Photoinduced Reduction of Novel Dual-Action Riboplatin Pt(IV) Prodrug.

Controlled photoreduction of Pt(IV) prodrugs is a challenging task due to the possibility of targeted light-controlled activation of anticancer agents without affecting healthy tissues. Also, a conjugation of photosensitizers and clinically used platinum drugs into one Pt(IV) prodrug allows combining photodynamic therapy and chemotherapy approaches into one molecule. Herein, we designed the cisplatin-based Pt(IV) prodrug Riboplatin with tetraacetylriboflavin in the axial position. A novel Pt(IV) prodrug is able to act both as a photodynamic therapy (PDT) agent through the conversion of ground-state 3O2 to excited-state 1O2 and as an agent of photoactivated chemotherapy (PACT) through releasing of cisplatin under gentle blue light irradiation, without the requirement of a reducing agent. The light-induced behavior of Riboplatin was investigated using an electrochemical sensor in MCF-7 tumor spheroids. Photocontrolled cisplatin release and ROS generation were detected electrochemically in real time. This appears to be the first confirmation of simultaneous photoactivated release of anticancer drug cisplatin and ROS from a dual-action Pt(IV) prodrug observed from the inside of living tumor spheroids.

Protein-sized nanozymes «artificial peroxidase¼ based on template catalytic synthesis of Prussian Blue.

We report on Prussian Blue based nanozymes, comparable in size with a natural enzyme peroxidase. Protein-sized nanoparticles have been synthesized in the course of reduction of ferric ion (Fe3+) and ferricyanide ([Fe(CN)6]3-) one-to-one mixture in reversed micelles (isooctane|AOT|water) used as templates. Aniline chosen as the best reductant for this aim has led to formation of composite (according to Raman spectroscopy) Prussian Blue - polyaniline nanoparticles. The protein-like size of the nanoparticles (∅ = 4 - 6 nm) has been confirmed by DLS and TEM imaging. Kinetic investigations of peroxidase-like activity in reversed micelles resulted in the catalytic rate constant belonging to the same size-dependence as regular bulk catalytically synthesized nanozymes (slope ≈ 2.6), allowing to denote the reported Prussian Blue nanoparticles synthesized in reversed micelles as nanozymes «artificial peroxidase¼. Hydrogen peroxide sensors made by dipping the suspension of the latter onto the electrode support, displayed two-fold higher sensitivity as compared to the Prussian Blue film-based ones. Protein-sized nanozymes «artificial peroxidase¼ would obviously provide an advantage over regular nanozymes in (bio)sensors and analytical kits.

Glucose test strips with the largest linear range made via single step modification by glucose oxidase-hexacyanoferrate-chitosan mixture.

We report on the amperometric second-generation glucose test strips with the linear calibration range covering blood glucose concentrations. Chitosan membrane was used for immobilization of both enzyme and mediator in a single step. Optimal chitosan concentration in membrane-forming mixture corresponds to the highest enzyme activity and dramatically improved mediator adsorption in final membrane. On one hand, the immobilized glucose oxidase activation with an increase of the chitosan polymer content in the membrane has been noticed. On the other hand, positively charged chitosan matrix retains mediator (hexacyanoferrate (III) ion) in the membrane, due to its negative charge. Additionally, the excessive adsorption of the mediator on screen-printed electrodes coated with membranes was proved by means of cyclic voltammetry and impedance spectroscopy. Glucose test strips have been elaborated via single-step modification of the sensor support with the enzyme-mediator-polymer mixture. Apparently, the record linear calibration range from 1 mM to 50 mM glucose was achieved recording amperometric response at 5th second after potential was applied. The elaborated test strips have been validated through analysis of standard serum and blood samples. In whole blood test strips keep 84-98% of their sensitivity in buffer solution.

Electrochemical Detection of a Novel Pt(IV) Prodrug with the Metronidazole Axial Ligand in the Hypoxic Area.

We report herein a Pt(IV) prodrug with metronidazole in axial positions Pt-Mnz. The nitroaromatic axial ligand was conjugated with a cisplatin scaffold to irreversibly reduce under hypoxic conditions, thereby retaining the Pt(IV) prodrug in the area of hypoxia. X-ray near-edge adsorption spectroscopy (XANES) on dried drug-preincubated tumor cell samples revealed a gradual release of cisplatin from the Pt-Mnz prodrug instead of rapid intracellular degradation. The ability of the prodrug to penetrate into three-dimensional (3D) spheroid cellular cultures was evaluated by a novel electrochemical assay via a platinum-coated carbon nanoelectrode, capable of single-cell measurements. Using a unique technique of electrochemical measurements in single tumor spheroids, we were able to both detect the real-time response of the axial ligand to hypoxia and establish the depth of penetration of the drug into the tumor model.

Pt(IV) Prodrugs with Non-Steroidal Anti-inflammatory Drugs in the Axial Position.

