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. 2025 Jul 31;10(31):34399–34413. doi: 10.1021/acsomega.5c02269

Benzimidazole-Coordinated Copper(II) Complexes as Effectual Chemotherapeutics against Malignancy

Ankita Saha †, Ananya Debnath †, Meena Chettri †, Rajani Kanta Mahato †, Dona Das ‡, Debanjan Sarkar ‡, Radhika Chaurasia §, Sankar Bhattacharyya ‡, Monalisa Mukherjee §,*, Bhaskar Biswas †,*
PMCID: PMC12355249  PMID: 40821591

Abstract

Cancer stands as the second-leading cause of global mortality, persistently representing a peril to human well-being. The challenges of drug insensitivity and resistance significantly impede advancements in cancer treatment, emphasizing the critical importance of developing innovative agents that specifically target malignant cells. Benzimidazole derivatives are a preferred choice in cancer therapy, and a variety of benzimidazole-based molecules have demonstrated incredible potential for anticancer therapeutic objectives. Albeit such advancements, there are certain pragmatic limitations, including low bioavailability, which results in insufficient plasma concentration levels, side effects, and toxicity that need to be addressed. In this quest to overcome the existing hurdles, we elucidate the synthesis, structural characterization, and substantial proliferative activity of two copper­(II) complexes bearing benzimidazole ligands. The ligands, 2-(thiophen-2-yl)-1-(thiophen-2-ylmethyl)-1H-benzo­[d]­imidazole (L) and 6-methyl-2-(thiophen-2-yl)-1-(thiophen-2-ylmethyl)-1H-benzo­[d]­imidazole (L′) were prepared by the coupling of thiophene-2-carboxaldehyde with o-phenylenediamine and 3,4-diaminotoluene, respectively, in water under an ambient condition. Both L and L′ react with Cu­(NO3)2·3H2O in methanol, producing the complexes, [Cu­(L)2(NO3)2] (complex 1) and [Cu2(L′)2(μ-CH3O)2]­(NO3)2 (complex 2), respectively. Both complexes exhibited solution-phase stability, as confirmed by mass spectral analysis. X-ray structural analysis divulges the mononuclear and dinuclear nature of complex 1 and complex 2, where Cu­(II) centers adopt a slightly distorted square planar geometry in both complexes. Energy framework analysis suggests the higher stability of complex 2 than complex 1, attributed to the more robust character of the dinuclear copper complex. Molecular docking studies for complex 1 and complex 2 against p53, BAX, BCL2, and PARP proteins suggest stable conformations for both complexes. The cell viability and cytotoxicity of the synthetic compounds were evaluated against mouse cancer cell lines, as well as human breast cancer cell lines. Cell cycle, apoptosis, caspase, and TUNEL assays have been carried out to unveil the cell proliferative screening mechanism for the synthetic compounds. The intercalative binding mode of the complexes for CT-DNA triggers the apoptosis of the tumor cells. Hence, we postulate that these compounds have the potential to broaden the arsenal of effective anticancer therapies.

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1. Introduction

Benzimidazoles have undergone a remarkable evolution during the last few decades, emerging as a pivotal class of N-heterocycles possessing a myriad of pharmacological properties. The escalating interest in benzimidazole compounds in diverse biological applications is underscored by their super stability, molecular durability, biocompatibility, and excellent medicinal properties. − A pivotal exemplar within the pantheon of benzimidazoles in nature is central to the fact that N-ribosyl-dimethylbenzimidazole axially coordinates with cobalt in vitamin B12. It is well-documented that benzimidazole derivatives stand out as potent inhibitors against various enzymes and possess significant medicinal properties, like antimicrobial, antidiabetic, antiparasitic, antitumor, antihistamine, and analgesic activities, including applications in some neurological, endocrinological, and ophthalmological drugs. It is critical to note that benzimidazole acts as a ligand toward transition metal ions, exhibiting diverse coordination modes characterized by stability toward heat and oxidizing agents. − Benzimidazole ligands impart structural resemblance to nucleobases like pyrimidine and purine types, making them more interesting in the field of biological research. For instance, 2-pyridyl-1H-benzimidazoles in coordination with copper­(II) and silver­(I) exhibit promising activity against various bacterial strains like Staphylococcus aureus, Pseudomonas aeruginosa, Salmonella typhi, Staphylococcus epidermidis, Shigella flexneri, as well as fungi like Candida albicans.

Benzimidazole–copper­(II) complexes have garnered increasing interest owing to their versatile coordination chemistry and potential applications in diverse biological areas, including antimicrobial, antitumoral, and other pharmacologically relevant activities. The benzimidazole unit acts as a robust N-donor ligand, forming stable chelates with Cu­(II) ions and enabling structural tunability through various donor group substituents. Numerous studies have highlighted the functional potential of these complexes. Recently, Hamali et al. reported that benzimidazole–Cu­(II) derivatives exhibit significant activity against A549 lung cancer cells, underscoring their promise in medicinal chemistry. Furthermore, DFT studies support these findings, revealing low binding energies and multiple interactions with the MMP-9 protein binding site, consistent with their observed in vitro antiproliferative effects. In addition, Cu­(II) coordination polymers with benzimidazole-based ligands have demonstrated notable redox activity. Likewise, two isostructural Cu­(II) complexes incorporating methoxy-functionalized benzimidazole ligands were synthesized and characterized, showing an enhanced catalytic oxidation rate of 2-aminophenol (2-AP). Despite these advances, there remains ample scope for the design of new benzimidazole–copper complexes with improved stability, novel coordination motifs, and enhanced functional performance.

For instance, Alajmi et al. reported the synthesis of a copper complex [Cu­(BnI)2] (a) based on a biologically active benzimidazole-derived ligand (BnI), which exhibited notably strong activity against breast cancer cells, with IC50 values comparable to those of the standard drug, cisplatin (Figure ). Another study conducted by Qiao et al. emphasized the biological evaluation of a mononuclear copper­(II) complex (b) with a tridentate 2-substituted benzimidazole ligand (BMA). This copper complex on human cervical carcinoma (HeLa) cells triggered the inhibition of cell proliferation in a time- and dose-dependent manner through an intrinsic mitochondrial apoptotic pathway. Further, Naveen et al. synthesized a structurally similar Cu­(II) complex (c) containing 2-thiophene-2-yl-1-thiophen-2-ylmethyl-1­(H)-benzimidazole. In vitro cytotoxicity of the synthesized complex was found to be very high compared with the standard, cisplatin. Another work by Paul et al. reported the synthesis and characterization of a benzimidazole-based ligand, HL and its dinuclear copper­(II) complex, CuL (d). The study confirmed that both HL and CuL effectively bind to DNA through partial intercalative/electrostatic interactions and were also found to interact with protein (BSA). Further, molecular docking studies were performed on both the ligand and the complex, suggesting that the copper­(II) complex was a more active drug than the ligand, as reflected by their K b values and IC50 [458 mM (HL) and 22 mM (CuL)] values toward the MCF-7 cancer cell line.

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Comparison of Cu–benzimidazole complexes, 1 and 2 with previously reported structures (a, b, c, d). −

Typically, cancer remains a formidable challenge to human health and has garnered substantial attention from researchers across diverse scientific disciplines due to the myriad manifestations of various types of cancer at different stages. − Among all cancer types, breast cancer is the second leading cancer among females, and for studying the therapeutics against breast cancer, MCF-7 (Michigan Cell Foundation-7) is the most widely used human breast cancer cell line (Figure ). It is expected that by 2040, the annual death cases due to breast cancer will reach 1 million each year. ,

Since the fortuitous discovery of cisplatin, renowned for its antiproliferative activity, a captivating avenue of research within medicinal chemistry, with metallodrugs emerging as potential anticancer drugs. Widely employed platinum-based drugs, like cisplatin and its derivatives carboplatin and oxaliplatin, have demonstrated efficacy against a spectrum of solid tumors, including mesothelioma, neuroblastoma, ovarian cancer, lung cancer, brain tumor, head and neck cancer, and testicular cancer. Despite their therapeutic success, platinum-based drugs have drawbacks, such as high costs, severe side effects including nephrotoxicity, hepatotoxicity, ototoxicity, and neurotoxicityas well as the development of drug resistance, which necessitates exploration into cost-effective biometal compounds to mitigate these issues while maintaining high efficacy. , In such a quest, benzimidazole and its derivatives have emerged as potential ligands for metal coordination. The structural ease of conjugation of π-systems, planar structure, the ability to mimick imidazole functionality in proteins, along with the existence of these ligand systems in biologically relevant vitamin B12 facilitate the application of these ligands as potential drugs for cancer treatment.

