CH₃)₃).

¹³C NMR: δ 173.47, 155.63, 139.41, 136.64, 133.33, 133.09,

131.10, 129.98, 129.41, 127.65, 126.05, 123.14, 120.30, 118.58,

111.28, 108.52, 58.21, 53.44, 52.40 (C1), 50.68 (C3), 29.65, 28.68,

23.52. MS: m/z 445 (M^þþ 4), m/z 443 (M^þþ 2), m/z 441 (M^þ,

100%). Elemental analysis: calculated (C₂₃H₂₁Cl₂N₃O₂) C, H, N.

CH_aH_b), 2.98-2.97 (m, 1H, CH_aH_b), 1.58 (s, 9H, C(CH₃)₃). ¹³

C

NMR: δ 172.34, 155.35, 136.58, 135.94, 134.27, 132.91, 132.54,

131.23, 129.58, 128.72, 128.18, 126.02, 123.04, 120.30, 118.53,

Article

(5S,11aS)-5-(3,4-Dichlorophenyl)-2-t-butyl-5,6,11,11a-tetra-

Journal of Medicinal Chemistry, 2011, Vol. 54, No. 2 505

3.43-3.39 (m, 2H, CH₂). MS: m/z 454 (M^þþ 4), m/z 452

(M^þþ 2), m/z 450 (M^þ), m/z 373 (100%). Elemental analysis:

calculated for (C₂₁H₁₇Cl₃N₂O₃) C, H, N.

hydro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione (27).

Yield 48%; yellow powder; mp 174-175 °C; R_f=0.36 (CH₂Cl₂).

IR (cm^-1):3361 (-NH-), 1704 (-CO-), 1624 (-CO-). ¹H

NMR: δ 8.89 (s, 1H, NH), 7.61-7.36 (m, 4H, Ar), 7.26-7.22 (m,

3H, Ar), 6.32 (s, 1H, CHPh), 4.21 (dd, 1H, J = 3.6/7.6 Hz,

CHC(O)N), 3.43 (dd, 1H, J=3.6/7.8 Hz, CH_aH_b), 3.03-3.00

(m, 1H, CH_aH_b), 1.58 (s, 9H, C(CH₃)₃). ¹³C NMR: δ 172.21,

155.78, 139.73, 136.80, 133.06, 132.42, 130.97, 129.12, 126.58,

126.12, 123.06, 120.33, 118.64, 111.24, 107.55, 58.26 (C1), 57.21,

55.55, 53.42 (C3), 52.37, 29.68, 28.71, 23.37. MS: m/z 445 (M^þþ

4), m/z 443 (M^þþ 2), m/z 441 (M^þ), m/z 442 (100%). Elemental

analysis: calculated for (C₂₃H₂₁Cl₂N₃O₂) C, H, N.

(1S,3R)Methyl 1-(2,4-Dichlorophenyl)-2-(2-chloroacetyl)-2,3,4,

9-tetrahydro-1H-β-carboline-3-carboxylate (32). Yield 80%; green

powder; mp 106-109 °C; R_f=0.37 (100% CH₂Cl₂). IR (cm^-1):

3370 (-NH-), 1739, 1668 (-CO-). ¹HNMR:δ8.08 (s, 1H, NH),

7.51 (d, 1H, J = 7.2, Ar), 7.28-7.11 (m, 6H, Ar), 6.57 (s, 1H,

CHPh), 5.30 (br s, 1H, CHCOOCH₃), 4.18-4.10 (m, 2H, COC-

H₂Cl), 3.64 (s, 3H, OCH₃), 3.53-3.49 (m, 2H, CH₂). MS: m/z 454

(M^þþ 4), m/z 452 (M^þþ 2), m/z 450 (M^þ), m/z 373 (100%).

Elemental analysis: calculated for (C₂₁H₁₇Cl₃N₂O₃) C, H, N.

(1S,3S)Methyl 1-(2,4-Dichlorophenyl)-2-(2-chloroacetyl)-2,3,4,

9-tetrahydro-1H-β-carboline-3-carboxylate (33). Yield 88%; white

powder; mp 215-218 °C; R_f= 0.52 (CH₂Cl₂). IR (cm^-1): 3315

(5R,11aS) 5-(3,4-Dichlorophenyl)-2-tert-butyl-5,6,11,11a-tet-

rahydro-1H-imidazo[1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione

(28). Yield 55%; yellow powder; mp 273-275 °C; R_f= 0.68

(CH₂Cl₂). IR (cm^-1): 3320 (-NH-), 1702 (-CO-), 1624

1

(-NH-), 1737 (-CO-), 1664 (-CO-). H NMR: δ 7.50 (brs,

1H, NH), 7.26-7.15 (m, 7H, Ar), 6.57 (s, 1H, CHPh), 4.52 (s, 1H,

CHCOOCH₃), 4.20-4.15 (m, 2H, COCH₂Cl), 3.63 (br s, 3H,

OCH₃), 3.49-3.48 (m, 2H, CH₂). MS: m/z 454 (M^þþ 4), m/z 452

(M^þþ 2), m/z 450 (M^þ, 100%). Elemental analysis: calculated for

(C₂₁H₁₇Cl₃N₂O₃) C, H, N.

1

(-CO-). H NMR: δ 7.80 (s, 1H, NH), 7.59 (d, 2H, J=8.2

Hz, Ar), 7.44-7.39 (m, 4H, Ar), 7.29-7.20 (m, 1H, Ar), 6.30 (s,

1H, CHPh), 4.14 (q, 1H, J=8.0 Hz, CHC(O)N), 3.48-3.44 (m,

1H, CH_aH_b), 2.88-2.85 (m, 1H, CH_aH_b), 1.61 (s, 9H, C(CH₃)₃).

¹³C NMR: δ 173.43, 155.61, 139.36, 136.61, 133.35, 133.12,

131.12, 129.99, 129.40, 127.67, 126.03, 123.16, 120.33, 118.57,

111.26 108.59, 58.19, 52.40 (C1), 50.70 (C3), 28.66, 23.51.; MS:

m/z 445 (M^þþ 4), m/z 443 (M^þþ 2), m/z 441 (M^þ, 100%), m/z

442 (100%). Elemental analysis: calculated for (C₂₃H₂₁Cl₂-

N₃O₂) C, H, N.

(1R,3S)Methyl 1-(2,4-Dichlorophenyl)-2-(2-chloroacetyl)-2,3,4,

9-tetrahydro-1H-β-carboline-3-carboxylate (34). Yield 75%; white

powder; mp 109-112 °C; R_f=0.37 (CH₂Cl₂). IR (cm^-1): 3347

1

(-NH-), 1739 (-CO-), 1668 (-CO-). H NMR: δ 7.80 (brs,

1H, NH), 7.39 (d, 1H, J=8.0 Hz, Ar), 7.18-7.12 (m, 6H, Ar), 6.54

(s, 1H, CHPh), 5.56 (d, 1H, J=7.0 Hz, CHCOOCH₃), 3.95-3.93

(m, 2H, COCH₂Cl), 3.72 (s, 3H, OCH₃), 3.31 (dd, 1H, J = 3.6/7.6

Hz, CH_aCH_b), 3.18 (dd, 1H, J=3.6/7.6 Hz, CH_aCH_b). MS: m/z

454 (M^þþ 4), m/z 452 (M^þþ 2), m/z 450 (M^þ), m/z 373 (100%).

Elemental analysis: calculated for (C₂₁H₁₇Cl₃N₂O₃) C, H, N.

(1R,3R)Methyl 1-(3,4-Dichlorophenyl)-2-(2-chloroacetyl)-2,3,4,

9-tetrahydro-1H-β-carboline-3-carboxylate (35). Yield 86%; white

powder; mp 283-285 °C; R_f=0.47 (CH₂Cl₂). IR (cm^-1): 3283

(-NH-), 1737 (-CO-), 1662 (-CO-). ¹H NMR: δ 8.9 (s, 1H,

NH), 7.60 (d, 2H, J=7.6 Hz, Ar), 7.32-7.16 (m, 5H, Ar), 6.83 (s,

1H, CHPh), 4.99 (d, 1H, J = 7.0 Hz, CHCOOCH₃), 4.35 (m,

COCH₂Cl), 4.22 (dd, 1H, J=3.6/7.6 Hz, CH_aCH_b), 3.70 (dd, 1H,

J=3.6/7.6 Hz, CH_aCH_b), 3.49 (s, 3H, OCH₃). MS:m/z 454 (M^þþ

4), m/z 452 (M^þþ 2), m/z 450 (M^þ, 100%). Elemental analysis:

calculated for (C₂₁H₁₇Cl₃N₂O₃) C, H, N.

