Supramolecular Coronation of Platinum(II) Complexes by Macrocycles: Structure, Relativistic DFT Calculations, and Biological Effects.

Platinum-based anticancer drugs are actively developed utilizing lipophilic ligands or drug carriers for the efficient penetration of biomembranes, reduction of side effects, and tumor targeting. We report the development of a supramolecular host-guest system built on cationic platinum(II) compounds bearing ligands anchored in the cavity of the macrocyclic host. The host-guest binding and hydrolysis process on the platinum core were investigated in detail by using NMR, MS, X-ray diffraction, and relativistic DFT calculations. The encapsulation process in cucurbit[7]uril unequivocally promotes the stability of hydrolyzed dicationic cis-[PtII(NH3)2(H2O)(NH2-R)]2+ compared to its trans isomer. Biological screening on the ovarian cancer lines A2780 and A2780/CP shows time-dependent toxicity. Notably, the reported complex and its ß-cyclodextrin (ß-CD) assembly achieve the same cellular uptake as cisplatin and cisplatin@ß-CD, respectively, while maintaining a significantly lower toxicity profile.

Sensitivity to Cisplatin in Head and Neck Cancer Cells Is Significantly Affected by Patient-Derived Cancer-Associated Fibroblasts.

Cancer-associated fibroblasts (CAFs) are one of the most abundant and critical components of the tumor stroma. CAFs can impact many important steps of cancerogenesis and may also influence treatment resistance. Some of these effects need the direct contact of CAFs and cancer cells, while some involve paracrine signals. In this study, we investigated the ability of head and neck squamous cell carcinomas (HNSCC) patient-derived CAFs to promote or inhibit the colony-forming ability of HNSCC cells. The effect of cisplatin on this promoting or inhibiting influence was also studied. The subsequent analysis focused on changes in the expression of genes associated with cancer progression. We found that cisplatin response in model HNSCC cancer cells was modified by coculture with CAFs, was CAF-specific, and different patient-derived CAFs had a different "sensitizing ratio". Increased expression of VEGFA, PGE2S, COX2, EGFR, and NANOG in cancer cells was characteristic for the increase of resistance. On the other hand, CCL2 expression was associated with sensitizing effect. Significantly higher amounts of cisplatin were found in CAFs derived from patients who subsequently experienced a recurrence. In conclusion, our results showed that CAFs could promote and/or inhibit colony-forming capability and cisplatin resistance in HNSCC cells via paracrine effects and subsequent changes in gene expression of cancer-associated genes in cancer cells.

Leachability of metals from waste incineration residues by iron- and sulfur-oxidizing bacteria.

Hazardous waste disposal via incineration generates a substantial amount of ashes and slags which pose an environmental risk due to their toxicity. Currently, these residues are deposited in landfills with loss of potentially recyclable raw material. In this study, the use of acidophilic bioleaching bacteria (Acidithiobacillus ferrooxidans, Acidithiobacillus thiooxidans, and Leptospirillum ferrooxidans) as an environmentally friendly, efficient strategy for the recovery of valuable metals from incineration residues was investigated. Zinc, Cobalt, Copper, and Manganese from three different incineration residues were bio-extracted up to 100% using A. ferrooxidans under ferrous iron oxidation. The other metals showed lower leaching efficiencies based on the type of culture used. Sulfur-oxidizing cultures A. ferrooxidans and A. thiooxidans, containing sulfur as the sole substrate, expressed a significantly lower leaching efficiency (up to 50%). According to ICP-MS, ashes and slags contained Fe, Zn, Cu, Mn, Cr, Cd, and Ni in economically attractive concentrations between 0.2 and 75 mg g-1. Compared to conventional hydrometallurgical and pyrometallurgical processes, our biological approach provides many advantages such as: the use of a limited amount of used strong acids (H2SO4 or HCl), recycling operations at lower temperatures (~30 °C) and no emission of toxic gases during combustion (i.e., dioxins and furans).

Comparison of Metal Nanoparticles (Au, Ag, Eu, Cd) Used for Immunoanalysis Using LA-ICP-MS Detection.

