Unveiling the anti-inflammatory potential of organoselenium Schiff bases: computational and in vitro studies

Saad Shaaban *ab, Tarek A. Yousef c, Hanan A Althikrallah a, Yasair S. Al-Faiyz a, Ibrahim Elghamry a, Marwa Sharaky d, Radwan Alnajjar ef and Ahmed A. Al-Karmalawy *gh
aDepartment of Chemistry, College of Science, King Faisal University, Al-Ahsa 31982, Saudi Arabia. E-mail: sibrahim@kfu.edu.sa
bDepartment of Chemistry, Faculty of Science, Mansoura University, 35516 Mansoura, Egypt
cDepartment of Chemistry, College of Science, Imam Mohammad Ibn Saud Islamic University (IMSIU), Riyadh 11623, Saudi Arabia
dCancer Biology Department, Pharmacology Unit, National Cancer Institute (NCI), Cairo University, Cairo, Egypt
eCADD Unit, PharmD, Faculty of Pharmacy, Libyan International Medical University, Benghazi, Libya
fDepartment of Chemistry, Faculty of Science, University of Benghazi, Benghazi, Libya
gDepartment of Pharmaceutical Chemistry, College of Pharmacy, The University of Mashreq, Baghdad 10023, Iraq. E-mail: akarmalawy@horus.edu.eg
hPharmaceutical Chemistry Department, Faculty of Pharmacy, Horus University–Egypt, New Damietta, 34518, Egypt

Received 7th October 2024 , Accepted 20th November 2024

First published on 20th November 2024


Abstract

Novel organoselenium (OSe) Schiff bases (3a–c and 5a–c) were designed and synthesized by a simple and convenient approach to obtain good to excellent yields (up to 87%). Their chemical structures were confirmed by different spectroscopic and spectrometric techniques (FT-IR, NMR, and MS). The anti-inflammatory potential of the synthesized OSe Schiff bases (3a–c and 5a–c) was assessed by the protein expression analysis of inflammation-related genes. Interestingly, the anti-inflammatory activity of OSe Schiff bases 3a–c and 5a–c was confirmed by the produced downregulation of the inflammatory proteins (COX-2, IL-6, and IL-1β) by (0.43, 0.52, 0.49, and 0.36), (0.65, 0.54, 0.66, and 0.50), and (0.50, 0.44, 0.56, and 0.50)-fold-change, respectively. Furthermore, the binding affinities of the newly designed candidates were examined against the COX-2 receptor using a molecular docking approach. The compounds showed promising binding scores and exhibited similar binding modes as the co-crystallized inhibitor of COX-2 as confirmed through 200 ns molecular dynamic (MD) simulations and MM-GBSA calculations. The DFT study analyzed the electronic properties of compounds 3a–c and 5a–c, revealing that compound 3c exhibited the lowest energy gap and highest stability, while compound 5a demonstrated the greatest electron affinity. Notably, this study showed that the synthesized OSe Schiff bases 3a–c and 5a displayed a significant anti-inflammatory activity, highlighting their potential in biomedical applications.


1. Introduction

Inflammation is a serious physiological process that is initiated by the immune system due to a response to damage or infection. It involves the secretion of a series of mediators, with its duration being influenced by a range of factors and substances that may possess either pro-inflammatory or anti-inflammatory properties.1–3 Inflammation is a key protective mechanism crucial for safeguarding the body and facilitating the restoration of tissues to their normal state.4,5 Prolonged inflammation can stimulate the production of growth factors and cytokines, resulting in DNA and tissue damage, which may contribute to the development of various diseases. Within this context, OS and inflammation play critical roles in the pathogenesis of numerous diseases.6,7 Both processes are interconnected and can exacerbate each other, leading to tissue damage and disease progression.8–10

Since its discovery by Berzelius in 1817, researchers have been bewildered by the exceptional behavior of selenium.11,12 It is a unique element with a dual nature. Initially considered as a toxic substance, it is now recognized as an essential micronutrient, playing a critical role in various biophysical processes across a wide range of living organisms.13,14 It belongs to the chalcogen (i.e., oxygen) group and serves a significant function in the immune system's defense process and prevention of various malignancies. As expected, Se deficiency is often linked with the development of various infectious and autoimmune illnesses.11,15 Se is also essential for several intracellular biochemical pathways.12 It is incorporated into the structure of various proteins known as selenoproteins, where it is present as a part of the amino acids selenocysteine (I) and selenomethionine (II) (Fig. 1) that play a central role in the removal of free radicals.13


image file: d4nj04376f-f1.tif
Fig. 1 Organoselenium agents with promising biological activities (I–IX).

In contrast to phosphorous, oxygen, and sulfur, Se has a larger size and lower electronegativity, which endows it with higher polarizability, electron-donor ability, and nucleophilicity.16 Furthermore, most of the OSe compounds usually exhibit better biological activities than their S analogs.17 The latter is due to the enhanced amphiphilicity and pharmacokinetics of some of the OSe compounds.18 Accordingly, Se has attracted the interest of many researchers owing to its unique optoelectronic properties. Therefore, it has recently been involved in different industries, including supercapacitors, rectifiers, sodium-ion batteries, photovoltaic cells, photographic auxiliary devices, and emitting and transmitting devices. Furthermore, organoselenium compounds (OSe) have exhibited much success in the management of several disorders (e.g., stroke, diabetes, and cancer), and several OSe agents are currently in different clinical trials. For instance, ebselen (III) is the most common selena heterocyclic compound.19 It manifested promising anti-inflammatory, antioxidant, and chemopreventive activities, and is being evaluated in clinical phase two as a possible anticancer and anti-hypo/manic drug in bipolar patients.20 Furthermore, ethaselen (IV) is another selena heterocyclic compound that is a potential antineoplastic drug, and is being evaluated in clinical phase one as a therapeutic agent for non-small cell lung tumours (Fig. 1).21,22

Moreover, selenocoxib-3 (V), a seleno derivative of the nonsteroidal anti-inflammatory drug celecoxib, has shown enhanced activity and less toxicity.23 1,2-Diphenyldiselane ((Ph)2Se2) (VI) and its derivatives share various antioxidant and inflammatory properties with ebselen, and they also manifested good mimicry of the glutathione peroxidase (GPx) and thioredoxin reductase enzymes.22,24,25 Similarly, the N-maleanilic-based OSe acid (VII), synthesized in our laboratory, exhibited excellent GPx-like and anti-apoptotic properties in oligodendrocytes.26,27 Additionally, our OSe pseudopeptides (VIII and IX) have shown interesting cytoprotective activities for oligodendrocytes (Fig. 1).28,29