(5R,11aR) 5-(2,6-Dichlorophenyl)-2-tert-butyl-5,6,11,11a-tet-

rahydro-1H-imidazo [1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione

(29). Yield 48%; orange-yellow powder; mp 245-248 °C; R_f=

0.46 (CH₂Cl₂). IR (cm^-1): 3356 (-NH-), 1762, 1698 (-CO-).

¹H NMR: δ 7.62 (s, 1H, NH), 7.55 (d, 1H, J=8.2 Hz, Ar), 7.48-

7.47 (m, 1H, Ar), 7.26-7.19 (m, 5H, Ar), 6.79 (s, 1H, CHPh),

4.27 (dd, 1H, J=3.6/7.6 Hz, CHC(O)N), 3.42-3.41 (dd, J =3.6/

7.6 Hz, 1H, CH_aH_b), 3.08-3.05 (m, 1H, CH_aH_b), 1.61 (s, 9H,

C(CH₃)₃). ¹³C NMR: δ 172.68, 154.73, 136.49, 134.80, 134.39,

132.58, 131.45, 130.29, 129.56, 129.28, 128.88, 125.93, 122.80,

120.16, 118.54, 111.09, 108.54, 58.03, 56.93 (C1), 53.44, 51.30

(C3), 28.76, 21.85. MS: m/z 445 (M^þþ 4), m/z 443 (M^þþ 2), m/z

441 (M^þ), m/z 385 (100%). Elemental analysis: calculated for

(C₂₃H₂₁Cl₂N₃O₂) C, H, N.

(5S,11aS) 5-(2,6-Dichlorophenyl)-2-tert-butyl-5,6,11,11a-tet-

rahydro-1Himidazo [1⁰,5⁰:1,6]pyrido[3,4-b]indole-1,3(2H)-dione

(30). Yield 45%; yellowish powder; mp 243-245 °C; R_f=0.44

(CH₂Cl₂). ¹H NMR: δ 7.64 (s, 1H, NH), 7.56 (d, 1H, J=8.2 Hz,

Ar), 7.49-7.48 (m, 1H, Ar), 7.30-7.17 (m, 5H, Ar), 6.80 (s, 1H,

CHPh), 4.27 (dd, 1H, J=3.6/7.6 Hz, CHC(O)N), 3.41 (dd, 1H,

J=3.6/7.6 Hz, CH_aH_b), 3.09-3.06 (m, 1H, CH_aH_b), 1.62 (s, 9H,

C(CH₃)₃). ¹³C NMR: δ 172.67, 154.73, 136.47, 135.43, 134.79,

134.38, 132.58, 131.47, 130.29, 129.57, 129.29, 128.89, 122.81,

120.17, 118.54, 111.09, 108.55, 58.03, 56.93 (C1), 51.30 (C3),

29.04, 28.75, 21.84. MS: m/z 445 (M^þþ 4), m/z 443 (M^þþ 2), m/

z 441 (M^þ), m/z 385 (100%). Elemental analysis: valculated for

(C₂₃H₂₁Cl₂N₃O₂) C, H, N.

General Procedures for the Preparation of Methyl 1-(2,6-

Dichlorophenyl)-2-(2-chloroacetyl)-2,3,4,9-tetrahydro-1H-β-

carboline-3-carboxylate. Chloroacetyl chloride (6.41 mmol) was

added dropwise to a well stirred solution of an appropriate

amount of β-carboline 1-10 (2.67 mmol) and NaHCO₃(0.271 g,

3.23 mmol) in CHCl₃(40 mL) under ice cooling. The mixture

was then left to stir under nitrogen atmosphere at room tem-

perature for 1 h. The mixture was diluted with CH₂Cl₂, washed

with a solution of NaHCO₃and brine, dried over Na₂SO₄, and

evaporated under reduced pressure.

(1S,3R)Methyl 1-(3,4-Dichlorophenyl)-2-(2-chloroacetyl)-2,3,4,

9-tetrahydro-1H-β-carboline-3-carboxylate (36). Yield 79%; white

powder; mp 163-165 °C; R_f= 0.22 (CH₂Cl₂). IR (cm^-1):3387

(-NH-), 1734(-CO-), 1663 (-CO-). ¹H NMR: δ 8.9 (s, 1H,

NH), 7.77 (d, 2H, J = 8.2 Hz, Ar), 7.62-7.42 (m, 2H, Ar), 7.26-

7.23 (m, 3H, Ar), 6.72 (s, 1H, CHPh), 4.99-4.97 (m, 1H, CHC-

OOCH₃), 4.36-4.34 (m, 2H, COCH₂Cl), 4.10 (s, 1H, CH_aCH_b),

3.75 (dd, 1H, J=3.6/7.6 Hz, CH_aCH_b), 3.49 (s, 3H, OCH₃). MS:

m/z 454 (M^þþ 4), m/z 452 (M^þþ 2), m/z 450 (M^þ, 100%).

Elemental analysis: calculated for (C₂₁H₁₇Cl₃N₂O₃) C, H, N.

(1S,3S)Methyl 1-(3,4-Dichlorophenyl)-2-(2chloroacetyl)-2,3,4,

9-tetrahydro-1H-β-carboline-3-carboxylate (37). Yield 84%; white

powder; mp 283-284 °C; R_f=0.45 (CH₂Cl₂). IR (cm^-1): 3281

1

(-NH-), 1736(-CO-), 1662 (-CO-). H NMR:δ 8.9 (s, 1H,

NH), 7.77 (d, 2H, J=8.0 Hz, Ar), 7.61(d, 2H,J=8.0 Hz, Ar), 7.32-

7.17 (m, 3H, Ar), 6.83 (s, 1H, CHPh), 4.98 (d, 1H, J=7.0 Hz, CHC-

OOCH₃), 4.36-4.34 (m, 2H, COCH₂Cl),4.21(d,1H,J =3.4/7.6 Hz,

CH_aCH_b), 3.70 (dd, 1H, J=3.4/7.6 Hz, CH_aCH_b), 3.46 (s, 3H,

OCH₃). MS: m/z 454 (M^þþ 4), m/z 452 (M^þþ 2), m/z 450 (M^þ,

100%). Elemental analysis: calculated for (C₂₁H₁₇Cl₃N₂O₃) C,H,N.

(1R,3S)Methyl 1-(3,4-Dichlorophenyl)-2(2-chloroacetyl)-2,3,4,

9-tetrahydro-1H-β-carboline-3-carboxylate (38). Yield 77%; white

powder; mp 162-165 °C; R_f=0.24 (CH₂Cl₂). IR (cm^-1): 3275

(-NH-), 1738, 1663 (-CO-). ¹H NMR: δ 7.9 (s, 1H, NH), 7.75

(d, 2H, J=7.6 Hz, Ar), 7.22-7.12 (m, 5H, Ar), 6.03 (s, 1H, CHPh),

4.16 (d, 1H, J = 7.0 Hz, CHCOOCH₃), 4.13-4.06 (m, 2H,

COCH₂Cl), 3.61-3.58 (m, 2H, CH₂), 3.47 (s, 3H, OCH₃). MS:

m/z 454 (M^þþ 4), m/z 452 (M^þþ 2), m/z 450 (M^þ, 100%).

Elemental analysis: calculated for (C₂₁H₁₇Cl₃N₂O₃) C, H, N.

(1R,3R)Methyl

1-(2,4-Dichlorophenyl)-2-(2-chloroacetyl)-

2,3,4,9-tetrahydro-1H-β-carboline-3-carboxylate (31). Yield 78%;

orange powder; mp 215-218 °C; R_f=0.53 (CH₂Cl₂). IR (cm^-1):

3294 (-NH-), 1731, 1666 (-CO-). ¹HNMR:δ7.50 (s, 1H, NH),

7.30-7.16 (m, 7H, Ar), 6.63 (s, 1H, CHPh), 4.53 (s, 1H, CHCO-

OCH₃), 4.20-4.15 (m, 2H, COCH₂Cl), 3.64 (s, 3H, OCH₃),

506 Journal of Medicinal Chemistry, 2011, Vol. 54, No. 2

Mohamed et al.