Immunochemical methods are used not only in clinical practice for the diagnosis of a wide range of diseases but also in basic and advanced research. Based on the unique reaction between the antibody and its respective antigens, it serves to specifically recognize target molecules in biological complex samples. Current methods of labelling antibodies with elemental labels followed by detection by inductively coupled plasma mass spectrometry (ICP-MS) allow detection of multiple antigens in parallel in a single analysis. Using the laser ablation (LA) modality (LA-ICP-MS), it is also possible to monitor the spatial distribution of biogenic elements. Moreover, the employment of metal nanoparticle-labeled antibodies expands the applicability also to molecular imaging by LA-ICP-MS. In this work, conjugates of model monoclonal antibody (DO-1, recognizing p53 protein) with various metal nanoparticles-based labels were created and utilized in dot-blot analysis in order to compare their benefits and disadvantages. Based on experiments with the p53 protein standard, commercial kits of gold nanoparticles proved to be the most suitable for the preparation of conjugates. The LA-ICP-MS demonstrated very good repeatability, wide linear dynamic range (0.1-14 ng), and limit of detection was calculated as a 1.3 pg of p53 protein.

A Clearance Period after Soluble Lead Nanoparticle Inhalation Did Not Ameliorate the Negative Effects on Target Tissues Due to Decreased Immune Response.

The inhalation of metal (including lead) nanoparticles poses a real health issue to people and animals living in polluted and/or industrial areas. In this study, we exposed mice to lead(II) nitrate nanoparticles [Pb(NO3)2 NPs], which represent a highly soluble form of lead, by inhalation. We aimed to uncover the effects of their exposure on individual target organs and to reveal potential variability in the lead clearance. We examined (i) lead biodistribution in target organs using laser ablation and inductively coupled plasma mass spectrometry (LA-ICP-MS) and atomic absorption spectrometry (AAS), (ii) lead effect on histopathological changes and immune cells response in secondary target organs and (iii) the clearance ability of target organs. In the lungs and liver, Pb(NO3)2 NP inhalation induced serious structural changes and their damage was present even after a 5-week clearance period despite the lead having been almost completely eliminated from the tissues. The numbers of macrophages significantly decreased after 11-week Pb(NO3)2 NP inhalation; conversely, abundance of alpha-smooth muscle actin (α-SMA)-positive cells, which are responsible for augmented collagen production, increased in both tissues. Moreover, the expression of nuclear factor κB (NF-κB) and selected cytokines, such as tumor necrosis factor alpha (TNFα), transforming growth factor beta 1 (TGFß1), interleukin 6(IL-6), IL-1α and IL-1ß , displayed a tissue-specific response to lead exposure. In summary, diminished inflammatory response in tissues after Pb(NO3)2 NPs inhalation was associated with prolonged negative effect of lead on tissues, as demonstrated by sustained pathological changes in target organs, even after long clearance period.

Selectively Oxidized Cellulose with Adjustable Molecular Weight for Controlled Release of Platinum Anticancer Drugs.

The synthesis of selectively oxidized cellulose, 2,3-dicarboxycellulose (DCC), is optimized for preparation of highly oxidized material for biological applications, which includes control over the molecular weight of the product during its synthesis. Conjugates of DCC and cisplatin simultaneously offer a very high drug binding efficiency (>90%) and drug loading capacity (up to 50 wt %), while retaining good aqueous solubility. The adjustable molecular weight of the DCC together with variances in drug feeding ratio allows to optimize cisplatin release profiles from delayed (<2% of cisplatin released during 6 h) to classical burst release with more than 60% of cisplatin released after 24 h. The release rates are also pH-dependent (up to 2 times faster release at pH 5.5 than at pH 7.4), which allows to exploit the acidic nature of tumor microenvironment. Extensive in vitro studies were performed on eight different cell lines for two cisplatin-DCC conjugates with different release profiles. In comparison with free cisplatin, both cisplatin-DCC conjugates demonstrated considerably lower cytotoxicity toward healthy cells. Conjugates with burst release profiles were found more effective against prostate cell lines, while DCC conjugates with slower release were more cytotoxic against ovarian and lung carcinoma cell lines. In vivo studies indicated a significantly longer survival rate, a reduction in tumor volume, and a higher accumulation of platinum in tumors of mice treated with the cisplatin-DCC conjugate in comparison to those treated by free cisplatin.