On the other hand, Schiff bases are widely recognized organic scaffolds with excellent coordination potential to various transition metals, and are therefore used extensively in pharmaceutical and medicinal research.30–33 They are easily accessible from the reaction of aldehydes/ketones with primary amines to afford the respective azomethines (–CH[double bond, length as m-dash]N–), and have shown a broad range of bioactivities (e.g., antimicrobial, antiviral, and anticancer).34–37 The latter is attributed to their ability to form hydrogen bonds with various enzymes and proteins, and therefore alter their activities. To this end, OSe-based Schiff bases have recently emerged as a distinct class of ligands that have both ‘soft’ and ‘hard’ donor sites, and act readily as hemilabile chelates.38–42

Very recently, we described OSe-based Schiff bases as potential inhibitors of the MPro, a protein involved in replicating SARS-CoV-2. Our results demonstrated that these agents hold promise as potential MPro inhibitors, and might be developed as antiviral drugs in the future.30 Computational calculation investigations suggested that these compounds hold promise as effective MPro inhibitors with favourable pharmacokinetic profiles and drug-like characteristics. Notably, these studies were limited to theoretical calculations, and no biological investigations were performed to evaluate the bioactivity of these OSe-based Schiff bases.

Moreover, these compounds have shown potential apoptosis induction in colorectal carcinoma via upregulation of P53, BAX, and caspases 3, 6, 8, and 9 and downregulation of MMP2, MMP9, and BCL-2, assuring the apoptotic potentials.43

As this is an enormously unknown group of compounds, nothing is known about their anti-inflammatory potential and their biological targets. Furthermore, there is ample evidence in the literature indicating that OSe compounds (diorganyl diselenides, for instance, and particularly, diphenyl diselenide) have a wide range of applications as anti-inflammatory agents, and some of these compounds have already entered clinical trials, e.g., ebselen and ethaselen.29,44–47 In continuation of our previous work, our objectives were expanded further to investigate the underlying anti-inflammatory activities of these compounds to determine whether these candidates may be useful for further medicinal studies in the future.

Encouraged by the promising anticancer and antiviral activities, and by the fact that some anticancer agents are (on occasion) good anti-inflammatory candidates, we herein aim to investigate the anti-inflammatory activities using inflammation-related proteins such as COX-2, IL-6, and IL-1β. Ultimately, a study using quantum chemical calculations was performed to understand the chemical properties of the synthesized compounds, which were subsequently compared with the experimental data.

1.1. Rationale of the work design

The diselenide (Se–Se) functionality has attracted much interest as it is often present among the metabolites of nutritional compounds, and it possesses unique redox-cycling properties.48 Accordingly, diselenides manifested good anti-inflammatory activities. Furthermore, they share common biochemical characteristics with ebselen, as they can readily react with the –SH groups (for instance, from the cysteine amino acid found in enzymes or proteins) to form selenosulfide (–Se–S–), selenol (–SeH), or disulfide (–S–S–).49,50 Moreover, (Ph)2Se2, the simplest diselenide compound, manifested diverse pharmacological properties, including antioxidant, neuroprotective, and anti-inflammatory activities.51

(Ph)2Se2 has many physicochemical restrictions arising from its low bioavailability, which hinders the administration of parenteral solutions. Furthermore, it has some toxicity problems (e.g., off-target inhibition of δ-aminolevulinate acid dehydratase) that hinder its pharmaceutical applications.52,53 Structural alterations, such as an attachment of extra functional groups or conjugation with a receptor binding affinity moiety, can be used to circumvent these challenges.54–57

Inspired by the above-proven scientific facts and based on the chemical structure of diphenyl diselenide as a lead compound, we tried to optimize the anti-inflammatory activity, and decrease the side effects and toxicities as much as possible. This was achieved by the addition of a p-NH2 group at each phenyl ring (2), which was then reacted with an appropriate aldehyde to form the corresponding Schiff base (3a–3c). Therefore, the N atom at each phenyl ring could act as a hydrogen bond acceptor to improve the receptor binding affinity. Moreover, the additional aromatic moiety at each side will improve the hydrophobic interactions within the receptor active site. Different substitutions have been attached to the aryl ring of the Schiff base moiety to investigate the impact on the activity of the designed Schiff bases-tethered OSe compounds (Fig. 2). This was done to obtain new anti-inflammatory candidates that would be superior to the parent diphenyl diselenide.


image file: d4nj04376f-f2.tif
Fig. 2 Design of the new Schiff base-tethered OSe compounds as promising anti-inflammatory candidates through lead optimization strategy.

The aminophenyl diselenide scaffold (2) was further converted to a mono-atomic selenium connected to a methyl group at one end, while keeping one aniline ring at the other side (4). After that, compound 4 was modified to a mono-atomic selenium linker between one Schiff base moiety and a methyl group (5a–5c) to further investigate the structure–activity relationship (Fig. 2).

2. Results and discussion

2.1. Chemistry

The synthesis of OSe compounds is increasingly growing, inspired by their promising chemopreventive and antioxidant activities.15,58 Accordingly, researchers' trials have recently been directed towards the development of efficient protocols to develop novel OSe compounds and explore their potential bioactivities.59,60 Despite the substantial progress in the preparation of these compounds, their development has often encountered several synthetic challenges. Among these are the use of toxic, sensitive, and expensive OSe starting materials (e.g., PhSeBr) and reagents (e.g., SeCl4, Se(SiMe3)2, KSeCN, SeF6, and SeOCl2).61–63 Therefore, the development of straightforward yet simple synthetic protocols using stable and less toxic building blocks is highly desired. Within this context, diaryl diselenides are frequently utilized as starting building blocks for the preparation of numerous OSe scaffolds.64 Diorganyl diselenides provide access to different OSe precursors, including organyl selenide halide via halogenation and selenenic, seleninic, and selenonic acids via oxidation, as well as various Se-based reactive species (e.g., RSe˙, RSe, and RSe+).46,63,65,66

Similarly, the potent medicinal properties (e.g., anti-inflammatory, anticancer, and chemopreventive) exhibited by Schiff bases have recently attracted our attention.67–69 It is presumed that a proper formulation of OSe-based hybrids might have enhanced the anti-inflammatory activity compared to their individual OSe or Schiff base precursors.