A commercially available antitumor drug, Abemaciclib, exemplifies the medical significance of a similar benzimidazole ligand moiety. A plethora of benzimidazole derivatives have been developed for various cancer types, each employing distinct mechanisms of action. Despite being a commonly used pharmacophore, the availability of target-specific benzimidazoles remains limited. One notable example is Glasdegib, an FDA-approved drug, which is a benzimidazole derivative recommended for the treatment of acute myeloid leukemia (AML). It functions by inhibiting the sonic hedgehog receptor smoothened (SMO). However, the clinical use of Glasdegib is not without challenges. Its efficacy can vary among AML patients, and the development of resistance is a concern. Additionally, side effects such as myelosuppression and gastrointestinal disturbances necessitate close monitoring and appropriate management. Benzimidazole derivatives exhibit a diverse array of pharmacological activities, encompassing antimicrobial, antiulcerative, antihypertensive, antiviral, anti-inflammatory, antiprotozoal, and anticancer activities. ,− Consequently, the inclusion of benzimidazole derivatives into metal complexes holds the potential toward significant enhancement of antiproliferative activities. Albeit such advancements, there are certain pragmatic limitations including low bioavailability, which results in insufficient plasma concentration levels, side effects, and toxicity, which need to be addressed. Copper is integral to various cellular functions, serving as a vital micronutrient and a crucial cofactor for many metalloenzymes. These metalloenzymes are involved in vital biological activities like mitochondrial metabolism (cytochrome c oxidase), as well as in cellular defense against reactive oxygen species (ROS), exemplified by superoxide dismutase (SOD). Furthermore, copper plays a crucial role in processes like angiogenesis and the migration of endothelial cells. However, elevated levels of copper have been associated with promoting tumor growth and metastasis. , Interestingly, copper complexes have demonstrated anticancer effects through multiple mechanisms, including intracellular copper accumulation, induction of cytotoxicity, activation of apoptosis inhibitor factor (XIAP), proteasome inhibition, DNA binding, and intercalation. ,

The literature survey underscores the profound potential of the copper­(II) complex with the ligand 2-[(1H-benzimidazol-2-ylimino)-methyl]-6-methoxyphenol against MCF-7 human breast cancer cells, as evidenced by the MTT method. Simultaneously, this complex has shown efficacy in relieving chronic pain in late-stage cancer patients, akin to nonsteroidal anti-inflammatory drugs. Leveraging the pharmacological values of benzimidazoles and their derivatives in the synthesis of various drugs, and considering the utility of cisplatin and other metallodrugs, this study elucidates the synthesis of two benzimidazole ligands (L and L′) and their copper complexes, complex 1 and complex 2 under ambient conditions. Thorough characterization via various spectral analyses, coupled with X-ray crystallography, validates the structural composition of the compounds. The comprehensive antiproliferative activity of the synthetic compounds has been explored against two mouse and one human breast cancer cell lines.

2. Result and Discussion

2.1. Synthesis, Spectroscopic Characterization, and Solution Phase Stability of the Copper­(II) Complexes

The benzimidazole ligands L and L′ were synthesized by the reaction of thiophene-2-carboxaldehyde with o-phenylenediamine and 3,4-diaminotoluene, respectively, in pure water under ambient conditions. Copper­(II) nitrate salts were reacted with L and L′ in a 1:2 ratio in MeOH to obtain the copper­(II) complexes, complex 1 and complex 2 (Scheme ).

1. Synthesis of Benzimidazole Ligands (L, L′) and Their Copper Complexes (Complex 1 and Complex 2).

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The structural formulation of the copper complexes was confirmed by FT-IR and UV–vis spectral analysis. The appearance of characteristic peaks at 1617 and 1621 cm–1 for complex 1 and complex 2, respectively, indicates the presence of the azomethine group of the benzimidazole unit. Meanwhile the peaks at 2931 and 2935 cm–1 correspond to the C–H stretching frequencies of the −CH2– group in complex 1 and complex 2, respectively (Figure S5). UV–vis spectral analysis of the complexes in DMSO–H2O (v/v; 1/20) at room temperature reveals electronic bands at 303 and 301 nm for complex 1 and complex 2, respectively, which may be attributed to the π → π* or n → π* electronic transition of ligand origin (Figure S6). The solution-phase stability of the synthetic complexes is also confirmed by high-resolution mass spectral analysis in a DMSO–H2O (v/v; 1/20) solution. Complex 1 and complex 2 display molecular ion peaks at m/z 780.0258 and 1430.0348, respectively, suggesting the molecular integrity of the complexes in the water–DMSO solvent mixture (Figures S7 and S8)

2.2. Crystal Structure Description

The X-ray structural analysis divulges that complex 1 and complex 2 crystallize in the monoclinic and triclinic systems with P21/n and P1̅ space groups, respectively. Complex 1 appears as a mononuclear copper complex with square planar geometry, while complex 2 exists as a doubly methoxo-bridged dinuclear copper complex adopting square planar geometry. The square plane in complex 1 is composed of two benzimidazole-N-coordinated ligand units and two nitrate ions (Figure a), while both the square planes for the Cu1 and Cu2 centers in complex 2 (Figure b) are formed by two N-coordinations from two different benzimidazole units and the coordination from two bridging methoxy–O ions. The ORTEP representations of complexes 1 and 2 are shown in Figure . X-ray refinement data are summarized in Table . Selected bond lengths and bond angles for complexes 1 and 2 are tabulated in Tables S1 and S2, respectively.

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Thermal ellipsoidal plots of (a) complex 1 and (b) complex 2 with 30% probability. H atoms are not displayed for the same purpose.

1. Crystallographic Data and Structure Refinement Parameters for the Copper Complexes.

Parameters Complex 1 Complex 2
Crystal data 2349926 2349925
Empirical formula C32H24CuN6O6S4 C70H62Cu2N10O8S8
Formula weight 784.426 1557.979
T (K) 273.15 273.15
Wavelength (Å) 0.71073 0.71073
Crystal system Monoclinic Triclinic
Space group P21/n P1̅
     
Unit cell dimensions    
a (Å) 9.8687(3) 15.587(4)
b (Å) 9.8848(3) 15.595(4)
c (Å) 17.0595(6) 18.235(5)
α (°) 90 101.398(5)
β (°) 96.754(1) 101.397(7)
γ (°) 90 114.016(6)
V (Å3) 1652.61(9) 3772.6(17)
Z 2 2
ρ (gm cm–3) 1.576 1.371
Absorption coefficient (mm–1) 0.969 0.844
F(000) 808.2 1614.3
Goodness-of-fit on F2 1.060 1.187
Reflections collected 32359 103141
Independent reflections 2848 13075
Final R indexes [I ≥ 2σ (I)] R1 = 0.0785, wR 2 = 0.2098 R1 = 0.0999, wR 2 = 0.2827
Largest diff. peak/hole/e Å–3 1.99/–1.56 2.64/–1.73
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The distortion in the square plane of complex 1 is higher than that of complex 2, as evidenced by the Cu­(II)-centric bond angles (Tables S1 and S2). This is probably due to the nearby presence of the second-O of the nitrate around the square plane, induced by some geometric restriction. The Cu–N distances in the complexes are closely matched, ranging from 1.970(4) to 1.981(5) Å. However, the Cu–O bond lengths of 2.186(7) Å in complex 1 are found to be relatively longer compared to the average Cu–O distance of 1.9165(2) Å in complex 2. Bond length analysis within the coordinated NO3 – in complex 1 shows a nearly equivalent N–O bond length for the coordinated oxygen (O1) of 1.250 Å, which may be compared with the other N3–O2/O3 bond lengths of 1.252/1.169 Å.

Notably, the introduction of the methyl group in the benzimidazole ligand played a pivotal role in controlling the steric factor. Without a methyl-based ligand–copper­(II) complex, the minimal steric crowding allows space for the coordination of bulky nitrate ions to the Cu­(II) centers, leading to the generation of a mononuclear structure. In contrast, the methyl-based benzimidazole ligand–copper complex introduces larger crowding, which facilitates the prevention of bulkier nitrate coordination to the copper center. In comparison with the structurally analogous, previously reported copper-chloro/azido complexes, the mononuclear complex shows a shorter Cu(1)–N(2) bond distance of 1.97 Å compared to the Cu(1)–N(1) bond distance of 1.99 Å, indicating better stability of our synthetic copper complex 1. In contrast, upon comparison with the structurally similar reported dinuclear copper complex, it is revealed that the methoxy bridge coordinates with the square planar Cu­(II) ion in an end-on bridging mode, leading to a perfect Cu2N2 square with a Cu···Cu distance of 3.015 Å, like the coordination environment of the reported dinuclear copper–benzimidazole complex. However, the synthetic dinuclear complex 2 holds two units of ligands at individual copper centers with syn conformation. Additionally, C–H···π interactions consolidate the formation of a stable dinuclear copper complex.

Energy framework analysis using Crystal Explorer 21.2 software with B3LYP/6-31G­(d,p) basis sets was carried out for both copper complexes, using TONTO software for the cluster environment within 3.8 Å surrounding a particular molecule of interest. The total interaction energy is expressed as E tot = k ele E′ele + k pol E′ pol + k disp E′ disp + k rep E′ rep, where the k values belong to the scale factors for benchmarked energy models. E′ele represents the electrostatic energy, E′pol represents the polarization energy, E′disp represents the dispersion energy, and E′rep represents the repulsive energy (Tables S3 and S4). To identify the type of interaction energies developed due to supramolecular interactions, we set the tube dimension factor 300, where a greater tube radius is directly proportional to stronger and more prominent interactions. Energy framework analysis of the copper complexes reveals the dominant role of dispersive energies and Coulombic energies in the total energy and stabilization of the supramolecular frameworks (Figure S9). It is noticed that the dispersive forces (−173.7 kJ/mol) contribute to a higher extent than the Coulombic energies (−80.0 kJ/mol) in complex 1, whereas the dispersive energy contributes to a lower extent (−1063.1 kJ/mol) than the Coulombic forces (−2482.86 kJ/mol) in complex 2 (Tables S3 and S4). X-ray crystallography also suggests the involvement of short-ranged noncovalent interactions in complex 2 over complex 1 attributing to the robustness of the structure in complex 2. The significant difference in the total energy between the complexes also confirms the higher stability of complex 2.

