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The clinical use of many potent anticancer agents is limited by their non-selective toxicity to healthy tissue. One of these examples is vorinostat (SAHA), a pan histone deacetylase inhibitor, which shows high cytotoxicity with limited discrimination for cancerous over healthy cells. In an attempt to improve tumor selectivity, we exploited the properties of cobalt(III) as a redox-active metal center through stabilization with cyclen and cyclam tetraazamacrocycles, masking the anticancer activity of SAHA and other hydroxamic acid derivatives to allow for the complex to reach the hypoxic microenvironment of the tumor. Biological assays demonstrated the desired low in vitro anticancer activity of the complexes, suggesting effective masking of the activity of SAHA. Once in the tumor, the bioactive moiety may be released through the reduction of the Co III center. Investigations revealed long-term stability of the complexes, with cyclic voltammetry and chemical reduction experiments supporting the design hypothesis of SAHA release through the reduction of the Co III prodrug. The results highlight the potential for further developing this complex class as novel anticancer agents by masking the high cytotoxicity of a given drug, however, the cellular uptake needs to be improved., (© 2024 The Authors. Chemistry - A European Journal published by Wiley-VCH GmbH.)

The cellular accumulation and the underlying mechanisms for the two ruthenium-based anticancer complexes [Ru II (cym)(HQ)Cl] 1 (cym = η 6 -p-cymene, HQ = 8-hydroxyquinoline) and [Ru II (cym)(PCA)Cl]Cl 2 (PCA = N-fluorophenyl-2-pyridinecarbothioamide) were investigated in HCT116 human colorectal carcinoma cells. The results showed that the cellular accumulation of both complexes increased over time and with higher concentrations, and that 2 accumulates in greater quantities in cells than 1. Inhibition studies of selected cellular accumulation mechanisms indicated that both 1 and 2 may be transported into the cells by both passive diffusion and active transporters, similar to cisplatin. Efflux experiments indicated that 1 and 2 are subjected to efflux through a mechanism that does not involve p-glycoprotein, as addition of verapamil did not make any difference. Exploring the influence of the Cu transporter by addition of CuCl 2 resulted in a higher accumulation of 1 and 2 whilst the amount of Pt detected was slightly reduced when cells were treated with cisplatin. Complexes 1 and 2 were further explored in zebrafish where accumulation and distribution were determined with ICP-MS and LA-ICP-MS. The results correlated with the in vitro observations and zebrafish treated with 2 showed higher Ru contents than those treated with 1. The distribution studies suggested that both complexes mainly accumulated in the intestines of the zebrafish., (© 2023. The Author(s), under exclusive licence to Society for Biological Inorganic Chemistry (SBIC).)

Intracellular accumulation studies are a key step in metallodrug development but often variable results are obtained. Therefore, we aimed here to investigate different protocols for efficient and reproducible lysis of cancer cells in terms of protein content in lysates and in cell uptake studies of the Ru anticancer complex [chlorido(8-oxyquinolinato)(η6-p-cymene)ruthenium(II)] ([Ru(cym)(HQ)Cl]). The physical lysis methods osmosis and sonication were chosen for comparison with chemical lysis with the radioimmunoprecipitation assay (RIPA) buffer. Based on the protein content and the total Ru accumulated in the lysates, the latter determined using inductively coupled plasma-mass spectrometry, RIPA buffer was the most efficient lysis method. Measurements of plastic adsorption blanks revealed that the higher Ru content determined in the RIPA buffer lysis samples may be due a higher amount of Ru extracted from the plastic incubation plates compared with osmosis and sonication. Overall, we found that the choice of lysis method needs to be matched to the information sought and we suggest the least disruptive osmosis method might be the best choice for labile drug-biomolecule adducts. Minimal differences were found for experiments aimed at measuring the overall cell uptake of the Ru complex., (© The Author(s) 2023. Published by Oxford University Press.)