Diaminodiphenyl diselenide (2) was extensively used as a dynamic selenylating reagent due to its straightforward preparation, simple handling, and stability.64 Therefore, it was used as the starting building block in our synthetic strategy (Scheme 1). Bis(4-aminophenyl)diselenide (2) was obtained by the alkaline hydrolysis of 4-selenocyanatoaniline 1. The reaction of OSe compound (2) with different carbonyl compounds, namely, 4-formylbromobenzaldhyde, 4-formylflourobenzaldhyde, and 2-nitrobenzaldehyde, proceeded with very good yields (up to 87%) to get the target Schiff bases (3a–c). Furthermore, the reduction of diaminodiphenyl diselenide 2 using NaBH4 and reaction with CH3I afforded 4-(methylselenyl)aniline (4) in 57% yield. Likewise, the condensation of 4 with various aldehydes, namely, formyl-2-naphthol, 4-formylflourobenzaldhyde, and 2-nitrobenzaldehyde, furnished the respective Schiff bases (5a–c) in good yields (up to 86%).


image file: d4nj04376f-s1.tif
Scheme 1 Psreparation of organoselenium compounds (2–5).

The structures of all the synthesized Schiff bases and their precursors were confirmed by the spectroscopic and spectrometric techniques (FT-IR, 1H-NMR,13C-NMR, and Ms). Therefore, the FT-IR of the target Schiff bases (3a–c and 5a–c) revealed the absence of the characteristic amino groups (NH2) signals of the precursors 2 and 4. Also, these were confirmed by the 1H-NMR spectra of compounds 3a–c and 5a–c, which confirmed the absence of the signals for the amino groups’ protons at δ 3.70 and 3.72 ppm for 2 and 4, respectively. Additionally, the appearance of the characteristic Schiff bases anil structure (–N[double bond, length as m-dash]C[H with combining low line]) singlet signal at δ 8.63, 8.63, 8.85, 9.65, 8.64 and 8.88 ppm for both 3a–c and 5a–c respectively. Furthermore, the 13CNMR spectra of the OSe Schiff bases 3a–c and 5a–c showed the characteristic (–N[double bond, length as m-dash][C with combining low line]H) signals at 160.75, 160.54, 157.91, 155.65, 159.56, and 156.67 ppm, respectively. Furthermore, the 1HNMR spectra of the OSe Schiff bases 5a–c showed distinct singlet signals for the CH3 protons (C[H with combining low line][3 with combining low line]Se) at 2.39, 2.39, and 2.40 ppm, respectively (Supplementary Materials, SI1, ESI).

2.2. Biological evaluation

2.2.1. Protein expression of the inflammation-related genes. To examine the anti-inflammatory effects of the Schiff base-tethered OSe compounds (3a, 3b, 3c, and 5a) on the HCT116 cancer cell line, their IC50 values (6.00, 5.00, 5.43, and 5.73 µg mL−1, respectively) were utilized.43 A protein expression analysis focusing on inflammation-related genes was conducted, evaluating the levels of COX-2, IL-6, and IL-1β in both treated and untreated cells. The goal was to detect any changes in protein expression and to understand the molecular mechanisms underlying the anti-inflammatory effects of compounds 3a, 3b, 3c, and 5a. Notably, treatment with compounds 3a, 3b, 3c, and 5a induced downregulation of inflammatory proteins. Accordingly, COX-2, IL-6, and IL-1β were downregulated by (0.43, 0.52, 0.49, and 0.36), (0.65, 0.54, 0.66, and 0.50), and (0.50, 0.44, 0.56, and 0.50)-fold-change, respectively, assuring the anti-inflammatory potential, as depicted in Fig. 3.
image file: d4nj04376f-f3.tif
Fig. 3 Anti-inflammatory potential of compounds 3a, 3b, 3c, and 5a against (A) COX-2, (B) IL-6, and (C) IL-1β.

Generally, the bifunctional Schiff bases (3a–3c) were found to be superior to the monofunctional one (5a), which may be attributed to their greater ability to form extra hydrogen bonds and hydrophobic interactions within the receptor pocket.

2.3. Computational studies

2.3.1. Molecular docking. The affinities of the newly designed candidates were examined against the COX-2 receptor using a molecular docking approach. Observing the binding site of the target COX-2 receptor (https://www.rcsb.org/structure/3LN1), it was noted that Leu338 and Ser339 are the essential amino acids responsible for the antagonistic activity of the COX-2 receptor.

Compounds 3a, 3b, 3c, and 5a were found to be superior COX-2 inhibitory candidates with −6.46, −5.95, −5.94, and −6.37 kcal mol−1 binding scores, compared to the co-crystallized inhibitor (−10.02 kcal mol−1). Compound 3a comprised two hydrogen bonds with Gln178 and Ser516. Compound 3b showed one hydrogen bond with Met508 and one pi-hydrogen bond with Ser339. Besides, compound 3c represented two pi-hydrogen interactions with Ser339 and His342. However, the 5a derivative formed only a pi–hydrogen interaction with Ser339 of the COX-2 binding site. Furthermore, the co-crystallized inhibitor of the COX-2 receptor bound both Ser339 and Leu338 with two hydrogen bonds (Fig. 4).


image file: d4nj04376f-f4.tif
Fig. 4 3D Interactions and orientations of 3a, 3b, 3c, 5a, and the co-crystallized inhibitor within the binding pocket of the COX-2 receptor (PDB ID: 3LN1).

Based on the above, similar binding modes and promising binding scores suggest that the examined candidates have potential inhibitory activity against COX-2, a potential target for induced anti-inflammatory activity.

2.3.2. Molecular dynamic simulation. Since molecular docking lacks the movement of the protein, its results are not reliable. Hence, molecular dynamic (MD) simulations were implemented for 200 ns to study the compounds that showed superior docking scores to the co-crystal inhibitor of COX-2. MD mimics the cell environment, where water is used as the explicit solvent, and the protein's charge is equalized with Na and Cl ions. Initially, the root mean square deviation (RMSD) of the Cα atoms in the protein backbone is tracked relative to its starting position and plotted against the simulation time (Fig. 5). As seen in Fig. 5, most proteins showed a stable RMSD. The conformation of the protein structure was moved by 3.00 Å at the beginning of the simulation due to relaxation, and it kept the relaxed conformation toward the end of the simulation without any visual fluctuation of the protein structure, which indicates great stability.
image file: d4nj04376f-f5.tif
Fig. 5 The RMSD of the Cα atoms in the protein backbone as a function of the simulation time.