Gold nanoparticles as labels for immunochemical analysis using laser ablation inductively coupled plasma mass spectrometry.

In this paper, we describe the labelling of antibodies by gold nanoparticles (AuNPs) with diameters of 10 and 60 nm with detection by laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS). Additionally, the AuNPs labelling strategy is compared with commercially available labelling reagents based on MeCAT (metal coded affinity tagging). Proof of principle experiments based on dot blot experiments were performed. The two labelling methods investigated were compared by sensitivity and limit of detection (LOD). The absolute LODs achieved were in the range of tens of picograms for AuNP labelling compared to a few hundred picograms by the MeCAT labelling.

DNA interaction with platinum-based cytostatics revealed by DNA sequencing.

The main mechanism of action of platinum-based cytostatic drugs - cisplatin, oxaliplatin and carboplatin - is the formation of DNA cross-links, which restricts the transcription due to the disability of DNA to enter the active site of the polymerase. The polymerase chain reaction (PCR) was employed as a simplified model of the amplification process in the cell nucleus. PCR with fluorescently labelled dideoxynucleotides commonly employed for DNA sequencing was used to monitor the effect of platinum-based cytostatics on DNA in terms of decrease in labeling efficiency dependent on a presence of the DNA-drug cross-link. It was found that significantly different amounts of the drugs - cisplatin (0.21 µg/mL), oxaliplatin (5.23 µg/mL), and carboplatin (71.11 µg/mL) - were required to cause the same quenching effect (50%) on the fluorescent labelling of 50 µg/mL of DNA. Moreover, it was found that even though the amounts of the drugs was applied to the reaction mixture differing by several orders of magnitude, the amount of incorporated platinum, quantified by inductively coupled plasma mass spectrometry, was in all cases at the level of tenths of µg per 5 µg of DNA.

Direct Analysis of Gold Nanoparticles from Dried Droplets Using Substrate-Assisted Laser Desorption Single Particle-ICPMS.

Single particle inductively coupled plasma mass spectrometry (SP-ICPMS) has been generally accepted as a powerful tool in the field of nanoanalysis. The method has usually been restricted to direct nanoparticle (NP) introduction using nebulization or microdroplet generation systems. In this work, AuNPs are introduced into ICPMS by substrate-assisted laser desorption (SALD) directly from a suitable absorbing plastic surface using a commercial ablation cell for the first time. In SALD, desorption of individual NPs is mediated using a frequency-quintupled Nd:YAG laser (213 nm) operated at a rather low laser fluence. Conditions including laser fluence, laser beam scan rate, and carrier gas flow rate were optimized in order to gain the highest AuNP transport efficiency and avoid AuNP disintegration within the laser irradiation. The method was demonstrated on a well-characterized reference material, 56 nm AuNPs with a transport efficiency of 61% and commercially available 86 nm AuNPs. Feasibility of our technique for NP detection and characterization is discussed here, and the results are compared with an established technique, nebulizer SP-ICPMS.

Joint forces of mass spectrometric techniques (ICP-MS and MALDI-TOF-MS) and fluorescence spectrometry in the study of platinum-based cytostatic drugs interactions with metallothionein MT2 and MT3.

Herby, the interaction of metallothioneins with commonly used Pt-based anticancer drugs - cisplatin, carboplatin, and oxaliplatin - was investigated using the combined power of elemental (i.e. LA-ICP-MS, CE-ICP-MS) and molecular (i.e. MALDI-TOF-MS) analytical techniques providing not only required information about the interaction, but also the benefit of low sample consumption. The amount of Cd and Pt incorporated within the protein was determined for protein monomers and dimer/oligomers formed by non-oxidative dimerization. Moreover, fluorescence spectrometry using Zn2+-selective fluorescent indicator - FluoZin3 - was employed to monitor the ability of Pt drugs to release natively occurring Zn from the protein molecule. The investigation was carried out using two protein isoforms (i.e. MT2, MT3), and significant differences in behaviour of these two isoforms were observed. The main attention was paid to elucidating whether the protein dimerization/oligomerization may be the reason for the potential failure of the anticancer therapy based on these drugs. Based on the results, it was demonstrated that the interaction of MT2 (both monomers and dimers) interacted with Pt drugs significantly less compared to MT3 (both monomers and dimers). Also, a significant difference between monomeric and dimeric forms (both MT2 and MT3) was not observed. This may suggest that dimer formation is not the key factor leading to the inactivation of Pt drugs.

