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A “click” synthetic approach toward first-in-class dinucleoside phosphate mimics possessing a luminescent dirhenium carbonyl linker instead of a phosphodiester entity is reported.

Benzannulation of a ditopic ligand affords a significant phosphorescence blue-shift and enhanced emission quantum yield of mono- and dinuclear Ir(iii) complexes. The complexes are excellent oxygen photosensitizers. A use in bioimaging is reported.

Mono- and dinuclear Pt(ii) complexes of a new ditopic ligand with a thiophene cyclometalating core are reported. The mononuclear complex shows dual emission of fluorescence and phosphorescence and is demonstrated as a dual channel bioimaging probe.

Nowadays, dermatophyte infections are relatively easy to cure, especially since the introduction of orally administered antifungals such as terbinafine and itraconazole. However, these drugs may cause side effects due to liver damage or their interactions with other therapeutics. Hence, the search for new effective chemotherapeutics showing antidermatophyte activity seems to be the urge of the moment. Potassium salts of N-acylhydrazinecarbodithioates are used commonly as precursors for the synthesis of biologically active compounds. They were also found to act as antimicrobials, including activity against Candida spp. strains. Keeping that in mind, the activity of a series of five potassium N-acylhydrazinecarbodithioates (1a-e) and their aminotriazole-thione derivatives (2a-e) was evaluated against a set of pathogenic, keratinolytic fungi, but also against some Gram-positive and Gram-negative bacteria. All tested compounds were found non-toxic for L-929 and HeLa cells, with the IC30 and IC50 values assessed in the MTT assay above 128 µg/mL. The compound 5-amino-3-(naphtalen-1-yl)-4,5-dihydro-1H-1,2,4-triazole-5-thione (2d) was found active against all fungal strains tested. Scanning Electron Microscopy (SEM) revealed inhibition of mycelium development of T. rubrum cultivated on nail fragments and treated with 2d 24 h after infection with fungal spores. TEM (Transmission Electron Microscopy) observation of mycelium treated with 2d showed ultrastructural changes in the morphology of hyphae. Finally, the RNA-seq analysis indicated that a broad spectrum of genes responded to stress induced by the 2d compound. In conclusion, the results obtained confirm the potential of N-acylhydrazinecarbodithioate derivatives for future use as promising leads for the development of new antidermatophyte agents.

Nonsteroidal anti-inflammatory drugs (NSAIDs) are inhibitors of cyclooxygenase enzyme (COX) and were found to have positive effects in reducing the risk of developing gynecological cancers. However, long-term administration of NSAIDs carries the risk of various side effects, including those in the digestive and circulatory systems. Therefore, there is a constant need to develop new NSAID derivatives. In this work, we investigated rhenium NSAIDs, comparing their effects on endometrial cancer cells with original NSAIDs, demonstrating the high activity of aspirin and indomethacin derivatives. The cytotoxic activity of rhenium derivatives against the Ishikawa and HEC-1A cancer cell lines was higher than that of the original NSAIDs. The IC

Immunotherapy has revolutionized cancer treatment in recent years. Although currently approved checkpoint inhibitors (CPIs) yield remarkable anti-tumoral responses in several cancer types, a substantial proportion of patients do not benefit from such therapies. Local activation of innate immune signaling pathways is a promising approach to overcome the immunosuppressive tumor microenvironment, induce anti-tumor immunity, and improve the efficacy of CPI therapies. Here, we assessed the mode of action and efficacy of the RNA-based innate immune stimulator CV8102 for local immunotherapy in preclinical models. Intratumoral (i.t.) administration of CV8102 activated innate immune responses in the tumor microenvironment and draining lymph nodes, resulting in a dose-dependent anti-tumoral response. Combining i.t. CV8102 with systemic anti-programmed death protein 1 (PD-1) treatment further enhanced anti-tumoral responses, inducing tumor infiltration and activation of CD8+ T cells. The resulting memory response prevented tumor growth in rechallenged animals and impaired the growth of non-injected distal tumors. Therefore, i.t. CV8102 delivery is a promising approach for local cancer immunotherapy, especially in combination with CPIs. Clinical testing of CV8102 is ongoing (NCT03291002).