2.3. Molecular Docking Analysis of Complexes 1 and 2

A molecular docking study of complex 1 and complex 2 was performed against p53, BAX, BCL2, and PARP proteins, and the binding energy is summarized in Table S5. The 3D versions of the protein–complex interactions (complex 1 and complex 2) with the respective proteins are shown in Figures S10 and S11. The binding energies of complexes 1 and 2 summarized in Table S5 were compared with a positive control drug, docetaxel. From this study, it was observed that complexes 1 and 2 strongly bound with p53, BAX, and BCL2 proteins at their binding sites. Figures S10 and S11 show that BCL2 (2W3L) interacts with complexes mainly via hydrophobic interactions and hydrogen bonds. The receptor molecule forms hydrophobic interactions via ASP62B, ARG66B, ARG68A, VAL118A, and TYR161B interacting residues, and the receptor molecule forms hydrogen bonds via ASN122A interacting residues. Thereafter, the interaction between BAX (4S0O) and complexes was examined, exhibiting hydrophobic interactions via GLU17B, LYS21B, ALA24B, LEU47B, VAL50B, and LEU141B, and hydrogen bonds via MET20B and ARG145B. Following this, the interactions of p53 (1TUP) with the complexes were also examined. It is evident that 1TUP interacts with the complexes by forming hydrophobic interactions via GLU171B, ASP186C, and LEU188C, hydrogen bonds via VAL172B, ARG196C, and ASN 235C, and salt bridges via GLU198C. The interaction of the complexes with PARP (4DQY) also conferred the hydrophobic interactions via TYR907C and LEU985C. The RMSD values for the proteins are as follows: p53 – 79.72 Å, BAX – 45.11 Å, BCL2 – 35.47 Å, and PARP – 26.45 Å. Molecular docking studies revealed that complex 1 shows relatively higher binding interactions with the active site of p53, BAX, and BCL2 compared to complex 2 with the respective proteins, suggesting their inhibitory effect against these proteins of the apoptotic pathway and showing potent anticancer activity.

2.4. Cell Viability and Cytotoxicity Assay of Complexes 1 and 2 against Normal Human PBMCs

MTT assay and hemolysis assay were performed to determine the cytotoxicity and cell viability of complexes 1 and 2 against normal human peripheral blood mononuclear cells (PBMCs) and normal human erythrocytes (RBCs). Negligible changes were observed in the cell viability percentages when concentrations ranging from 0 to 80 μM/mL of complex 1 and complex 2 were incubated with normal human PBMCs. Normal red blood cells were exposed to various concentrations (0–80 μM/mL) of complex 1 and complex 2 for 1 h, and the hemolysis percentage at these doses was nonsignificant compared to 0.1% Triton-treated RBCs (positive control). Therefore, the dose-dependence study suggests the nontoxic concentrations of the complexes up to 80 μM/mL on normal human cells (Figure ).

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(A,B) MTT assay of normal human PBMCs. Normal human PBMCs were exposed to various concentrations of complex 1 and complex 2 (1 μM, 5 μM, 10 μM, 20 μM, 40 μM, and 80 μM) and incubated for 24 h in 37 °C. Graph showing that there is no significant change in live cell percentage of normal human PBMCs treated with complex 1 and complex 2 compared to control, (C, D) Hemolysis assay of normal RBC cells. A negligible amount of hemolysis was found with the application of complex 1 and complex 2 against normal RBCs.

2.5. Dose Kinetics of Complexes 1 and 2 against Sarcoma-180, 4T1, and MCF-7

After examining cell viability, the cytotoxic effect of complexes 1 and 2 was tested against Sarcoma-180, 4T1, and MCF-7 cell lines by the MTT assay. For this, different concentrations of complex 1 and complex 2 were added to each cell line, along with a positive control (methotrexate, 10 μM) and a vehicle control (DMSO, 10 μM) and incubated for 24 h. The IC50 values of complex 1 and complex 2 against Sarcoma-180 and 4T1 cell lines were determined to be 63.33 μM, 49.16 μM, and 61.90 μM, and 49.31 μM, respectively. For the MCF-7 cell line, IC50 values of complex 1 and complex 2 were estimated to be 66.91 μM and 66.40 μM, respectively. From this study, it is revealed that both complex 1 and complex 2 have good cytotoxic effects against mouse and human cancer cells (Figures and ).

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A–D show the MTT assay of two mouse cancer cells; Sarcoma-180 and 4T1. Sarcoma-180 and 4T1 cells were exposed to various concentrations (1 μM, 5 μM, 10 μM, 20 μM, 40 μM, and 80 μM) of complex 1 and complex 2 and incubated for 24 h at 37 °C. Graph showing the live Sarcoma-180 and 4T1 cell percentage upon treatment with different concentrations of (A, B) complex 1 and (C, D) complex 2, compared with control cells. (E,F) Interpretation of the IC50 concentration of complex 1 and complex 2.

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(A, B) Concentration-dependent cytotoxic effect of 1 and 2 on MCF-7 cells. After treatment with different concentrations of 1 and 2, cells were incubated for 24 h. Graph showing the cell percentage of live MCF-7 cells upon treatment with different doses of 1 and 2 compared to the control. (C) Explication of the IC50 concentration of 1 and 2 against MCF-7 cells.

2.6. Effect of Complex 1 and Complex 2 on the MCF-7 Cell Cycle Distribution toward the Hypodiploid (Sub G0/G1) Stage

Following up on the cytotoxic effects of the synthetic complexes, we aimed to understand the cell cycle distribution of the copper complexes. One of the main characteristics of cancer initiation and progression is the dysregulation of the cell cycle stages. To study the different cell cycle stages, propidium iodide is widely used, which is a fluorogenic component that binds to the hydrogen bonds of DNA. The total amount of nuclear DNA changes depending on the stage of the cell cycle, mainly at different cell cycle checkpoints. The total emission of fluorescence generated by propidium iodide (PI) is proportional to the cellular DNA amount; thus, flowcytometric single-cell DNA-PI fluorescence measurement indicates the proportion of cells at a particular cell cycle stage.

In our study, we applied different doses of complex 1 and complex 2 to MCF-7 cells and incubated them for 24 h, along with control MCF-7 cells, vehicle control-treated cells, and positive control-treated cells. In the case of control cancer cells, the percentage of the hypoploid stage (indicating DNA damage and dead cells) was found to be 4.22%. But after the application of different concentrations (40, 60, and 80 μM) of complex 1 and complex 2, the percentage of the hypoploid stage significantly increased to 15.1%, 17.1%, and 28.4% for complex 1 and 14.9%, 23.5%, and 32.5% for complex 2, respectively. There was no significant change observed in the hypoploid stage after the application of the vehicle control, and in the case of the positive control, the percentage of the hypoploid stage was found to be 21.9% (Figure ).

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(A) Cell cycle assay of MCF-7 cells treated various concentrations of (40 μM/mL, 60 μM/mL, and 80 μM/mL) of complex 1 and complex 2 for 24 h. Proliferation of MCF-7 cells was strongly suppressed by complex 1 and complex 2 in a dose-dependent manner. (B) Graphical representation of the distribution of cells in different stages of the cell cycle.

2.7. Complex 1 and Complex 2 Induced Cellular Apoptosis in MCF-7 Cells in a Dose-Dependent Manner

Although the cell cycle assay of the copper complexes suggests an increase in hypoploid cells, which ensures copper­(II) complex-induced cell death, the cells cannot differentiate whether the death is apoptotic or necrotic. To check the proapoptotic role of complex 1 and complex 2 against MCF-7 cells, we performed the terminal deoxynucleotidyl transferase (bromo deoxy uridine or BrdU) nick end labeling or TUNEL assay. This assay allows the incorporation of fluorescently labeled BrdU at the 3′ end of nicked DNA, which is characteristic of apoptotic cell death, as apoptosis induces regular nicks in DNA causing DNA fragmentation. We found that complex 1 and complex 2 dose-dependently increased the TUNEL-positive cells in the case of the MCF-7 cell line (Figure ).

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TUNEL assay of MCF-7 cells showing the fragmented DNA via apoptosis. MCF-7 cells were treated with various concentrations of complex 1 and complex 2 and incubated for 24 h. Then, all the processes were done according to the manufacturer’s instructions followed by flow cytometric analysis. The dot plots and graph showing the percentage of TUNEL positive cells (which were stained with FITC tagged anti-BrdU antibody) were increased in a dose-dependent manner.

2.8. Complexes 1 and 2 Induced Early Apoptosis in MCF-7 Cells in a Dose-Dependent Manner

During the apoptosis process, a membrane phospholipid called phosphatidyl serine translocates toward the outer membrane of the dying cells. This translocation of phosphatidyl serine is an early indicator of the apoptosis pathway. Notably, Annexin V mainly binds with this phosphatidyl serine, thereby presenting conclusive evidence of an apoptotic event. From this study, we determined that complex 1 and complex 2 significantly increased apoptotic cell death after the application of different concentrations in the case of MCF-7 cells. Upon treatment with complex 1 and complex 2 at concentrations of 40 μM, 60 μM, and 80 μM, the percentage of Annexin V-positive apoptotic cells increased to 10.45%, 24.17%, and 24.52% in the case of complex 1 and 18.97%, 29.64%, and 33.32% in the case of complex 2 from 5.03% compared to control MCF-7 cells (Figure ).