Unveiling the nanotoxicological aspects of Se nanomaterials differing in size and morphology.

Although the general concept of nanotechnology relies on exploitation of size-dependent properties of nanoscaled materials, the relation between the size/morphology of nanoparticles with their biological activity remains not well understood. Therefore, we aimed at investigating the biological activity of Se nanoparticles, one of the most promising candidates of nanomaterials for biomedicine, possessing the same crystal structure, but differing in morphology (nanorods vs. spherical particles) and aspect ratios (AR, 11.5 vs. 22.3 vs. 1.0) in human cells and BALB/c mice. Herein, we report that in case of nanorod-shaped Se nanomaterials, AR is a critical factor describing their cytotoxicity and biocompatibility. However, spherical nanoparticles (AR 1.0) do not fit this statement and exhibit markedly higher cytotoxicity than lower-AR Se nanorods. Beside of cytotoxicity, we also show that morphology and size substantially affect the uptake and intracellular fate of Se nanomaterials. In line with in vitro data, in vivo i.v. administration of Se nanomaterials revealed the highest toxicity for higher-AR nanorods followed by spherical nanoparticles and lower-AR nanorods. Moreover, we revealed that Se nanomaterials are able to alter intracellular redox homeostasis, and affect the acidic intracellular vesicles and cytoskeletal architecture in a size- and morphology-dependent manner. Although the tested nanoparticles were produced from the similar sources, their behavior differs markedly, since each type is promising for several various application scenarios, and the presented testing protocol could serve as a concept standardizing the biological relevance of the size and morphology of the various types of nanomaterials and nanoparticles.

Zinc recovery from bioleachate using a microbial electrolysis cell and comparison with selective precipitation.

Metal recycling is essential for strengthening a circular economy. Microbial leaching (bioleaching) is an economical and environmentally friendly technology widely used to extract metals from insoluble ores or secondary resources such as dust, ashes, and slags. On the other hand, microbial electrolysis cells (MECs) would offer an energy-efficient application for recovering valuable metals from an aqueous solution. In this study, we investigated a MEC for Zn recovery from metal-laden bioleachate for the first time by applying a constant potential of -100 mV vs. Ag/AgCl (3 M NaCl) on a synthetic wastewater-treating bioanode. Zn was deposited onto the cathode surface with a recovery efficiency of 41 ± 13% and an energy consumption of 2.55 kWh kg-1. For comparison, Zn recovery from zinc sulfate solution resulted in a Zn recovery efficiency of 100 ± 0% and an energy consumption of 0.70 kWh kg-1. Furthermore, selective metal precipitation of the bioleachate was performed. Individual metals were almost completely precipitated from the bioleachate at pH 5 (Al), pH 7 (Zn and Fe), and pH 9 (Mg and Mn).

Optimized biogenic sulfuric acid production and application in the treatment of waste incineration residues.

The biological leaching of metals from different waste streams by bacteria is intensively investigated to address metal recycling and circular economy goals. However, usually external addition of sulfuric acid is required to maintain the low pH optimum of the bacteria to ensure efficient leaching. Extremely acidophilic Acidithiobacillus spp. producing sulfuric acid and ferric iron have been investigated for several decades in the bioleaching of metal-containing ores. Their application has now been extended to the extraction of metals from artificial ores and other secondary sources. In this study, an optimized process for producing biogenic sulfuric acid from elemental sulfur by two sulfur-oxidizing species, A. thiooxidans and A. caldus and their combinations, was investigated in batch and stirred tank experiments. Using a combined culture of both species, 1.05 M and 1.4 M biogenic sulfuric acid was produced at 30 °C and 6% elemental sulfur in batch and semi continuous modes, respectively. The acid produced was then used to control the pH in a heap bioleaching system in which iron- and sulfur-oxidizing A. ferrooxidans was applied to biologically leach metals from waste incineration residuals. Metals like Cu, Ni, Al, Mn, and Zn were successfully recovered by 99, 93, 84, 77 and 68%, respectively within three weeks of heap bioleaching. Overall, a potential value recovery of incorporated metals >70% could be achieved. This highlights the potential and novelty of applying extremely acidophilic sulfur-oxidizing bacteria for cheap and efficient production of biogenic sulfuric acid and its use in pH control.