In the present study, we continue our work related to the synthesis of 1,8-naphthalimide and carborane conjugates and the investigation of their anticancer activity and DNA-binding ability. For this purpose, a series of 4-carboranyl-1,8-naphthalimide derivatives, mitonafide, and pinafide analogs were synthesized using click chemistry, reductive amination, amidation, and Mitsunobu reactions. The calf thymus DNA (ct-DNA)-binding properties of the synthesized compounds were investigated by circular dichroism (CD), UV–vis spectroscopy, and thermal denaturation experiments. Conjugates 54–61 interacted very strongly with ct-DNA (∆Tm = 7.67–12.33 °C), suggesting their intercalation with DNA. They were also investigated for their in vitro effects on cytotoxicity, cell migration, cell death, cell cycle, and production of reactive oxygen species (ROS) in a HepG2 cancer cell line as well as inhibition of topoisomerase IIα activity (Topo II). The cytotoxicity of these eight conjugates was in the range of 3.12–30.87 µM, with the lowest IC50 value determined for compound 57. The analyses showed that most of the conjugates could induce cell cycle arrest in the G0/G1 phase, inhibit cell migration, and promote apoptosis. Two conjugates, namely 60 and 61, induced ROS production, which was proven by the increased level of 2′-deoxy-8-oxoguanosine in DNA. They were specifically located in lysosomes, and because of their excellent fluorescent properties, they could be easily detected within the cells. They were also found to be weak Topo II inhibitors.

Ferrocenylated glycol nucleic acid (Fc-GNA) components are rarely studied in the field of xeno nucleic acid (XNA) chemistry. As an attempt to contribute to XNA chemistry, in the present article we report a seven-step synthesis of the first semicanonical dinucleoside containing the Fc-GNA nucleoside linked to the adenosine nucleoside with a phosphodiester bond. First, the nucleoside-bearing ethynylferrocenyl moiety in the C5 position of the uracil nucleobase was obtained. In the following steps, the nucleoside was transformed into the phosphoramidite intermediate that in turn was reacted with N6-benzoyl-2′,3′-O-isopropylideneadenosine to afford the target dinucleoside phosphate with 47% yield. The newly obtained Fc-GNA nucleoside is redox-active, and on the basis of this property (function), it belongs to a new class of functional GNA (fun-GNA) nucleosides. The electrochemistry of the Fc-GNA nucleoside, dinucleoside phosphate, and ferrocenyl furanopyrimidone nucleoside that was obtained as an undesired byproduct of Fc-GNA nucleoside synthesis was investigated by cyclic voltammetry (CV). The CV result showed the presence of a one-electron ferrocenyl-centered redox wave in each case. The half-wave potentials of the Fc-GNA nucleoside and dinucleoside phosphate were 89 and 99 mV, respectively, against the FcH/FcH+ couple. Finally, the activity of the newly obtained Fc-GNA components was studied against the nontumorigenic mouse L929 and human cervix adenocarcinoma HeLa cells. The synthesized compounds showed no cytotoxic activity against the tested cell lines. K.K. thanks the National Science Center in Cracow, Poland (grant OPUS UMO-2018/29/B/ST5/00055), for financial support. Crystallographic measurements were carried out at the Biological and Chemical Research Centre, University of Warsaw, established within the project cofinanced by European Union from the European Regional Development Fund under the Operational Programme Innovative Economy, 2007–2013. The X-ray diffraction data were collected at the Core Facility for Crystallographic and Biophysical Research to support the development of medicinal products sponsored by the Foundation for Polish Science (FNP).

Antibiotic-resistant microorganisms causing a life-threatening infection pose a serious challenge for modern science. The rapidly growing number of incidents for which the use of standard antibiotics is ineffective forces us to develop new alternative methods of killing microorganisms. Antimicrobial proteins and peptides (AMPs) can be promising candidates to solve this problem. Colicin-M is one of the representatives of this group and is naturally produced byEscherichia coliacting on other closely related bacterial strains by disrupting their outer cell membrane. This bacteriocin has huge potential as a potent antimicrobial agent, especially, since it was recognized by the FDA as safe. In this work, we present the expression of colicin M in model transgenicNicotiana tabacumL. plants. We demonstrate that purified colicin retains its antibacterial activity against the controlEscherichia colistrains and clinical isolates ofEscherichia coliandKlebsiella pneumoniae. Our results also show that plant-derived ColM is not toxic for L929 and HeLa cell line, which allows us to suppose that plant-based expression could be an alternative production method of such important proteins.