Ruthenium piano-stool complexes have been explored for their anticancer activity and some promising compounds have been reported. Herein, we conjugated a derivative of plecstatin-1 to peptides in order to increase their cancer cell targeting ability. For this purpose, plecstatin-1 was modified at the arene ligand to introduce a functional amine handle ( 3 ), which resulted in a compound that showed similar activity in an in vitro anticancer activity assay. The cell-penetrating peptide TAT 48-60 , tumor-targeting neurotensin 8-13 , and plectin-targeting peptide were functionalized with succinyl or β-Ala-succinyl linkers under standard solid-phase peptide synthesis (SPPS) conditions to spatially separate the peptide backbones from the bioactive metal complexes. These modifications allowed for conjugating precursor 3 to the peptides on resin yielding the desired metal-peptide conjugates (MPCs), as confirmed by high-performance liquid chromatography (HPLC), NMR spectroscopy, and mass spectrometry (MS). The MPCs were studied for their behavior in aqueous solution and under acidic conditions and resembled that of the parent compound plecstatin-1. In in vitro anticancer activity studies in a small panel of cancer cell lines, the TAT-based MPCs showed the highest activity, while the other MPCs were virtually inactive. However, the MPCs were significantly less active than the small molecules plecstatin-1 and 3 , which can be explained by the reduced cell uptake as determined by inductively coupled plasma MS (ICP-MS). Although the MPCs did not display potent anticancer activities, the developed conjugation strategy can be extended toward other metal complexes, which may be able to utilize the targeting properties of peptides.

Pt(terpyridine) complexes are well-known DNA intercalators. The introduction of an NHC co-ligand rendered such a complex highly antiproliferative in cancer cells compared to its chlorido derivative. Despite the high potency, zebrafish embryos tolerated the compound well, especially compared to cisplatin. DNA interaction studies support a mode of action related to intercalation.

A new [PdPtL 4 ] 4+ heterobimetallic cage containing hydrazone linkages has been synthesised using the sub-component self-assembly approach. 1 H and DOSY nuclear magnetic resonance (NMR) spectroscopy and electrospray ionisation mass spectrometry (ESIMS) data were consistent with the formation of the [PdPtL 4 ] 4+ architecture. The cage was stimulus-responsive and could be partially disassembled and reassembled by the addition of dimethylaminopyridine (DMAP) and p -tolenesulfonic acid (TsOH), respectively. Additionally, the stability of the hydrazone cage against hydrolysis in the presence of water and nucleophilic decomposition in the presence of guest molecules was compared to a previously synthesised imine-containing [PdPtL 4 ] 4+ cage. It was established that the hydrazone linkage was more resistant to hydrolysis. Furthermore, the host-guest (HG) chemistry with a series of drug and drug-like molecules was examined. The hydrazone cage was shown to interact with cisplatin while the smaller imine cage was shown to interact with 5-fluorouracil and oxaliplatin in CD 3 CN. No HG interactions were observed in the more polar d 6 -DMSO. In vitro antiproliferative activity studies demonstrated both cages were active against the cancer cell lines tested and displayed half-maximal inhibitory (IC 50 ) values in the range of 25-35 μM. Most [PdPtL 4 ] 4+ -drug mixtures tested had higher IC 50 values than the hosts. However, the [PdPtL 4 ] 4+ cages, and [PdPtL 4 ] 4+ :drug mixtures were less cytotoxic than the well established anticancer drugs cisplatin, oxaliplatin and 5-fluorouracil.

The substitution of phenyl rings in established drugs with ferrocenyl moieties has been reported to yield compounds with improved biological activity and alternative modes of action, often involving the formation of reactive oxygen species (ROS). Translating this concept to N-heterocyclic carbene (NHC) complexes, we report here organometallics with a piano-stool structure that feature di- or tridentate ligand systems. The ligands impacted the cytotoxic activity of the NHC complexes, but the coordination modes seemed to have a limited influence, which may be related to the propensity of forming the same species in solution. In general, the stability of the complexes in an aqueous environment and their reactivity to selected biomolecules were largely dominated by the nature of the metal center. While the complexes promoted the formation of ROS, the levels did not correlate with their cytotoxic activity. However, the introduction of ferrocenyl moieties had a significant impact on the antiproliferative potency of the complexes and, in particular, some of the ferrocenyl-functionalized compounds yielded IC 50 values in the low μM range.