Subsequently, the RMSDs of the ligands within the protein's active site were tracked relative to their initial positions, and plotted as a function of simulation time (Fig. 6). Fig. 6 depicts the ligand behaviour inside the active site through simulation time. Compound 3a showed the best stability among the ligands with an RMSD of 3.00 Å throughout the simulation time. Compound 3b, on the other hand, was the most unstable compound. At the beginning of the simulation, its RMSD jumped to around 7.00 Å before it settled down to 3.00 Å at around 20–40 ns, then reached 5.00 Å again at 50 ns and kept fluctuating up and down between 3.00 and 7.00 throughout the simulation. Compound 3c was also stable; it moved by 4.00 Å at the beginning of the simulation and held this position until the end of the simulation. Compound 5c also showed similar behaviour to compound 3c, where it moved gradually until it reached 4.5 Å at 40 ns, and held its position toward the end of the simulation. Finally, the co-crystal showed the best stability with a solid RMSD of less than 2.00 Å during the simulation trajectories.


image file: d4nj04376f-f6.tif
Fig. 6 The RMSDs of the ligands 3a, 3b, 3c, 5a, and co-crystal within the active site of the protein as a function of simulation time (ns).

Furthermore, compounds 3a, 3c, and 5a were subject to residue decomposition analysis, and their interactions were studied in detail. As depicted in Fig. 7 and Table 1, most compounds formed hydrophobic interactions, such as π–π and vdW interactions. Compound 3a formed hydrophobic interactions with Ala502, Phe504, Val509, and His75, along with two water-bridge H-bonds toward Ser339 and Tyr341. Compound 3c forms a strong hydrophobic interaction with Phe504, Try371, and Val509, along with 30% H-bond toward Gln178. Finally, compound 5a forms multiple weak hydrophobic interactions, as seen in Fig. 7.


image file: d4nj04376f-f7.tif
Fig. 7 Protein–ligand contacts for (A) 3a, (B) 3c, and (C) 5a during the simulation time are represented as bars, which present the percentage of interactions as interaction fractions.
Table 1 Prime MM-GBSA energies for ligands in complex with the COX-2 protein (in kcal mol−1)
Compounds ΔG Bind Coulomb Covalent H-bond Lipo Packing Solv_GB vdW
Coulomb: Coulomb energy; Covalent: covalent binding energy; H-bond: hydrogen-bonding energy; Lipo: lipophilic energy; Packing: pi–pi packing energy; Solv_GB: generalized born electrostatic solvation energy; vdW: van der Waals energy.
3a −58.66 −0.78 1.89 −0.01 −24.29 −2.72 22.44 −55.18
3b −64.70 −9.72 2.32 −0.20 −25.63 −1.50 23.10 −53.07
3c −88.60 −1.04 1.91 −0.26 −34.20 −5.26 17.47 −67.22
5a −53.62 −3.70 1.07 −0.32 −21.86 −0.94 13.74 −41.62
Co-crystal −60.08 −19.65 0.95 −1.82 −18.60 −0.95 22.30 −42.32


Notably, the MM-GBSA calculations clarified that the examined compounds showed comparable ΔG binding energies to the co-crystallized inhibitor, especially compounds 3b and 3c (−64.70 and −88.60) kcal mol−1, which exceeded that of the co-crystallized inhibitor (−60.08 kcal mol−1), respectively (Table 1).

2.3.3. DFT studies. The optimized structures of the synthesized compounds 3a, 3b, 3c, and 5a were generated using the DMOL3 program within the Materials Studio package, with the resulting structures depicted in Fig. 8. The values of the highest occupied molecular orbital (HOMO) and lowest unoccupied molecular orbital (LUMO) were analyzed to evaluate charge transfer within the molecules. The energy gap (EHOMOELUMO) offers insights into the compounds' chemical reactivity and kinetic stability. The HOMO acts as the electron donor, while the LUMO functions as the electron acceptor. Taken together, they are known as frontier molecular orbitals (FMOs). By examining the HOMO and LUMO energy levels, various other chemical properties70 were calculated, including ionization potentials (I), electron affinities (A), chemical softnesses (S), chemical hardnesses (η), electronic chemical potentials (μ), electronegativities (χ), and the global electrophilicity index (ω).70–72 The calculated values for these properties are summarised in Table 2.
image file: d4nj04376f-f8.tif
Fig. 8 Optimized structures and spatial electron distribution of HOMO and LUMO of compounds 3a, 3b, 3c, and 5a.
Table 2 E HOMO, ELUMO, and molecular descriptors of compounds 3a, 3b, 3c, and 5a
Molecule E HOMO (eV) E HOMO (eV) ΔE (eV) I (eV) A (eV) η (eV) S (eV) χ (eV) μ (eV) ω (eV)
3a −4.25 −2.74 1.51 4.25 2.74 3.50 0.66 0.76 −3.50 8.09
3b −4.16 −2.76 1.40 4.16 2.76 3.46 0.71 0.70 −3.46 8.55
3c −4.74 −3.37 1.37 4.74 3.37 4.06 0.73 0.69 −4.06 12.00
5a −4.20 −2.18 2.02 4.20 2.18 3.19 0.50 1.01 −3.19 5.04


Smaller gaps between the EHOMO and ELUMO indicate greater chemical stability and facilitate charge transfer, enhancing biological activities. In this study, the energy gaps for the synthesized compounds 3a, 3b, 3c, and 5a were ranked as follows: 5a > 3a > 3b > 3c. Consequently, compound 3c, with the lowest energy gap (Egap = 1.37 eV), is the most stable candidate.

The ionization potential (I) of a compound is influenced by the HOMO energy, with higher HOMO energies correlating with enhanced electron donation. In this analysis, compound 3b exhibited the lowest ionization potential and the highest HOMO energy (I = 4.16 eV), indicating its strongest electron-donating capability among the compounds. Conversely, the electron affinity (A) of the compounds is associated with LUMO energy, where lower LUMO energies correspond to better electron acceptance. Here, compound 5a displayed the highest electron affinity (A = 2.18 eV) and the lowest LUMO energy, suggesting that it is an effective electron acceptor.