ntroduction: Lower limb amputation is a surgery performed as a last resort, when all other therapeutic options have been exhausted. The duration of treatment lasts from a few to several months and depends on the extent of amputation, the patientas overall health and the course of the stump healing process. Materials and methods A retrospective analysis was performed using the database of the General and Vascular Surgery Ward of the Nikolay Pirogov Regional Specialist Hospital in Łodź. Patients who underwent lower limb amputation at the transfemoral level in 2017 were analyzed. 92 patients undergoing surgery were qualified for the study. Patients were divided into two groups: those with no healing complications and those with stump healing complications. Medical records of both groups were analyzed for risk factors for impaired healing. The obtained data were subjected to statistical analysis. Results Patients with impaired stump healing most often had minimal bleeding and higher ASA scores compared to patients without healing complications. No differences between the two groups were found for the remaining parameters. Patients with complications needed an average of 28 days to heal the wound and spent an average of 40 days in hospital, compared to 14 and 21 days, respectively, for patients without complications. The percentage of deaths in the group of patients with complications was also significantly higher (35%) than in the group of patients without complications (5%). Conclusions Statistically significant factors increasing the risk of impaired stump healing include high ASA scale and minimal muscle bleeding during surgery. Patients who experienced this complication are at greater risk of prolonged hospitalization and death in the postoperative period. This study showed statistically significant risk factors for impaired stump healing following amputation and confirmed the negative impact of this complication on the length of hospitalization and risk of death.

Compounds targeting bacterial topoisomerases are of interest for the development of antibacterial agents. Our previous studies culminated in the synthesis and characterization of small-molecular weight thiosemicarbazides as the initial prototypes of a novel class of gyrase and topoisomerase IV inhibitors. To expand these findings with further details on the mode of action of the most potent compounds, enzymatic studies combined with a molecular docking approach were carried out, the results of which are presented herein. The biochemical assay for 1-(indol-2-oyl)-4-(4-nitrophenyl) thiosemicarbazide (4) and 4-benzoyl-1-(indol-2-oyl) thiosemicarbazide (7), showing strong inhibitory activity against Staphylococcus aureus topoisomerase IV, confirmed that these compounds reduce the ability of the ParE subunit to hydrolyze ATP rather than act by stabilizing the cleavage complex. Compound 7 showed better antibacterial activity than compound 4 against clinical strains of S. aureus and representatives of the Mycobacterium genus. In vivo studies using time-lapse microfluidic microscopy, which allowed for the monitoring of fluorescently labelled replisomes, revealed that compound 7 caused an extension of the replication process duration in Mycobacterium smegmatis, as well as the growth arrest of bacterial cells. Despite some similarities to the mechanism of action of novobiocin, these compounds show additional, unique properties, and can thus be considered a novel group of inhibitors of the ATPase activity of bacterial type IIA topoisomerases.

Nonsteroidal anti-inflammatory drugs (NSAIDs) are inhibitors of cyclooxygenase enzyme (COX) and were found to have positive effects in reducing the risk of developing gynecological cancers. However, long-term administration of NSAIDs carries the risk of various side effects, including those in the digestive and circulatory systems. Therefore, there is a constant need to develop new NSAID derivatives. In this work, we investigated rhenium NSAIDs, comparing their effects on endometrial cancer cells with original NSAIDs, demonstrating the high activity of aspirin and indomethacin derivatives. The cytotoxic activity of rhenium derivatives against the Ishikawa and HEC-1A cancer cell lines was higher than that of the original NSAIDs. The IC50 after 24-h incubation of Ishikawa and HEC-1A were 188.06 µM and 394.06 µM for rhenium aspirin and 228.6 µM and 1459.3 µM for rhenium indomethacin, respectively. At the same time, IC50 of aspirin and indomethacin were 10,024.42 µM and 3295.3 µM for Ishikawa, and 27,255.8 µM and 5489.3 µM for HEC-1A, respectively. Moreover, these derivatives were found to inhibit the proliferation of both cell lines in a time- and state-dependent manner. The Ishikawa cell proliferation was strongly inhibited by rhenium aspirin and rhenium indomethacin after 72-h incubation (*** = p < 0.001), while the HEC-1A proliferation was inhibited by the same agents already after 24-h incubation (*** = p < 0.001). Furthermore, the ROS level in the mitochondria of the tested cells generated in the presence of rhenium derivatives was higher than the original NSAIDs. That was associated with rhenium indomethacin exclusively, which had a significant effect (*** = p < 0.001) on both Ishikawa and HEC-1A cancer cells. Rhenium aspirin had a significant effect (*** = p < 0.001) on the mitochondrial ROS level of Ishikawa cells only. Overall, the research revealed a high potential of the rhenium derivatives of aspirin and indomethacin against endometrial cancer cells compared with the original NSAIDs.