Metal complexes bind to a wide variety of biomolecules and the control of the reactivity is essential when designing anticancer metallodrugs with a specific mode of action in mind. In this study, we used the highly cytotoxic compound [RuII(cym)(8-HQ)Cl] (cym = η6-p-cymene, 8-HQ = 8-hydroxyquinoline), the more inert derivative [RuII(cym)(8-HQ)(PTA)](SO3CF3) (PTA = 1,3,5-triaza-7-phosphaadamantane), and [RuII(cym)(PCA)Cl]Cl (PCA = pyridinecarbothioamide) as a complex with a different coordination environment about the Ru center and investigated their stability, interactions with proteins, and behavior in medium (αMEM) and human serum by capillary zone electrophoresis. The developed method was found to be robust and provides a quick and low-cost technique to monitor the interactions of such complexes with biomolecules. Each complex was found to behave very differently, emphasizing the importance of the choice of ligands and demonstrating the applicability of the developed method. Additionally, the human serum albumin binding site preference of [RuII(cym)(8-HQ)Cl] was investigated through displacement studies, revealing that the compound was able to bind to both sites I and site II, and the type of adducts formed with transferrin was determined by mass spectrometry., (© The Author(s) 2022. Published by Oxford University Press.)

The number of donor atoms available on peptides that can competitively coordinate to metal centers renders the site-selective generation of advanced metal-peptide conjugates in high purity a challenging venture. Herein, we present a transmetalation-based synthetic approach on solid support in which an imidazolium pro-ligand can be used to selectively anchor a range of transition metal half-sandwich complexes onto peptides in the presence of multiple coordinative motifs. Amenable to solid support, a range of N-terminus and/or lysine conjugated metal-peptide conjugates were obtained in high purity after cleavage from the resin. The metalated peptides were evaluated for their anticancer properties against human cancer cell lines. While no cytotoxic activity was observed, this platform has the potential to i) provide a pathway to site-selective peptide labelling, ii) be explored as a biorthogonal handle and/or iii) generate a new strategy for ligand design in transition metal catalysts., (© 2021 Wiley-VCH GmbH.)

Redox-active Cu(II) complexes are able to form reactive oxygen species (ROS) in the presence of oxygen and reducing agents. Recently, Faller et al. reported that ROS generation by Cu(II) ATCUN complexes is not as high as assumed for decades. High complex stability results in silencing of the Cu(II)/Cu(I) redox cycle and therefore leads to low ROS generation. In this work, we demonstrate that an exchange of the α-amino acid Gly with the β-amino acid β-Ala at position 2 (Gly2→β-Ala2) of the ATCUN motif reinstates ROS production ( • OH and H 2 O 2 ). Potentiometry, cyclic voltammetry, EPR spectroscopy and DFT simulations were utilized to explain the increased ROS generation of these β-Ala2-containing ATCUN complexes. We also observed enhanced oxidative cleavage activity towards plasmid DNA for β-Ala2 compared to the Gly2 complexes. Modifications with positively charged Lys residues increased the DNA affinity through electrostatic interactions as determined by UV/VIS, fluorescence, and CD spectroscopy, and consequently led to a further increase in nuclease activity. A similar trend was observed regarding the cytotoxic activity of the complexes against several human cancer cell lines where β-Ala2 peptide complexes had lower IC 50 values compared to Gly2. The higher cytotoxicity could be attributed to an increased cellular uptake as determined by ICP-MS measurements., (© 2021 The Authors. Chemistry - A European Journal published by Wiley-VCH GmbH.)

Multimetallic complexes have been shown in several examples to possess greater anticancer activity than their monometallic counterparts. The increased activity has been attributed to altered modes of action. We herein report the synthesis of a series of heterodimetallic compounds based on a ditopic ligand featuring 2-pyridylimine chelating motifs and organometallic half-sandwich moieties. The complexes were characterized by a combination of 1 H NMR spectroscopy, electrospray ionization mass spectrometry, elemental analysis and single crystal X-ray diffraction. Investigations into the stability of representative complexes in DMSO- d 6 and 10% DMSO- d 6 /D 2 O revealed the occurrence of solvent-chlorido ligand exchange. Proliferation assays in four human cancer cell lines showed that the Os-Rh complex possessed minimal activity, while all other complexes were inactive., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Green, Steel, Riisom, Hanif, Söhnel, Jamieson, Wright, Crowley and Hartinger.)