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(A) Determination of apoptosis by Annexin V/PI double staining method. MCF-7 cells were treated with 40 μM/mL, 60 μM/mL, and 80 μM/mL doses of complex 1 and complex 2 and incubated for 24 h and compared with cancer control cells. (B) Graph showing a gradual increase of annexin-positive cells upon treatment with different concentrations of complex 1 and complex 2.

2.9. Complexes 1 and 2 Induced the Caspase Enzyme Activity in MCF-7 Cells

Caspase enzymes play a vital role in generating apoptosis. Caspases, a group of serine proteases, are broadly grouped as executioner caspases and initiator caspases. Two pathways are involved in the generation of initiator caspases, which are known as the extrinsic and intrinsic pathways. Both pathways then activate the executioner caspases, which in turn induce the downstream process of the cell death mechanism. To check the level of total active caspases generated in complex 1- and complex 2-treated MCF-7 cells, we applied a pan-caspase inhibitor, called z-VAD-FMK as a negative control, along with control cancer cells and selected concentrations of complex 1- and complex 2-treated cancer cells. A fluorogenic pan-caspase substrate FITC-VAD-FMK was used, which binds with all the different caspase family members, and fluorescence was captured using confocal imaging. Our choice of the pan-caspase substrate is prompted by the fact that MCF-7 lacks functional executioner caspase; MCF-7 cells do not express caspase-3, and apoptosis can be induced by other executioner caspases such as caspase-6 or caspase-7. Here, we found that the level of caspase increased significantly after the application of complex 1 and complex 2 in a dose-dependent manner (Figure ).

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(A) Confocal imaging of caspase assay showing that complex 1 and complex 2 increase the expression of caspase in a dose-dependent manner. Blue channel: Hoechst staining of nuclei, green channel: caspase FITC staining, overlay of blue and green channel image. (B) Graphical representation of the gray value of pan-caspase, which indicates that complex 1 and 2 significantly increases the production of caspase in MCF-7 cells.

2.10. Analysis of Change in CT-DNA Conformation

Various metal-based drugs primarily target DNA. The CD spectral analysis of complex 1 and complex 2 provides valuable evidence on the binding mode of metal complexes with DNA. Typically, the CD spectrum of the CT-DNA exhibits signature positive and negative bands at 275 and 245 nm, respectively. The positive band is attributed to the stacking of heterogeneous bases, and the negative band is correlated with the right-handed helicity of B-DNA. In general, the molecules’ groove binding and electrostatic interaction show less or no perturbation on the base stacking and helicity. At the same time, intercalation increases the intensities of both the positive and negative bands. , Increasing the concentration of complex 1 and complex 2 (Figure S12) increases the intensities of both the negative and positive bands of CT-DNA. This increased intensity in the negative band suggests that the complex can unwind the DNA helix and reduce its stability to a certain extent. Moreover, the increased intensity in the positive band indicates that the binding disturbs the right-handed helicity of DNA and induces certain conformational changes of the secondary structure within the DNA molecule, such as the conversion from a more B-like to a more C-like structure. These conformational changes are in the order of complex 2 > complex 1.

3. Conclusions

This research study provides an in-depth account of the synthesis, spectroscopic analysis, and structural characterization of newly formulated substituted benzimidazole ligands. Both synthetic complexes exhibit solution-phase durability in a water–DMSO mixture. Additionally, it explores two copper­(II) complexes that differ significantly in structure but incorporate the aforementioned ligands (L and L′). X-ray structural analysis reveals that the complexes adopt square planar geometries. Molecular docking studies attribute the stable protein-complex conformation to both complexes. MTT assay and hemolysis assay were conducted to assess the cytotoxicity and cell viability of complexes 1 and 2 against normal human PBMC and normal human RBC cells. Our findings indicate that concentrations up to 80 μM/mL for both complexes did not exhibit any toxic effects on normal human cells. The MTT assay was employed to evaluate the cytotoxic impact of 1 and 2 on mouse cancer cells, specifically Sarcoma-180 and the 4T1 mouse cancer cell line, as well as the human breast cancer cell line MCF-7. Both compounds demonstrated cytotoxic effects against mouse cancer cells and exhibited significant activity against both mouse and human breast cancer cells. Our study indicates that the application of different concentrations of complex 1 and complex 2 significantly enhanced apoptotic cell death, particularly in the case of MCF-7 cells. Upon exposure to complex 1 and complex 2 at concentrations of 40 μM, 60 μM, and 80 μM, Annexin V-positive apoptotic cells proportionally increased to 10.45%, 24.17%, and 24.52% for complex 1 and 18.97%, 29.64%, and 33.32% for complex 2, compared to the control MCF-7 cells with 5.03%. To assess the total caspase activity induced by complex 1 and complex 2 in the treated MCF-7 cells, we employed a pan-caspase inhibitor, z-VAD-FMK, as a negative control, along with control cancer cells and MCF-7 cells treated with selected concentrations of complex 1 and complex 2. The experimental outcomes confirmed a significant dose-dependent increase in the percentage of caspase activity following the application of complex 1 and complex 2. Taken together, this study provides insights into the activation of executioner caspases, initiating the downstream processes of the cell death mechanism and acting as potential candidates for breast cancer therapy. Complex 1 and complex 2 interact with DNA via intercalation, inducing helix unwinding and a transition from a B-like structure to a more C-like structure. Complex 2 exhibits stronger binding and greater conformational alterations than complex 1. Hence, these cost-effective synthetic compounds might be a new source of anticancer agents, paving a new horizon for benzimidazole-based anticancer drug development toward precision medicine.

4. Experimental Section

4.1. Preparation of the Benzimidazole Ligands and Copper­(II) Complexes

4.1.1. Chemicals, Solvents, and Starting Materials

O-Phenylenediamine ≥98.0% (Sigma-Aldrich, USA), >98.0% pure thiophene-2-aldehyde (Spectrochem, India), >97.0% 3,4-diaminotoluene (Sigma-Aldrich, USA), and copper­(II) nitrate trihydrate (Merck, India) were purchased from their respective outlets. All the chemicals used were of analytical grade.

4.1.2. Synthesis of the Benzimidazole Ligands

The benzimidazole ligands, 2-(thiophen-2-yl)-1-(thiophen-2-ylmethyl)-1H-benzo­[d]­imidazole (L) and 6-methyl-2-(thiophen-2-yl)-1-(thiophen-2-ylmethyl)-1H-benzo­[d]­imidazole (L′) were synthesized following the reported synthetic protocol. Aromatic amines, o-phenylenediamine (1 mmol, 0.108 g)/3,4-diaminotoluene (1 mmol, 0.123 g), were individually placed in 15 mL test tubes. Thiophene-2-aldehyde (2 mmol, 0.224 g) was added separately to each amine, along with 2 mL of water. These solution mixtures were dipped into an oil bath and heated at 75 °C. A controlled airflow of nearly 300 bubbles/min was maintained by immersing a syringe into the reaction mixture, using an aquarium air pump. This solution was left under continuous stirring for 12 h at 75 °C, followed by the addition of ethyl acetate (3 × 3 mL) to the organic part, which was then collected and dried over sodium sulfate for 2–3 h. Thereafter, the solvent was evaporated, and the resulting product was purified by column chromatography using ethyl acetate–hexane (15%) as the eluent and silica gel as the stationary phase.

Yield of L: 0.295 g (84%). Anal. Calc. for C16H12N2S2 (L): C, 64.83; H, 4.08; N, 9.45; found: C, 64.84; H, 4.11; N, 9.42; 1H NMR (DMSO-d 6, Figure S1): δ 7.74–7.72 [1H, Ph-H], 7.74–7.72 [3H, H-2-substituted thio], δ 7.41–7.40 [1H, Ph-H], 7.31–7.24 [3H, H-2-substituted thio], δ 7.03–6.97 [1H, Ph-H], δ 6.96–6.95 [1H, Ph-H], δ 5.94 [2H, CH 2]. 13C­{1H} NMR (DMSO-d 6, Figure S3): δ 147.20 (C in imidazole ring), δ 142.86–111.29 (PhC and ThioC), δ 43.50 (CH2); IR (KBr, cm–1): 3102 (νAr–C–H), 1622 (νCN); UV–vis (λmax, nm): 236, 303.

Yield of L′: 0.306 g (83%). Anal. Calc. for C17H14N2S2 (L′): C, 65.77; H, 4.55; N, 9.02; found: C, 65.75; H, 4.57; N, 9.04; 1H NMR (DMSO-d 6, Figure S2): δ 7.71–7.69 [1H, Ph-H], δ 7.50–7.49 [1H, Ph-H ], δ 7.44–7.43­[1H,Ph-H], δ 7.26–7.24 [1H, H-2-substituted thio], δ 7.16 [3H, H-2-substituted thio], 7.14–7.12 [1H, H-2-substituted thio], δ 6.87–6.86 [1H, H-2-substituted thio], δ 5.68 [2H, CH 2], δ 2.48 [3H, CH 3]. 13C­{1H}­(DMSO-d 6, Figure S4): δ 147.15 (H–C in imidazole ring), δ 141.11–109.43 (PhC and ThioC), δ 44.05 (CH2), δ 21.93–21.61­(CH3); IR (KBr, cm–1): 3103 (νAr–C–H), 1623 (νCN); UV–vis (λmax, nm): 232, 308.