The knowledge pertaining to the chemistry and biological activity of glycol nucleic acid (GNA) components, like nucleosides and nucleotides, is still very limited. Herein we report on the preparation of the uracil nucleoside (1) and nucleotide ester GNA (2). The compounds are functionalised with a luminescent phenanthrenyl group. In DMSO, 1 and 2 are brightly fluorescent, with emission maxima at 390 nm, nanosecond decay times (0.6 and 0.75 ns, respectively), and quantum yields of ca. 0.2. In the solid phase, they show excimeric emission, with maxima at 495 nm (1) and 432 nm (2), and decay times of 3.7 ns (1) and 2.9 ns (2). The anticancer activity of the GNA components, as well as gemcitabine hydrochloride, used as a reference drug, were examined in vitro against human cancer HeLa and Ishikawa cells, as well as against normal L929 cells, using a battery of biochemical assays. Furthermore, biodistribution imaging studies were carried out in HeLa cells, with luminescence confocal microscopy, which showed that the compounds localized mainly in the lipophilic cellular compartments. Nucleoside (1) and nucleotide ester (2) features two different anticancer activity profiles. At 24 h of treatment, the nucleoside acts mainly as a toxin and induces necrosis in HeLa cells, whereas the nucleotide ester exhibits pro-apoptotic activity. At longer treatment times (72 h), the nucleoside and the reference, gemcitabine hydrochloride, featured almost identical signs of anticancer activity, such as S-phase cell cycle arrest, proliferation inhibition, and apoptosis induction. In view of this data, one can hypothesize that despite the structural differences, the newly obtained phenanthrenyl GNA nucleoside (1) and gemcitabine may share a common mechanism of anticancer activity in HeLa cancer cells. The GNA components were also examined as antiplasmodial agents against Plasmodium falciparum, in vitro. Nucleoside (1) was found to be more potent than nucleotide (2), displaying activity in the low micromolar range. Furthermore, both phenanthrene derivatives were found to display resistance indices at least 9-fold lower than chloroquine diphosphate (CQDP).

International journal of molecular sciences 22(5), 2772 (2021). doi:10.3390/ijms22052772, We synthesized a series of novel 3-carboranyl-1,8-naphthalimide derivatives, mitonafide and pinafide analogs, using click chemistry, reductive amination and amidation reactions and investigated their in vitro effects on cytotoxicity, cell death, cell cycle, and the production of reactive oxygen species in a HepG2 cancer cell line. The analyses showed that modified naphthalic anhydrides and naphthalimides bearing ortho- or meta-carboranes exhibited diversified activity. Naphthalimides were more cytotoxic than naphthalic anhydrides, with the highest IC50 value determined for compound 9 (3.10 ��M). These compounds were capable of inducing cell cycle arrest at G0/G1 or G2M phase and promoting apoptosis, autophagy or ferroptosis. The most promising conjugate 35 caused strong apoptosis and induced ROS production, which was proven by the increased level of 2���-deoxy-8-oxoguanosine in DNA. The tested conjugates were found to be weak topoisomerase II inhibitors and classical DNA intercalators. Compounds 33, 34, and 36 fluorescently stained lysosomes in HepG2 cells. Additionally, we performed a similarity-based assessment of the property profile of the conjugates using the principal component analysis. The creation of an inhibitory profile and descriptor-based plane allowed forming a structure���activity landscape. Finally, a ligand-based comparative molecular field analysis was carried out to specify the (un)favorable structural modifications (pharmacophoric pattern) that are potentially important for the quantitative structure���activity relationship modeling of the carborane���naphthalimide conjugates., Published by Molecular Diversity Preservation International, Basel