(1R,3R)Methyl 1-(2,6-Dichlorophenyl)-2-(2-chloroacetyl)-2,3,4,

9-tetrahydro-1H-β-carboline-3-carboxylate (39). Yield 76%; brown

powder; mp 147-150 °C; R_f=0.44 (100% CH₂Cl₂). IR (cm^-1):

3304 (-NH-), 1739, 1666 (-CO-). ¹H NMR: δ 8.98 (s, 1H, NH),

7.75-7.15 (m, 7H, Ar), 6.90 (s, 1H, CHPh), 4.99 (s 1H,

CHCOOCH₃), 4.25-4.22 (m, 2H, COCH₂Cl), 3.67 (s, 3H, OCH₃),

3.37-3.16 (m, 2H, CH₂).MS:m/z454 (M^þþ 4), m/z452 (M^þþ 2),

m/z 450 (M^þ), m/z 375 (100%). Elemental analysis: calculated for

(C₂₁H₁₇Cl₃N₂O₃) C,H,N.

30%; white powder; mp 301-303 °C; R_f=0.23 (CH₂Cl₂/MeOH

98:2). IR (cm^-1): 3305 (-NH-), 1665 (-CO-), 1624 (-CO-).

¹H NMR: δ 8.09 (s, 1H, NH), 7.54 (d, 2H, J=8.0 Hz, Ar), 7.31

(d, 2H, J =8.0 Hz, Ar), 7.26-7.08 (m, 3H, Ar), 6.32 (s, 1H,

CHPh), 4.24 (dd, 1H, J=3.6/7.2 Hz, CH_aH_b), 4.14 (d, 1H, J=

7 Hz, CHC(O)N), 4.07 (d, 2H, J=7 Hz, CH₂C(O)N), 3.47 (dd,

1H, J=3.6/7.2 Hz, CH_aH_b), 3.01 (s,3H, NCH₃). ¹³C NMR: δ

165.86, 162.97, 135.61, 135.01, 133.70, 131.75, 130.25, 129.04,

128.15, 127.15, 126.14, 123.08, 120.31, 118.43, 111.33, 109.23,

52.95 (C1), 51.39, 49.34 (C3), 33.76, 27.16. MS: m/z 417 (M^þþ

4), m/z 415 (M^þþ 2), m/z 413 (M^þ), m/z 414 (100%). Elemental

analysis: calculated for (C₂₁H₁₇Cl₂N₃O₂) C, H, N.

(1S,3S)Methyl 1-(2,6-Dichlorophenyl)-2-(2-chloroacetyl)-2,3,4,

9-tetrahydro-1H-β-carboline-3-carboxyl (40). 78%; brown pow-

der; mp 153-156 °C. R_f=0.42 (100% CH₂Cl₂). IR (cm^-1): 3309

1

(6R,12aR) 6-(2,4-Dichlorophenyl)-2-ethyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (45). Yield

34%; white powder; mp 203-207 °C; R_f=0.36 (CH₂Cl₂/MeOH

98:2). IR (cm^-1): 3444 (-NH-), 1663, 1628 (-CO-). ¹H NMR:

δ 8.12 (s, 1H, NH), 7.59-7.11 (m,7H, Ar), 6.66 (s, 1H, CHPh),

4.38 (dd, 1H, J=3.6/7.2 Hz, CHC(O)N), 4.27-3.78 (m, 3H,

CH₂C(O)N þ CH_aH_b), 3.44-3.40 (m, 1H, CH_aH_b), 3.25-3.23

(m, 2H, NCH₂), 1.23 (t, 3H, J=7.2 Hz, CH₃). ¹³C NMR: δ

166.57, 165.53, 165.35, 138.83, 136.26, 133.61, 132.04, 129.42,

128.25, 127.20, 122.83, 120.21, 118.56, 111.30, 106.58, 56.27

(C1), 53.42 (C3), 49.56, 41.30, 27.00, 24.12, 12.00. MS: m/z 431

(M^þþ 4), m/z 429 (M^þþ 2), m/z 427 (M^þ), m/z 57 (100%).

Elemental analysis: calculated for (C₂₂H₁₉Cl₂N₃O₂) C, H, N.

(6S,12aR) 6-(2,4-Dichlorophenyl)-2-ethyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (46). Yield

33%; white powder; mp 227-230 °C; R_f=0.37 (CH₂Cl₂/MeOH

98:2). IR (cm^-1): 3394(-NH-), 1663, 1627 (-CO-). ¹H NMR:

δ 8.32 (s, 1H, NH), 7.54-7.09 (m,7H, Ar), 6.72(s, 1H, CHPh),

4.21 (dd, 1H, J=3.6/7.2 Hz, CHC(O)N), 4.12-3.99 (m, 3H,

CH₂C(O)N þ CH_aH_b), 3.57-3.49 (m, 1H, CH_aH_b), 3.47-3.37

(m, 2H, NCH₂), 1.22 (t, 3H, J=7.2 Hz, CH₃). ¹³C NMR: δ

165.39, 163.36, 136.36, 135.57, 135.03, 133.68, 131.84, 130.20,

129.12, 127.14, 126.15, 122.99, 120.23, 118.40, 111.38, 109.11,

52.99 (C1), 49.38 (C3), 48.80, 40.87, 27.04, 11.71. MS: m/z 431

(M^þþ 4), m/z 429 (M^þþ 2), m/z 427 (M^þ), m/z 57 (100%).

Elemental analysis: calculated for (C₂₂H₁₉Cl₂N₃O₂) C, H, N.

(6S,12aS) 6-(2,4-Dichlorophenyl)-2-ethyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (47). Yield

33%; white powder; mp 205-208 °C; R_f= 0.35 (CH₂Cl₂/

MeOH 98:2). IR (cm^-1): 3316 (-NH-), 1670 (-CO-), 1663

(-CO-). ¹H NMR: δ 8.21 (s, 1H, NH), 7.57 (d, 2H, J = 7.6 Hz,

Ar), 7.52 (d, 2H, J = 7.6 Hz, Ar), 7.39-7.10 (m, 3H, Ar), 6.62 (s,

1H, CHPh), 4.37 (dd, 1H, J = 3.6/7.2 Hz, CHC(O)N),

4.11-4.04 (m, 3H, CH₂C(O)N þ CH_aH_b), 3.41-3.39 (m, 1H,

CH_aH_b), 2.99-2.96 (m, 2H, NCH₂), 1.22 (t, 3H, J = 7.2 Hz,

CH₃). ¹³C NMR: δ 165.56, 163.36, 156.59, 131.84, 131.28,

130.21, 129.42, 128.26, 127.24, 122.83, 120.25, 120.21, 118.57,

111.37, 109.16, 106.59, 56.29 (C1), 53.57 (C3), 40.96, 30.95,

27.02. MS: m/z 430 (M^þþ 4), m/z 428 (M^þþ 2, 100%), m/z 426

(M^þ). Elemental analysis: calculated for (C₂₂H₁₉Cl₂N₃O₂)

C, H, N.

(-NH-), 1737, 1666 (-CO-). H NMR: δ 8.98 (s, 1H, NH),

7.30-7.15 (m, 7H, Ar), 6.90 (s, 1H, CHPh), 4.99 (s, 1H, CHC-

OOCH₃), 4.25-4.21 (m, 2H, COCH₂Cl), 3.68 (s, 3H, OCH₃),

3.22-3.17 (m, 2H, CH₂). MS: m/z 454 (M^þþ 4), m/z 452 (M^þþ

2), m/z 450 (M^þ), m/z 375 (100%). Elemental analysis: calcu-

lated for (C₂₁H₁₇Cl₃N₂O₃) C, H, N.

General Procedures for the Preparation of 2-Alkyl 6-(Aryl)-

2,3,6,7,12,12a-hexahydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-

1,4-dione. An appropriate amount of methyl amine (2.8 mmol)

was added to a solution of the appropriate amount of chloro-

acetyl derivative (1.4 mmol) in methanol (25 mL). The mixture

was heated to reflux under nitrogen atmosphere for 16 h. It was

then cooled to room temperature and evaporated to dryness under

reduced pressure. The residue was then dissolved in CH₂Cl₂, the

organic layer was washed with water and dried over Na₂SO₄,

filtered, and evaporated under reduced pressure. The crude prod-

uct was then purified using column chromatography.

(6R,12aR) 6-(2,4-Dichlorophenyl)-2-methyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (41). Yield

38%; white powder; mp 222-225 °C; R_f=0.24 (CH₂Cl₂/MeOH

98:2). IR (cm^-1): 3307 (-NH-), 1665, 1624 (-CO-). ¹H NMR:

δ 8.07 (s, 1H, NH), 7.58-7.17 (m, 7H, Ar), 6.64 (s, 1H, CHPh),

4.25 (dd, 1H, J=3.6/7.6 Hz, CHC(O)N), 4.23-4.07 (m, 3H,

CH₂C(O)N þ CH_aH_b), 3.47 (dd, 1H, J=3.4/7.6 Hz, CH_aH_b),

3.01 (s,3H, NCH₃). ¹³C NMR: δ 165.86, 162.96, 136.31, 135.61,

135.01, 131.76, 130.25, 129.05, 127.16, 126.14, 123.08, 120.31,

118.43, 111.33, 109.24, 56.45 (C1), 52.94, 51.39 (C3), 49.33,

33.34, 27.16. MS: m/z 417 (M^þþ 4), m/z 415 (M^þþ 2), m/z

413 (M^þ), m/z 57 (100%). Elemental analysis: calculated for

(C₂₁H₁₇Cl₂N₃O₂) C, H, N.