We report herein the design, synthesis, and biological investigation of a series of novel Pt(IV) prodrugs with non-steroidal anti-inflammatory drugs naproxen, diclofenac, and flurbiprofen, as well as these with stearic acid in the axial position. Six Pt(IV) prodrugs 5-10 were designed, which showed superior antiproliferative activity compared to cisplatin as well as an ability to overcome tumor cell line resistance to cisplatin. By tuning the drug lipophilicity via variation of the axial ligands, the most potent Pt(IV) prodrug 7 was obtained, with an enhanced cellular accumulation of up to 153-fold that of cisplatin and nanomolar cytotoxicity both in 2D and 3D cell cultures. Pt2+ species were detected at different depths of MCF-7 spheroids after incubation with Pt(IV) prodrugs using a Pt-coated carbon nanoelectrode. Cisplatin accumulation in vivo in the murine mammary EMT6 tumor tissue of BALB/c mice after Pt(IV) prodrug injection was proved electrochemically as well. The drug tolerance study on BALB/c mice showed good tolerance of 7 in doses up to 8 mg/kg.

Novel Copper-Containing Cytotoxic Agents Based on 2-Thioxoimidazolones.

A series of 73 ligands and 73 of their Cu+2 and Cu+1 copper complexes with different geometries, oxidation states of the metal, and redox activities were synthesized and characterized. The aim of the study was to establish the structure-activity relationship within a series of analogues with different substituents at the N(3) position, which govern the redox potentials of the Cu+2/Cu+1 redox couples, ROS generation ability, and intracellular accumulation. Possible cytotoxicity mechanisms, such as DNA damage, DNA intercalation, telomerase inhibition, and apoptosis induction, have been investigated. ROS formation in MCF-7 cells and three-dimensional (3D) spheroids was proven using the Pt-nanoelectrode. Drug accumulation and ROS formation at 40-60 µm spheroid depths were found to be the key factors for the drug efficacy in the 3D tumor model, governed by the Cu+2/Cu+1 redox potential. A nontoxic in vivo single-dose evaluation for two binuclear mixed-valence Cu+1/Cu+2 redox-active coordination compounds, 72k and 61k, was conducted.

Constant Potential Amperometric Flow-Injection Analysis of Ions and Neutral Molecules Transduced by Electroactive (Conductive) Polymers.

We first report on constant potential (dc) amperometric flow-injection analysis (FIA) transduced by electroactive (conductive) polymers. Amperometric response is caused by the polymer recharging in order to maintain the electrode potential at a constant level when (i) ions are crossing the film|solution interface and polarizing electrode|film interface or (ii) ions or neutral molecules are specifically interacting with the polymer recharging it. The response under constant solution flow is a current peak and in flow-injection mode is a couple of current peaks directed opposite of the first sharp, analytically valuable peak. In both constant flow and flow-injection regimes, the peak current is dependent on analyte concentrations; obviously, the FIA mode provides more advantageous analytical characteristics. Constant potential amperometric flow-injection analysis is shown for boronate- and sulfate-functionalized polyanilines as well as for Prussian Blue, a member of the inorganic polymer family. As a proof of concept, the successful dc amperometric detection of lactate in human sweat with boronate-functionalized polyaniline has been shown. The proposed approach would revolutionize the field of conductive/electroactive polymer-supported ion sensing with the introduction of reliable and robust amperometry as a valuable alternative to existing potentiometry.

Nonenzymatic Sensor for Lactate Detection in Human Sweat.

For noninvasive diagnostics of hypoxia, we propose the nonenzymatic sensor based on screen-printed structures with the working surface modified in course of electropolymerization of 3-aminophenylboronic acid (3-APBA) with imprinting of lactate. Impedimetric sensor allows lactate detection in the range from 3 mM to 100 mM with the detection limit of 1.5 mM; response time is 2-3 min. Sensor sensitivity remains unchanged within 6 months of storage unpacked in dry state at a room temperature, which is unachievable for enzyme based devices. Analysis of human sweat with poly(3-APBA) based sensor is possible due to (i) much higher lactate content compared to other polyols and (ii) high sensor selectivity (Klactateglucose < 3 × 10-2). Successful detection of lactate in human sweat by means of the poly(3-APBA) based sensor has been confirmed using the highly specific reference method based on lactate oxidase enzyme (correlation coefficient r > 0.9). The attractive performance characteristics of poly(3-APBA) based enzyme-free sensors justify their future use for noninvasive clinical analysis and sports medicine.

Reagentless polyol detection by conductivity increase in the course of self-doping of boronate-substituted polyaniline.

We report on the novel reagentless and label-free detection principle based on electroactive (conducting) polymers considering sensors for polyols, particularly, saccharides and hydroxy acids. Unlike the majority of impedimetric and conductometric (bio)sensors, which specific and unspecific signals are directed in the same way (resistance increase), making doubtful their real applications, the response of the reported system results in resistance decrease, which is directed oppositely to the background. The mechanism of the resistance decrease is the polyaniline self-doping, i.e., as an alternative to proton doping, an appearance of the negatively charged aromatic ring substituents in polymer chain. Negative charge "freezing" at the boron atom is indeed a result of complex formation with di- and polyols, specific binding. Changes in Raman spectra of boronate-substituted polyaniline after addition of glucose are similar to those caused by proton doping of the polymer. Thermodynamic data on interaction of the electropolymerized 3-aminophenylboronic acid with saccharides and hydroxy acids also confirm that the observed resistance decrease is due to polymer interaction with polyols. The first reported conductivity increase as a specific signal opens new horizons for reagentless affinity sensors, allowing the discrimination of specific affinity bindings from nonspecific interactions.