Additionally, the electronic chemical potential (μ) reflects the system's energy sensitivity to changes in electron count. In this study, compound 3c showed a more negative electronic chemical potential value (μ = −4.06 eV). Chemical hardness, which measures the resistance to charge transfer, was determined for the synthesized compounds in the following order: 3c > 3b > 3a > 5a. Chemical softness, being the inverse of hardness, follows the opposite trend.

Throughout this study, compounds 3a, 3b, 3c, and 5a exhibited the highest anti-inflammatory activity compared to the other derivatives. Furthermore, the spatial distribution of electrons in the HOMO and LUMO was calculated, clearly demonstrating the electron donation capabilities. The spatial electron distribution for each compound (3a, 3b, 3c, and 5a) is depicted in Fig. 8.

3. Conclusions

Inspired by diphenyl diselenide as a lead compound, we tried to optimize its anti-inflammatory activity and decrease the side effects and toxicities as much as possible. Accordingly, two different series of OSe tethered Schiff bases (3a–3c and 5a–5c) were synthesized starting from diaminodiphenyl diselenide (2). Compounds 3a, 3b, 3c, and 5a induced downregulation of the inflammatory proteins (COX-2, IL-6, and IL-1β) by (0.43, 0.52, 0.49, and 0.36), (0.65, 0.54, 0.66, and 0.50), and (0.50, 0.44, 0.56, and 0.50)-fold-change, respectively, assuring their anti-inflammatory potential. Moreover, based on the molecular docking results, the similar binding modes and promising binding scores suggest that the examined candidates have potential inhibitory activity against COX-2, a potential target for the induced anti-inflammatory activity. A 200 ns MD simulation clarified that compound 3a showed the best stability among the ligands with an RMSD of 3.00 Å throughout the simulation time. Moreover, the MM-GBSA calculations clarified that the examined compounds showed comparable ΔG binding energies to the co-crystallized inhibitor, especially compounds 3b and 3c (−64.70 and −88.60) kcal mol−1, which exceeded that of the co-crystallized inhibitor (−60.08 kcal mol−1), respectively. The synthesized compounds 3a, 3b, 3c, and 5a also exhibited distinct electronic properties that correlated with their stability and biological activities. Compound 3c, with the lowest energy gap, demonstrated the highest stability, while compound 5a showed excellent electron affinity, making it a promising electron acceptor. Briefly, compounds 3a, 3b, 3c, and 5a exhibited prominent anti-inflammatory potential, and could be considered as promising lead candidates for further optimization as well. Notably, the bifunctional Schiff bases (3a–3c) were found to be superior to the monofunctional one (5a), which may be attributed to its greater ability to form extra hydrogen bonds and hydrophobic interactions within the receptor pocket. Accordingly, it is highly recommended to synthesize more diverse bifunctional Schiff bases to study their structure-anti-inflammatory relationships.

4. Materials and methods

4.1. Chemistry

4.1.1. Synthesis of the Schiff bases tethered OSe. The Schiff bases-tethered OSe compounds were synthesized according to our reported literature method (see detailed Experimental procedures in the Supplementary materials, SI1, ESI).24,30,34,40

4.2. Biological evaluation

4.2.1. Protein expression of the inflammation-related genes. To investigate the anti-inflammatory activity of the most active compounds (3a, 3b, 3c, and 5a) using their IC50 values (6.00, 5.00, 5.43, and 5.73 µg mL−1, respectively)43 on the HCT116 cancer cell line, a protein expression analysis was carried out for inflammation-related genes. This analysis encompassed the assessment of protein expression levels for COX-2, IL-6, and IL-1β in both the cells treated with compounds 3a, 3b, 3c, and 5a and the untreated cells (Supplementary materials, SI2, ESI). The objective was to identify any alterations in the expression of these proteins and gain insights into the molecular mechanisms responsible for the anti-inflammatory effects induced by the examined Schiff bases-tethered OSe compounds.

4.3. Computational studies

4.3.1. Molecular docking. The target candidates (3a–c and 5a–c) were docked against the COX-2 receptor. This was done using the AutoDock Vina,39,40,73 and the selected poses were visualized using the PyMOL software.74 The chemical structures of the compounds were sketched in ChemDraw, transferred to the drug design program, corrected, and energy-minimized.75 The target protein structure was obtained from the PDB (https://www.rcsb.org/structure/3LN1), 3D hydrogenated, corrected for errors, and energy-minimized.76,77 Finally, the examined candidates were docked against the target COX-2 receptor, and the most promising compounds were selected for further investigation based on the scores and binding modes as well.78
4.3.2. Molecular dynamic simulation and MM-GBSA calculations. The Desmond package (Schrödinger LLC)79,80 was used to perform the molecular dynamic simulation at 200 ns77,81 for the examined complexes of (3a, 3b, 3c, 5a, and co-crystal)-3LN1. The detailed method was explained in the Supplementary materials, SI3 (ESI). Moreover, the thermal_mmgbsa.py python script (Schrödinger LLC) was applied to investigate the molecular mechanics of generalized born surface area (MM-GBSA) energies.82,83 The detailed method was represented in the Supplementary materials, SI4 (ESI).
4.3.3. DFT studies. The DMOL3 program was utilized within the Materials Studio package for cluster calculations (see detailed experimental procedures in the Supplementary Materials, SI5, ESI).

Author contributions

Conceptualization and supervision: Saad Shaaban and Ahmed A. Al-Karmalawy; data curation, validation, visualization, and methodology: Saad Shaaban, Tarek A. Yousef, Hanan A Althikrallah, Yasair S. Al-Faiyz, Ibrahim Elghamry, Marwa Sharaky, Radwan Alnajjar, Ahmed A. Al-Karmalawy; writing – review & editing: Saad Shaaban, Tarek A. Yousef, Hanan A Althikrallah, Yasair S. Al-Faiyz, Radwan Alnajjar, Ahmed A. Al-Karmalawy. Additionally, the authors approved the final version of the manuscript.

Data availability

The data supporting this article have been included in the main manuscript and the supplementary materials.

Conflicts of interest

The authors declare no conflict of interest.

Acknowledgements

The authors acknowledge the Deanship of Scientific Research, Vice Presidency for Graduate Studies and Scientific Research, King Faisal University, Saudi Arabia for supporting this research by Grant No. KFU242337.

