key: cord-0998722-6thqahu3 authors: Ali, Arif; Sepay, Nasim; Afzal, Mohd; Sepay, Nayim; Alarifi, Abdullah; Shahid, M.; Ahmad, Musheer title: Molecular designing, crystal structure determination and in silico screening of copper(II) complexes bearing 8-hydroxyquinoline derivatives as anti-COVID-19 date: 2021-02-24 journal: Bioorg Chem DOI: 10.1016/j.bioorg.2021.104772 sha: d0f3d0458c21dc919da81f8b4f71fa4fdd42d96f doc_id: 998722 cord_uid: 6thqahu3 The pandemic by COVID-19 is hampering everything on the earth including physical and mental health, daily life and global economy. At the moment, there are no defined drugs, while few vaccines are available in the market to combat SARS-CoV-2. Various molecules, mainly organic, designed and tested for the virus but they are unable to preformed accordingly. In this work we designed two copper complexes from the ligands analogues with chloroquine and hydroxychloroquine. We have synthesized and characterized them with the help of multiple spectroscopy and X-ray crystallography. The complexes are screened through in silico method with the chloroquine and hydroxychloroquine and found the target protein and mechanism of action of these organic molecules as well as complexes. The results indicate that the balanced hydrophobic and polar groups in the complexes favor their binding in the active site of the viral ADP-ribose-1 monophosphatase enzyme over the parent organic molecules. Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is the cause of COVID-19 pandemic [1] . The transmission stages of the virus are asymptomatic, moderate, extreme and critical. The major symptoms associated with these infections are high fever, pneumonia, encephalopathy, dry cough and dyspnea and in severe cases, it may causes heart failure and kidney injury [2] . Presently, there are no defined drugs, although few vaccines are available in the market with a hope to combat SARS-CoV-2 deadly viral infection. It is well know that the viral proteases have long been shown to be effective targets of antiviral therapies. This is the reason why most of the antiviral drugs have been used for the COVID-19 treatment at an early stage of diagnosis [3] . Navarro et al. reported that the incorporation of metal ions into organic pharmacophores displayed better anti-malarial activity than chloroquine, against the chloroquine-susceptible strain which compliment the molecular diversity created by purely organic scaffolds [4] . Quinoline moiety based anti-malarial drug chloroquine and hydroxychloroquine have been approved for the Food and Drug Administration (FDA) for the treatment of COVID-19 [5] . These medicines are only use for emergency purposes because of their some dose related toxic effect in critical cases [5] . However, these medicines are safe if taken within clinically instructed dose limits. Therefore, development of new advanced molecules is on demand. There are many medical problems which is not accessible to organic therapeutic agents and here medicinal inorganic chemistry provides a new opening. Metal-based drugs offer potential benefits over the various non-steroidal anti-inflammatory drugs (NSAIDs), broad-spectrum fluoroquinolone and ornidazole drugs and possess modified pharmacological and toxicological properties, exemplifying the role of the metal ions [6] [7] [8] . This is because of the fact that metal can coordinate ligands in a particular three dimensional configuration and geometries, thermodynamic and kinetic properties. Thus allowing: (i) synergistic effects of the ligands and the coordination residue, (ii) the tailoring/tuning of the molecule to recognize and interact with specific biological target, (iii) protection from the enzymatic degradation of the drug and (iv) superior delivery of pharmacophore through cell membranes and hence, pharmacokinetics profile of the drug (absorption, distribution and excretion etc) may get altered upon metal ion complexation [9] . Among the transition metals, copper is a physiologically important endogenous metal ion and cofactor of several enzymes involved in oxidative metabolism, ceruloplasmine, superoxide dismutase, ascorbic acid oxidase and tyrosinase etc that carry out fundamental biological functions, important in cellular growth and development [10] . Some time it has been observed that metal complex of drugs molecules or allied ligands performed more efficiently than that of the compound itself. The Mn 2+ complex of curcumin (a well-known anti-bacterial) show strong activity against multiple strain of both gram-positive and gram-negative bacteria [11] . A large number of copper complex has also been successfully tested in various diseases including cancer [12] . Recently, Cu 2+ -curcumin [13] and Cu 2+ -ornidazole [6] complex has efficiently been designed to target nucleic acid. Recently, several authors have brought interest in copper well prior to SARS-CoV-2 viral activities has been significant, since the existing outburst of the epidemic continues to develop, and there is no vaccine or drugs are presently accessible, the alternative is now to make the immune system competent to fight somehow against the SARS-CoV-2 [14] . It has been recognized that copper can boost the host's immune system response against pathogens via cell-mediated. immunity such as T helper cells, B cells, neutrophils natural killer (NK) cells, and macrophages [15] . All the above facts are prompted us to design some compounds for COVID-19 with the help of structural knowledge of chloroquine and hydroxychloroquine (HCQ). In this case, the designed organic molecule should be capable of coordination with Cu 2+ to increase activity. In recent time, computer aided drug designing is a reliable and mostly used methodology [16] [17] [18] [19] . This technology can be utilized to find COVID-19 target protein for the designed Cu-complexes and explore the structure activity relationship of them. Synthesis of these molecules through a simple and cost-effective method are also very challenging task. Reagent-grade 2-(bromomethyl) benzonitrile and 8-hydroxyquinoline were purchased from Sigma Aldrich and used as received, Sodium borohydride, transition metal salt Co(NO 3 ) 2 .6H 2 O was acquired from Aldrich and used as received. The solvents and anhydrous K 2 CO 3 (Merck), potassium iodide, NaOH were obtained from Thermo Fisher scientific, India and used as without purification. All the solvents were purified prior to use following standard methods. Microanalyses for the compounds were performed using a CE-440 elemental analyzer (Exeter Analytical Inc.). Infrared spectra were obtained (KBr disk, 400-4000 cm -1 ) on a Perkin-Elmer Model 1320 spectrometer. ESI mass spectra were recorded on a WATERS Q-TOF Premier mass spectrometer. The NMR ( 1 H and 13 C-NMR) analysis were run on a Bruker Avance-300 spectrometer in DMSO-d 6 at room temperature. Thermal gravimetric analysis of complex 1 and 2 was determined using Mettler Toledo thermogravimetric analyzer (TGA) with built in gas controller (TGA2 SF/1100) and fitted with XP1U TGA balance (ultra-micro balance), in the temperature range 25-800 ºC under nitrogen atmosphere at a heating rate of 10 °C/min. The ligand L1 was synthesized according to the procedure reported earlier [20] as; 8hydroxyquinoline (2g, 13.7mmol) and dry K 2 CO 3 (5g, 36.17mmol) were mixed in a round-bottom flask under an inert atmosphere, and acetonitrile (60 mL) added to the above mixture. The mixture was allowed to stirring for 60 minutes at 90 °C . The mixture was treated with 2-(bromomethyl) benzonitrile (2.69g, 13.7mmol), and the resulting solution refluxed for 24h. After completion of this period, the solution was allowed to cool to room temperature and the mixture was poured slowly in ice cold water (100 mL) with constant stirring to give a white muddy solid precipitate that was collected by filtration and dried under vacuum. Block-shaped pale yellow crystals of L1 were collected from the slow evaporation of ethanolic solution after 2 weeks. Yield The ligand L2 was achieved by hydrolyzing L1 (2g, 7.68 mmol) by 6(N) NaOH solution (100 mL) under refluxing condition for 24h. The resulting solution was allowed to cool 5 0 C and acidified with the 6(N) HCl to obtained light pale yellow solid. This compound was collected by filtration A mixture containing L1 (0.02g), L2 (0.02g) and 8-hydroxyquinoline (Hq) (0.02g) was dissolved in 3mL methanol in a round bottom flask and added Cu(NO 3 ) 2 ·3H 2 O (0.08g) dissolved in 2mL water solution dropwise, then added 3 drops of NH 3 solution and allow to stirrer for 6h at 80 0 C. The resulting solution was cool down and filtered out, then kept to slow evaporation at room temperature. After 2-3 week green block shape crystals were obtained and washed with cold ethanol, which was suitable for data collection of single crystal X-ray. Yield: 75%. Melting Point: The diffraction data was collected with a Bruker SMART APEX CCD diffractometer using monochromatic Mo-Kα radiation (λ =0.71073 Å) at 100(2) K. The linear absorption coefficients, and the anomalous dispersion corrections were mentioned from the International Tables for X-ray crystallography [21] . The data integration and reduction were worked out with SAINT software [22] . Empirical absorption correction was applied to the collected reflections with SADABS [23] . Using Olex2 [24] , the structure was solved with the olex2.solve [25] structure solution program using Charge Flipping and refined with the