Recently, nanomaterials synthesized ecologically using microorganisms have attracted much interest. In the present study, the ability of Gloeophyllum striatum to synthesize silver nanoparticles is described for the first time. Nanoparticles were formed in an eco-friendly extracellular manner and characterized by UV-Vis, FT-IR, MADLS and SEM techniques. The obtained nanoparticles showed excellent activity against gram-positive and gram-negative bacteria. The MIC values for gram-negative bacteria were 15 µM, while for gram-positive strains they reached 30 µM. The haemolytic and cytotoxic activities of the synthesized nanoparticles towards mammalian cells were also determined. The addition of AgNPs at the concentrations above 30 µM caused 50% haemolysis of red blood cells after they 24-hour incubation. A decrease in the viability of fibroblasts by over 50% was also found in the samples treated with nanoparticles at the concentrations above 30 µM. The ecotoxicological risk of silver nanoparticles was assessed using A. franciscana and D. magna crustaceans as well as L. sativum plants. The EC50 values for A. franciscana and D. magna were 61.97 and 0.275 µM, respectively. An about 20% reduction in the length of L. sativum shoots and roots was noted after the treatment with AgNPs at the concentration of 100 µM.

In this paper, we report the Yb(OTf)3‐mediated etherification reaction that allowed obtaining R, R and S, R isomers of ferrocenyl glycol nucleic acid (Fc‐GNA) nucleosides from their corresponding stereo‐defined (S, R) precursor. The R, R absolute configuration of the chiral carbon atoms in one of the Fc‐nucleoside isomers was assigned by single‐ crystal X‐ray diffraction. Density functional theory calculations showed that the stereochemical outcome of the reaction can be explained by an exo attack of ethylene glycol on the pro‐R‐ or pro‐S‐ configured α‐ferrocenyl carbenium ion intermediate. The R, R isomer was transformed into the corresponding nucleotide diethyl ester which is, to the best of our knowledge, the first Fc‐GNA nucleotide ever reported. The obtained compounds feature reversible one‐electron oxidation of the ferrocenyl portion of their molecular structures. Anticancer activity studies showed that the (S, R) nucleoside was the most active against HeLa and Ishikawa cancer cells, while it did not show any activity against nontumorigenic L929 cells. The compound induced apoptosis in HeLa cells at IC50 = 66.0 μM concentration upon 72 h of treatment.

A dinuclear iridium(III) complex IrIr shows dual emission consisting of near infrared (NIR) phosphorescence (λmax = 714 nm, CH2Cl2, T = 300 K) and green fluorescence (λmax = 537 nm). The NIR emission stems from a triplet state (T1) localized on the ditopic bridging ligand (3LC). Because of the dinuclear molecular structure, the phosphorescence efficiency (ΦPL = 3.5%) is high compared to those of other known red/NIR-emitting iridium complexes. The weak fluorescence stems from the lowest excited singlet state (S1) of 1LC character. The occurrence of fluorescence is ascribed to relatively slow intersystem crossing (ISC) from state S1 (1LC) to the triplet manifold. The measured ISC rate corresponds to a time constant τISC of 2.1 ps, which is an order of magnitude longer than those usually found for iridium complexes. This slow ISC rate can be explained in terms of the LC character and large energy separation (0.57 eV) of the respective singlet and triplet excited states. IrIr is internalized by live HeLa cells as evidenced by confocal luminescence microscopy.

Ten aminophosphonates bearing 2-nitrofuran moiety and two N-aryl 5-nitro-furfuralaldimines, namely, N-(2-nitrofurfurylidene)-p-toluidine and N-(2-nitrofurfurylidene)-p-anisidine, were tested in aspect of their antibacterial action. O,O′-diphenyl derivatives were found inactive, while O,O′-dimethyl and O,O′-diethyl derivatives were revealed to act moderately efficiently against clinical isolates of S. aureus, especially against methicillin-resistant (MRSA) strains. A high activity against these strains was found for N-(4-methylphenyl)-5-nitrofurfuralaldimine and N-(4-methoxyphenyl)-5-nitro-furfuralaldimine, but they showed the cytotoxicity at a dangerous level.