References

  1. L. Abdulkhaleq, M. Assi, R. Abdullah, M. Zamri-Saad, Y. Taufiq-Yap and M. Hezmee, Vet. World, 2018, 11, 627 CrossRef CAS .
  2. Y. T. Eloutify, R. A. El-Shiekh, K. M. Ibrahim, A. R. Hamed, A. A. Al-Karmalawy, A. A. Shokry, Y. H. Ahmed, B. Avula, K. Katragunta, I. A. Khan and M. R. Meselhy, Inflammopharmacology, 2023, 31, 859–875 CrossRef CAS .
  3. S. A. Antar, N. A. Ashour, M. E. Marawan and A. A. Al-Karmalawy, Int. J. Mol. Sci., 2023, 24, 4004 CrossRef CAS PubMed .
  4. R. Medzhitov, Nature, 2008, 454, 428–435 CrossRef CAS PubMed .
  5. A. A. Alzain, F. A. Elbadwi, A. A. Al-Karmalawy, R. Elhag, W. Osman and R. A. Mothana, Open Chem., 2023, 21 DOI:10.1515/chem-2023-0161 .
  6. S. S. Savant, S. Sriramkumar and H. M. O’Hagan, Cancers, 2018, 10, 251 CrossRef PubMed .
  7. S. A. Antar, A. M. Mahmoud, W. Abdo, C. Gad and A. A. Al-Karmalawy, Pharm. Sci., 2023, 29(4), 397–416 CrossRef CAS .
  8. S. Rizwan, P. ReddySekhar and B. MalikAsrar, Antioxid. Redox Signaling, 2014, 20(7) DOI:10.1089/ars.2012.5149 .
  9. W. Rehman, M. K. Baloch, A. Badshah and S. Ali, J. Chin. Chem. Soc., 2005, 52, 231–236 CrossRef CAS .
  10. J. Zhang, L. Wang, A. Zhong, G. Huang, F. Wu, D. Li, M. Teng, J. Wang and D. Han, Dyes Pigm., 2019, 162, 590–598 CrossRef CAS .
  11. G. Genchi, G. Lauria, A. Catalano, M. S. Sinicropi and A. Carocci, Int. J. Mol. Sci., 2023, 24, 2633 CrossRef CAS PubMed .
  12. K. Kalimuthu, C. K. Keerthana, M. Mohan, J. Arivalagan, J. Christyraj, M. A. Firer, M. H. A. Choudry, R. J. Anto and Y. J. Lee, J. Cell. Biochem., 2022, 123, 532–542 CrossRef CAS PubMed .
  13. D. E. Handy, J. Joseph and J. Loscalzo, Nutrients, 2021, 13, 3238 CrossRef CAS PubMed .
  14. D. Radomska, R. Czarnomysy, D. Radomski, A. Bielawska and K. Bielawski, Nutrients, 2021, 13, 1649 CrossRef CAS .
  15. S. Shaaban, H. Ba-Ghazal, Y. S. Al-Faiyz, A. A. Al-Karmalawy, N. Amri and I. Youssef, Tetrahedron, 2024, 133957 CrossRef CAS .
  16. M. J. Maroney and R. J. Hondal, Free Radicals Biol. Med., 2018, 127, 228–237 CrossRef CAS .
  17. C. W. Nogueira, N. V. Barbosa and J. B. Rocha, Arch. Toxicol., 2021, 95, 1179–1226 CrossRef CAS PubMed .
  18. J. B. Rocha, C. S. Oliveira and P. A. Nogara, Organoselenium Compounds in Biology and Medicine, 2017, pp. 342–376 Search PubMed .
  19. N. Astrain-Redin, I. Talavera, E. Moreno, M. J. Ramirez, N. Martinez-Saez, I. Encio, A. K. Sharma, C. Sanmartin and D. Plano, Antioxidants, 2023, 12(1), 139 CrossRef CAS PubMed .
  20. A. L. Sharpley, C. Williams, A. A. Holder, B. R. Godlewska, N. Singh, M. Shanyinde, O. MacDonald and P. J. Cowen, Psychopharmacology, 2020, 237, 3773–3782 CrossRef CAS PubMed .
  21. L. Wang, Z. Yang, J. Fu, H. Yin, K. Xiong, Q. Tan, H. Jin, J. Li, T. Wang, W. Tang, J. Yin, G. Cai, M. Liu, S. Kehr, K. Becker and H. Zeng, Free Radical Biol. Med., 2012, 52, 898–908 CrossRef CAS PubMed .
  22. M. S. S. Adam, A. M. Abu-Dief, M. Makhlouf, S. Shaaban, S. O. Alzahrani, F. Alkhatib, G. S. Masaret, M. A. Mohamed, M. Alsehli and N. M. El-Metwaly, Polyhedron, 2021, 201, 115167 CrossRef CAS .
  23. D. Desai, I. Sinha, K. Null, W. Wolter, M. A. Suckow, T. King, S. Amin and R. Sinha, Int. J. Cancer, 2010, 127, 230–238 CrossRef CAS PubMed .
  24. S. Shaaban, A. Negm, M. A. Sobh and L. A. Wessjohann, Adv. Anticancer Agents Med. Chem., 2016, 16, 621–632 CrossRef CAS .
  25. S. Shi, K. Li, J. Peng, J. Li, L. Luo, M. Liu, Y. Chen, Z. Xiang, P. Xiong and L. Liu, Biomed. Pharmacother., 2022, 149, 112828 CrossRef CAS PubMed .
  26. M. Cherkaoui-Malki, S. Shaaban, M. Tahri-Joutey, A. Elshobaky, F.-E. Saih, D. Vervandier-Fasseur, C. Jacob, B. Nasser, N. Latruffe and P. Andreoletti, Multidisciplinary Digital Publishing Institute Proceedings, 2019, 11, 33 Search PubMed.
  27. S. Shaaban, D. Vervandier-Fasseur, P. Andreoletti, A. Zarrouk, P. Richard, A. Negm, G. Manolikakes, C. Jacob and M. Cherkaoui-Malki, Bioorg. Chem., 2018, 80, 43–56 CrossRef CAS PubMed .
  28. S. Shaaban, A. Zarrouk, D. Vervandier-Fasseur, Y. S. Al-Faiyz, H. El-Sawy, I. Althagafi, P. Andreoletti and M. Cherkaoui-Malki, Arabian J. Chem., 2021, 14(4), 103051 CrossRef CAS .