(6S,12aR) 6-(2,4-Dichlorophenyl)-2-methyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (42). Yield

35%; white powder; mp 305-308 °C; R_f=0.24 (CH₂Cl₂/MeOH

98:2). IR (cm^-1): 3305 (-NH-), 1665, 1624 (-CO-). ¹H NMR:

δ 8.27 (s, 1H, NH), 7.51-7.06 (m, 7H, Ar), 6.75 (s, 1H, CHPh),

4.23 (dd, 1H, J=3.4/7.2 Hz, CHC(O)N), 4.13 -4.01 (m, 3H,

CH₂C(O)N þ CH_aH_b), 3.47 (dd, 1H, J=3.4/7.2 Hz, CH_aH_b),

3.00 (s,3H, NCH₃). ¹³C NMR: δ 165.88, 162.99, 136.35, 135.59,

135.00, 133.71, 131.79, 130.21, 129.06, 127.15, 123.03, 120.26,

118.41, 111.37, 109.14,53.44, 52.92 (C1), 51.38, 49.37 (C3),

33.33, 27.17. MS: m/z 417 (M^þþ 4), m/z 415 (M^þþ 2), m/z

413 (M^þ), m/z 57 (100%). Elemental analysis: calculated for

(C₂₁H₁₇Cl₂N₃O₂) C, H, N.

(6R,12aS) 6-(2,4-Dichlorophenyl)-2-ethyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (48). Yield

30%; white powder; mp 225-228 °C; R_f=0.34 (CH₂Cl₂/MeOH

98:2). IR (cm^-1): 3287 (-NH-), 1665 (-CO-), 1658 (-CO-).

¹H NMR: δ 8.34 (s, 1H, NH), 7.52 (d, 2H, J=7.6 Hz, Ar), 7.43

(d, 2H, J =7.6 Hz, Ar), 7.30-7.16 (m, 3H, Ar), 6.72 (s, 1H,

CHPh), 4.21 (dd, 1H, J=3.6/7.2 Hz, CHC(O)N), 4.07-3.99 (m,

3H, CH₂C(O)N þ CH_aH_b), 3.48-3.43 (m, 1H, CH_aH_b), 2.99-

2.96 (m, 2H, NCH₂), 1.18 (t, 3H, J=7.2 Hz, CH₃). ¹³C NMR: δ

165.38, 136.35, 136.34, 135.57, 135.02, 133.66, 131.82, 130.19,

129.10, 127.13, 122.99, 121.30, 120.23, 118.40, 111.36, 109.09,

52.98 (C1), 49.37 (C3), 48.80, 33.80, 27.03. MS: m/z 430 (M^þþ

4), m/z 428 (M^þþ 2, 100%), m/z 426 (M^þ). Elemental analysis:

calculated for (C₂₂H₁₉Cl₂N₃O₂) C, H, N.

(6S,12aS) 6-(2,4-Dichlorophenyl)-2-methyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (43). Yield

33%; white powder; mp 223-225 °C; R_f=0.22 (CH₂Cl₂/MeOH

98:2). IR (cm^-1): 3314 (-NH-), 1702 (-CO-), 1664 (-CO-).

¹H NMR: δ 8.12 (s, 1H, NH), 7.55 (d, 2H, J=7 Hz, Ar), 7.40 (d,

2H, J=7 Hz, Ar), 7.28-7.10 (m, 3H, Ar), 6.34 (s, 1H, CHPh),

4.09-4.02 (m, 1H, CHC(O)N), 4.12-3.94 (m, 2H, CH₂C(O)N),

3.81 (dd, 1H, J = 3.4/7.2 Hz, CH_aH_b), 3.25-3.22 (m, 1H,

CH_aH_b), 3.09 (s,3H, NCH₃). ¹³C NMR δ 166.13, 165.99,

138.84, 136.30, 133.61, 131.27, 129.43, 128.25, 127.24, 125.84,

122.87, 120.23, 118.59, 111.33, 111.26, 106.62, 56.23 (C1), 53.63,

53.45 (C3), 51.89, 33.76. MS: m/z 417 (M^þþ 4), m/z 415 (M^þþ

2), m/z 413 (M^þ),

(6R,12aR) 6-(3,4-Dichlorophenyl)-2-ethyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (49). Yield

35%; white powder; mp 298-300 °C; R_f=0.33 (CH₂Cl₂/MeOH

(6R,12aS) 6-(2,4-Dichlorophenyl)-2-methyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (44). Yield

Article

Journal of Medicinal Chemistry, 2011, Vol. 54, No. 2 507

98:2). IR (cm^-1):3317 (-NH-), 1673 (-CO-), 1662 (-CO-).

¹H NMR: δ 7.93 (s, 1H, NH), 7.57-7.54 (m, 2H, Ar), 7.42-7.38

(m, 2H, Ar), 7.27-7.16 (m, 3H, Ar), 7.01 (s, 1H, CHPh), 4.26

(dd, 1H, J=3.6/7.2 Hz, CHC(O)N), 4.12-3.98 (m, 3H, CH₂C-

(O)N þ CH_aH_b), 3.60 (dd, 1H, J=3.6/7.2 Hz, CH_aH_b), 3.01-

2.96 (m, 2H, NCH₂), 1.20 (t, 3H, J=7.2 Hz, CH₃). MS:m/z 430

(M^þþ 4), m/z 428 (M^þþ 2, 100%), m/z 426 (M^þ). Elemental

analysis: calculated (C₂₂H₁₉Cl₂N₃O₂) C, H, N.

(6S,12aR) 6-(3,4-Dichlorophenyl)-2-ethyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (50). Yield

28%; yellow powder; mp 92-95 °C; R_f=0.33 (CH₂Cl₂/MeOH

98:2). IR (cm^-1): 3330 (-NH-), 1659 (-CO-), 1663 (-CO-).

¹H NMR: δ 8.52 (s, 1H, NH), 7.57-7.55 (m, 2H, Ar), 7.43-7.40

(m, 2H, Ar), 7.28-7.12 (m, 3H, Ar), 6.72 (s, 1H, CHPh), 3.97-

3.93 (m, 3H, CH₂C(O)N þ CH_aH_b), 3.25 (dd, 1H, J=3.6/7.2 Hz,

CHC(O)N), 3.17 (dd, 1H, J=3.6/7.2 Hz, CH_aH_b), 3.01-2.96

(m, 2H, NCH₂), 1.24 (t, 3H, J=7.2 Hz, CH₃). MS: m/z 430

(M^þþ 4), m/z 428 (M^þþ 2, 100%), m/z 426 (M^þ). Elemental

analysis: calculated for (C₂₂H₁₉Cl₂N₃O₂) C, H, N.

130.24, 129.14, 127.16, 126.17, 123.05, 120.28, 118.42, 111.34,

109.23, 56.44 (C1), 53.07 (C3), 49.36, 45.74, 28.58, 27.07, 19.93,

13.79. MS: m/z 459 (M^þþ 4), m/z 457 (M^þþ 2), m/z 455 (M^þ),

m/z 57 (100%).Elemental analysis: calculated for (C₂₄H₂₃Cl₂-

N₃O₂) C, H, N.

(6S,12aR) 6-(2,4-Dichlorophenyl)-2-butyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (56). Yield

38%; white powder; mp 203-206 °C; R_f=0.52 (CH₂Cl₂/MeOH

98:2). IR (cm^-1): 3326 (-NH-), 1668, 1651 (-CO-). ¹H NMR:

δ 8.28 (s, 1H, NH), 7.52-7.07 (m, 7H, Ar), 6.73 (s, 1H, CHPh),

4.22-4.20 (dd, 1H, CHC(O)N), 4.11-4.03 (m, 2H, CH₂C-

(O)N), 3.45-3.43 (dd, 1H, CHaH_b), 3.28-3.24 (m, 1H, CH_aH_b),

2.99-2.96 (t, 2H, NCH₂), 1.59-1.51 (m, 2H, CH₂CH₂CH₃),

1.37-1.32 (m, 2H, CH₂CH₃), 0.97-0.94 (t, 3H, CH₃). ¹³C

NMR: δ 165.57, 163.37, 136.31, 135.57, 135.01, 133.64, 131.81,

130.20, 129.11, 127.13, 126.14, 123.00, 120.24, 118.39, 111.35,

109.16, 53.03 (C1), 49.37 (C3), 49.33, 45.72, 28.55, 27.05, 19.90,

13.77. MS: m/z 459 (M^þþ 4), m/z 457 (M^þþ 2), m/z 455 (M^þ),

m/z 57.1 (100%). Elemental analysis: calculated for (C₂₄H₂₃Cl₂-

N₃O₂) C, H, N.

