Computational Evaluation of Novel Schiff Base Complexes as Anti Dengue and Anti-Cancer Agent

Introduction

Research on interactions of the metal complexes especially Co(II), Ni(II) and Cu(II) with DNA helps to get  a  immense knowledge about development of novel chemotherapeutics and medicines [1-4].  The interactions of metal complexes with DNA usually cause damage in cancer cells [5-7]. Dengue virus basically belongs to Flavivirus genus and is a member of Flaviviridae family [8,9]. The major source of the dengue virus development is human and monkey. Usually dengue affected humans have severe flu, fever, body pain like symptoms and body temperature almost reaches 40°C which makes human to be critical. Severe headache, flushing of face and rashes in skin are the extreme symptoms of Dengue fever and this severe condition is said to be Dengue Hemorrhagic Fever (DHF) and Dengue Shock Syndrome (DSS) [10,11]. It is not possible to do the all experiment on the interaction of drugs towards the targeting molecule in low cost. Docking is the only way to give the corresponding exact results of newly synthesized drugs towards the targeting molecule [12-18]. Auto dock vina and discovery studio is a useful tool to study the interaction between the ligand and the target molecule. This software helps to determine the binding mode, binding affinity and also the exact stable configuration of the ligand, which binds with the receptor [19-22].  Using this docking study, we can understand the coordination mode and the binding nature. It will help to design a good drug for the disease [23-26]. In this present work, a neoteric Schiff base ligand, 4-(4-chlorobenzylideneamino)-N-(4-(4-chlorobenzylideneamino) phenyl)benzamide (L1) and Co(II), Ni(II) and Cu(II)  complexes of L1 were synthesized. The synthesized ligand and its complexes were treated with human DNA topoisomerase I (PDB: 1SC7) and Dengue NS3 protease-helicase (PDB ID: 2VBC) by docking.

Materials and Experimental Methods

All the chemicals which were used in the present work were purchased from commercial sources and were used without further purification. 4,4′-diaminobenzanilide,  p-chlorobenzaldehyde, cobalt chloride hexahydrate (CoCl₂. 6H₂O), nickel chloride hexahydrate  (NiCl₂. 6H₂O) and copper chloride dihydrate  (CuCl₂. 6H₂O) were bought from Sigma Aldrich, USA and utilized as received. Solvents used in the present research were purchased from Merck and were used without further purification.

¹H-NMR spectrum of the synthesized Schiff base ligand L1 was recorded in DMSO (d₆) and TMS is used as an internal standard on a Bruker Advance DRX 300 FT-NMR instrument. Vibrational spectra of the ligand and complexes were recorded in JASCO/FT-IR410 spectrometer in the range of 4000-400 cm^-1 at 16 scans/min. Potassium bromide disc method was employed for sample preparation. Electronic spectra of the complexes dissolved in DMSO were recorded using Perkin Elmer Lambda-25 UV-Vis. spectrophotometer in the range of 200-800 nm. The Electron Ionization Mass spectra of the ligand was recorded using JEOL DX-303 EI mass spectrometer, and X- band EPR spectra of the copper complex in DMSO was recorded on Varian E-4 X-band spectrometer using DPPH as the g-marker at room temperature at Indian Institute of Technology, Madras, India.

Experimental

Synthesis of 4-(4-chlorobenzylideneamino)-N-(4-(4-chlorobenzylideneamino)phenyl) benzamide (L1)

A hot solution of 1.136 g (5 mmol) 4,4′-diaminobenzanilide in 10 mL methanol was added drop wise to a hot stirring solution of 1.4057g (10 mmol) p-chlorobenzaldehyde in10 mL methanol. The above mixture was stirred under reflux condition for 5 hours (Scheme-1).  A yellow colour solid mass appeared was filtered, washed with dimethyl ether several times and dried in in vacuo at room temperature.