  29. S. Shaaban, M. A. Arafat, H. E. Gaffer and W. S. Hamama, Der Pharma Chem., 2014, 6, 186–193 Search PubMed .
  30. S. Shaaban, A. Abdou, A. G. Alhamzani, M. M. Abou-Krisha, M. A. Al-Qudah, M. Alaasar, I. Youssef and T. A. Yousef, Life, 2023, 13, 912 CrossRef CAS PubMed .
  31. S. K. Verma, R. Verma, K. Rakesh and D. C. Gowda, Eur. J. Med. Chem. Rep., 2022, 6, 100087 CAS .
  32. K. Rakesh, R. Suhas and D. C. Gowda, Int. J. Pept. Res. Ther., 2019, 25, 227–234 CrossRef CAS .
  33. Y. Jiang, K. Rakesh, N. S. Alharbi, H. Vivek, H. Manukumar, Y. Mohammed and H.-L. Qin, Bioorg. Chem., 2019, 89, 103015 CrossRef CAS .
  34. W. S. Hamama, G. G. El-Bana, S. Shaaban, O. Habib and H. H. J. R. A. Zoorob, RSC Adv., 2016, 6, 24010–24049 RSC .
  35. K. Rakesh, C. Shantharam and H. Manukumar, Bioorg. Chem., 2016, 68, 1–8 CrossRef CAS PubMed .
  36. K. Rakesh, H. Vivek, H. Manukumar, C. Shantharam, S. N. A. Bukhari, H.-L. Qin and M. Sridhara, RSC Adv., 2018, 8, 5473–5483 RSC .
  37. S.-M. Wang, G.-F. Zha, K. Rakesh, N. Darshini, T. Shubhavathi, H. Vivek, N. Mallesha and H.-L. Qin, MedChemComm, 2017, 8, 1173–1189 RSC .
  38. W. I. Mortada, S. Shaaban, H. A. Althikrallah, M. Alaasar, H. A. Alshwyeh and A. H. Ragab, J. Food Compos. Anal., 2024, 106358 CrossRef CAS .
  39. H. M. Abd El-Lateef, S. Shaaban, K. Shalabi and M. M. Khalaf, J. Taiwan Inst. Chem. Eng., 2022, 133, 104258 CrossRef .
  40. S. Shaaban, M. S. S. Adam and N. M. El-Metwaly, J. Mol. Liq., 2022, 363, 119907 CrossRef CAS .
  41. K. Rakesh, H. Manukumar and D. C. Gowda, Bioorg. Med. Chem. Lett., 2015, 25, 1072–1077 CrossRef CAS PubMed .
  42. L. Kang, X.-H. Gao, H.-R. Liu, X. Men, H.-N. Wu, P.-W. Cui, E. Oldfield and J.-Y. Yan, Mol. Diversity, 2018, 22, 893–906 CrossRef CAS PubMed .
  43. S. Shaaban, M. M. Hammouda, A. H. Althikrallah, Y. J. Al Nawah, H. Ba-Ghazal, M. Sharaky, S. H. Abulkhair and A. A. Al-Karmalawy, Curr. Med. Chem., 2024, 31, 1–15 CrossRef PubMed .
  44. C. Gallo-Rodriguez and J. B. Rodríguez, Synthesis, 2024, 56(15), 2295–2315 CrossRef CAS .
  45. C. Gallo-Rodriguez and J. B. Rodriguez, ChemMedChem, 2024, e202400063 CrossRef CAS .
  46. J. M. Sonego, S. I. de Diego, S. H. Szajnman, C. Gallo-Rodriguez and J. B. Rodriguez, Chem. – Eur. J., 2023, 29, e202300030 CrossRef CAS .
  47. P. Zhang, S. Zhang, H. Hu, T. Hu, K. Shi, Y. Xu, G. Xu, H. Hu and S. Pan, Food Biosci., 2024, 58, 103651 CrossRef CAS .
  48. M. Álvarez-Pérez, W. Ali, M. A. Marć, J. Handzlik and E. Domínguez-Álvarez, Molecules, 2018, 23, 628 CrossRef PubMed .
  49. N. V. Barbosa, C. W. Nogueira, P. A. Nogara, A. F. de Bem, M. Aschner and J. B. Rocha, Metallomics, 2017, 9, 1703–1734 CrossRef CAS PubMed .
  50. L. P. Wolters and L. Orian, Curr. Org. Chem., 2016, 20, 189–197 CrossRef CAS .
  51. R. M. Rosa, D. J. Moura, A. C. R. e Silva, J. Saffi and J. A. P. Henriques, Mutat. Res., Genet. Toxicol. Environ. Mutagen., 2007, 631, 44–54 CrossRef CAS PubMed .
  52. O. d R. A. Junior, E. Antônio, R. M. Mainardes and N. M. Khalil, J. Trace Elem. Med. Biol., 2017, 39, 176–185 CrossRef PubMed .
  53. C. W. Nogueira and J. B. Rocha, J. Braz. Chem. Soc., 2010, 21, 2055–2071 CrossRef CAS .
  54. E. M. Abbass, A. A. Al-Karmalawy, M. Sharaky, M. Khattab, A. Y. A. Alzahrani and A. I. Hassaballah, Bioorg. Chem., 2024, 142, 106936 CrossRef CAS .
  55. M. A. E. Mourad, A. Abo Elmaaty, I. Zaki, A. A. E. Mourad, A. Hofni, A. E. Khodir, E. M. Aboubakr, A. Elkamhawy, E. J. Roh and A. A. Al-Karmalawy, J. Enzyme Inhib. Med. Chem., 2023, 38, 2205043 CrossRef PubMed .
  56. A. A. Al-Karmalawy, M. Rashed, M. Sharaky, H. S. Abulkhair, M. M. Hammouda, H. O. Tawfik and M. A. Shaldam, Eur. J. Med. Chem., 2023, 115661,  DOI:10.1016/j.ejmech.2023.115661 .
  57. A. A. Al-Karmalawy, M. H. A. Mousa, M. Sharaky, M. A. E. Mourad, A. M. El-Dessouki, A. O. Hamouda, R. Alnajjar, A. A. Ayed, M. A. Shaldam and H. O. Tawfik, J. Med. Chem., 2023, 67(1), 492–512 CrossRef PubMed .
  58. S. Shaaban, K. T. Abdullah, K. Shalabi, T. A. Yousef, O. K. Al Duaij, G. M. Alsulaim, H. A. Althikrallah, M. Alaasar, A. S. Al-Janabi and A. M. Abu-Dief, Appl. Organomet. Chem., 2024, e7712 CrossRef CAS .