Two new di(2,2'-bipyridine) ligands, 2,6-bis([2,2'-bipyridin]-5-ylethynyl)pyridine ( L1 ) and bis(4-([2,2'-bipyridin]-5-ylethynyl)phenyl)methane ( L2 ) were synthesized and used to generate two metallosupramolecular [Fe 2 ( L ) 3 ](BF 4 ) 4 cylinders. The ligands and cylinders were characterized using elemental analysis, electrospray ionization mass spectrometry, UV-vis, 1 H-, 13 C and DOSY nuclear magnetic resonance (NMR) spectroscopies. The molecular structures of the [Fe 2 ( L ) 3 ](BF 4 ) 4 cylinders were confirmed using X-ray crystallography. Both the [Fe 2 ( L1 ) 3 ](BF 4 ) 4 and [Fe 2 ( L2 ) 3 ](BF 4 ) 4 complexes crystallized as racemic ( rac ) mixtures of the ΔΔ (P) and ΛΛ (M) helicates. However, 1 H NMR spectra showed that in solution the larger [Fe 2 ( L2 ) 3 ](BF 4 ) 4 was a mixture of the rac -ΔΔ/ΛΛ and meso -ΔΛ isomers. The host-guest chemistry of the helicates, which both feature a central cavity, was examined with several small drug molecules. However, none of the potential guests were found to bind within the helicates. In vitro cytotoxicity assays demonstrated that both helicates were active against four cancer cell lines. The smaller [Fe 2 ( L1 ) 3 ](BF 4 ) 4 system displayed low μM activity against the HCT116 (IC 50 = 7.1 ± 0.5 μM) and NCI-H460 (IC 50 = 4.9 ± 0.4 μM) cancer cells. While the antiproliferative effects against all the cell lines examined were less than the well-known anticancer drug cisplatin, their modes of action would be expected to be very different., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Lisboa, Riisom, Vasdev, Jamieson, Wright, Hartinger and Crowley.)

Ruthenium-based complexes have attracted attention as promising anticancer candidates due to their lower general toxicity than the widely used platinum drugs. The complex [Ru II (η 6 -p-cymene)(8-oxyquinolinato)Cl] 1 has shown significant cytotoxic activity in cancer cells, independent of the cellular uptake. In an attempt to rationalize this finding, we investigated the fate of 1 in cells as well as developed an analysis method for 1 and its derivatives based on molecular mass spectrometry. The methods were applied for the analysis of cell medium and HCT116 human colorectal cancer cell lysate samples. The amount of Ru detected in cell lysate after treatment with 1 by ICP-MS was virtually time-independent, while the Ru content in the cell pellet increased significantly over the course of 24 h. The compound was still detectable by LC-ESI-TOF-MS after 24 h in cell lysate as its [1- Cl] + ion that may be formed directly from 1 or after a chlorido-aqua ligand exchange reaction facilitated in the cytosol.

Cisplatin is a widely used chemotherapeutic drug. Due to severe side effects and intrinsic or acquired resistance, there is a great interest in developing new platinum-based anticancer agents and a need for robust and validated analytical methods for determination of platinum accumulation in biological samples. A validated ICP-MS method for quantification of total carbon and platinum in cell samples is presented, applicable for cellular drug accumulation studies of platinum-based drugs, enabling estimation of drug accumulation while simultaneously determining carbon to monitor the sample digestion efficiency. Adequate precision (RSD <6%), accuracy and sensitivity were achieved for carbon and platinum determinations. Limits of detection were 0.9-3.0 mg/L for carbon and 0.11-0.50 μg/L for platinum. Determination of platinum by ICP-MS in cell samples digested applying either open-vessel or microwave-assisted acid digestion produced similar concentrations, although the residual carbon content in the sample solutions were significantly higher following open-vessel acid digestion compared to microwave-assisted acid digestion. Experiments showed that the residual carbon content after acid digestion did not have an influence on determination of total platinum by ICP-MS., (Copyright © 2018 Elsevier B.V. All rights reserved.)