1
Supporting information
Design, synthesis, molecular docking and biological evaluation of N-methyl-3-nitro4-(nitromethyl)-4H-chromen-2-amine derivatives as potential anticancer agents
A. Parthiban1, J. Muthukumaran2, A. Moushumi Priya3, S. Jayachandran3,*, R. Krishna2,*
and H. Surya Prakash Rao1,*
1
Department of Chemistry, School of Physical, Chemical and Applied Sciences,
Pondicherry University, Puducherry – 605 014.
2
Centre for Bioinformatics, School of Life Sciences, Pondicherry University, Puducherry
– 605 014.
3
Department of Biotechnology, School of Life Sciences, Pondicherry University,
Puducherry - 605 014.
*
Corresponding authors
H. Surya Prakash Rao, Department of Chemistry, School of Physical, Chemical and
Applied
Sciences,
Pondicherry
University,
Puducherry
–
605
014,
Email:
hspr.che@pondiuni.edu.in, Tel: +91-413-2654411, : +91-413-2656230.
S. Jayachandran, Department of Biotechnology, School of Life Sciences, Pondicherry
University, Puducherry – 605 014, Email: sr999jayachandran@gmail.com, Tel : +91413-2654428, Fax: +91-413-2655715
R. Krishna, Centre for Bioinformatics, Pondicherry University, Puducherry – 605 014,
Email: krishstrucbio@gmail.com, Tel: +91 413 2654580, Fax: +91 413 2655211
2
Contents:
IR (KBr), 1H NMR (400 MHz, CDCl3),
13
C NMR (100 MHz, CDCl3) and
DEPT-135 NMR (100 MHz, CDCl3) spectrum of N-methyl-3-nitro-4(nitromethyl)-4H-chromen-2-amine 8a.
IR (KBr), 1H NMR (400 MHz, CDCl3),
S5
13
C NMR (100 MHz, CDCl3) and
DEPT-135 NMR (100 MHz, CDCl3) spectrum of 6-chloro-N-methyl-3nitro-4-(nitromethyl)-4H-chromen-2-amine 8b.
IR (KBr), 1H NMR (400 MHz, CDCl3),
13
S7
C NMR (100 MHz, CDCl3) and
DEPT-135 NMR (100 MHz, CDCl3) spectrum of 6-bromo-N-methyl-3nitro-4-(nitromethyl)-4H-chromen-2-amine 8c.
S9
IR (KBr), 1H NMR (400 MHz, DMSO-d6 + CCl4, 1:1), 13C NMR (100 MHz
(DMSO-d6 + CCl4, 1:1) and DEPT-135 NMR (100 MHz (DMSO-d6 + CCl4,
1:1)
spectrum
of
chromen-2-amine 8d.
6,8-dichloro-N-methyl-3-nitro-4-(nitromethyl)-4HS11
IR (KBr), 1H NMR (400 MHz, DMSO-d6 + CCl4, 1:1), 13C NMR (100 MHz
(DMSO-d6 + CCl4, 1:1) and DEPT-135 NMR (100 MHz (DMSO-d6 + CCl4,
1:1) spectrum of 6-nitro-N-methyl-3-nitro-4-(nitroethyl)-4H-chromen-2amine 8e.
1
S13
H NMR (400 MHz, DMSO-d6 + CCl4, 1:1) and 13C NMR (100 MHz (DMSO-d6 + CCl4,
1:1) spectrum of 6-chloro-N,7-dimethyl-3-nitro-4-(nitromethyl)-4H-chromen-2-amine 8f.
S15
3
IR (KBr), 1H NMR (400 MHz, CDCl3),
13
C NMR (100 MHz, CDCl3) and
DEPT-135 NMR (100 MHz, CDCl3) spectrum of 8-methoxy-N-methyl-3nitro-4-(nitromethyl)-4H-chromen-2-amine 8g.
IR (KBr), 1H NMR (400 MHz, CDCl3),
13
S16
C NMR (100 MHz, CDCl3) and
DEPT-135 NMR (100 MHz, CDCl3) spectrum of 6-methoxy-N-methyl-3nitro-4-(nitromethyl)-4H-chromen-2-amine 8h.
IR (KBr), 1H NMR (400 MHz, CDCl3 + CCl4, 1:1),
S18
13
C NMR (100 MHz,
CDCl3 + CCl4, 1:1) and DEPT-135 NMR (100 MHz, CDCl3 + CCl4, 1:1)
spectrum
of
6-ethoxy-N-methyl-3-nitro-4-(nitromethyl)-4H-chromen-2-
amine 8i.