Purpose To define the dose–response relationship for initial and residual pATM and γH2AX foci and temporal response of pATM foci in fibroblasts of 4 hyper-radiosensitivity (HRS)-positive cancer patients and 8 HRS-negative cancer patients and answer the question regarding the role of DNA double-strand break (DSB) recognition and repair in the mechanism of HRS. Methods and Materials The cells were irradiated with single doses (0.1-4 Gy) of 6-MV X rays. The number of initial and residual pATM and γH2AX foci was assessed 1 hour and 24 hours after irradiation, respectively. Kinetics of DSB recognition and repair was estimated by pATM foci assay after irradiation with 0.2 and 2 Gy. Results Hyper-radiosensitivity response (confirmed by the induced-repair model) was clearly evident for residual pATM and γH2AX foci in fibroblasts of HRS-positive patients but not in fibroblasts of HRS-negative patients. Significantly less DSB was recognized by pATM early (10-30 minutes) after irradiation with 0.2 Gy in HRS-positive compared with HRS-negative fibroblasts. Conclusions The present results provide evidence for the role of DSB recognition by pATM and repair in the mechanism of HRS and seem to support the idea of nucleo-shuttling of the pATM protein to be involved in HRS response.

Farmed chinchillas continue to have very low fertility and prolificacy, although the reproductive potential of females is much higher and can be exploited. An increasing number of studies deal with the specific nature of reproduction in this species, with particular focus on female fertility. However, there is a lack of analyses and studies of males. Therefore, this study has attempted to determine the impact of the season of the year on the reproductive activity of male chinchillas and on the results of their use for breeding in the period from January 1991 to December 1996. The analysis was conducted on farmed animals on one of the largest farms in Poland. The farm was established in 1991 with an initial foundation stock of about 150 females and 25 males. Breeding was based on animals imported from a reputable farm in Germany, from which modern breeding technologies were adopted as well. Polygamous breeding with 6 females per male is used. Sexual activity was varied between years. Taking into account the entire foundation stock, an average of 8.15 litters and 14.77 pups were obtained per male per year. Analysis of five selected males revealed higher values: the mean number of litters ranged from 7.13 to 11.8 and the litter size ranged from 1.9 to 2.59. Thus, better utilization of males for breeding is possible on chinchilla farms. The system of keeping a male with many females at the same time enables strict selection of males.

This work presents the results of an analysis of concentration levels of selected trace metals (Pb, Zn, Cu, Cd) in plants and soil near the main roads of the Warsaw agglomeration. The aim of the study was to assess the degree of contamination by selected trace elements of plants growing at different distances from the road (directly at the roadside and 50 meters away from the road) and to examine the relationship between the concentration of the tested parameters in the soil and their content in the plant. It was found that the lack of clear differentiation in the levels of all heavy metals in plants depending on the distance from the road was associated with the possibility of considerable pollutant migration. In addition, it was found that dandelion usually reveals higher concentrations of heavy metals than meadow-grass.

The synthesis, characterization and biological activity of six bioorganometallic conjugates of ciprofloxacin with ferrocenyl, ruthenocenyl and cymantrenyl entities are described. Their antimicrobial activities were investigated against Gram-positive bacteria, Gram-negative bacteria and bloodstream forms of Trypanosoma brucei. Furthermore, the morphological changes of bacterial cells upon treatment with the conjugates were examined by scanning electron microscopy. In addition, the cytotoxicity of the conjugates against tumor and normal mammalian cells was also investigated. The results showed that conjugation of an organometallic moiety can significantly enhance the antimicrobial activity of the antibiotic ciprofloxacin drug. It was found that N-alkyl cymantrenyl and ruthenocenyl ciprofloxacin conjugates were the most effective derivatives although other conjugates also showed significant antimicrobial activity. The increase in the antimicrobial activity was most likely due to two independent mechanisms of action. The first mechanism is due to the bacterial topoisomerase inhibitory activity of ciprofloxacin while the second mechanism can be attributed to the generation of reactive oxygen species caused by the organometallic moiety. The presence of two modes of action enables the conjugates to kill bacteria in their stationary growth phase and to overcome the drug resistance of S. aureus strains. In addition, the conjugates showed promising selectivity toward bacterial and parasitic cells over mammalian cells.