(6S,12aS) 6-(2,4-Dichlorophenyl)-2-butyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (57). Yield

34%; white powder; mp 140-143 °C; R_f=0.51 (CH₂Cl₂/MeOH

98:2). IR (cm^-1): 3366 (-NH-), 1654 (-CO-), 1633 (-CO-).

¹H NMR: δ 8.07 (s, 1H, NH), 7.57-7.55 (m, 2H, Ar), 7.32-7.13

(m, 5H, Ar), 6.87 (s, 1H, CHPh), 4.30-4.25 (d, 1H, CHC(O)N),

3.55-3.53 (m, 2H, CH₂C(O)N), 3.32-3.29 (dd, 1H, CHaH_b),

2.91-2.87 (m, 1H, CH_aH_b), 2.17-2.16 (t, 2H, NCH₂), 1.50-

1.46 (m, 2H, CH₂CH₂CH₃), 1.36-1.30 (m, 2H, CH₂CH₃),

0.93-0.89 (t, 3H, CH₃). ¹³C NMR: δ 171.22, 169.44, 136.34,

135.77, 132.78, 130.58, 130.50, 128.06, 126.82, 122.61, 121.70,

119.89, 118.89, 111.11, 110.60, 57.44 (C1), 54.44 (C3), 52.29,

39.02, 31.55, 30.95, 24.61, 20.10, 13.77. MS: m/z 458 (M^þþ 4),

m/z 456 (M^þþ 2, 100%), m/z 454 (M^þ). Elemental analysis:

calculated for (C₂₄H₂₃Cl₂N₃O₂) C, H, N.

(6R,12aS) 6-(2,4-Dichlorophenyl)-2-butyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (58). Yield

35%; yellow powder; mp 207-210 °C; R_f=0.51 (CH₂Cl₂/MeOH

98:2). IR (cm^-1): 3399 (-NH-), 1699 (-CO-), 1653 (-CO-).

¹H NMR: δ 8.44-8.41 (d, 1H, NH), 7.52-7.50 (d, 2H, Ar),

7.42-7.45 (d, 1H, Ar), 7.35-7.12 (m, 3H, Ar), 7.06-7.01 (d, 1H,

Ar), 6.70 (d, 1H, CHPh), 5.31 (s, 1H, CHC(O)N), 4.21-4.19

(dd, 1H, CHaH_b), 4.08-3.98 (m, 2H, CH₂C(O)N), 2.99-2.95

(m, 1H, CH_aH_b), 2.17-2.16 (t, 2H, NCH₂), 1.57-1.54 (m, 2H,

CH₂CH₂CH₃), 1.36-1.32 (m, 2H, CH₂CH₃), 0.97-0.93 (t, 3H,

CH₃). ¹³C NMR: δ 171.21, 165.59, 136.35, 135.00, 133.66,

131.84, 130.18, 129.13, 127.13, 126.15, 122.96, 120.20, 118.39,

111.37, 109.09, 53.45, 53.02 (C1), 49.39 (C3), 49.34, 45.73, 30.95,

29.70, 19.93, 13.79. MS: m/z 458 (M^þþ 4), m/z 456 (M^þþ 2,

100%), m/z 454 (M^þ). Elemental analysis: calculated for

(C₂₄H₂₃Cl₂N₃O₂) C, H, N.

(6R,12aR) 6-(2,4-Dichlorophenyl)-2-tert-butyl-2,3,6,7,12,12a-

hexahydropyrazino [1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (59).

Yield 32%; white powder; mp 140-142 °C; R_f=0.65 (CH₂Cl₂/

MeOH 98:2). IR (cm^-1): 3309 (-NH-), 1724, 1666(-CO-). ¹H

NMR: δ 8.15 (s, 1H, NH), 7.54-7.12 (m, 7H, Ar), 6.62 (s, 1H,

CHPh), 4.29-4.25 (dd, 1H, CHC(O)N), 4.21-3.96 (m, 3H,

CH₂C(O)N þ CH_aH_b), 3.71-3.69 (dd, 1H, CH_aH_b), 1.52

(s,9H, C(CH₃)₃). ¹³C NMR: δ 167.55, 167.12, 164.12, 138.84,

136.00, 133.66, 129.43, 128.28, 127.35, 127.15, 122.79, 120.24,

118.53, 111.32, 109.39, 106.67, 58.15 (C1), 57.12, 53.90 (C3),

53.25, 47.19, 27.96, 26.82, 24.11. MS: m/z 459 (M^þþ 4), m/z 457

(M^þþ 2), m/z 455 (M^þ), m/z 57 (100%). Elemental analysis:

calculated for (C₂₄H₂₃Cl₂N₃O₂) C, H, N.

(6S,12aS) 6-(3,4-Dichlorophenyl)-2-ethyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (51). Yield

28%; yellow powder; mp 298-301 °C; R_f= 0.34 (CH₂Cl₂/

MeOH 98.2). IR (cm^-1):3325 (-NH-), 1712 (-CO-), 1663

1

(-CO-). H NMR: δ 8.19 (brs, 1H, NH), 7.54-7.53 (m, 2H,

Ar), 7.40-7.32 (m, 2H, Ar), 7.26-7.19 (m, 3H, Ar), 6.70 (s, 1H,

CHPh), 4.24 (dd, 1H, J=3.6/7.6 Hz, CHC(O)N), 4.16-3.98 (m,

3H, CH₂C(O)N þ CH_aH_b), 3.55 (dd, 1H, J=3.6/7.6 Hz, CH_aH_b),

2.95-2.94 (m, 2H, NCH₂), 1.19 (t, 3H, J=7.2 Hz, CH₃). MS:m/z

430 (M^þþ 4), m/z 428 (M^þþ 2, 100%), m/z 426 (M^þ).

Elemental analysis: calculated for (C₂₂H₁₉Cl₂N₃O₂) C, H, N.

(6R,12aS) 6-(3,4-Dichlorophenyl)-2-ethyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (52). Yield

34%; white powder; mp 94-96 °C; R_f= 0.35 (CH₂Cl₂/MeOH

98:2). IR (cm^-1): 3337 (-NH-), 1710 (-CO-), 1665 (-CO-).

¹H NMR: δ 7.93 (s, 1H, NH), 7.57-7.54 (m, 2H, Ar), 7.42-7.38

(m, 2H, Ar), 7.28-7.19 (m, 3H, Ar), 7.01 (s, 1H, CHPh), 4.26

(dd, 1H, J=3.6/7.6 Hz, CHC(O)N), 4.12-3.98 (m, 3H, CH₂C-

(O)N þ CH_aH_b), 3.55 (dd, 1H, J=3.6/7.6 Hz, CH_aH_b), 3.01-

2.96 (m, 2H, NCH₂), 1.22 (t, 3H, J=7.2 Hz, CH₃). MS: m/z 430

(M^þþ 4), m/z 428 (M^þþ 2, 100%), m/z 426 (M^þ). Elemental

analysis: calculated for (C₂₂H₁₉Cl₂N₃O₂) C, H, N.

(6R,12aR) 6-(2,6-Dichlorophenyl)-2-ethyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (53). Yield

30%; yellowish white powder; mp 110-112 °C; R_f=0.36 (CH₂Cl₂/

MeOH 98:2). IR (cm^-1): 3446 (-NH-), 1670, 1630(-CO-). ¹H

NMR: δ 9.00 (s, 1H, NH), 7.57-7.37 (m,7H, Ar), 6.97 (s, 1H,

CHPh), 4.22 (dd, 1H, J=3.6/7.6 Hz, CHC(O)N), 3.78-3.65 (m,

3H, CH₂C(O)N þ CH_aH_b), 3.59-3.55 (m, 1H, CH_aH_b), 3.25-

3.23 (m, 2H, NCH₂), 1.25 (t, 3H, J=7.2 Hz, CH₃). MS: m/z 431

(M^þþ 4), m/z 429 (M^þþ 2), m/z 427 (M^þ), m/z 57 (100%).

Elemental analysis: calculated for (C₂₂H₁₉Cl₂N₃O₂) C, H, N.