Scheme 1: Synthesis of Schiff base Ligand L1 Click here to View scheme

 

Synthesis of Cobalt(II), Nickel(II) and Copper (II) complexes of L1

To the hot stirring solution of the 0.9447 g (2 mmol) Schiff base ligand L1 in 20 mL of methanol, the corresponding metal(II) chlorides [CoCl₂.6H₂O (0.237 g, 1 mmol), NiCl₂.6H₂O (0.2379 g,  1 mmol) and CuCl₂.2H₂O (0.170 g,  1 mmol)] in 20 mL of methanol were added, stirred under reflux for 6 hours (Scheme-2). Then the product obtained was filtered, washed and dried in vacuo at room temperature.

Scheme 2: Synthesis of Schiff base metal complexes   Click here to View figure

 

Molecular Docking Study

Human‐DNA–Topo‐I complex (PDBID: 1SC7) [27] and Dengue NS3 protease-helicase (PDB ID: 2VBC) [28] were obtained from Protein Data Bank (http://www.rcsb.org/pdb). Metal complexes were converted into PDB format by using Mercury software [29]. The ‘receptor’ (DNA/Dengue) and ‘ligand’ (metal complexes) files were docked using AutoDock Tools. The hetero atoms such as water molecules and excess ligand were removed. The polar hydrogen atoms and Kollman charges were added to the receptor (DNA/Dengue) molecule to investigate the binding nature. The binding which was enclosed in a box have number of grid points in x × y × z directions of 40 × 40 × 40 and a grid spacing of 0.35 Å. The docking studies were conducted using AutoDockTools (ADT) version 1.5.4 and finalized by Auto Dock vina program [30]. The docked structures were exposed using Discovery studio software.

Results and Discussion

¹H-NMR Spectral Studies and Electron Impact Mass Spectral analysis

In the ¹H NMR spectrum of the ligand L1, which is provided in the Figure 1 there is a signal at 9.61 ppm attributed to amido proton of benzanilide group present in the ligand L1 [31]. The signal for azomethine proton appeared as a small peak at 8 ppm [32]. The multiplet in the region 6.05-7.55 ppm represents the protons present in the aromatic moiety.

The Electron Ionization mass spectrum of the Schiff base ligand L1 is given in the Figure 2. The molecular ion M⁺ peak is present at 470.9364, which is corresponding to the molecular weight of the ligand L1. In addition there is an (M⁺ + 2) isotopic peak at 472.9053. The peaks at 367.9976, 326.0706, 311.0307, 241.8728, 207.0274, 152.0043 and 91.0319 correspond to various fragments of the ligand.

Figure 1: 1H-NMR Spectra of L1 Click here to View figure

 

Figure 2: EI- Mass spectra of L1 Click here to View figure

 

FT-IR Spectral Studies

The FT-IR spectrum of the Schiff base ligand L1 and its complexes were represented in the Figure 3. The FT-IR spectra of the ligand L1 exhibits a band at 1592 cm^-1, which is attributed to imino stretching frequency of the Schiff base ligand [33]. The strong peak at 3354 cm^-1 is the characteristic N-H stretching frequency of the secondary amide group of 4,4ʹ-diaminobenzanilide part of the ligand. To study the binding mode of the ligand to the metal in the complexes, the IR spectrum of the free ligand and its complexes were compared. The band near 1592 cm^-1 is shifted to lower frequencies in complexes. This clearly indicates the coordination of the imino-nitrogen of the ligand to the metal center [34]. Further the IR spectra of the complexes show some new sharp signals in the region 467 cm^-1, 437 cm^-1 and 465 cm^-1 for CoL1₂, NiLl₂ and CuLl₂ complexes respectively, which corresponds to metal-nitrogen stretching formed by the coordination of imino nitrogen of the ligand with the metal centers.