The conjugation of organometallic groups to current β-lactam antibiotics is a field of increasing study due to the ability of certain organometallic groups to enhance the antibiotic potency of these drugs. Herein, we report the antibacterial properties of two metallocenyl (ferrocenyl and ruthenocenyl) 7-aminocephalosporanic acid (7-ACA) antibiotic conjugates. Continuing a trend we found in our previous studies, the ruthenocenyl conjugate showed greater antibacterial activity than its ferrocenyl counterpart. Compared with the previously published 7-aminodesacetoxycephalosporanic acid (7-ADCA) conjugates, the 3-acetyloxymethyl group significantly improved the compounds' activity. Furthermore, the Rc-7-ACA compound was more active against clinical Staphylococcus aureus isolates than the ampicillin reference. Noticeably, neither of the two new compounds showed an undesirable toxic effect in HeLa and L929 cells at the concentrations at which they displayed strong antibacterial effects. The antibacterial activity of the two metallocenyl 7-ACA derivatives was further confirmed by scanning electron microscopy (SEM). SEM micrographs showed that bacteria treated with metallocenyl 7-ACA derivatives feature cell wall damage and morphology changes. Using a CTX-M-14 β-lactamase competition assay based on nitrocefin hydrolysis, we showed that the Rc-7-ACA bound more favorably to CTX-M-14 than its ferrocenyl counterpart, again confirming the superiority of the ruthenocenyl moiety over the ferrocenyl one in interacting with proteins. We also report a 1.47 A resolution crystal structure of Rc-7-ACA in complex with the CTX-M-14 E166A mutant, an enzyme sharing a similar active site configuration with penicillin-binding proteins, the molecular target of β-lactam antibiotics. These results strengthen the case for the antibacterial utility of the Rc and Fc groups.

ntroduction: Lower limb amputation is a surgery performed as a last resort, when all other therapeutic options have been exhausted. The duration of treatment lasts from a few to several months and depends on the extent of amputation, the patientapos;s overall health and the course of the stump healing process.A retrospective analysis was performed using the database of the General and Vascular Surgery Ward of the Nikolay Pirogov Regional Specialist Hospital in Łódź. Patients who underwent lower limb amputation at the transfemoral level in 2017 were analyzed. 92 patients undergoing surgery were qualified for the study. Patients were divided into two groups: those with no healing complications and those with stump healing complications. Medical records of both groups were analyzed for risk factors for impaired healing. The obtained data were subjected to statistical analysis.Patients with impaired stump healing most often had minimal bleeding and higher ASA scores compared to patients without healing complications. No differences between the two groups were found for the remaining parameters. Patients with complications needed an average of 28 days to heal the wound and spent an average of 40 days in hospital, compared to 14 and 21 days, respectively, for patients without complications. The percentage of deaths in the group of patients with complications was also significantly higher (35%) than in the group of patients without complications (5%).Statistically significant factors increasing the risk of impaired stump healing include high ASA scale and minimal muscle bleeding during surgery. Patients who experienced this complication are at greater risk of prolonged hospitalization and death in the postoperative period. This study showed statistically significant risk factors for impaired stump healing following amputation and confirmed the negative impact of this complication on the length of hospitalization and risk of death.

Glycol nucleic acids (GNA) are synthetic genetic-like polymers with an acyclic three-carbon propylene glycol phosphodiester backbone. Here, synthesis, luminescence properties, circular dichroism (CD) spectra, and confocal microscopy speciation studies of (R,S) and (S,R) pyrenyl-GNA (pyr-GNA) nucleosides are reported in HeLa cells. Enantiomerically pure nucleosides were obtained by a Sharpless asymmetric dihydroxylation reaction followed by semi-preparative high-performance liquid chromatography (HPLC) separation using Amylose-2 as the chiral stationary phase. The enantiomeric relationship between stereoisomers was confirmed by CD spectra, and the absolute configurations were assigned based on experimental and theoretical CD spectra comparisons. The pyr-GNA nucleosides were not cytotoxic against human cervical (HeLa) cancer cells and thus were utilized as luminescent probes in the imaging of these cells with confocal microscopy. Cellular staining patterns were identical for both enantiomers in HeLa cells. Compounds showed no photocytotoxic effect and were localized in the lipid membranes of the mitochondria, in cellular vesicles and in other lipid cellular compartments. The overall distribution of the pyrene and pyrenyl-GNA nucleosides inside the living HeLa cells differed, since the former compound gives a more granular staining pattern and the latter a more diffuse one.