(6S,12aS) 6-(2,6-Dichlorophenyl)-2-ethyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (54). Yield

28%; yellowish white powder; mp 112-115 °C; R_f=0.34 (CH₂Cl₂/

MeOH 98:2). ¹H NMR: δ 8.99 (s, 1H, NH), 7.52-7.12 (m,7H,

Ar), 6.20 (s, 1H, CHPh), 4.18-4.15 (m, 1H, CHC(O)N), 4.07-

4.02 (m, 3H, CH₂C(O)N þ CH_aH_b), 3.83-3.73 (m, 1H, CH_aH_b),

3.28-3.24 (m, 2H, NCH₂), 1.32 (t, 3H, J=7.2 Hz, CH₃). MS: m/

z 431 (M^þþ 4), m/z 429 (M^þþ 2), m/z 427 (M^þ), m/z 57 (100%).

Elemental analysis: calculated for (C₂₂H₁₉Cl₂N₃O₂) C, H, N.

(6R,12aR) 6-(2,4-Dichlorophenyl)-2-butyl-2,3,6,7,12,12a-hexa-

hydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (55). Yield

37%; white powder; mp 140-142 °C; R_f=0.52 (CH₂Cl₂/MeOH

98:2). IR (cm^-1): 3285 (-NH-), 1662, 1654 (-CO-). ¹H NMR:

δ 8.15 (s, 1H, NH), 7.54-7.08 (m, 7H, Ar), 6.73 (s, 1H, CHPh),

4.23-4.21 (dd, 1H, CHC(O)N), 4.12-4.04 (m, 2H, CH₂C-

(O)N), 3.48-3.46 (dd, 1H, CHaH_b), 3.29-3.25 (m, 1H, CH_aH_b),

2.99-2.96 (t, 2H, NCH₂), 1.59-1.55 (m, 2H, CH₂CH₂CH₃),

1.38-1.34 (m, 2H, CH₂CH₃), 0.97-0.94 (t, 3H, CH₃). ¹³C

NMR: δ 165.58, 163.39, 136.32, 135.87, 135.04, 133.66, 131.82,

(6S,12aR) 6-(2,4-Dichlorophenyl)-2-tert-butyl-2,3,6,7,12,12a-

hexahydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (60).

Yield 30%; white powder; mp 200-202 °C; R_f=0.64 (CH₂Cl₂/

MeOH 98:2). IR (cm^-1): 3399 (-NH-), 1741, 1663 (-CO-).

¹H NMR: δ 8.27(s, 1H, NH), 7.52-7.06 (m, 7H, Ar), 6.70 (s, 1H,

CHPh), 4.24-4.20 (m, 1H, CHC(O)N), 4.15-3.97 (m, 3H,

508 Journal of Medicinal Chemistry, 2011, Vol. 54, No. 2

Mohamed et al.

CH₂C(O)N þ CH_aH_b), 3.44-3.40 (dd, 1H, CH_aH_b), 1.51 (s,9H,

C(CH₃)₃). ¹³C NMR: δ 166.73, 164.12, 136.36, 135.54, 133.65,

131.81, 130.19, 129.11, 127.14, 126.22, 122.95, 120.20, 118.40,

111.35, 109.30, 55.05(C1), 53.89, 53.45 (C3), 52.84, 49.25, 46.63,

27.96, 27.68, 26.83. MS: m/z 459 (M^þþ 4), m/z 457 (M^þþ 2), m/

z 455 (M^þ), m/z 57 (100%). Elemental analysis: calculated for

(C₂₄H₂₃Cl₂N₃O₂) C, H, N.

(6S,12aS) 6-(2,4-Dichlorophenyl)-2-tert-butyl-2,3,6,7,12,12a-

hexahydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (61).

Yield 32%; yellow powder; mp 145-148 °C; R_f=0.62 (CH₂Cl₂/

MeOH 98:2). IR (cm^-1): 3320 (-NH-), 1710 (-CO-), 1665

(-CO-). ¹H NMR: δ 8.95 (s, 1H, NH), 7.53-7.51 (m, 1H, Ar),

7.46-7.42 (d, 2H, Ar), 7.27-7.12 (m, 4H, Ar), 6.62 (s, 1H,

CHPh), 4.31-4.25 (dd, 1H,CHC(O)N), 3.99-3.95 (dd, 1H,

CHaH_b), 3.46-3.42 (m, 2H, CH₂C(O)N), 2.96-2.92 (m, 1H,

CH_aH_b), 1.48 (s, 9H, C(CH₃)₃). ¹³C NMR: δ 167.55, 167.12,

166.72, 164.12, 131.81, 130.21, 129.42, 128.27, 127.14, 120.23,

118.52, 118.40, 111.25, 109.36, 106.67, 58.15 (C1), 57.12, 53.89

(C3), 53.26, 47.19, 46.63, 27.96, 24.10. MS: m/z 458 (M^þþ 4), m/

z 456 (M^þþ 2, 100%), m/z 454 (M^þ). Elemental analysis:

calculated for (C₂₄H₂₃Cl₂N₃O₂) C, H, N.

Yield 36%; yellow powder; mp 95-97 °C; R_f=0.61 (CH₂Cl₂/

MeOH 98:2). IR (cm^-1): 3300 (-NH-), 1695 (-CO-), 1662

(-CO-). ¹H NMR: δ 8.89 (s, 1H, NH), 7.70-7.65 (d, 1H, Ar),

7.45-7.42 (d, 1H, Ar), 7.29-7.14 (m, 5H, Ar), 6.80 (s, 1H,

CHPh), 3.97-3.93 (dd, 1H, CHC(O)N), 3.73-3.26 (m, 3H,

CH₂C(O)N þ CH_aH_b), 3.26-3.25 (dd, 1H, CH_aH_b), 1.26 (s,

9H, C(CH₃)₃). MS: m/z 459 (M^þþ 4), m/z 457 (M^þþ 2), m/z 455

(M^þ), m/z 375 (100%). Elemental analysis: calculated for

(C₂₄H₂₃Cl₂N₃O₂) C, H, N.

(6R,12aR) 6-(2,6-Dichlorophenyl)-2-tert-butyl-2,3,6,7,12,12a-

hexahydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (67).

Yield 26%; yellowish white powder; mp 99-103 °C; R_f=0.61

(CH₂Cl₂/MeOH 98:2). IR (cm^-1): 3317 (-NH-), 1730, 1685

(-CO-). ¹H NMR: δ 8.99 (s, 1H, NH), 7.58-7.15 (m, 7H, Ar),

6.31 (s, 1H, CHPh), 4.32-4.31 (m, 1H, CHC(O)N), 4.17-4.07

(m, 3H, CH₂C(O)N þ CH_aH_b), 3.92-3.85 (dd, 1H, CH_aH_b),

1.27 (s,9H, C(CH₃)₃). MS: m/z 459 (M^þþ 4), m/z 457 (M^þþ 2),

m/z 455 (M^þ), m/z 57 (100%). Elemental analysis: calculated for

(C₂₄H₂₃Cl₂N₃O₂) C, H, N.

(6S,12aS) 6-(2,6-Dichlorophenyl)-2-tert-butyl-2,3,6,7,12,12a-

hexahydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (68).

Yield 23%; yellowish-white powder; mp 90-92 °C; R_f=0.63

(CH₂Cl₂/MeOH 98:2). IR (cm^-1): 3320 (-NH-), 1731, 1690

(-CO-). ¹H NMR: δ 8.99 (s, 1H, NH), 7.60-7.13 (m, 7H, Ar),

6.24 (s, 1H, CHPh), 4.11-4.09 (m, 1H, CHC(O)N), 4.07-3.80

(m, 3H, CH₂C(O)N þ CH_aH_b), 3.39-3.32 (dd, 1H, CH_aH_b),

1.26 (s,9H, C(CH₃)₃). MS: m/z 459 (M^þþ 4), m/z 457 (M^þþ 2),

m/z 455 (M^þ), m/z 57 (100%). Elemental analysis: calculated for

(C₂₄H₂₃Cl₂N₃O₂) C, H, N.

Biology. Phosphodiesterase Assay. Human recombinant PDE

5A (12.5 units/mL), 10 units/mL PDE3B, 400 units/mL PDE4B,

or 50 units/mL PDE11A (BPS Biosciences) were added to the

wells of black 96-well nonbinding plates. Immediately, the pro-

tein was treated with compound or vehicle control. Enzyme was

incubated with drug for 30 min prior to addition of 50 nM

TAMRA-cGMP and/or 50 nM fluorescein-cAMP (Molecular

Devices) wereaddedtoeach assay well. The plates wereincubated

for 1.5h at 30 °C. After incubation, immobilizedmetal affinity for

phosphochemicals (IMAP)-based fluorescence polarization (FP)

phosphodiesterase evaluation assay (Molecular Devices) binding

reagent was added to each well, and the plates were incubated for

an additional 30 min at 30 °C. FP was measured according to

manufacturer’s specifications usingaBiotekSynergy4platereader.

