Search

This review delves into recent advancements in the field of nitro(het)aromatic bioreductive agents tailored for hypoxic environments. These compounds are designed to exploit the low-oxygen conditions typically found in solid tumors, making them promising candidates for targeted cancer therapies. Initially, this review focused on their role as gene-directed enzyme prodrugs, which are inert until activated by specific enzymes within tumor cells. Upon activation, these prodrugs undergo chemical transformations that convert them into potent cytotoxic agents, selectively targeting cancerous tissue while sparing healthy cells. Additionally, this review discusses recent developments in prodrug conjugates containing nitro(het)aromatic moieties, designed to activate under low-oxygen conditions within tumors. This approach enhances their efficacy and specificity in cancer treatment. Furthermore, this review covers innovative research on using nitro(het)aromatic compounds as fluorescent probes for imaging hypoxic tumors. These probes enable non-invasive visualization of low-oxygen regions within tumors, providing valuable insights for the diagnosis, treatment planning, and monitoring of therapeutic responses. We hope this review will inspire researchers to design and synthesize improved compounds for selective cancer treatment and early diagnostics.

(1) Background: The aim of the work is the evaluation of in vitro antiproliferative and pro-apoptotic activity of four benzimidazole derivatives containing colchicine-like and catechol-like moieties with methyl group substitution in the benzimidazole ring against highly invasive breast cancer cell line MDA-MB-231 and their related impairment of tubulin dynamics. (2) Methods: The antiproliferative activity was assessed with the MTT assay. Alterations in tubulin polymerization were evaluated with an in vitro tubulin polymerization assay and a docking analysis. (3) Results: All derivatives showed time-dependent cytotoxicity with IC 50 varying from 40 to 60 μM after 48 h and between 13 and 20 μM after 72 h. Immunofluorescent and DAPI staining revealed the pro-apoptotic potential of benzimidazole derivatives and their effect on tubulin dynamics in living cells. Compound 5d prevented tubulin aggregation and blocked mitosis, highlighting the importance of the methyl group and the colchicine-like fragment. (4) Conclusions: The benzimidazole derivatives demonstrated moderate cytotoxicity towards MDA-MB-231 by retarding the initial phase of tubulin polymerization. The derivative 5d containing a colchicine-like moiety and methyl group substitution in the benzimidazole ring showed potential as an antiproliferative agent and microtubule destabilizer by facilitating faster microtubule aggregation and disrupting cellular and nuclear integrity.

Three new molecular complexes (phen) 3 (2-amino-Bz) 2 (H + )(BF 4 - )·3H 2 O 5 , (phen) 3 (2-amino-5(6)-methyl-Bz) 2 (H + )(BF 4 - )·H 2 O 6 , and (phen)(1-methyl-2-amino-Bz)(H + )(BF 4 - ) 7 , were prepared by self-assembly of 1,10-phenanthroline (phen) and various substituted 2-aminobenzimidazoles. Confirmation of their structures was established through spectroscopic methods and elemental analysis. The X-ray diffraction analysis revealed that the crystal structure of 7 is stabilized by the formation of hydrogen bonds and short contacts. In addition, the molecular geometry and electron structure of molecules 5 and 6 were theoretically evaluated using density functional theory (DFT) methods. According to the DFT B3LYP/6-311+G* calculations, the protonated benzimidazole (Bz) units act as NH hydrogen bond donors, binding two phenanthrolines and a BF 4 - ion. Non-protonated Bz unit form hydrogen bonds with the N-atoms of a third molecule phen. The molecular assembly is held together by π-π stacking between benzimidazole and phenanthroline rings, allowing for N-atoms to associate with water molecules. The complexes were tested in vitro for their tumor cell growth inhibitory effects on prostate (PC3), breast (MDA-MB-231 and MCF-7), and cervical (HeLa) cancer cell lines using MTT-dye reduction assay. The in vitro cytotoxicity analysis and spectrophotometric investigation in the presence of ct-DNA, showed that self-assembled molecules 5 - 7 are promising DNA-binding anticancer agents warranting further in-depth exploration.