Figure 3: FT-IR Spectra of Schiff base ligand L1 and its complexes   Click here to View figure

 

Electronic Spectral Analysis

The electronic spectra of complexes Co(L1)₂, Ni(L1)₂ and Cu(L1)₂ are recorded in the range of 200 nm-800 nm in DMSO and were  provided in Figure 4. The spectral details below 350 nm are similar for all the complexes and they are corresponding to the transitions (π-π^* and n- π^*) of benzene ring and non-bonding electrons present on the nitrogen of azomethine group in the Schiff base complexes [35]. In the electronic spectra of the cobalt complex, the band around 380 nm is assigned to LMCT [36] transition while the d-d band observed in the lower energy region around 430 nm is assigned to the combination of  ²B_1g ⟶ ¹A_1g and  ¹B_1g ⟶ ²E_g transitions. This transition represents a square planar geometry for cobalt complex [37-38]. In the electronic spectrum of the nickel complex, the band around 360 nm is assigned to LMCT transition. Additionally a broad band observed in the lower energy region around 370-410 nm is assigned to d-d (¹A_1g ⟶ ¹B_1g)   transition. This transition represents the square planar geometry for nickel complex [39]. The electronic spectrum of copper complex shows d-d band at 450 nm, which is attributed to the combination of ²B_1g ⟶ ²E_g and ²B_1g ⟶ ²B_2g transitions. This give the evidence for the distorted square planar geometry for d⁹ Cu(II) system [40].

Figure 4: UV-Vis Spectra of Schiff base complexes     Click here to View figure

 

EPR Spectral Analysis

The EPR spectra of copper complex give useful information in studying the metal ion environment. In the EPR spectrum of the copper complex Cu(L1)₂ there is an isotropic signal, without any hyperfine splitting, with g_iso = 2.12 as shown in the Figure 5. When comparing the g_iso value of the copper complex with the g value of free electron (2.0023) indicate that there is an increase in the covalent nature of the bonding between the metal ion and the ligand molecule [41]. Isotropic lines are usually appeared due to the occupancy of the unpaired electron in a degenerate orbital in square planar geometry.

Figure 5: EPR Spectrum of Cu(L1)2     Click here to View figure

 

Molecular Docking with Human DNA Topoisomerase I

Human DNA topoisomerase I was the selective targeting area for synthesizing the anticancer drug [42]. The molecular docking of Co (II), Ni (II) and Cu(II) complexes were performed to determine the binding affinity value and the selected binding residue, along with the sterically suitable conformations. The low value of the binding energy shows the more effective binding affinity between the ‘receptor’(DNA) and the ‘ligand’(complex) molecules. The various conformations of docked molecular complexes were analyzed in terms of binding energy, hydrogen bonding and hydrophobic interaction between receptors and the acceptor. More negative value of the relative binding values suggests that the interaction between the DNA and ligand is so strong, due to the extended aromatic ring. Phenyl ring has higher free binding energy which gives a better binding affinity value comparing with a compound containing bipyridyl ring. In these work mononuclear complexes give better results towards the HDNA. The binding energy values of the Co(II), Ni(II) and Cu(II) complexes were respectively -16.8, -10.4 and -15.1 kcal mol⁻¹ towards human DNA topoisomerase I. This shows that Cobalt(II) and Copper(II) molecule easily bind with the DNA helix and the Nickel(II) complex preferred to bind with the outermost protein’s amino acid residue. The binding values were shown in the table 1, 2 and 3 and the docked images were shown in Figure 6.

Figure 6: (a), (c) and (e) demonstrate the binding of Co(II), Ni(II) and Cu(II) complexes with active site of human DNA topoisomerase I, and (b), (d) and (f) demonstrate the binding of Co(II), Ni(II) and Cu(II) complexes with selective Nucleotide of human DNA topoisomerase I Click here to View figure

 

Molecular Docking with NS3 Protease-Helicase

NS3 protease-helicase (dengue virus) is a very important target area which should be docked. Cobalt(II), Nickel(II) and Copper(II) complexes exhibit very low binding energy value, it means that complexes having very high binding affinity towards the NS3 protease-helicase. The values of the binding energy in various levels are present in table 1, 2 & 3.The distance between the selected receptor to targeted molecule is also low. The binding energy values of the Co(II), Ni(II) and Cu(II) complexes were -12.4, -13.7 and -13.0 kcal mol⁻¹respectively. The binding interactions were shown in fig.7. From the theoretical point of view these complexes are considered to be a good anti-dengue drug.