Cell Cultures. MDA-MB-231 breast tumor cells were obtained

from ATCC. They were grown under standard cell culture condi-

tions at 37 °C in a humidified atmosphere with 5% CO₂. Cells were

grown in RPMI 1640 supplemented with 5% fetal bovine serum,

2 mM glutamine, 100 units/mL penicillin, 100 units/mL strepto-

mycin, and 0.25 μg/mL amphotericin. Cells were harvested at 70-

90% confluence with trypsin/EDTA and used immediately. Cell

count and viability was determined by Trypan blue staining

followed by hemocytometry. Only cultures displaying >95% cell

viability were used for experiments.

(6R,12aS) 6-(2,4-Dichlorophenyl)-2-tert-butyl-2,3,6,7,12,12a

exahydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (62).

Yield 25%; yellow powder; mp 198-200 °C; R_f=0.61 (CH₂Cl₂/

MeOH 99.5:0.5). IR (cm^-1): 3387 (-NH-), 1741 (-CO-), 1663

1

(-CO-). H NMR: δ 8.11-8.07 (d, 1H, NH), 7.51-7.49 (m,

2H, Ar), 7.27-7.09 (m, 5H, Ar), 6.58 (s, 1H, CHPh), 5.29-5.28

(dd, 1H, CHC(O)N), 3.66-3.61 (dd, 1H, CHaH_b), 3.45-3.42

(m, 2H, CH₂C(O)N), 2.96-2.91 (dd, 1H, CH_aH_b), 1.07 (s, 9H,

C(CH₃)₃). ¹³C NMR: δ 173.39, 171.76, 167.34, 131.56, 129.61,

129.42, 128.93, 128.88, 128.01, 127.04, 125.92, 120.05, 119.91,

118.32, 111.24, 105.97, 104.65, 55.95 (C1), 53.44 (C3), 50.51,

46.00, 28.63, 24.05. MS: m/z 458 (M^þþ 4), m/z 456 (M^þþ 2,

100%), m/z 454 (M^þ). Elemental analysis: calculated for

(C₂₄H₂₃Cl₂N₃O₂) C, H, N.

(6R,12aR) 6-(3,4-Dichlorophenyl)-2-tert-butyl-2,3,6,7,12,12a-

hexahydropyrazino[1⁰,2⁰:1,6]pyrido [3,4-b]indole-1,4-dione (63).

Yield 36%; white powder; mp 299-303 °C; R_f=0.62 (CH₂Cl₂/

MeOH 98:2). IR (cm^-1):3387 (-NH-), 1695 (-CO-), 1652

(-CO-). ¹H NMR: δ 7.96 (s, 1H, NH), 7.55-7.53 (d, 2H, Ar),

7.43-7.31 (d, 2H, Ar), 7.28-7.00 (m, 3H, Ar), 6.71 (s, 1H,

CHPh), 4.35-4.28 (dd, 1H, CHC(O)N), 4.66-4.60 (dd, 1H,

CH_aH_b), 4.14-4.00 (m, 3H, CH₂C(O)N þ CH_aH_b), 1.27 (s, 9H,

C(CH₃)₃). MS: m/z 459 (M^þþ 4), m/z 457 (M^þþ 2), m/z 455

(M^þ, 100%). Elemental analysis: calculated for (C₂₄H₂₃Cl₂-

N₃O₂) C, H, N.

(6S,12aR) 6-(3,4-Dichlorophenyl)-2-tert-butyl-2,3,6,7,12,12a-

hexahydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (64).

Yield 35%; yellow powder; mp 93-96 °C; R_f=0.62 (CH₂Cl₂/

MeOH 98:2). IR (cm^-1):3377 (-NH-), 1662 (-CO-), 1653

(-CO-). ¹H NMR: δ 8.01 (s, 1H, NH), 7.57-7.55 (d, 2H, Ar),

7.43-7.41 (d, 2H, Ar), 7.29-7.14 (m, 3H, Ar), 6.78 (s, 1H,

CHPh), 3.97-3.93 (dd, 1H, CHC(O)N), 3.73-3.26 (m, 3H,

CH₂C(O)N þ CH_aH_b), 3.26-3.25 (dd, 1H, CH_aH_b), 1.27 (s,

9H, C(CH₃)₃). MS: m/z 459 (M^þþ 4), m/z 457 (M^þþ 2), m/z 455

(M^þ, 100%). Elemental analysis: calculated for (C₂₄H₂₃Cl₂-

N₃O₂) C, H, N.

(6S,12aS) 6-(3,4-Dichlorophenyl)-2-tert-butyl-2,3,6,7,12,12a-

hexahydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (65).

Yield 38%; yellow powder; mp 298-301 °C; R_f=0.62 (CH₂Cl₂/

MeOH 98:2). IR (cm^-1): 3322 (-NH-), 1715, 1663 (-CO-).

¹H NMR: δ 7.87 (s, 1H, NH), 7.55-7.54 (d, 2H, Ar), 7.40-7.33

(d, 2H, Ar), 7.27-7.01 (m, 3H, Ar), 6.61 (s, 1H, CHPh), 4.30-

4.25 (dd, 1H, CHC(O)N), 4.14-4.00 (m, 3H, CH₂C(O)N þ

CH_aH_b), 4.60-4.57 (dd, 1H, CH_aH_b), 1.29 (s, 9H, C(CH₃)₃).

MS: m/z 459 (M^þþ 4), m/z 457 (M^þþ 2), m/z 455 (M^þ), m/z

375 (100%). Elemental analysis: calculated for (C₂₄H₂₃Cl₂-

N₃O₂) C, H, N.

Growth Assays. Tissue culture treated microtiter 96-well plates

were seeded at a density of 5000 cells/well. The plates were

incubated for 18-24 h prior to any treatment. All test compounds

were solubilized in 100% DMSO and diluted with media to obtain

a final DMSO concentration of 0.1%. Cells were dosed, incubated

at 37 °C, and cell viability measured 72 h after treatment by the Cell

Titer Glo Assay (Promega), which is a luminescent assay that is an

indicator of live cells as a function of metabolic activity and ATP

content. The assay was performed according to manufacturer’s

specifications. Luminescence was measured by a Perkin-Elmer

Victor multilabel plate reader.

Experimental Design and Data Analysis. Drug effects on tumor

cell growth and PDE activity were measured and potency was

expressed by an IC₅₀value (50% inhibitory concentration). For

growth assays, the IC₅₀value was determined by testing a range of

eight concentrations with quadruple replicates per concentration.

For enzyme assays, the IC₅₀value was determined by testing a

(6R,12aS) 6-(3,4-Dichlorophenyl)-2-tert-butyl-2,3,6,7,12,12a-

hexahydropyrazino[1⁰,2⁰:1,6]pyrido[3,4-b]indole-1,4-dione (66).

Article

Journal of Medicinal Chemistry, 2011, Vol. 54, No. 2 509

range of 10 concentrations with quadruple replicates per concen-

tration. Dose response curves were analyzed using PrismTM 4

software (GraphPad) to calculate IC₅₀values using a four para-

meter logistic equation.

inhibits growth and induces apoptosis of human breast tumor cells

by phosphodiesterase 5 inhibition, elevation of cyclic GMP and

activation of protein kinase G. Mol. Cancer Ther. 2009, 8, 3331–

3340.

(7) Abadi, A. H.; Abouel-Ella, D. A.; Ahmed, N. S.; Gary, B. D.;

Thaiparambil, J. T.; Tinsley, H. N.; Keeton, A. B.; Piazza, G. A.

Synthesis of novel tadalafil analogues and their evaluation as

phosphodiesterase inhibitors and anticancer agents. Arzneim.

Forsch. 2009, 59, 415–421.

(8) Jenkins, P. R.; Wilson, J.; Emmerson, D.; Garcia, M. D.; Smith,

M. R.; Gray, S. J.; Britton, R. G.; Mahale, S.; Chaudhuri, B.

Design, synthesis and biological evaluation of new tryptamine and

tetrahydro-beta-carboline-based selective inhibitors of CDK4.

Bioorg. Med. Chem. 2008, 16, 7728–7739.