Thienopyrimidines are structural analogs of quinazolines, and the creation of new 2-alkyl derivatives of ethyl 4-aminothienopyrimidine-6-carboxylates for the study of their anti-proliferative properties is of great pharmacological interest. Some 2-alkyl-4-amino-thieno[2,3- d ]pyrimidines 2 - 5 were synthesized, and their cyto- and phototoxicity against BALB 3T3 cells were established by an in vitro 3T3 NRU test. The obtained results indicate that the tested compounds are not cytotoxic or phototoxic, and that they are appropriate to be studied for their anti-proliferative and anti-tumor properties. The anti-proliferative potential of the compounds was investigated on MCF-7 and MDA-MB-231 cancer cells, as well as a MCF-10A cell line (normal human mammary epithelial cells). The most toxic to MCF-7 was thienopyrimidine 3 with IC 50 13.42 μg/mL (IC 50 0.045 μM), followed by compound 4 (IC 50 28.89 μg/mL or IC 50 0.11 μM). The thienopyrimidine 4 revealed higher selectivity to MCF-7 and lower activity (IC 50 367 μg/mL i.e., 1.4 μM) than compound 3 with MCF-10A cells. With respect to MDA-MB-231 cells, ester 2 manifested the highest effect with IC 50 52.56 μg/mL (IC 50 0.16 μM), and 2-ethyl derivative 4 revealed IC 50 62.86 μg/mL (IC 50 0.24 μM). It was estimated that the effect of the substances on the cell cycle progression was due to cell cycle arrest in the G2 stage for MDA-MB-231, while arrest in G1 was detected for the estrogen (ER)-positive MCF-7 cell line. The tested compound's effects on the change of the zeta potential in the tumorigenic cells utilized in this study were determined. The calculation which we performed of the physicochemical properties and pharmacokinetic parameters influencing the biological activity suggested high intestinal absorption, as well as drug-likeness., Competing Interests: The authors declare no conflict of interest.

[1,3,5]Triazino[1,2- a ]benzimidazole-2-amines bearing heterocyclic moiety in 4-position were synthesized. The compounds were characterized by elemental analysis, IR, 1 H-NMR, 13 C-NMR, and HRMS spectroscopy. The molecular geometry and electron structure of these molecules were theoretically studied using density functional theory (DFT) methods. The molecular structure of the synthesized fused triazinobenzimidazole was confirmed to correspond to the 3,4-dihydrotriazinobenzimidazole structure through the analysis of spectroscopic NMR data and DFT calculations. The antinematodic activity was evaluated in vitro on isolated encapsulated muscle larvae (ML) of Trichinella spiralis . The results showed that the tested triazinobenzimidazoles exhibit significantly higher efficiency than the conventional drug used to treat trichinosis, albendazole, at a concentration of 50 μg/mL. The compound 3c substituted with a thiophen-2-yl moiety exhibited the highest anthelmintic activity, with a larvicidal effect of 58.41% at a concentration of 50 μg/mL after 24 h of incubation. Following closely behind, the pyrrole analog 3f demonstrated 49.90% effectiveness at the same concentration. The preliminary structure-anti- T. spiralis activity relationship (SAR) of the analogues in the series was discussed. The cytotoxicity of the benzimidazole derivatives against two normal fibroblast cells (3T3 and CCL-1) and two cancer human cell lines (MCF-7 breast cancer cells and chronic myeloid leukemia cells AR-230) was evaluated using the MTT-dye reduction assay. The screening results indicated that the compounds showed no cytotoxicity against the tested cell lines. An in silico study of the physicochemical and pharmacokinetic characteristics of the novel synthesized fused triazinobenzimidazoles showed that they were characterized by a significant degree of drug-likeness and optimal properties for anthelmintic agents.

1H-benzimidazol-2-yl hydrazones with varying hydroxy and methoxy phenyl moieties were designed. Their effect on tubulin polymerization was evaluated in vitro on porcine tubulin. The compounds elongated the nucleation phase and slowed down the tubulin polymerization comparably to nocodazole. The possible binding modes of the hydrazones with tubulin were explored by molecular docking at the colchicine binding site. The anticancer activity was evaluated against human malignant cell lines MCF-7 and AR-230, as well as against normal fibroblast cells 3T3 and CCL-1. The compounds demonstrated a marked antineoplastic activity in low micromolar concentrations in both screened in vitro tumor models. The most active were the trimethoxy substituted derivative 1i and the positional isomers 1j and 1k , containing hydroxy and methoxy substituents: they showed IC 50 similar to the reference podophyllotoxin in both tumor cell lines, accompanied with high selectivity towards the malignantly transformed cells. The compounds exerted moderate to high ability to scavenge peroxyl radicals and certain derivatives- 1l containing metha -hydroxy and para -methoxy group, and 1b-e with di/trihydroxy phenyl moiety, revealed HORAC values high or comparable to those of well-known phenolic antioxidants. Thus the 1H-benisimidazol-2-yl hydrazones with hydroxy/methoxy phenyl fragments were recognized as new agents exhibiting promising combined antioxidant and antineoplastic action.