Figure 7: (a) (c) and (e) demonstrate the binding of Co(II), Ni(II) and Cu(II) complexes with active site of NS3 protease-helicase, and (b), (d) and (f) demonstrate the binding of Co(II), Ni(II) and Cu(II) complexes with selective Amino acid residue  of NS3 protease-helicase Click here to View figure

 

Table1: Binding interactions of Co(II) complex with human DNA topoisomerase I and NS3 protease-helicase

Mode	Affinity (kcal/mol)		Distance from best mode
	Human DNA topoisomerase I	NS3 protease-helicase
			Human DNA topoisomerase I		NS3 protease-helicase
			rmsdl.b.             rmsdu.b.		rmsdl.b.       rmsdu.b.
1	-16.8	-12.4	0	0	0	0
2	-16.5	-12.3	3.771	9.455	2.459	10.883
3	-16.2	-12.2	0.522	8.076	4.393	8.79
4	-15.6	-12.2	4.114	8.625	1.405	9.713
5	-15.4	-12	4.148	9.703	4.273	8.645
6	-14.6	-12	15.172	21.499	0.596	7.523
7	-14.5	-11.5	4.101	9.898	4.684	8.329
8	-14.4	-11.3	15.451	22.1	3.498	8.79
9	-14.4	-11.1	15.378	21.17	12.94	18.217

 

Table 2: Binding interactions of Ni(II) complex with human DNA topoisomerase I and NS3 protease-helicase

Mode	Affinity (kcal/mol)		Distance from best mode
	Human DNA topoisomerase I	NS3 protease-helicase
			Human DNA topoisomerase I		NS3 protease-helicase
			rmsdl.b.              rmsdu.b.		rmsdl.b.                rmsdu.b.
1	-10.4	-13.7	0.000	0.000	0.000	0.000
2	-9.9	-13.4	1.841	7.560	1.573	7.663
3	-9.7	-12.2	5.281	12.004	31.948	38.354
4	-9.5	-12.0	5.377	11.101	32.727	37.825
5	-9.1	-11.9	1.954	2.421	1.7333	7.947
6	-8.8	-11.7	4.640	10.252	32.315	37.338
7	-8.7	-11.6	7.369	13.017	46.385	51.726
8	-8.6	-11.4	2.349	9.610	20.661	24.252
9	-8.3	-11.2	5.571	12.245	32.817	39.072

 

Table 3: Binding interactions of Cu(II) complex with human DNA topoisomerase I and NS3 protease-helicase

Mode	Affinity (kcal/mol)		Distance from best mode
	Human DNA topoisomerase I	NS3 protease-helicase
			Human DNA topoisomerase I		NS3 protease-helicase
			rmsdl.b.       rmsdu.b.		rmsdl.b.     rmsdu.b.
1	-15.1	-13.0	0.000	0.000	0.000	0.000
2	-14.6	-12.3	1.229	1.495	3.964	9.445
3	-14.2	-12.2	10.544	18.244	2.039	9.414
4	-14.2	-12.0	3.599	11.470	4.001	9.745
5	-14.0	-11.8	3.636	10.036	3.795	10.126
6	-13.6	-11.6	1.725	9.229	32.154	38.142
7	-13.1	-11.5	0.956	7.945	48.576	58.949
8	-13.0	-11.5	14.412	18.972	3.690	10.115
9	-13.0	-11.4	10.661	17.927	4.031	10.573

 

Conclusion

A new Schiff base ligand which contains several phenyl rings and its Co(II), Ni(II) and Cu(II) complexes were synthesized. The formation of the ligand and its metal complexes were confirmed by various spectroscopic techniques. The docking studies were carried out using synthesized metal complexes with human DNA topoisomerase I (PDB: 1SC7) and Dengue NS3 protease-helicase (PDB ID: 2VBC) using Auto Dock vina and Discovery studio software. The binding energy values of the Co(II), Ni(II) and Cu(II) complexes were respectively -12.4, -13.7 and -13.0 kcal mol⁻¹ towards NS3 protease-helicase and -16.8, -10.4 and -15.1 kcal mol⁻¹ towards human DNA topoisomerase I, which showed that these types of compounds can be act as an potential Anti-Dengue and Anticancer agent.