(9) Leteurtre, F.; Madalengoitia, J.; Orr, A.; Guzi, T. J.; Lehnert, E.;

Macdonald, T.; Pommier, Y. Rational design and molecular effects

of a new topoisomerase II inhibitor, azatoxin. Cancer Res. 1992, 52,

4478–4483.

(10) Sunder-Plassmann, N.; Sarli, V.; Gartner, M.; Utz, M.; Seiler, J.;

Huemmer, S.; Mayer, T. U.; Surrey, T.; Giannis, A. Synthesis and

biological evaluation of new tetrahydro-beta-carbolines as inhibi-

tors of the mitotic kinesin Eg5. Bioorg. Med. Chem. 2005, 13, 6094–

6111.

(11) Doggrell, S. A. Comparison of clinical trials with sildenafil,

vardenafil and tadalafil in erectile dysfunction. Expert Opin.

Pharmacother. 2005, 6, 75–84.

(12) Duan, H.; Zheng, J.; Lai, Q.; Liu, Z.; Tian, G.; Wang, Z.; Li, J.;

Shen, J. 2-Phenylquinazolin-4(3H)-one, a class of potent PDE5

inhibitors with high selectivity versus PDE6. Bioorg. Med. Chem.

Lett. 2009, 19, 2777–2779.

(13) Al-Shaiji, T. F.; Brock, G. B. Phosphodiesterase type 5 inhibitors

for the management of erectile dysfunction: preference and adher-

ence to treatment. Curr. Pharm. Des. 2009, 15, 3486–3495.

(14) Sung, B. J.; Hwang, K. Y.; Jeon, Y. H.; Lee, J. I.; Heo, Y. S.; Kim,

J. H.; Moon, J.; Yoon, J. M.; Hyun, Y. L.; Kim, E.; Eum, S. J.;

Park, S. Y.; Lee, J. O.; Lee, T. G.; Ro, S.; Cho, J. M. Structure of the

catalytic domain of human phosphodiesterase 5 with bound drug

molecules. Nature 2003, 425, 98–102.

Molecular Modeling. The crystal structure of tadalafil in

complex with PDE5 (PDB 1UDU) was prepared for docking

using Molecular Operating Environment software (MOE)

2009.10 (CCG). Addition of hydrogens and assignment

of correct protonation states of all residues at physiological

pH = 7.4 was performed using the Protonate3D module in

MOE. The compound structures were drawn using ChemSketch.

We employed a Pipeline Pilot (SciTegic Inc.) workflow to

systematically name and ionize the compounds (pH=7.4). In

the same workflow, we set a chiral flag for each compound

using the ChemAxon Standardizer component to keep the

molecules in their correct stereoconfigurations during later

steps. We then employed the wash and minimization procedures in

MOE to prepare the final compound library for docking.

The docking procedure was performed with GOLD v3.2

(CCDC). For all docking runs, we used default parameters.

The genetic algorithm was set to automatic mode at 200%

search effiency. The binding site was defined by specifying the

coordinates of the cocrystallized ligand tadalafil. The active site

˚

radius setting remained at the default value of 10 A with cavity

detection enabled.

Three independent docking experiments were done for all

compounds, employing the three implemented scoring functions

in GOLD (GoldScore, ChemScore, and ASP) to drive the

genetic algorithm (GA). In each docking experiment, a total

of 10 GA runs per compound were conducted, resulting in 10

binding poses for each compound per experiment. Thus, we

obtained a total of 30 poses per compound.

All poses generated with one scoring function were then

rescored in GOLD utilizing the remaining two scoring functions.

Further rescoring of all poses was performed with DrugScore^CSD

and DrugScore^PDBusing the DrugScore Online service v0.9

(http://pc1664.pharmazie.uni - marburg.de/drugscore/). Finally,

a consensus score was calculated amounting to the arithmetic

mean of the normalized Goldscore, Chemscore, ASP, DrugScor-

e^CSD, and DrugScore^PDBvalues. The depicted binding modes

represent the docking pose with the optimal consensus score.

(15) Ke, H.; Wang, H. Crystal structures of phosphodiesterases and

implications on substrate specificity and inhibitor selectivity. Curr.

Top. Med. Chem. 2007, 7, 391–403.

(16) Zoraghi, R.; Francis, S. H.; Corbin, J. D. Critical amino acids in

phosphodiesterase-5 catalytic site that provide for high-affinity

interaction with cyclic guanosine monophosphate and inhibitors.

Biochemistry 2007, 46, 13554–13563.

(17) Whaley, W. M.; Govindachari, T. R. The Pictet-Spengler synth-

esis of tetrahydroisoquinolines and related compounds. Org. Re-

act. 1951, 6, 151.

(18) Ungemach, F.; DiPierro, M.; Weber, R.; Cook, J. M. Stereospecific

Synthesis of trans-1,3-Disubstituted-l,2,3,4-tetrahydro-β-carbo-

lines. J. Org. Chem. 1981, 46, 164–168.

Supporting Information Available: Results from elemental

analyses for compounds 1-68 and the NOESY spectra for

compounds 1 and 2. This material is available free of charge

via the Internet at http://pubs.acs.org.

(19) Cox, E. D.; Cook, J. M. The Pictet-Spengler condensation: a new

direction for an old reaction. Chem. Rev. 1995, 95, 1797–1842.

(20) Czerwinski, K.; Deng, L.; Cook, J. Mechanism driven trans

stereospecificity in the Pictet-Spengler reaction. Stereospecific

fomation of trans-1,2,3-trisubstituted-tetrahydro-β-carbolines by

condensation of N_b-diphenymethyl tryptophan isopropyl esters

with aldehydes. Tetrahedron Lett. 1992, 48, 4721–4724.

(21) Pulka, K.; Kulis, P.; Tymecka, D.; Frankiewicz, L.; Wilczek, M.;

Kozminski, M.; Misicka, A. Diastereoselective Pictete-Spengler

condensation of tryptophan with R-amino aldehydes as chiral

carbonyl components. Tetrahedron Lett. 2008, 64, 1506–1514.

(22) Sandrin, J.; Soerens, D.; Cook, J. M. Carbon-13 NMR of 1,3-

Disubstituted 1,2,3,4-Tetrahydro-β-Carbolines. Heterocycles

1976, 4, 1249–1255.

(23) Ungemach, F.; Soerens, D.; Weber, R.; DiPierro, M.; Campos, O.;

Mokry, P.; Cook, J. M.; Silverton, J. V. General Method for the

Assignment of Stereochemistry of 1,3-Disubstituted 1,2,3,4-Tetra-

hydro-β-Carbolines by Carbon-13 Spectroscopy. J. Am. Chem.

Soc. 1980, 102, 6976–6984.

(24) Zoraghi, R.; Corbin, J. D.; Francis, S. H. Phosphodiesterase-5

Gln817 is critical for cGMP, vardenafil, or sildenafil affinity: its

orientation impacts cGMP but not cAMP affinity. J. Biol. Chem.

2006, 281, 5553–5558.

References

(1) Conti, M.; Beavo, J. Biochemistry and physiology of cyclic nucleo-

tide phosphodiesterases: essential components in cyclic nucleotide

signaling. Annu. Rev. Biochem. 2007, 76, 481–511.

(2) Omori, K.; Kotera, J. Overview of PDEs and their regulation. Circ.

Res. 2007, 16, 309–327.

(3) Ghofrani, H. A.; Osterloh, I. H.; Grimminger, F. Sildenafil: from

angina to erectile dysfunction to pulmonary hypertension and

beyond. Nature Rev. Drug Discovery 2006, 5, 689–702.

(4) Masson, P.; Lambert, S. M.; Brown, M.; Shabsigh, R. PDE-5

inhibitors: current status and future trends. Urol. Clin. North Am.

2005, 32, 511–525.

(5) Watanabe, H.; Ohashi, K.; Takeuchi, K.; Yamashita, K.; Yokoyama,

T.; Tran, Q. K.; Satoh, H.; Terada, H.; Ohashi, H.; Hayashi, H.

Sildenafil for primary and secondary pulmonary hypertension. Clin.

Pharmacol. Ther. 2002, 71, 398–402.

(6) Tinsley, H. N.; Gary, B. D.; Keeton, A. B.; Zhang, W.; Abadi,

A. H.; Reynolds, R. C.; Piazza, G. A. Sulindac sulfide selectively

Article Doi

Synthesis and molecular modeling of novel tetrahydro-β-carboline derivatives with phosphodiesterase 5 inhibitory and anticancer properties

DOI: 10.1021/jm100842v

Source and publish data:

Authors:

Article abstract of DOI:10.1021/jm100842v

Full text of DOI:10.1021/jm100842v

Products guided by the article

R&D Labs maybe for 1262447-27-7

Relevant to this article

Hot Product