olex2.refine [25] refinement package using Gauss-Newton minimisation. All hydrogen atoms were located in difference Fourier maps in the structures and refined isotropically. All non-H atoms were refined anisotropically. The crystal and refinement data are collected in Table S1 for ligands L1 and L2 and Table S2 for complexes 1 and 2, respectively. The selected bond distances and angles for ligands are given in Table S3 (for L1) and Table S4 (for L2) and Table S5 (for complex 1) and Table S6 (for complex 2). Energy minimized structure of chloroquinoline and HCQ, L1 and L2 which was obtained from The chloroquine and hydroxychloroquine are differ by an alkyl chain hydroxyl group (Fig. 1) . For protein binding of ligand, CH-π interaction play a vital role and it is also found in polar carbohydrate protein interactions [26] . Therefore, the 4-aminoquinoline moiety of these molecules is key part for the protein binding. However, the part is not suitable for ligation with metal. For this reason we choose 8-hydroxyquinoline. Both the chloroquine and hydroxychloroquine contain nine carbon side chain with hetero atoms. Considering these feature we replaced the side chain Table S7 . Crystal structure of ligand L1 and L2 was confirmed by the crystallographic data which data shows ligands crystallized in monoclinic, tetragonal crystal system with P2 1 /c and I4 1 /a space group, respectively. The asymmetric unit of both ligands have been shown non-planar structure with dihedral angle 79.570(32) 0 and 14.032(54) 0 for L1 and L2, respectively. 8-hydroxyquinoline moiety has planar geometry and 2-(Bromomethyl)benzonitrile is non-planar and hence, L1 crystallized as non-planar geometry. However, in L2, hydrolysis of nitrile group to convert into carboxylate group is approaching to planar molecule ( Fig. 2a and b) . Ligand L1 and L2 are comprises by same organic moieties with different functionality of groups. Ligand L1 is stabilized via hydrogen bonding (C17-N2…H10b= 2.669Å, C15-H15…N1= 2.691 Å, C17-N2…H6= 2.657 Å) and weak interaction such as π-π, C-H…π, give a arabesque like pattern along b-axis ( Fig. 3a and b) . Similarly, in ligand L2 is also stabilized via strong hydrogen bonding and other weak interaction such as O2-H2…O1= 2.111, C11-H11…O2= 2.639 Å, C12-H12…O1= 2.286Å hydrogen bonding, C-H…π (C17-H17…C13= 2.717 Å) and C3-H3…H16-C16= 2.346 Å weak interaction and form a herringbone(45 0 ) like supramolecular framework along c-axis ( Fig. 4a and b) . Mixing of Cu(NO 3 ) 2 ·3H 2 O, 8-hydroxyquinoline, L1 and L2 in 1:1:1:1 ratio under appropriate conditions, it give a pale yellow solid of complex 1 and by omitting L2 with 1:1:2 ratio under suitable reaction conditions the mixture produce the complex 2 (Fig. 5a) . Both the complexes were finally characterized with the help of single crystal X-ray crystallography. It is important to know that the drug-likeliness properties of all the aforesaid compounds and complexes before any screening. The results are summarized in the The RNA of SARS-CoV-2 translate two classes of proteins structural proteins and non-structural proteins. The first category consist with nucleocapsid, envelope, membrane and trimeric spike proteins. These proteins help to build and maintain the structure and shape of the virus play most important role in the infection process. Whereas the second category includes sixteen proteins which come in to existence from polyprotein PP1a and PP1b. They are also very essential for viral metabolism processes like translation. Therefore, any one of these proteins can be the target for all these compounds (chloroquine, hydroxychloroquine, L1, L2, complex 1 and 2). 6W02) and 9 (PDB: 6W4B) for the screening of these compounds. We have screened all the six compounds with the above mentioned seven viral proteins and the data associated to the binding energy is summarized in the unit. Therefore, the study shows that the complex 2 can be act as an anti-COVID-19 agent through the inhibiting the ADP-ribose-1 monophosphatase and main protease enzymes. However, in the case of HCQ, the side-chain hydroxyl group involved in the hydrogen bonding ( Fig. S15a and S15b). The ligand with aromatic moiety increases the hydrophobicity after ligation with Cu 2+ which may be favor to bind at the hydrophobicity loving active site of the protein. It is clear from the large green surface on the molecular electro static potential (MEP) which indicates that both the complexes are sufficiently hydrophobic in nature ( Fig. S13a and S13b). In the case of complex 1, the D22, G48, V49, A52, A129 and F156 amino acid residues are in the contact (Fig. 6a) . For complex 2, the total number interacting amino acids are higher with respect to the complex 1 and these amino