4.1.3. Synthesis of Copper­(II) Complexes

A methanolic solution containing Cu­(NO3)2·3H2O (0.241 g, 1 mmol) was added dropwise into a methanolic solution of L (0.688 g, 2 mmol), and a methanolic solution containing Cu­(NO3)2·3H2O (0.241 g, 1 mmol) was added dropwise into a methanolic solution of L′ (0.620 g, 2 mmol) separately. The resulting reaction mixtures instantaneously turned green, which were further stirred for 20 min. The reaction solution was then allowed to undergo slow evaporation. After a week, green-colored microcrystalline products corresponding to complex 1 and complex 2 were separated and dried over silica gel for subsequent use.

Yield of complex 1: 0.1310 g (∼77.0% metal salt-based) Anal. for C32H24N6O6S4Cu: C, 49.00; H, 3.60; N, 10.71; found: C, 49.01; H, 3.55; N, 10.73. IR (KBr pellet, cm–1; Figure S5): 3023, 2931­(νCH2), 1617 (νCN); UV–vis (1 × 10–4 M, λmax(abs), nm, DMSO-H2O; Figure S6): 235, 303; HRMS (DMSO–H2O, m/z; Figure S7): 780.0258.

Yield of complex 2: 0.1850 g (∼76.80% metal salt-based) Anal. for C70H62N10O8S8Cu2: C, 53.79; H, 4.51; N, 8.96; found: C, 53.85; H, 4.47; N, 8.99; IR (KBr pellet, cm–1; Figure S5): 3025, 2935­(νCH2), 1621­(νCN); UV–vis (1 × 10–4 M, λmax(abs), nm, DMSO–H2O; Figure S6): 235, 301; HRMS (DMSO–H2O, m/z; Figure S8): 1430.0348.

4.2. Physical Measurements

FT-IR spectra of the L and L′ and copper complexes were recorded using an FTIR-8400S SHIMADZU spectrometer in the range of 400–3600 cm–1. 1H and 13C NMR spectra of L and L′ were obtained on a Bruker Avance 400 MHz spectrometer in DMSO-d 6 at 298 K. Steady-state absorption and other spectral data were recorded with a HITACHI U-2910 spectrophotometer. A PerkinElmer 2400 CHN microanalyzer was employed to carry out the elemental analyses of the compounds. Mass spectral measurements were performed with a Q-TOF micro quadrupole mass spectrometer.

4.3. X-Ray Structural Studies and Refinement

A Rigaku XtaLAB mini diffractometer equipped with a Mercury 375R (2 × 2 bin mode) CCD detector was employed to collect X-ray diffraction data for the copper­(II) complexes, with graphite monochromated Mo–Kα radiation (λ=0.71073 Å) at 293 K for both complex 1 and complex 2 using ω scans. The data were reduced, and the space group was determined using CrysAlisPro 1.171.39.35c and 1.171.39.7f. The crystal structures were solved by the dual-space method, and the space group was redetermined using SHELXT-2015. The crystallographic data were refined by full-matrix least-squares procedures using the SHELXL-2015 software package through the OLEX2 suite.

4.4. Molecular Docking

The complex structures were prepared using ChemDraw software, optimized, and converted into PDB files using the open Babel software. The complex structures were processed into PDBQT files using AutoDock4 software, and Gasteiger charges were added to the complexes. After completion of the docking procedure, the .dlg files of the complexes–protein structures were converted into .pdbqt files using AutoDock tools. Thereafter, with the help of PyMol software, these complexes are converted to the PDB format and also visualized by PyMol software. To analyze the interacting residues between the complexes and the receptor molecules, the Protein–Ligand Interaction Profiler (PLIP) (https://plip-tool.biotec.tu-dresden.de/plip-web/plip/index) is used.

The three-dimensional structures of BAX (4S0O), BCL2 (2W3L), p53 (1TUP), and PARP (4DQY) were downloaded from the RCSB Protein Data Bank (RCSB.org). The 3D structures of the complexes were generated using Discovery Studio. Using AutoDock 4.2 and MGL (Molecular Graphics Laboratory) Tools, the protein moiety was cleaned by deleting water and other heteroatoms, followed by adding polar hydrogens and Kollman charges. The related receptor moiety was saved in PDBQT format. Then, we performed ligand–protein docking using AutoDock 4.2 software. Here, we have used the Lamarckian genetic algorithm and performed ten runs. For postdocking analysis, the best docking poses were selected on the basis of their binding free energy and visualized in the Discovery Studio visualizer.

4.5. Materials and Methods for Biological Activities

Sarcoma-180 and 4T1, mouse cancer cell lines, and MCF-7, a human breast cancer cell line, were obtained from NCCS (National Centre for Cell Science, Pune, India). Human peripheral blood mononuclear cells and human red blood cells (R407-0050) were purchased from Himedia and Rockland Immunochemicals, respectively. High-glucose Dulbecco’s modified Eagle’s medium (DMEM), 10% fetal bovine serum (Himedia, India), and 1% penicillin and streptomycin mixture (Sigma-Aldrich, Germany) were used for the maintenance and culture of MCF-7 cells. The cell line was maintained at 37 °C in an incubator with 5% CO2. All of the experiments were performed in doubly deionized water. Escherichia coli (MTCC 1302, Gram-negative) was purchased from the Microbial Type Culture Collection, CSIR-IMTech, Chandigarh, India.

4.6. MTT Assay for Dose Kinetics, Cell Viability, and Hemolysis Assay

3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was performed to determine the cell viability and cytotoxicity of complex 1 and complex 2. The MTT reagent, possessing a positive charge and a lipophilic structure, facilitates penetration across both the cell membrane and the inner mitochondrial membrane. Sarcoma-180, 4T1, and MCF-7 cells were seeded at 1 × 104 cells/well in a 96-well cell culture plate and incubated overnight in a CO2 incubator at 37 °C to promote cell adhesion. Then, various concentrations of complex 1 and complex 2 (1 μM/mL, 5 μM/mL, 10 μM/mL, 20 μM/mL, 40 μM/mL, and 80 μM/mL) were added to these cells. DMSO and methotrexate served as the vehicle control and positive control, respectively. The cells were further incubated in a 5% CO2 incubator for 24 h at 37 °C. MTT reagent (Himedia) was added to the cells in each well and allowed to react for 2 h at 37 °C. When the MTT reagent was reduced to purple formazan crystals by metabolically active cells, to dissolve the formazan crystals, a solubilization solution was introduced. ELISA reader measured the absorbance of homogenized MTT-formazans at 540 nm. The toxicity of complex 1 and complex 2 against normal human PBMCs was checked by the MTT assay. ,

Based on the protocol of Henkelman et al. a hemolysis assay was performed, and the human RBC was taken at a concentration of 1 × 109/mL and incubated with different concentrations of these compounds at 37 °C for 1 h. After that, the samples were centrifuged, and 300 μL of supernatant was taken in a 96-well cell culture plate, and absorbance was read at 540 nm.

4.7. Cell Cycle Assay

Different stages of the cell cycle were assessed with flow cytometry. MCF-7 cells were plated (2 × 105 cells/well) on 24-well culture plates and incubated overnight so that the cells could adhere to the wells. Different concentrations of complex 1 and complex 2 (40 μM/mL, 60 μM/mL, and 80 μM/mL) were added to the cells and incubated in a 5% CO2 incubator for 24 h at 37 °C. DMSO was used as the vehicle control, and methotrexate was used as the positive control. The cell cycle assay was analyzed by flow cytometric. After 24 h of incubation, the cells were retrieved and centrifuged at 3000 rpm for 5 min. To fix and permeabilize the cells simultaneously, chilled methanol was applied to the cells and kept at −20 °C for 5 min. Then, these cells were diluted with PBS containing 50 μg/mL RNase-A and kept overnight at 4 °C. After that, cells were stained with propidium iodide (PI) (concentration was 10 μg/mL), and the fluorescence emission was acquired by a flow cytometer (CytoFLEX, Beckman Coulter, US) and analyzed with FlowJo software.

4.8. Apoptosis Assay

As per the manufacturer’s protocol, Annexin V was applied to determine the occurrence of cell death via apoptosis. MCF-7 cells (2 × 105 cells/well) were taken and allowed to adhere in a 24-well cell culture plate and then incubated with different concentrations (40 μΜ/mL, 60 μM/mL, and 80 μM/mL) of complex 1 and complex 2 applied to the cells along with control cancer cells, DMSO (vehicle control)-treated and methotrexate (positive control)-treated cells for 24 h at 37 °C. Then, the cells were collected and diluted in 1× binding buffer. The cells were then centrifuged at 2000 rpm for 10 min. The pellets were then stained with Annexin V-FITC (FITC Annexin V Apoptosis Detection Kit, BD Pharmingen, USA) and incubated for 30 min. Then, propidium iodide was introduced to the cells, and they were analyzed by a flow cytometer (CytoFLEX, Beckman Coulter, US, FlowJo software).

4.9. Caspase Assay

To perform the caspase assay, we first adhered MCF-7 cells on coverslips (105 cells/well) overnight at 37 °C in a CO2 incubator. Then, 60 μM and 80 μM concentrations of complex 1 and complex 2 were introduced to the cells and incubated for 24 h, along with control cancer cells. DMSO was used as the vehicle control. Z-VAD-FMK, a pan-caspase inhibitor was added to the cells as a negative control. After that, the fluorogenic caspase substrate FITC-VAD-FMK was added to the cells and incubated for 1 h and counter-stained with Hoechst for visualization of the nucleus. Then, the images were captured using a confocal microscope (Leica DMi8, Leica Biosystems).