  59. S. Shaaban, H. A. Althikrallah, A. Negm, A. Abo Elmaaty and A. A. Al-Karmalawy, RSC Adv., 2024, 14, 18576–18587 RSC .
  60. S. Shaaban, K. Shalabi, T. A. Yousef, M. Abou-Krisha, A. A. Alanazi, H. A. Althikrallah, M. Alaasar, A. M. Abu-Dief and A. S. Al-Janabi, J. Taiwan Inst. Chem. Eng., 2024, 165, 105766 CrossRef CAS .
  61. H. E. Gaffer, M. R. Elgohary, H. A. Etman and S. Shaaban, Pigm. Resin Technol., 2017, 46, 210–217 CrossRef .
  62. B. F. Abdel-Wahab and S. Shaaban, Synthesis, 2014, 1709–1716 CrossRef .
  63. S. Shaaban, M. A. Arafat and W. S. Hamama, ARKIVOC, 2014, 2014, 470–505 Search PubMed .
  64. A. D. Sonawane, R. A. Sonawane, M. Ninomiya and M. Koketsu, Dalton Trans., 2021, 50, 12764–12790 RSC .
  65. P. N. Makhal, A. Nandi and V. R. Kaki, ChemistrySelect, 2021, 6, 663–679 CrossRef CAS .
  66. S. Shaaban, H. M. A. El-Lateef, M. M. Khalaf, M. Gouda and I. Youssef, Polymers, 2022, 14, 2208 CrossRef CAS PubMed .
  67. K. Wang, J. Yin, J. Chen, J. Ma, H. Si and D. Xia, Phytomedicine, 2024, 128, 155258 CrossRef CAS PubMed .
  68. J. He, X. Feng, Y. Liu, Y. Wang, C. Ge, S. Liu and Y. Jiang, Biomed. Pharmacother., 2024, 177, 117163 CrossRef CAS .
  69. L. Zhao, Y. Weng, X. Zhou and G. Wu, Org. Lett., 2024, 26(22), 4835–4839 CrossRef CAS PubMed .
  70. S. Shaaban, H. Ferjani, H. M. Abd El-Lateef, M. M. Khalaf, M. Gouda, M. Alaasar and T. A. Yousef, Front. Chem., 2022, 10, 961787 CrossRef CAS .
  71. S. Shaaban, H. Ferjani, I. Althagafi and T. Yousef, J. Mol. Struct., 2021, 1245, 131072 CrossRef CAS .
  72. S. Shaaban, H. Ferjani, T. Yousef and M. Abdel-Motaal, J. Inorg. Organomet. Polym. Mater., 2022, 32, 1878–1890 CrossRef CAS .
  73. R. Huey, G. M. Morris and S. Forli, The Scripps Research Institute Molecular Graphics Laboratory, 2012, vol. 10550, p. 1000 Search PubMed .
  74. S. Yuan, H. S. Chan and Z. Hu, Wiley Interdiscip. Rev.: Comput. Mol. Sci., 2017, 7, e1298 Search PubMed .
  75. D. Elebeedy, I. Badawy, A. A. Elmaaty, M. M. Saleh, A. Kandeil, A. Ghanem, O. Kutkat, R. Alnajjar, A. I. Abd El Maksoud and A. A. Al-karmalawy, Comput. Biol. Med., 2022, 141, 105149 CrossRef CAS PubMed .
  76. A. A. Elmaaty, K. M. Darwish, A. Chrouda, A. A. Boseila, M. A. Tantawy, S. S. Elhady, A. B. Shaik, M. Mustafa and A. A. Al-karmalawy, ACS Omega, 2022, 7, 875–899 CrossRef CAS PubMed .
  77. R. R. Ezz Eldin, M. A. Saleh, M. H. Alotaibi, R. K. Alsuair, Y. A. Alzahrani, F. A. Alshehri, A. F. Mohamed, S. M. Hafez, A. A. Althoqapy, S. K. Khirala, M. M. Amin, A. H. AbdElwahab, M. S. Alesawy, A. A. Elmaaty and A. A. Al-Karmalawy, J. Enzyme Inhib. Med. Chem., 2022, 37, 1098–1119 CrossRef CAS PubMed .
  78. A. M. El-Naggar, A. M. A. Hassan, E. B. Elkaeed, M. S. Alesawy and A. A. Al-Karmalawy, Bioorg. Chem., 2022, 123, 105770 CrossRef CAS PubMed .
  79. S. Release, Maestro-Desmond Interoperability Tools, Schrödinger, New York, NY, 2017 Search PubMed .
  80. M. M. Hammoud, M. Khattab, M. Abdel-Motaal, J. Van der Eycken, R. Alnajjar, H. Abulkhair and A. A. Al-Karmalawy, J. Biomol. Struct. Dyn., 2022, 1–18 CrossRef PubMed .
  81. R. M. El-Masry, A. A. Al-Karmalawy, R. Alnajjar, S. H. Mahmoud, A. Mostafa, H. H. Kadry, S. M. Abou-Seri and A. T. Taher, New J. Chem., 2022, 46, 5078–5090 RSC .
  82. A. A. Al-Karmalawy, R. Alnajjar, A. A. Elmaaty, F. A. Binjubair, S. T. Al-Rashood, B. S. Mansour, A. Elkamhawy, W. M. Eldehna and K. A. Mansour, J. Biomol. Struct. Dyn., 2023, 1–13,  DOI:10.1080/07391102.2023.2240419 .
  83. A. Abo Elmaaty, A. A. Al-Karmalawy, M. S. Nafie, M. M. Shamaa, I. Zaki, R. Alnajjar and M. Y. Zakaria, Int. J. Pharm., 2023, 640, 122980 CrossRef CAS PubMed .

Footnote

Electronic supplementary information (ESI) available: SI1. Synthesis of the organoselenium compounds; SI2. Anti-inflammatory markers assay (enzyme-linked immunosorbent assay); SI3. Molecular dynamic simulations; SI4. MD trajectory analysis and prime MM-GBSA calculations; SI5. DFT studies. See DOI: https://doi.org/10.1039/d4nj04376f

This journal is © The Royal Society of Chemistry and the Centre National de la Recherche Scientifique 2025
Click here to see how this site uses Cookies. View our privacy policy here.