Acknowledgement

The authors are thankful to Indian Institute of Technology, Madras, India for EI-mass and EPR spectral analysis and also Dr. S. Gnanavel, Research and Project Centre for Chemical and Biological Science for docking study, Salem, India.

References

1. Sigman, D. S.; Mazumder, A.; Perrin, D. M. Chem. Rev. 1993, 93(6), 2295-2316
   CrossRef
2. Jamieson, E. R.; Lippard, S. J. Chem. Rev. 1999, 99(9), 2467-2498
   CrossRef
3. Liu, H. K.; Sadler, P. J. Acc. Chem. Res. 2011, 44(5), 349-359
   CrossRef
4. Leung, M. H. M.; Harada, T.; Kee, T. W. Curr. Pharm. Des. 2013, 19(11), 2070-2083
5. Alagesan, M.; Bhuvanesh, N. S. P.; Dharmaraj, N. Dalton Trans. 2014, 43(16), 6087-6099
   CrossRef
6. Shao, J.; Ma, Z. Y.; Li, A.; Liu, Y. H.; Xie, C. Z.; Qiang, Z. Y.; Xu, J. Y. J. Inorg. Biochem. 2014, 136, 13-23
   CrossRef
7. Mjos, K. D.; Orvig, C. Chem. Rev. 2014, 114(8), 4540-4563
   CrossRef
8. Wichapong, K.; Nueangaudom, A.; Pianwanit, S.; Sippl, W.; Kokpol, S. Trop. Biomed. 2013, 30(3), 388–408
9. Murphy, F.A.; Mahy, B.W.J.; van Regenmortel, M.H.V. Encycl. Virol. 2008, 140–148
10. Mastrangelo, E.; Milani, M.; Bollati, M.; Selisko, B.; Peyrane, F.; Pandini, V.; Sorrentino, G.; Canard, B.; Konarev, P.V.; Svergun, D.I.; de Lamballerie, X.; Coutard, B.; Khromykh, A.A.; Bolognesi, M. J. Mol. Biol. 2007, 372(2), 444–455
    CrossRef
11. Byrd, C.M.; Grosenbach, D.W.; Berhanu, A.; Dai, D.; Jones, K.F.; Cardwell, K.B.;  Schneider, C.; Yang, G.; Tyavanagimatt, S.; Harver, C.; Wineinger, K. A.; Page, J.; Stavale, E.; Stone, M. A.; Fuller, K. P.; Lovejoy, C.; Leeds, J. M.; Hruby, D. E.; Jordan, R. .Antimicrob. Agents Chemother. 2013, 57(4), 1902–1912
    CrossRef
12. Noble, C.G.; She, C.C.; Chao, A.T.; Shi P.Y. J. Vir. 2012, 86(1), 438–446
    CrossRef
13. Madhavi Sastry, G.; Adzhigirey, M.; Day, T.; Annabhimoju, R.; Sherman, W. J. Comput. Aided Mol. Des., 2013, 27 (3), 221–234
    CrossRef
14. Bas, D.C.; Rogers, D. M.; Jensen, J. H. Proteins Struct. Funct. Genet. 2008,73(3), 765–783
    CrossRef
15. Li, H.; Robertson, A. D.; Jensen, J.H. Proteins. 2005, 61 (4), 704–721
    CrossRef
16. Irwin, J.J.; Shoichet, B.K. J. Chem. Inf. Model. 2005, 45 (1), 177–182
    CrossRef
17. Greenwood, J. R.; Calkins, D.; Sullivan, A.P.; Shelley, J. C. J. Comput. Aided Mol. Des. 2010, 24 (6-7), 591–604
    CrossRef
18. Greco, M. N.; Hawkins, M. J.; Powell, E. T.; Almond, H. R.; Garavilla, L.; Hall, J.; Minor, L. K.; Wang, Y.; Corcoran, T.W.; Cera, E. D.; Cantwell, A. M.; Savvides, S.N.; Damiano, B.P.; Maryanoff , B. E. J. Med .Chem. 2007, 50(8), 1727–1730
    CrossRef