IR (KBr), 1H NMR (400 MHz, CDCl3 + CCl4, 1:1),
S20
13
C NMR (100 MHz,
CDCl3 + CCl4, 1:1) and DEPT-135 NMR (100 MHz, CDCl3 + CCl4, 1:1)
spectrum of 6-methyl-N-methyl-3-nitro-4-(nitroethyl)-4H-chromen-2-amine
8j.
IR (KBr), 1H NMR (400 MHz, CDCl3 + CCl4, 1:1),
S22
13
C NMR (100 MHz,
CDCl3 + CCl4, 1:1) and DEPT-135 NMR (100 MHz, CDCl3 + CCl4, 1:1)
spectrum of 6-ethyl-N-methyl-3-nitro-4-(nitromethyl)-4H-chromen-2-amine
8k.
S24
Two-dimensional ligand interaction diagram of nine 4H-chromene derivatives with Bcl-2
Protein
S27
Two-dimensional ligand interaction diagram of nine 4H-chromene derivatives with Bclw Protein
S32
Two-dimensional ligand interaction diagram of nine 4H-chromene derivatives with BclxL Protein
S37
4
Table 1 Molecular docking results of nine 4H-chromene derivatives with anti-apoptotic
Bcl-2, Bcl-w and Bcl-xL Proteins
S42
Table 2 Lipinski’s Values for two 4H-chromene derivatives 8h and 8i
S45
Table 3 Predicted ADMET properties of two 4H-chromene derivatives 8h and 8i
[Abbreviation: - IS: Intestinal Absorption, AS: Aqueous Solubility, BBB: Blood Brain
Barrier, HT: Hepato toxicity, PPB: Plasma Protein Binding]
S46
5
IR (KBr) spectrum of N-methyl-3-nitro-4-(nitromethyl)-4H-chromen-2-amine 8a.
400 MHz (CDCl3) 1H NMR spectrum of N-methyl-3-nitro-4-(nitromethyl)4H-chromen-2-amine 8a.
6
100 MHz (CDCl3) 13C NMR spectrum of N-methyl-3-nitro-4-(nitromethyl)4H-chromen-2-amine 8a.
DEPT-135 NMR (100 MHz, CDCl3) spectrum of N-methyl-3-nitro-4-(nitromethyl)-4Hchromen-2-amine 8a.
7
IR (KBr) spectrum of 6-chloro-N-methyl-3-nitro-4-(nitromethyl)-4H-chromen-2-amine
8b.
400 MHz (CDCl3) 1H NMR spectrum of 6-chloro-N-methyl-3-nitro-4-(nitromethyl)-4Hchromen-2-amine 8b.
8
100 MHz (CDCl3) 13C NMR spectrum of 6-chloro-N-methyl-3-nitro-4-(nitromethyl)-4Hchromen-2-amine 8b.
DEPT-135 NMR (100 MHz, CDCl3) spectrum of 6-chloro-N-methyl-3-nitro-4(nitromethyl)-4H-chromen-2-amine 8b.
9
IR (KBr) spectrum of 6-bromo-N-methyl-3-nitro-4-(nitromethyl)-4H-chromen-2-amine
8c
400 MHz (CDCl3) 1H NMR spectrum of 6-bromo-N-methyl-3-nitro-4-(nitromethyl)-4Hchromen-2-amine 8c.
10
100 MHz (CDCl3) 13C NMR spectrum of 6-bromo-N-methyl-3-nitro-4-(nitromethyl)-4Hchromen-2-amine 8c.
DEPT-135 NMR (100 MHz, CDCl3) spectrum 6-bromo-N-methyl-3-nitro-4(nitromethyl)-4H-chromen-2-amine 8c
11
IR (KBr) spectrum of 6,8-dichloro-N-methyl-3-nitro-4-(nitromethyl)-4H-chromen-2amine 8d.
400 MHz (DMSO-d6 + CCl4, 1:1) 1H NMR spectrum of 6,8-dichloro-N-methyl-3-nitro-4(nitromethyl)-4H-chromen-2-amine 8d.
12
100 MHz (DMSO-d6 + CCl4, 1:1) 13C NMR spectrum of 6,8-dichloro-N-methyl-3-nitro4-(nitromethyl)-4H-chromen-2-amine 8d.
DEPT-135 NMR (100 MHz, CDCl3) spectrum of 6,8-dichloro-N-methyl-3-nitro-4(nitromethyl)-4H-chromen-2-amine 8d.
13
IR (KBr) spectrum of 6-nitro-N-methyl-3-nitro-4-(nitroethyl)-4H-chromen-2-amine 8e.