4.10. Terminal dUTP Nick-End Labeling (TUNEL) Assay

To study the effect of complex 1 and complex 2 on the DNA integrity of MCF-7 cells, we performed the Terminal dUTP nick-end labeling (TUNEL) assay by using a TUNEL assay kit. First, 2 × 105 cells were seeded in a 24-well cell culture plate along with the most effective concentrations of complex 1 and complex 2 and incubated for 24 h. DMSO (10 μL/mL) was used as the vehicle control, while the positive control and the negative control were provided along with the TUNEL assay kit and prepared as per the manufacturer’s instructions. After incubation, the cells were collected and fixed with 1× fixation buffer (4% paraformaldehyde) for 15 min. Subsequently, the cells were centrifuged at 2000 rpm for 10 min, and the supernatants were discarded. Then, chilled 70% ethanol was added to each sample and incubated overnight at −20 °C. Then, all the samples were incubated with DNA labeling solution (BrdUTP, TdT enzyme, and reaction buffer) for 60 min at 37 °C. Then, all these samples were stained with Alexa Fluor-488 conjugated Anti-BrdU antibody and incubated for 30 min. The cells were then washed twice with PBS, PI/RNase was added to each sample, and the cells were analyzed using flow cytometry (Cytoflex, Beckman Coulter, USA).

4.11. CD Spectropolarimetry

The circular dichroism (CD) spectra were recorded on a JASCO J815 spectropolarimeter (JASCO International Co. Ltd., Hachioji, Japan) equipped with a JASCO temperature controller (model PFD-425L/15) interfaced with an HP PC at 20 ± 0.5 °C using instrument parameters reported previously.

CD spectra of CT-DNA (50 mM) before and after the addition of the copper complexes (10–50 mM) were recorded in phosphate buffer. Each sample solution was scanned in the range of 200–320 nm, and its CD spectra were generated after averaging three scans and subtracting the buffer background.

Supplementary Material

ao5c02269_si_001.pdf (853KB, pdf)
ao5c02269_si_002.cif (769KB, cif)
ao5c02269_si_003.cif (2.6MB, cif)
ao5c02269_si_004.pdf (98.5KB, pdf)
ao5c02269_si_005.pdf (115KB, pdf)

Acknowledgments

A.S. sincerely thanks the Department of Science and Technology for the fellowship under the INSPIRE scheme (2022/IF220637). M.C. thanks the UGC for providing the SJSGC fellowship (202223-UGCES-22-GE-WES-F-SJSGC-6410). M.M. and R.C. are also thankful to the Department of Science and Technology (DST/WOS-A/CS-106/2021) for their funding.

Supplementary crystallographic data for complex 1 and complex 2 are available free of charge from the Director, CCDC, 12 Union Road, Cambridge, CB2 1EZ, UK (fax: + 44–1223–336033; E-mail: deposit@ccdc.cam.ac.uk or website: http://www.ccdc.cam.ac.uk) upon request, quoting deposition numbers CCDC 2349926 and 2349925, respectively.

The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acsomega.5c02269.

  • Experimental details, mass spectra, FT-IR, UV–vis, and a table of crystallographic parameters (PDF)

  • CIF of complex 1 (CIF)

  • CIF of complex 2 (CIF)

  • checkCIF/PLATON report of thio (PDF)

  • checkCIF/PLATON report of thio1 (PDF)

A.S.: conceptualization, formal analysis, investigation; A.D.: conceptualization, formal analysis, investigation; M.C.: formal analysis, methodology, investigation; R.K.M.: formal analysis, methodology, investigation; D.D.: formal analysis, validation, investigation; D.S.: formal analysis, validation; R.C.: formal analysis, validation; S.B.: formal analysis, validation, investigation; M.M.: conceptualization, writing-reviewing and editing, supervision; B.B.: conceptualization, writing-reviewing and editing, supervision.

The authors declare no competing financial interest.