UV-Induced fingerprint spectroscopy.

Here, we present the potential analytical applications of photochemistry in combination with fluorescence fingerprinting. Our approach analyzes the fluorescence of samples after ultraviolet light (UV) treatment. Especially in presence of metal ions and thiol-containing compounds, the fluorescence behavior changes considerably. The UV-induced reactions (changes) are unique to a given sample composition, resulting in distinct patterns or fingerprints (typically in the 230-600 nm spectral region). This method works without the need for additional chemicals or fluorescent probes, only suitable diluent must be used. The proposed method (UV fingerprinting) suggests the option of recognizing various types of pharmaceuticals, beverages (juices and wines), and other samples within only a few minutes. In some studied samples (e.g. pharmaceuticals), significant changes in fluorescence characteristics (mainly fluorescence intensity) were observed. We believe that the fingerprinting technique can provide an innovative solution for analytical detection.

Mapping of MeLiM melanoma combining ICP-MS and MALDI-MSI methods.

Here we developed a powerful tool for comprehensive data collection and mapping of molecular and elemental signatures in the Melanoma-bearing Libechov Minipig (MeLiM) model. The combination of different mass spectrometric methods allowed for detail investigation of specific melanoma markers and elements and their spatial distribution in tissue sections. MALDI-MSI combined with HPLC-MS/MS analyses resulted in identification of seven specific proteins, S100A12, CD163, MMP-2, galectin-1, tenascin, resistin and PCNA that were presented in the melanoma signatures. Furthermore, the ICP-MS method allowed for spatial detection of zinc, calcium, copper, and iron elements linked with the allocation of the specific binding proteins.

Determination of aluminium in groundwater samples by GF-AAS, ICP-AES, ICP-MS and modelling of inorganic aluminium complexes.

The paper presents the results of aluminium determinations in ground water samples of the Miocene aquifer from the area of the city of Poznan (Poland). The determined aluminium content amounted from <0.0001 to 752.7 µg L(-1). The aluminium determinations were performed using three analytical techniques: graphite furnace atomic absorption spectrometry (GF-AAS), inductively coupled plasma atomic emission spectrometry (ICP-AES) and inductively coupled plasma mass spectrometry (ICP-MS). The results of aluminium determinations in groundwater samples for particular analytical techniques were compared. The results were used to identify the ascent of ground water from the Mesozoic aquifer to the Miocene aquifer in the area of the fault graben. Using the Mineql+ program, the modelling of the occurrence of aluminium and the following aluminium complexes: hydroxy, with fluorides and sulphates was performed. The paper presents the results of aluminium determinations in ground water using different analytical techniques as well as the chemical modelling in the Mineql+ program, which was performed for the first time and which enabled the identification of aluminium complexes in the investigated samples. The study confirms the occurrence of aluminium hydroxy complexes and aluminium fluoride complexes in the analysed groundwater samples. Despite the dominance of sulphates and organic matter in the sample, major participation of the complexes with these ligands was not stated based on the modelling.

Determination of Renal Distribution of Zinc, Copper, Iron, and Platinum in Mouse Kidney Using LA-ICP-MS.