400 MHz (DMSO-d6 + CCl4, 1:1) 1H NMR spectrum of 6-nitro-N-methyl-3-nitro-4(nitroethyl)-4H-chromen-2-amine 8e.
14
100 MHz (DMSO-d6 + CCl4, 1:1) 13C NMR spectrum of 6-nitro-N-methyl-3-nitro-4(nitroethyl)-4H-chromen-2-amine 8e.
DEPT-135 NMR (100 MHz, CDCl3) spectrum of 6-nitro-N-methyl-3-nitro-4(nitroethyl)-4H-chromen-2-amine 8e.
15
400 MHz (DMSO-d6 + CCl4, 1:1) 1H NMR spectrum of 6-chloro-N,7-dimethyl-3-nitro-4(nitromethyl)-4H-chromen-2-amine 8f.
100 MHz (DMSO-d6 + CCl4, 1:1) 13C NMR spectrum of 6-chloro-N,7-dimethyl-3-nitro4-(nitromethyl)-4H-chromen-2-amine 8f.
16
IR (KBr) spectrum of 8-methoxy-N-methyl-3-nitro-4-(nitromethyl)-4H-chromen-2-amine
8g.
400 MHz (CDCl3) 1H NMR spectrum of 8-methoxy-N-methyl-3-nitro-4(nitromethyl)-4H-chromen-2-amine 8g.
17
100 MHz (CDCl3) 13C NMR spectrum of 8-methoxy-N-methyl-3-nitro-4-(nitromethyl)4H-chromen-2-amine 8g.
DEPT-135 NMR (100 MHz, CDCl3) spectrum of 8-methoxy-N-methyl-3-nitro-4(nitromethyl)-4H-chromen-2-amine 8g.
18
IR (KBr) spectrum of 6-methoxy-N-methyl-3-nitro-4-(nitromethyl)-4H-chromen-2-amine
8h.
400 MHz (CDCl3) 1H NMR spectrum of 6-methoxy-N-methyl-3-nitro-4-(nitromethyl)4H-chromen-2-amine 8h.
19
100 MHz (CDCl3) 13C NMR spectrum of 6-methoxy-N-methyl-3-nitro-4-(nitromethyl)4H-chromen-2-amine 8h.
DEPT-135 NMR (100 MHz, CDCl3) spectrum of 6-methoxy-N-methyl-3-nitro-4(nitromethyl)-4H-chromen-2-amine 8h.
20
IR (KBr) spectrum of 6-ethoxy-N-methyl-3-nitro-4-(nitromethyl)-4H-chromen-2-amine
8i.
400 MHz (CDCl3 + CCl4, 1:1) 1H NMR spectrum of 6-ethoxy-N-methyl-3-nitro-4(nitromethyl)-4H-chromen-2-amine 8i.
21
100 MHz (CDCl3 + CCl4, 1:1) 13C NMR spectrum of 6-ethoxy-N-methyl-3-nitro-4(nitromethyl)-4H-chromen-2-amine 8i.
DEPT-135 NMR (100 MHz, CDCl3) spectrum of 6-ethoxy-N-methyl-3-nitro-4(nitromethyl)-4H-chromen-2-amine 8i.
22
IR (KBr) spectrum of 6-methyl-N-methyl-3-nitro-4-(nitroethyl)-4H-chromen-2-amine 8j.
400 MHz (CDCl3 + CCl4, 1:1) 1H NMR spectrum of 6-methyl-N-methyl-3-nitro-4(nitroethyl)-4H-chromen-2-amine 8j.
23
100 MHz (CDCl3 + CCl4, 1:1) 13C NMR spectrum of 6-methyl-N-methyl-3-nitro-4(nitroethyl)-4H-chromen-2-amine 8j.
DEPT-135 NMR (100 MHz, CDCl3) spectrum of 6-methyl-N-methyl-3-nitro-4(nitroethyl)-4H-chromen-2-amine 8j.
24
IR (KBr) spectrum of 6-ethyl-N-methyl-3-nitro-4-(nitromethyl)-4H-chromen-2-amine 8k.
400 MHz (CDCl3 + CCl4, 1:1) 1H NMR spectrum of 6-ethyl-N-methyl-3-nitro-4(nitromethyl)-4H-chromen-2-amine 8k.
25
100 MHz (CDCl3 + CCl4, 1:1) 13C NMR spectrum of 6-ethyl-N-methyl-3-nitro-4(nitromethyl)-4H-chromen-2-amine 8k.
DEPT-135 NMR (100 MHz, CDCl3) spectrum of 6-ethyl-N-methyl-3-nitro-4(nitromethyl)-4H-chromen-2-amine 8k.