References

  1. Boiani M., Gonzalez M.. Imidazole and Benzimidazole Derivatives as Chemotherapeutic Agents. Mini-Rev. Med. Chem. 2005;5(4):409–424. doi: 10.2174/1389557053544047. [DOI] [PubMed] [Google Scholar]
  2. Mudi P. K., Mahanty A. K., Kotakonda M., Prasad S., Bhattacharyya S., Biswas B.. A Benzimidazole Scaffold as a Promising Inhibitor against SARS-CoV-2. J. Biomol. Struct. Dyn. 2023;41(5):1798–1810. doi: 10.1080/07391102.2021.2024448. [DOI] [PubMed] [Google Scholar]
  3. Narasimhan B., Sharma D., Kumar P.. Benzimidazole: A Medicinally Important Heterocyclic Moiety. Med. Chem. Res. 2012;21(3):269–283. doi: 10.1007/s00044-010-9533-9. [DOI] [Google Scholar]
  4. Mudi P. K., Mahato R. K., Verma H., Panda S. J., Purohit C. S., Silakari O., Biswas B.. In Silico Anti-SARS-CoV-2 Activities of Five-Membered Heterocycle-Substituted Benzimidazoles. J. Mol. Struct. 2022;1261:132869. doi: 10.1016/j.molstruc.2022.132869. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Yerragunta V., Gayathri R., Srujana S., Srujana S., Devi R., Divya A.. Benzimidazole Derivatives and Its Biological Importance: A Review. PharmaTutor. 2014;2(3):109–113. [Google Scholar]
  6. Patil A., Ganguly S., Surana S.. A Systematic Review of Benzimidazole Derivatives as an Antiulcer Agent. Rasayan J. Chem. 2008;1(3):447–460. [Google Scholar]
  7. Mudi P. K., Mahato R. K., Joshi M., Shit M., Choudhury A. R., Das H. S., Biswas B.. Copper­(II) Complexes with a Benzimidazole Functionalized Schiff Base: Synthesis, Crystal Structures, and Role of Ancillary Ions in Phenoxazinone Synthase Activity. Appl. Organomet. Chem. 2021;35(6):1–12. doi: 10.1002/aoc.6211. [DOI] [Google Scholar]
  8. Küçükbay H., Günal S., Orhan E., Durmaz R.. Synthesis and Antimicrobial Activities of Some Transition Metal Benzimidazole Complexes. Asian J. Chem. 2010;22(9):7376–7382. [Google Scholar]
  9. Aghatabay N. M., Neshat A., Karabiyik T., Somer M., Haciu D., Dülger B.. Synthesis, Characterization and Antimicrobial Activity of Fe­(II), Zn­(II), Cd­(II) and Hg­(II) Complexes with 2,6-Bis­(Benzimidazol-2-Yl) Pyridine Ligand. Eur. J. Med. Chem. 2007;42(2):205–213. doi: 10.1016/j.ejmech.2006.09.023. [DOI] [PubMed] [Google Scholar]
  10. Ülküseven B., Tavman A., Ötük G. S.. Characterization and Antimicrobial Activity of D8–10 Metal Complexes of Some 2-Substituted-1H-Benzimidazoles. Met. Based Drugs. 1999;6(3):163–167. doi: 10.1155/MBD.1999.163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Rogalewicz B., Czylkowska A.. Recent Advances in the Discovery of Copper­(II) Complexes as Potential Anticancer Drugs. Eur. J. Med. Chem. 2025;292:117702. doi: 10.1016/j.ejmech.2025.117702. [DOI] [PubMed] [Google Scholar]
  12. Hernández-Romero D., Rosete-Luna S., López-Monteon A., Chávez-Piña A., Pérez-Hernández N., Marroquín-Flores J., Cruz-Navarro A., Pesado-Gómez G., Morales-Morales D., Colorado-Peralta R.. First-Row Transition Metal Compounds Containing Benzimidazole Ligands: An Overview of Their Anticancer and Antitumor Activity. Coord. Chem. Rev. 2021;439:213930. doi: 10.1016/j.ccr.2021.213930. [DOI] [Google Scholar]
  13. Hamali M. A., Roney M., Dubey A., Uddin M. N., Zulkifli N. A., Fasihi Mohd Aluwi M. F., Musa M., Tajuddin A. M., Kassim K.. Cu­(II) Complexes Based on Benzimidazole Ligands: Synthesis, Characterization, DFT, Molecular Docking & Bioactivity Study. Future Med. Chem. 2024;16(23):2535–2546. doi: 10.1080/17568919.2024.2419353. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Alhomaidan L. M., Tar H., Alnafisah A. S., Aroua L. M., KouKi N., Alminderej F. M., Lalevee J.. Copper II Complexes Based on Benzimidazole Ligands as a Novel Photoredox Catalysis for Free Radical Polymerization Embedded Gold and Silver Nanoparticles. Polymers. 2023;15(5):1289. doi: 10.3390/polym15051289. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gawande M. B., Goswami A., Felpin F. X., Asefa T., Huang X., Silva R., Zou X., Zboril R., Varma R. S.. Cu and Cu-Based Nanoparticles: Synthesis and Applications in Catalysis. Chem. Rev. 2016;116:3722–3811. doi: 10.1021/acs.chemrev.5b00482. [DOI] [PubMed] [Google Scholar]
  16. Alajmi M. F., Hussain A., Rehman M. T., Khan A. A., Shaikh P. A., Khan R. A.. Design, Synthesis, and Biological Evaluation of Benzimidazole-Derived Biocompatible Copper­(II) and Zinc­(II) Complexes as Anticancer Chemotherapeutics. Int. J. Mol. Sci. 2018;19(5):1492. doi: 10.3390/ijms19051492. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Qiao X., Ma Z. Y., Shao J., Bao W. G., Xu J. Y., Qiang Z. Y., Lou J. S.. Biological Evaluation of a Cytotoxic 2-Substituted Benzimidazole Copper­(II) Complex: DNA Damage, Antiproliferation and Apoptotic Induction Activity in Human Cervical Cancer Cells. BioMetals. 2014;27(1):155–172. doi: 10.1007/s10534-013-9696-1. [DOI] [PubMed] [Google Scholar]
  18. Naveen P., Jain R., Kalaivani P., Shankar R., Dallemer F., Prabhakaran R.. Unpredicted Formation of Copper­(II) Complexes Containing 2-Thiophen-2-Yl-1-Thiophen-2-Ylmethyl-1: H -Benzoimidazole and Their Most Promising in Vitro Cytotoxicity in MCF-7 and HeLa Cell Lines over Cisplatin. New J. Chem. 2017;41(17):8885–8898. doi: 10.1039/C7NJ01273J. [DOI] [Google Scholar]
  19. Paul A., Gupta R. K., Dubey M., Sharma G., Koch B., Hundal G., Hundal M. S., Pandey D. S.. Potential Apoptosis Inducing Agents Based on a New Benzimidazole Schiff Base Ligand and Its Dicopper­(Ii) Complex. RSC Adv. 2014;40(78):41228–41236. doi: 10.1039/c4ra08680e. [DOI] [Google Scholar]
  20. Mahato S., Meheta N., Kotakonda M., Joshi M., Shit M., Choudhury A. R., Biswas B. S.. Synthesis, Structure, Polyphenol Oxidase Mimicking and Bactericidal Activity of a Zinc-Schiff Base Complex. Polyhedron. 2021;194:114933. doi: 10.1016/j.poly.2020.114933. [DOI] [Google Scholar]
  21. Mahato R. K., Mahanty A. K., Paul S., Gopal V., Perumalsamy B., Balakrishnan G., Ramasamy T., Dharumadurai D., Biswas B.. Catalytic Oxidative Coupling of O-Phenylenediamine, in-Vitro Antibacterial and Antitumor Activities of a Gold­(III)-Bipyridine Complex. J. Mol. Struct. 2021;1223:129264. doi: 10.1016/j.molstruc.2020.129264. [DOI] [Google Scholar]
  22. Mahato S., Meheta N., Kotakonda M., Joshi M., Ghosh P., Shit M., Choudhury A. R., Biswas B.. Ligand Directed Synthesis of a Unprecedented Tetragonalbipyramidal Copper (II) Complex and Its Antibacterial Activity and Catalytic Role in Oxidative Dimerisation of 2-Aminophenol. Appl. Organomet. Chem. 2020;34(11):e5935. doi: 10.1002/aoc.5935. [DOI] [Google Scholar]
  23. Sarkar D., Mandal B., Pramanik A., Haldar A., Das D., Ganguly R., Mandal D., Bhattacharyya S.. Synthesis, Structure, and Mechanism of Action of a Unique Mixed-Ligand Nickel (II)-Complex of Tetra-Dentate Phenol-Based Ligand with Di-Aquo Co-Ligands with Significant pro-Apoptotic and Anti-Metastatic Effect against Malignant Human Cancer Cells. Inorg. Chim. Acta. 2024;569:122136. doi: 10.1016/j.ica.2024.122136. [DOI] [Google Scholar]
  24. Ahmmed E., Sarkar D., Mondal A., Saha N. C., Bhattacharyya S., Chattopadhyay P.. A New Metal-Free Benzorhodol-Based Photoluminophore Selective for Carbon Monoxide Detection Applicable in Both in Vitro and in Vivo Bioimaging. Anal. Methods. 2022;14(33):3196–3202. doi: 10.1039/D2AY00835A. [DOI] [PubMed] [Google Scholar]
  25. Dasari S., Bernard Tchounwou P.. Cisplatin in Cancer Therapy: Molecular Mechanisms of Action. Eur. J. Pharmacol. 2014;740:364–378. doi: 10.1016/j.ejphar.2014.07.025. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. De Gooijer C. J., Baas P., Burgers J. A.. Current Chemotherapy Strategies in Malignant Pleural Mesothelioma. Transl. Lung Cancer Res. 2018;7(5):574–583. doi: 10.21037/tlcr.2018.04.10. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Lee Y. T., Tan Y. J., Oon C. E.. Benzimidazole and Its Derivatives as Cancer Therapeutics: The Potential Role from Traditional to Precision Medicine. Acta Pharm. Sin. B. 2023;13(2):478–497. doi: 10.1016/j.apsb.2022.09.010. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. du Bois A., Lück H. J., Meier W., Adams H. P., Möbus V., Costa S., Bauknecht T., Richter B., Warm M., Schröder W., Olbricht S., Nitz U., Jackisch C., Emons G., Wagner U., Kuhn W., Pfisterer J., von Maillot K., Lange W., Berg D., Schlicht E., Peterseim H., Elling D., Öney T., Zimmermann V., Renziehausen K., Rohrmann G., Bach H. J., Müller H., Jäger W., Mickan H., Ackermann R. H., Wernicke K., Czygan P. J., Schulze-Tollert J., Becker H. J., Nast J., Kramb P., Kröner M., Petru E., Carstensen M., Müller W., Zippel H. H., Hilfrich J., Herchenhein W., Mesrogli M., Schneider A., Deutsch G., Klöck F. K., Maurer W., Sünter S., Göppinger A., Strigl R., Schuhmann R., Kühndel K., Fischer D., Leißner C., Peters F., Niedner W., Peschke K. H., Silz T., Schwörer D., Meinerz W., Kramer D., Richter P., Steichele D. F., Krieger P., Lange M., Beck T., Friese K., Rother D., Heilmann L., Dietel J., Petri E., Meyer-Grohbrügge J., Jovanovic V., Robke K., Merkle E., Göretzlehner G., Hanker J. P., Karg C., Burkert W., Grüneberger A., Flachsenberg S.. A Randomized Clinical Trial of Cisplatin/Paclitaxel versus Carboplatin/Paclitaxel as First-Line Treatment of Ovarian Cancer. J. Natl. Cancer Inst. 2003;95(17):1320–1330. doi: 10.1093/jnci/djg036. [DOI] [PubMed] [Google Scholar]