19. Iijima, K.; Katada, J.; Yasuda, E.; Uno, I.; Hayashi, Y. J. Med. Chem. 1999, 42(2), 312–323
    CrossRef
20. Aoyama, Y.; Uenaka, M.; Kii, M.; Tanaka, M.; Konoike, T.; Hayasaki- Kajiwara, Y.; Naya, N.; Nakajima, M. Bioorg. Med. Chem., 2001, 9(11), 3065–3075
    CrossRef
21. Aoyama, Y.; Uenaka, M.; Konoike, T.; Hayasaki-Kajiwara, Y.; Naya, N.; Nakajima, M. Bioorg. Med. Chem. Lett. 2001, 11(13), 1691–1694
    CrossRef
22.  Aoyama, Y.; Konoike ,T.; Kanda, A.; Naya, N.; Nakajima, M. Bioorg. Med. Chem.Lett. 2001, 11(13), 1695–1697
    CrossRef
23. Maruoka, H.; Muto, T.; Tanaka, T.; Imajo, S.; Tomimori, Y.; Fukudaa, Y.; Nakatsuka, T. Bioorg. Med. Chem. Lett. 2007, 17(12), 3435–3439
    CrossRef
24. Wawer, M.; Bajorath, J. J .Chem. Inf.  Model. 2010, 50(8), 1395–1409
    CrossRef
25. Thangapandian, S.; John, S.; Sakkiah, S.; Lee, K. L. J. Chem. Inf. Model. 2011, 51(1), 33–44
    CrossRef
26. Lie, M. A.; Thomsen, R.; Pedersen, C. N. S.; Schiøtt B.; Christensen, M. H. J. Chem. Inf. Model. 2011, 51(4), 909–917
    CrossRef
27. Bujacz, A. ActaCrystallogr. D. 2012, 68(10), 1278-1289
    CrossRef
28. Mirza, S.B., Salmas R.E., Fatmi M.Q. and Durdagi S.J, mol graph model., 2016,  66, 99-107.
29. Macrae, C. F.; Edgington, P. R.; McCabe, P.; Pidcock, E.; Shields, G. P.; Taylor, R.; Towler, M.; van de Streek, J. J. Appl. Crystallogr. 2006, 39, 453-457
    CrossRef
30. Trott, O.; Olson, A. J. J. Comput. Chem. 2010, 31(2), 455-461
31. Abdulghani, A. J.; Mohuee, S.K. J Chem Bio Phy Sci. 2016, 6(2), 579-595
32. Patel, K. V.; Bhattacharya, P. K. Ind. J. Chem. 1982, 21(12), 1110-1112
33. Singh, D. P.; Vandna Malik; Krishan Kumar; Chetan Sharma ; Aneeja, K.R. Spectrochim. A. 2010, 76(1), 45-49
    CrossRef
34. Raman, N.; Sobha, S. Spectrochim. Acta A. 2012, 85(1), 223–234
    CrossRef
35. Ramesh, R.; Maheswaran, S. J.Inorg.Biochem. 2003, 96(4), 457-462
    CrossRef
36. Ortiz, B.; Park, S. M. Bull. korean Chem.Soc. 2000, 21(4), 405- 411
37. Shakir, M.; Nasam, O.S.M.; Mohamed A. K.; Varkey, S.P. Polyhedron. 1996, 15 (8), 1283- 1287
    CrossRef
38. Chem, L. S.; Cummings, S.C. Inorg. Chem. 1978, 17 (9), 2358-2361
    CrossRef
39. Kirchhoff, J. R.; GamacheJr, R. E.; Blaskie, M. W.; Del Paggio, A. A.; Lengel, R. K.; McMillin, D.R. Inorg. Chem. 1983, 22 (17), 2380-2384
    CrossRef
40. Natarajan, C.; Tharmaraj, P.; Murugesan, R. J. Coord. Chem. 1992, 26(3), 205-213
    CrossRef
41. Raman, N.; Dhaveedhu Raja, J.; Shakthivel, A. J. Chem. Sci. 2007, 119(4), 303-310
    CrossRef
42. Pommier, Y. Nat. Rev. Cancer., 2006, 6(10), 789-802
    CrossRef

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