26
Molecular docking studies of 4H-chromene derivatives (8a-8g, 8j, 8k) with antiapoptotic Bcl-2, Bcl-w and Bcl-xL proteins
Binding mode analysis of nine 4H-chromene derivatives with Bcl-2 protein:
Molecular docking results of nine 4H-chromene derivatives (8a-8g, 8j, 8k) with antiapoptotic Bcl-2 protein revealed that the residue R107 makes cation-π interaction with
4H-chromenes 8b, 8c, 8j and 8k as HA 14-1. Similarly, the residues S205, E136, R139
and G145 form HBs with ligand molecules 8a, 8g, 8d, 8e & 8k. In addition, the ligands
also showed VdW and polar/charge interactions with anti-apoptotic Bcl-2 protein. The
glide docking score of nine 4H-chromene derivatives with Bcl-2 protein was given in
Table 1.
Binding mode analysis of nine 4H-chromene derivatives with Bcl-w protein:
Similar to HA 14-1, compounds 8e and 8f makes hydrogen bonding interaction with R55
residue of Bcl-w protein with the bond distances of 2.0 Å and 2.1 Å. In addition,
compounds 8a, 8b, 8c, 8d, 8e, 8f, 8g, and 8j were involved in formation of HB with
residues R94, G153 and L149 of the protein. Moreover, the R94 and F52 residues of Bclw were engaged in the formation of cation-π and π-π interactions with ligand molecules
(8d & 8k) as HA 14-1. The docking score, VdW, polar/charge interactions of nine 4Hchromene derivatives with anti-apoptotic Bcl-w protein was presented in Table 1.
Binding mode analysis of nine 4H-chromene derivatives with Bcl-xL protein:
The binding of Bcl-xL protein with ligand molecules was stabilized by the presence of
VdW, polar/charge, cation-π and π-π interactions. Here, the residues Y195 and Y101 of
Bcl-xL protein were involved in the formation of π-π (8c) and cation-π (8d & 8f)
interactions with ligand molecules. Other molecular interactions of ligand molecules with
Bcl-xL protein was given in Table 2.
In summary, all nine 4H-chromene derivatives with target proteins show the similar
binding profile as reference molecule, which is evident from their in silico interaction
results. The two-dimensional ligand interaction diagrams and molecular docking results
are presented here.
27
Two-dimensional ligand interaction diagram of nine 4H-chromene derivatives (8a8g, 8j, 8k) Bcl-2 Protein.
The purple circle represents the amino acid residues participate in hydrogen bonding,
electrostatic or polar interactions and the Van der Waals interactions are represented by
green circles. Cation-π interactions are showed as orange line and hydrogen bonds are
displayed as dotted arrows
1.4H-chromene 8a:
2. 4H-chromene 8b:
28
3. 4H-chromene 8c:
4. 4H-chromene 8d:
29
5. 4H-chromene 8e:
6. 4H-chromene 8f:
30
7. 4H-chromene 8g:
8. 4H-chromene 8j:
31
9. 4H-chromene 8k:
32
Two-dimensional ligand interaction diagram of nine 4H-chromene derivatives (8a8g, 8j, 8k) with Bcl-w Protein.
1.4H-chromene 8a:
2. 4H-chromene 8b:
33
3. 4H-chromene 8c:
4. 4H-chromene 8d:
34
5. 4H-chromene 8e:
6. 4H-chromene 8f:
35
7. 4H-chromene 8g:
8. 4H-chromene 8j:
36
9. 4H-chromene 8k:
37
Two-dimensional ligand interaction diagram of nine 4H-chromene derivatives(8a8g, 8j, 8k) and with Bcl-xL Protein.