acids are I23, A38, V49, L126, A129, A154, V155 and L160 (Fig. 6b) . It is very interesting to note that the mode of binding is only CH-π interactions between aromatic parts of the complexes and amino acids ( Fig. S15a and S15b). It should be mention here that the topological polar surface area (TPSA ; Table S7 ), the sum of surfaces of polar atoms in a molecule, is high in complex 2 with respect to complex 1. The complex 2 is slightly more polar with hydrophobic surface than complex 1. Therefore, an amphiphilic molecule 2 is good protein binder and hydrophobicity is not only the factor to it. In summary of this work, it can be state that we successfully designed and synthesized two copper complexes against COVID-19. In this work, we designed copper complexes with the help of structural information of the chloroquine and HCQ. We compared the drug likeness and pharmacophysical properties of the complexes with the organic molecules. Among various important structural and non-structural viral protein, we recognize the ADP-ribose-1 monophosphatase enzyme as a protein host for these investigated molecules. Complex 2 show strong binding affinity for the protein. Therefore, considering all the results, it is clear that the Complex 2 have enormous potential as an anti-COVID-19 agent. There are no conflicts of interest. The species Severe acute respiratory syndrome-related coronavirus: classifying 2019-nCoV and naming it SARS-CoV-2 COVID-19 pathophysiology: A review Current drugs with potential for treatment of COVID-19: A literature review Metalchloroquine derivatives as possible anti-malarial drugs: evaluation of anti-malarial activity and mode of action Does zinc supplementation enhance the clinical efficacy of chloroquine/hydroxychloroquine to win today's battle against COVID-19? In situ reactivity of electrochemically generated nitro radical anion on Ornidazole and its monomeric Cu(II) complex with nucleic acid bases and calf thymus DNA Structural and some medicinal characteristics of the copper(II)-hydroxychloroquine complex Novel copper(II) complex of N-Propyl-norfloxacin and 1,10-Phenanthroline with enhanced antileukemic and DNA nuclease activities Release studies from smart hydrogels as carriers for piroxicam and copper(II)-oxicam complexes as antiinflammatory and anti-cancer drugs. X-ray structures of new copper(II)-piroxicam andisoxicam complex molecules The many "faces" of copper in medicine and treatment Multitargeting antibacterial activity of a synthesized Mn 2+ complex of curcumin on grampositive and gram-negative bacterial strains Advances in copper complexes as anticancer agents Spectroscopy driven DFT computation for a structure of the monomeric Cu 2+ -curcumin complex and thermodynamics driven evaluation of its binding to DNA: Pseudo-binding of curcumin to DNA COVID-19: The Potential role of copper and N-acetylcysteine (NAC) in a combination of candidate antiviral treatments against SARS-CoV-2 Is copper beneficial for COVID-19 patients? Computer-aided drug design against spike glycoprotein of SARS-CoV-2 to aid COVID-19 treatment COVID-19: CADD to the rescue Role of computer-aided drug design in modern drug discovery A computer-aided drug design approach to predict marine druglike leads for SARS-CoV-2 main protease inhibition Evaluation of catacholase mimicking activity and apoptosis in human colorectal carcinoma cell line by activating mitochondrial pathway of copper(II) complex coupled with 2-(quinolin-8-yloxy)(methyl)benzonitrile and 8-hydroxyquinoline International Tables for X-Ray Crystallography SAINT, version 6.02; Bruker AXS: Madison, WI Empirical Absorption Correction Program OLEX2 : a complete structure solution, refinement and analysis program The anatomy of a comprehensive constrained, restrained refinement program for the modern computing environment -Olex2 dissected For TOC TOC synopsis: Inhibition of ADP-ribose-1 monophosphatase enzyme of COVID-19 by a copper complex can inhibit coronavirus infection. For TOC TOC synopsis: Inhibition of ADP-ribose-1 monophosphatase enzyme of COVID-19 Highlights  New quinoline based Cu(II)-complexes were designed and synthesized as anti-Covid-19  In silico screening revealed that complexes display excellent SARS CoV-2 M pro inhibitory activity Complex 2 is a good protein binder than 1 due to more polar with hydrophobic surface  Complex 2 have enormous potential as an anti-COVID-19 agent The Department of Chemistry, Jadavpur University has gratefully acknowledged for research Supplementary crystallographic data for ligands L1 and L2 and complex 1 and 2, respectively. ☒ The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.☐The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:There are no conflicts of interest.