Molecular Docking Study for Binding Affinity of 2H-thiopyrano[2,3-b]quinoline Derivatives against CB1a

Quinoline-based molecules are major constituents in natural products, active pharmacophores, and have excellent biological activities. Using 2H-thiopyrano[2,3-b]quinoline derivatives and CB1a protein (PDB ID: 2IGR), the molecular docking study has been revealed in this article. The study of in silico molecular docking analysis of such derivatives to determine the binding affinity, residual interaction, and hydrogen bonding of several 2H-thiopyrano[2,3-b]quinolines against CB1a is reported here. The current work demonstrated that 2H-thiopyrano[2,3-b]quinoline derivatives could be effective antitumor agents to produce potent anticancer medicines in the near future.


Introduction
Cancer is marked by the uncontrolled, rapid, and pathological growth of improperly altered cells and used in a broader sense. Despite considerable advancements in cancer therapy in recent times, cancer remains the world's second biggest cause of death, trailing only cardiovascular (CVS) diseases [1]. Chemotherapeutic drug resistance is still a major issue in the fght against cancer; in addition, chemotherapy is further hampered by a lack of selectivity. Anticancer medications, in general, kill both healthy and cancerous cells, and they frequently have major side efects. Many eforts have been undertaken to fnd new chemotherapeutic medicines with low side efects and to develop safe and efective strategies to treat this disease [2]. Among all cancer types, cervical cancer has the world's fourth most frequent malignancy among women, with the third highest fatality rate [3]. Cervical cancer alone accounts for about 12% of all cancers in women worldwide, according to a WHO (World Health Organization) report [4], and it is more common in developing nations. Until now, chemotherapeutic medication treatment for cervical cancer has been associated with a poor prognosis and a slew of negative side efects [5]. Terefore, scientists are working for the development of new safer chemicals having potent anticancer activity. For the identifcation of efcient cellular targets [6], early detection is required which improves the efcacy of the available chemotherapeutic approaches in the feld of cancer research [7,8].
Te quinoline core is a signifcant structural unit in many naturally occurring compounds [26][27][28][29], with interesting biological activity, and many conventional medications. Quinoline core having substitutions at diferent positions has shown substantial anticancer efcacy against a variety of targets, including topoisomerase I [30], tubulin [31], protein kinase [32], and so on. Some well-known quinolone-based anticancer medicines have been approved by the FDA (Food and Drug Administration) e.g., camptothecin, irinotecan, topotecan, and others. Furthermore, few such quinoline-based anticancer drugs are still in clinical testing phase such as bosutinib, lenvatinib, cabozantinib, and farnesyltransferase inhibitors (tipifarnib) [33].
Several well-known compounds having 2H-thiopyrano [2,3-b]quinolines can be synthesized in a highly efcient manner and create one C-S and one C-C bond with a high extraction economy, likely via a Michael addition, intramolecular aldol-dehydration domino process. It is common knowledge that combining pharmacophore units with different biological activities results in a new hybrid entity with higher biological processes and efcacy than the parent medications. Based on the previous discoveries, unique bioactive hybrid compounds based on quinoline having better anticancer efcacy have been developed nowadays.
In this article, we have reported the binding afnity and several interactions of compounds 1-4 (as shown in Figure 2) with the anticancer-based protein CB1a (CB1a, a novel anticancer peptide derived from natural antimicrobial peptide cecropin B: PDB ID: 2IGR). Te values of binding afnity for 1-4 lies between −5.3 and −6.1 Kcal/ mol. Te interaction also shows that compound 4 shows highest binding afnity (−6.1 Kcal/mol) as compared to other derivatives of thiopyrano [2,3-b]quinolines. For the docking study, we used several software packages such as AutoDock vina 4, discovery studio, and protein-ligand interaction profler. Tese software packages help in the study and analysis of docking position, docking size, binding afnity, energy range as well as exhaustiveness. Several amino acids are attached with ligands (thiopyrano
Singh et al. revealed the synthesis of thiopyrano[2,3-(b)] quinoline derivatives by using 3-formyl-quinoline-2-thione derivative and acrylonitrile in the presence of cheap base triethylamine (TEA) and dimethyl formamide (DMF) at room temperature [36]. It is a rapid and efcient one-pot reaction for the synthesis of 3-cyanothiopyrano[2,3-(b)] quinoline derivatives (Scheme 2). Te whole reaction was synthesized very easy by the Domino Michael addition followed by cyclization. Kumar