  29. Giaccone G., Splinter T. A., Debruyne C., Kho G. S., Lianes P., Van Zandwijk N., Pennucci M. C., Scagliotti G., Van Meerbeeck J., Van Hoesel Q., Curran D., Sahmoud T., Postmus P. E.. Randomized Study of Paclitaxel-Cisplatin versus Cisplatin-Teniposide in Patients with Advanced Non-Small-Cell Lung Cancer. J. Clin. Oncol. 1998;16(6):2133–2141. doi: 10.1200/JCO.1998.16.6.2133. [DOI] [PubMed] [Google Scholar]
  30. Jeon J., Lee S., Kim H., Kang H., Youn H., Jo S., Youn B., Kim H.. Revisiting Platinum-based Anticancer Drugs to Overcome Gliomas. Int. J. Mol. Sci. 2021;22(10):5111. doi: 10.3390/ijms22105111. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Park S. J., Ye W., Xiao R., Silvin C., Padget M., Hodge J. W., Van Waes C., Schmitt N. C.. Cisplatin and Oxaliplatin Induce Similar Immunogenic Changes in Preclinical Models of Head and Neck Cancer. Oral Oncol. 2019;95:127–135. doi: 10.1016/j.oraloncology.2019.06.016. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Chovanec M., Cheng L.. Advances in Diagnosis and Treatment of Testicular Cancer. Bmj. 2022;379:e070499. doi: 10.1136/bmj-2022-070499. [DOI] [PubMed] [Google Scholar]
  33. Yao X., Panichpisal K., Kurtzman N., Nugent K.. Cisplatin Nephrotoxicity: A Review. Am. J. Med. Sci. 2007;334(2):115–124. doi: 10.1097/MAJ.0b013e31812dfe1e. [DOI] [PubMed] [Google Scholar]
  34. Florea A. M., Büsselberg D.. Cisplatin as an Anti-Tumor Drug: Cellular Mechanisms of Activity, Drug Resistance and Induced Side Effects. Cancers. 2011;3(1):1351–1371. doi: 10.3390/cancers3011351. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Hu J., Guo Y., Zhao J., Zhang J.. In Vitro Antitumor Activity of Novel Benzimidazole-Based Cu­(II) Complexes. Bioorg. Med. Chem. 2017;25(20):5733–5742. doi: 10.1016/j.bmc.2017.08.053. [DOI] [PubMed] [Google Scholar]
  36. Kim E. S.. Abemaciclib: First Global Approval. Drugs. 2017;77(18):2063–2070. doi: 10.1007/s40265-017-0840-z. [DOI] [PubMed] [Google Scholar]
  37. Niu Z. X., Wang Y. T., Sun J. F., Nie P., Herdewijn P.. Recent Advance of Clinically Approved Small-Molecule Drugs for the Treatment of Myeloid Leukemia. Eur. J. Med. Chem. 2023;261:115827. doi: 10.1016/j.ejmech.2023.115827. [DOI] [PubMed] [Google Scholar]
  38. Thompson D. L., Moore D. C.. Glasdegib: A Novel Hedgehog Pathway Inhibitor for Acute Myeloid Leukemia. J. Adv. Pract. Oncol. 2020;11(2):196–200. doi: 10.6004/jadpro.2020.11.2.8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Patil A., Ganguly S., Surana S.. Synthesis and Antiulcer Activity of 2- [5-Substituted-1- H- Benzo (d) Imidazol-2-Yl Sulfinyl] Methyl-3-Substituted Quinazoline-4- (3 H) Ones. J. Chem. Sci. 2010;122(3):443–450. doi: 10.1007/s12039-010-0052-5. [DOI] [Google Scholar]
  40. Wang P., Zheng G. J., Wang Y. P., Wang X. J., Wei H. G., Xiang W. S.. Highly Practical and Cost-Efficient Synthesis of Telmisartan: An Antihypertensive Drug. Tetrahedron. 2012;68(11):2509–2512. doi: 10.1016/j.tet.2012.01.056. [DOI] [Google Scholar]
  41. Walia R., Hedaitullah M., Naaz S. F., Iqbal K., Lamba H. S.. Benzimidazole Derivatives – an Overview. Int. J. Res. Pharm. Chem. 2011;1(3):565–574. [Google Scholar]
  42. Tahlan S., Kumar S., Narasimhan B.. Pharmacological Significance of Heterocyclic 1H-Benzimidazole Scaffolds: A Review. BMC Chem. 2019;13(1):1–21. doi: 10.1186/s13065-019-0625-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Hordyjewska A., Popiołek Ł., Kocot J.. The Many “Faces” of Copper in Medicine and Treatment. BioMetals. 2014;27(4):611–621. doi: 10.1007/s10534-014-9736-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Urso E., Maffia M.. Behind the Link between Copper and Angiogenesis: Established Mechanisms and an Overview on the Role of Vascular Copper Transport Systems. J. Vasc Res. 2016;52(3):172–196. doi: 10.1159/000438485. [DOI] [PubMed] [Google Scholar]
  45. Nasulewicz A., Mazur A., Opolski A.. Role of Copper in Tumour Angiogenesis - Clinical Implications. J. Trace Elem. Med. Biol. 2004;18(1):1–8. doi: 10.1016/j.jtemb.2004.02.004. [DOI] [PubMed] [Google Scholar]
  46. Geraki K., Farquharson M. J., Bradley D. A.. Concentrations of Fe, Cu and Zn in Breast Tissue: A Synchrotron XRF Study INSTITUTE OF PHYSICS PUBLISHING PHYSICS IN MEDICINE Concentrations of Fe, Cu and Zn in Breast Tissue: A Synchrotron XRF Study. Phys. Med. Biol. 2002;47:2327–2339. doi: 10.1088/0031-9155/47/13/310. [DOI] [PubMed] [Google Scholar]
  47. Singh N. K., Kumbhar A. A., Pokharel Y. R., Yadav P. N.. Anticancer Potency of Copper­(II) Complexes of Thiosemicarbazones. J. Inorg. Biochem. 2020;210:111134. doi: 10.1016/j.jinorgbio.2020.111134. [DOI] [PubMed] [Google Scholar]
  48. Tabassum S., Asim A., Khan R. A., Hussain Z., Srivastav S., Srikrishna S., Arjmand F.. Chiral Heterobimetallic Complexes Targeting Human DNA-Topoisomerase Iα. Dalton Trans. 2013;42(48):16749–16761. doi: 10.1039/c3dt51209f. [DOI] [PubMed] [Google Scholar]
  49. Mackenzie C. F., Spackman P. R., Jayatilaka D., Spackman M. A.. CrystalExplorer Model Energies and Energy Frameworks: Extension to Metal Coordination Compounds, Organic Salts, Solvates and Open-Shell Systems. IUCrJ. 2017;4:575–587. doi: 10.1107/S205225251700848X. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Lavrik I., Golks A., Krammer P. H.. Death Receptor Signaling. J. Cell Sci. 2005;118(2):265–267. doi: 10.1242/jcs.01610. [DOI] [PubMed] [Google Scholar]
  51. Khazaei S., Esa N. M., Ramachandran V., Hamid R. A., Pandurangan A. K., Etemad A., Ismail P.. In Vitro Antiproliferative and Apoptosis Inducing Effect of Allium Atroviolaceum Bulb Extract on Breast, Cervical, and Liver Cancer Cells. Front. Pharmacol. 2017;8(JAN):1–16. doi: 10.3389/fphar.2017.00005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Slee E. A., Adrain C., Martin S. J.. Executioner Caspase-3, −6and −7 Perform Distinct, Non-Redundant Roles during the Demolition Phase of Apoptosis. J. Biol. Chem. 2001;276(10):7320–7326. doi: 10.1074/jbc.M008363200. [DOI] [PubMed] [Google Scholar]
  53. Pal A., Biswas B., Mitra M., Rajalakshmi S., Purohit C. S., Hazra S., Kumar G. S., Nair B. U., Ghosh R.. DNA Binding and Cleavage Activity by a Mononuclear Iron­(II)­Schiff Base Complex: Synthesis and Structural Characterization. J. Chem. Sci. 2013;125(9):1161–1168. doi: 10.1007/s12039-013-0473-z. [DOI] [Google Scholar]
  54. Shahabadi N., Kashanian S., Darabi F.. DNA Binding and DNA Cleavage Studies of a Water Soluble Cobalt­(II) Complex Containing Dinitrogen Schiff Base Ligand: The Effect of Metal on the Mode of Binding. Eur. J. Med. Chem. 2010;45(9):4239–4245. doi: 10.1016/j.ejmech.2010.06.020. [DOI] [PubMed] [Google Scholar]
  55. CrysAlisPro Data Collection and Processing Software for Agilent X-Ray Diffractometers, Technologies UK Ltd: Yarnton, Oxford, UK, 2014; Vol: 44; pp. 1–53. [Google Scholar]
  56. Sheldrick G. M.. SHELXT - Integrated Space-Group and Crystal-Structure Determination. Acta Crystallogr., Sect. A. 2015;71(1):3–8. doi: 10.1107/S2053273314026370. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Dolomanov O. V., Bourhis L. J., Gildea R. J., Howard J. A. K., Puschmann H.. OLEX2: A Complete Structure Solution, Refinement and Analysis Program. J. Appl. Crystallogr. 2009;42(2):339–341. doi: 10.1107/S0021889808042726. [DOI] [Google Scholar]
  58. Notarte, K. I. R. ; Quimque, M. T. J. ; Macaranas, I. T. ; Khan, A. ; Pastrana, A. M. ; Villaflores, O. B. ; Arturo, H. C. P. ; Pilapil, D. Y. H. ; Tan, S. M. M. ; Wei, D.-Q. , et al. Flavonoids from Uvaria Alba Attenuate LPS-Induced Inflammatory Responses by Suppressing NF-ΚB Activation in RAW 264.7 Macrophages: In Silico and in Vitro Perspectives. ChemRxiv. 2022. [Google Scholar]
  59. Deb J., Majumder J., Bhattacharyya S., Jana S. S.. A Novel Naproxen Derivative Capable of Displaying Anti-Cancer and Anti-Migratory Properties against Human Breast Cancer Cells. BMC Cancer. 2014;14(1):567. doi: 10.1186/1471-2407-14-567. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. Kis B., Pavel I. Z., Avram S., Moaca E. A., Herrero San Juan M., Schwiebs A., Radeke H. H., Muntean D., Diaconeasa Z., Minda D., Oprean C., Bojin F., Dehelean C. A., Soica C., Danciu C.. Antimicrobial Activity, in Vitro Anticancer Effect (MCF-7 Breast Cancer Cell Line), Antiangiogenic and Immunomodulatory Potentials of Populus Nigra L. Buds Extract. BMC Complement. Med. Ther. 2022;22(1):74. doi: 10.1186/s12906-022-03526-z. [DOI] [PMC free article] [PubMed] [Google Scholar]
  61. Henkelman S., Rakhorst G., Blanton J., van Oeveren W.. Standardization of Incubation Conditions for Hemolysis Testing of Biomaterials. Mater. Sci. Eng. 2009;29(5):1650–1654. doi: 10.1016/j.msec.2009.01.002. [DOI] [Google Scholar]
  62. Bhadra K., Maiti M., Kumar G. S.. Molecular Recognition of DNA by Small Molecules: AT Base Pair Specific Intercalative Binding of Cytotoxic Plant Alkaloid Palmatine. Biochim. Biophys. Acta, Gen. Subj. 2007;1770(7):1071–1080. doi: 10.1016/j.bbagen.2007.03.001. [DOI] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

ao5c02269_si_001.pdf (853KB, pdf)
ao5c02269_si_002.cif (769KB, cif)
ao5c02269_si_003.cif (2.6MB, cif)
ao5c02269_si_004.pdf (98.5KB, pdf)
ao5c02269_si_005.pdf (115KB, pdf)

Data Availability Statement

Supplementary crystallographic data for complex 1 and complex 2 are available free of charge from the Director, CCDC, 12 Union Road, Cambridge, CB2 1EZ, UK (fax: + 44–1223–336033; E-mail: deposit@ccdc.cam.ac.uk or website: http://www.ccdc.cam.ac.uk) upon request, quoting deposition numbers CCDC 2349926 and 2349925, respectively.

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