The main dose-limiting side effect of cisplatin is nephrotoxicity. The utilization of cisplatin is an issue of balancing tumour toxicity versus platinum-induced nephrotoxicity. In this study, we focused on intraorgan distribution of common essential trace elements zinc, copper, and iron in healthy mouse kidneys and distribution of platinum after cisplatin treatment. Renal distribution in 12 nontreated Nu-Nu mice (males) was assessed by laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS). Furthermore, 9 Nu-Nu mice were treated with cisplatin. The order of elements concentration in kidneys was as follows: Fe > Zn > Cu. All three metals showed the higher concentrations at the cortex and medulla (28.60, 3.35, and 93.83 µg/g for Zn, Cu, and Fe, respectively) and lower concentration at the pelvis and the urinary tract (20.20, 1.93, and 62.48 µg/g for Zn, Cu, and Fe, respectively). No statistically significant difference between cortex and medulla was observed for these elements. After platinum treatment, the concentration of platinum in kidneys was enhanced more than 60-times, p < 0.001. Platinum significantly showed the highest accumulation in cortex (2.11 µg/g) with a gradient distribution. Platinum was less accumulated in medulla and pelvis than in cortex, and the lowest accumulation occurred in the urinary tract (1.13 µg/g). Image processing has been successfully utilized to colocalize metal distribution using LA-ICP-MS and histological samples images.

Metallothionein dimerization evidenced by QD-based Förster resonance energy transfer and capillary electrophoresis.

Herein, we report a new simple and easy-to-use approach for the characterization of protein oligomerization based on fluorescence resonance energy transfer (FRET) and capillary electrophoresis with LED-induced detection. The FRET pair consisted of quantum dots (QDs) used as an emission tunable donor (emission wavelength of 450 nm) and a cyanine dye (Cy3), providing optimal optical properties as an acceptor. Nonoxidative dimerization of mammalian metallothionein (MT) was investigated using the donor and acceptor covalently conjugated to MT. The main functions of MTs within an organism include the transport and storage of essential metal ions and detoxification of toxic ions. Upon storage under aerobic conditions, MTs form dimers (as well as higher oligomers), which may play an essential role as mediators in oxidoreduction signaling pathways. Due to metal bridging by Cd2+ ions between molecules of metallothionein, the QDs and Cy3 were close enough, enabling a FRET signal. The FRET efficiency was calculated to be in the range of 11-77%. The formation of MT dimers in the presence of Cd2+ ions was confirmed by MALDI-MS analyses. Finally, the process of oligomerization resulting in FRET was monitored by CE, and oligomerization of MT was confirmed.

Oxidized polysaccharides for anticancer-drug delivery: What is the role of structure?

Study provides an in-depth analysis of the structure-function relationship of polysaccharide anticancer drug carriers and points out benefits and potential drawbacks of differences in polysaccharide glycosidic bonding, branching and drug binding mode of the carriers. Cellulose, dextrin, dextran and hyaluronic acid have been regioselectively oxidized to respective dicarboxylated derivatives, allowing them to directly conjugate cisplatin, while preserving their major structural features intact. The structure of source polysaccharide has crucial impact on conjugation effectiveness, carrier capacity, drug release rates, in vitro cytotoxicity and cellular uptake. For example, while branched structure of dextrin-based carrier partially counter the undesirable initial burst release, it also attenuates the cellular uptake and the cytotoxicity of carried drug. Linear polysaccharides containing ß-(1→4) glycosidic bonds and oxidized at C2 and C3 (cellulose and hyaluronate) have the best overall combination of structural features for improved drug delivery applications including potentiation of the cisplatin efficacy towards malignances.

Ferrocenes as new anticancer drug candidates: Determination of the mechanism of action.

Chemotherapy plays an essential role in the management of cancer worldwide. However, it is a non-specific treatment limited by major drawbacks, thus identification and testing of new promising molecular structures representing potential drug candidates are urgently needed. In this work, ferrocene complexes as potential antitumor drugs that display cytotoxicity in low micromolar concentrations against ovarian cancer cells A2780 and SK-OV-3 were investigated to identify their mode of action. Their mechanism of cellular accumulation was studied using differential pulse voltammetry and inductively coupled plasma - mass spectrometry. Their mode of cell death induction was determined by changes in the mitochondrial membrane potential, production of reactive oxygen species and by Annexin V staining. Transferrin receptors were identified as key mediators of intracellular accumulation of ferrocenes and the extent of cellular uptake reflected the anticancer activity of individual compounds. Functional analysis revealed activation of intrinsic apoptosis as a dominant mechanism leading to regulated cell death induced in ovarian cancer cells by ferrocenes. Ferrocenes represent a group of promising sandwich organometallic complexes exerting cytotoxic activity. We suggest their application not only as standalone chemotherapeutics but also as modifying substituents of known drugs to improve their antitumor effects.