Molecular Docking Studies
Cecropins are a category of antimicrobial peptides abundantly available in Hyalophora cecropia's immunological hemolymph. Te N-terminal regions of native cecropins include basic residues, while the C-terminal parts have hydrophobic residues. Cecropin B (CB) has the most potent antibacterial properties and to examine the efects on cells and synthetic liposomes, CB derivatives cecropin B1 (CB1) and cecropin B3 (CB3) were developed. CB1 was made by replacing the C-terminal segment with CB's N-terminal sequence, while CB3 was made by replacing the Nterminal segment with CB's C-terminal sequence. CB and its equivalents have been shown in previous research to break membranes, and some of them can destroy cancer cells.
Preliminary SAR interpreting has been conducted using various methods such as molecular docking modelling and docking studies, where the interpretability is considerably better explicit. In this approach, pharmacophore techniques can provide a signifcant beneft and a clearer look at the structural elements that contribute to the structure activity relationship (SAR) [38]. In this study, pharmacophore generation was discovered using AutoDock vina 4 (Te Scripps Research institute) [39] and discovery studio.
Te scoring of ligands (binding afnity, ligand internal energy, and distance) is described in the result and discussion section. Using the default settings, the "proteinligand interaction profler" [40] calculated the binding affnity of ligands in stable ligand-protein complexes required for ligand binding with the receptor.
Te molecular docking study shows that the value of dielectric constant is −0.1465 and the binding spacing is 0.375. Te default setting of exhaustiveness is 8, and the RMSD values are calculated relative to the best mode and use only movable heavy atoms. Two variants of RMSD metrics are provided, rmsd/lb (RMSD lower bound) and rmsd/ub (RMSD upper bound). In addition to this, the energy of binding modes in the output is 4. Te "protein-ligand interaction profler" (PLIP) was used to calculate the interaction of protein and ligands in the stable ligand-protein complex required for ligand binding to the receptor.   Based on molecular docking study of four compounds having thiopyrano[2,3-b]quinoline core 1-4 against CB1a (PDB ID: 2IGR). Te docking result is given in Table 1.
In general, the IC 50 of CB1a, CB, melittin, magainin II, as well as the cancer chemotherapeutic agent were determined using cytotoxicity assays on cancer and noncancer cells. CB1a has a stronger cytotoxic activity (lower IC 50 values) against leukaemia cells and stomach cancer than CB and magainin II. CB1a and CB are highly efective towards cancerous cells but not normal human cells. CB1a has an 8fold and 2-3-fold higher cytotoxic activity than CB against AGS and leukaemia cell lines, respectively. As a result, molecular docking study of the synthesized compounds was performed in the current study to explore their binding pattern with a unique anticancer peptide derived from the natural antimicrobial peptide cecropin B (CB1).
Because of its excellent selectivity, CB1a can be developed as a powerful anticancer drug. CB1a > CB1 > CB > CB3 is the  order of anticancer activity of CB analogues against tumor cell lines [41]. Terefore, for this study, CB1a, a novel anticancer peptide derived from natural antimicrobial peptide cecropin B, was used. Te synthetic compounds (1-4) were found to have binding afnity from −5.3 to −6.1 kcal/mol (Table 1) with the best result achieved using compound 4 (−6.1 kcal/ mol). Te hydrogen bond, residual interaction, and pi-pi interaction of the four compounds were summarized in (Table 1).
Te compounds ( Figure 6. Te in-silico interaction of compound 4 results matches the in vitro analysis of the synthesized compounds against the structure of CB1a, which shows high binding afnity and good activity as compared to other derivatives since the value is −6.1 kcal/mol.

Conclusion
Te current work demonstrates a useful study to fnd the possible efective antitumor agents having thiopyrano [2,3-b]quinoline derivatives, which could produce potent anticancer medicines in near future. To study the interaction of ligand and protein, several software packages were used in this work such as AutoDock vina 4, discovery studio, and protein-ligand interaction profler (PLIP). Te molecular docking of compounds having thiopyrano[2,3-b]quinoline core and its derivatives against CB1a (PDB ID: 2IGR) were studied in this article. Te in silico molecular docking study of these compounds reveals that these frameworks have potential bioactivity having high binding afnity, adequate residual interaction, and hydrogen bonding interaction against the protein CB1a. Te binding afnity value for thiopyrano [2,3-b]quinoline and its derivatives 1-4 is