Novel 4-amino-thieno[2,3-d]pyrimidine-6-carboxylates substituted at the second position were prepared by cyclocondensation of 2-amino-3-cyano-thiophene and aryl nitriles in an acidic medium. The design of the target compounds was based on structural optimization. The derivatives thus obtained were tested in vitro against human and mouse cell lines. The examination of the compound effects on BLAB 3T3 and MFC-10A cells showed that they are safe, making them suitable for subsequent experiments to establish their antitumor activity. The photoirritancy factor of the compounds was calculated. Using the MTT test, the antiproliferative activity to MCF-10A, MCF-7 and MDA-MB-231 cell lines was estimated. The best antiproliferative effect in respect to the MCF-7 cell line revealed compound 2 with IC 50 4.3 ± 0.11 µg/mL (0.013 µM). The highest selective index with respect to MCF-7 cells was shown by compound 3 (SI = 19.3), and to MDA-MB-231 cells by compound 2 (SI = 3.7). Based on energy analysis, the most stable conformers were selected and optimized by means of density functional theory (DFT). Ligand efficiency, ligand lipophilicity efficiency and the physicochemical parameters of the target 4-amino-thienopyrimidines were determined. The data obtained indicated that the lead compound among the tested substances is compound 2 .

Novel biocompatible compounds that stabilize proteins in solution are in demand for biomedical and/or biotechnological applications. Here, we evaluated the effect of six ionic liquids, containing mono- or dicholinium [Chol] 1or2 cation and anions of charged amino acids such as lysine [Lys], arginine [Arg], aspartic acid [Asp], or glutamic acid [Glu], on the structure, thermal, and storage stability of the Rapana thomasiana hemocyanin (RtH). RtH is a protein with huge biomedicinal potential due to its therapeutic, drug carrier, and adjuvant properties. Overall, the ionic liquids (ILs) induce changes in the secondary structure of RtH. However, the structure near the Cu-active site seems unaltered and the oxygen-binding capacity of the protein is preserved. The ILs showed weak antibacterial activity when tested against three Gram-negative and three Gram-positive bacterial strains. On the contrary, [Chol][Arg] and [Chol][Lys] exhibited high anti-biofilm activity against E. coli 25213 and S. aureus 29213 strains. In addition, the two ILs were able to protect RtH from chemical and microbiological degradation. Maintained or enhanced thermal stability of RtH was observed in the presence of all ILs tested, except for RtH-[Chol] 2 [Glu].

The development of ionic liquids based on active pharmaceutical ingredients (API-ILs) is a possible solution to some of the problems of solid and/or hydrophobic drugs such as low solubility and bioavailability, polymorphism and an alternative route of administration could be suggested as compared to the classical drug. Here, we report for the first time the synthesis and detailed characterization of a series of ILs containing a cation amino acid esters and anion ketoprofen (KETO-ILs). The affinity and the binding mode of the KETO-ILs to bovine serum albumin (BSA) were assessed using fluorescence spectroscopy. All compounds bind in a distance not longer than 6.14 nm to the BSA fluorophores. The estimated binding constants (KA) are in order of 10 5 L mol -1 , which is indicative of strong drug or IL-BSA interactions. With respect to the ketoprofen-BSA system, a stronger affinity of the ILs containing l-LeuOEt, l-ValOBu, and l-ValOEt cation towards BSA is clearly seen. Fourier transformed infrared spectroscopy experiments have shown that all studied compounds induced a rearrangement of the protein molecule upon binding, which is consistent with the suggested static mechanism of BSA fluorescence quenching and formation of complexes between BSA and the drugs. All tested compounds were safe for macrophages.

Camphor and borneol are wildly distributed in the essential oils of medicinal plants from various parts of the World. Our study has been carried out to evaluate the effect of these two bicyclic monoterpenes on rat thymocytes. Camphor and borneol at concentrations of 0.5 and 5 µg/mL did not induce significant toxicity on the immune system cells, while a significant increase of thymocyte viability was detected when cells were incubated with 50 µg/mL of camphor. A significant increase of cell viability was similarly detected when thymocytes were cultivated with borneol at concentrations of 0.5 and 5 µg/mL. The role of camphor and borneol in reactive oxygen species (ROS) production and mitochondrial membrane potential (MMP) disturbances in rat thymocytes as well as their potential mechanism(s) of action were also discussed.