1.4H-chromene 8a:
2. 4H-chromene 8b:
38
3. 4H-chromene 8c:
4. 4H-chromene 8d:
39
5. 4H-chromene 8e:
6. 4H-chromene 8f:
40
7. 4H-chromene 8g:
8. 4H-chromene 8j:
41
9. 4H-chromene 8k:
42
Table 1
Proteins
Ligands
G Score
H-bonds
Van der Waals interactions
Bcl-2
4H-chromene 8j
-4.910
-
4H-chromene 8e
-4.849
E136(2.1),
R139(2.3)
L201, V148, A100, D103,
F104, F198, Y202, R107
V133, A149, F104
4H-chromene 8d
-4.803
E136(2.1)
V133, A149, F104
4H-chromene 8b
-4.776
-
4H-chromene 8k
-4.766
G145(2.5)
4H-chromene 8g
-4.765
S205(2.5)
Y108, F104, A149, F198,
V148, Y202, A100, D103,
R107
L201, Y202, F198, D103,
A100, V148, F104, A149,
R107
W144, F198, V148, A100,
F104, Y202
4H-chromene 8c
-4.646
-
4H-chromene 8a
-4.558
S205(2.0)
4H-chromene 8f
-4.464
-
4H-chromene 8a
-5.123
R94(2.0)
Bcl-w
Y108, F104, D103, A100,
Y202, F198, V148, G145,
R107
A100, F104, F198, W144,
V148
R107, F112, Y108, F104,
A149, L137, G145
W92, F56, F52, V96, A97,
V100, L63, T59
Charge/
Polar
interaction
N143, W144,
G145
N143, G145,
R146, D140,
L137, E136,
R139
L137, R146,
R139, D140,
G145, E136
G145
Cation-π
interaction
π-π
interaction
R107(6.9)
-
-
-
-
-
R107(6.2)
-
W144, N143,
G145
R107(6.8)
-
G145, R107,
D103, L201,
S205
N143
-
-
R107(6.3)
-
Y202, L201,
G145, D103.
R107, S205
N143, R146
-
-
R55, G93, R94
-
-
43
Bcl-xL
4H-chromene 8e
-5.107
4H-chromene 8j
-5.055
R94(2.0),
R55(1.9)
R94(2.0)
V96, F52, V100, A97, L63,
T59, F56, W92, G153
G93, F56, L63, T59, A97, F52,
W92, V96
F146, V96, F52, W92, R55,
L63, T59, V100, F56
R55, W92, F56, T59, L63,
A97, V100, V96, F52
V100, A97, V96, F52, L63,
T59, F56, W92
Y150, F52, L149, V96, W92
G93, N91, R94, R55
R55, R94
-
-
4H-chromene 8g
-5.014
R94(2.0)
A97, G93, R94
-
-
4H-chromene 8k
-5.006
-
G93, R94, N91
-
F52(6.0)
4H-chromene 8b
-4.917
R94(2.0)
-
-
R55(5.5),
R94(6.9)
-
A97, G93, V96, F146, G153,
E51, W92, F52
V96, F52, V100, A97, L63,
T59, F56, W92
R139, L130, A142, A104,
L108, F105, F97, Y101
F105, F97, L130, Y101, R139,
V126, A142, A104, L108,
S145, F146
L130, S145, V126, F146,
L108, F97, A104, A142
E129, V126, F146, L108,
F105, A142, L130, F97, A104
W137, F191, N197, E92,
V141, Y101, F97
N91, G93, R55,
R94
D152, R159,
G93, R94, R55,
G153
Y150, L149,
R55
N91, G93, R55,
R94
V126, S145,
F146
-
4H-chromene 8d
-4.639
G153(2.2)
4H-chromene 8f
-4.846
4H-chromene 8c
-4.613
L149(2.3),
R55(2.1)
R94(2.1)
-
-
-
-
4H-chromene 8b
-5.388
-
F146(6.5)
-
4H-chromene 8a
-5.397
-
-
-
4H-chromene 8f
-5.135
-
E129, F105,
Y101
-
Y101(4.4)
-
4H-chromene 8d
-5.101
-
Y101(5.2)
-
4H-chromene 8c
-5.075
-
A93, G138,
E96, R100,
Y195
Y195, Y101,
G138, W137,
N136
R100, Y101,
-
Y195(4.0)
4H-chromene 8j
-5.055
-
F191, V141, A93, F97, R139
-
-
4H-chromene 8e
-4.990
-
F191, V141, E96
-
-
-
44
4H-chromene 8g
-4.988
-
4H-chromene 8k
-4.980
-
F146, S145, L108, F97, A104,
Y101,
E98, Y101, F97, N136, F146,
L108, V126, A142, R139,
G138
R139, G138,
N136, W137,
Y195, A93,
F97
F105, L130,
E129, V126
F105, A104,
E129, L130
-
-
-
-
45
Table 2.
Ligands
Molecular
Weight
4H-chromene 8h
4H-chromene 8i
295.25
309.27
Hydrogen
Bond
Donor
1
1
Hydrogen
Bond Acceptor
ALogP
9
9
1.509
1.858
46
Table 3.
Ligands
4H-chromene 8h
4H-chromene 8i
IA
0
0
AS
3
3
BBB
0
0
Cyt P450 2D6
0
0
HT
0
0
PPB
0
0