Four fac-[Re(CO)3(phen)(L)] complexes with phen = 1,10-phenanthroline and L = a non-steroidal anti-inflammatory drug (NSAID) carboxylate ligand were obtained from the reaction of fac-[ReCl(CO)3(phen)] with ligands such as aspirin, (S)-(+)-ibuprofen, (S)-(+)-naproxen and indomethacin. Single-crystal X-ray diffraction analysis revealed a pseudo-octahedral coordination of the rhenium atom whereas the NSAID carboxylates act as monodentate ligands. The complexes show orange luminescence in polar solvents. Of the four compounds obtained, only fac-[Re(CO)3(phen)(aspirin)] exhibited activity against HeLa human cancer cells while it was inactive against the non-tumorigenic mouse L929 cell line. After uptake and dissociation in HeLa cells, the fac-[Re(CO)3(phen)(aspirin)] complex produces the luminescent fac-[Re(CO)3(phen)]+ cation, which predominantly accumulates in the mitochondria but does not penetrate to the nucleus, as shown by confocal microscopy. Once generated, the fac-[Re(CO)3(phen)]+ cation reversibly binds to hen egg white lysozyme (HEWL) while the aspirin ligand holds the ability to inhibit the COX-2 enzyme. The anticancer activity of the fac-[Re(CO)3(phen)(aspirin)] complex could be linked to ROS production, and cell cycle disturbance followed by triggering an apoptotic pathway of cell death.

This paper presents a report of the synthesis and chemical transformations of anthracene-thymine and 4-thiothymine derivatives. In a reaction of 1-(anthracen-9-yl)-1-hydroxy-3-(N1-thyminyl)-propane with ethylene glycol either an elimination or substitution product is preferentially formed depending on reaction conditions. In the presence of Yb(OTf)3, the elimination product was the major one; while in the absence of Yb(OTf)3, the substitution reaction product prevailed. Thus, six fluorescent nucleic base derivatives were obtained. The three anthracene-thymine derivatives (2, 3, and 4) show strong fluorescence with emission quantum yields from 65% to about 100% in solution at ambient temperature. The emission of anthracene-thymine 1 and two anthracene-thiothymines is weaker due to the carbonyl substituents and the sulphur atom of thiothymine. Tests of antiproliferative activity on human cancer HeLa cells performed at 24 h and 5 h treatment times, respectively, revealed no detectable toxicity at the latter interval. Thus, confocal microscopy imaging studies were carried out on HeLa cells at 5 h incubation time. All six compounds under study were found in the interior of the cells, albeit the intensity of the compounds’ luminescence stemming from the interiors was different. All compounds localised in the lipid membranes of cytoplasmic compartments in general follow the same staining pattern. In addition, compounds 3 and 6 were also detected in the nucleus. Notably, no signs of photocytotoxicity or photobleaching in the timespan of imaging studies were observed. The new fluorescent nucleic base derivatives represent attractive leading structures for the development of preferably more selective luminescent probes in the future.

Fourfac-[Re(CO)3(phen)(L)] complexes with phen = 1,10-phenanthroline and L = a non-steroidal anti-inflammatory drug (NSAID) carboxylate ligand were obtained from the reaction offac-[ReCl(CO)3(phen)] with ligands such as aspirin, (S)-(+)-ibuprofen, (S)-(+)-naproxen and indomethacin. Single-crystal X-ray diffraction analysis revealed a pseudo-octahedral coordination of the rhenium atom whereas the NSAID carboxylates act as monodentate ligands. The complexes show orange luminescence in polar solvents. Of the four compounds obtained, onlyfac-[Re(CO)3(phen)(aspirin)] exhibited activity against HeLa human cancer cells while it was inactive against the non-tumorigenic mouse L929 cell line. After uptake and dissociation in HeLa cells, thefac-[Re(CO)3(phen)(aspirin)] complex produces the luminescentfac-[Re(CO)3(phen)]+cation, which predominantly accumulates in the mitochondria but does not penetrate to the nucleus, as shown by confocal microscopy. Once generated, thefac-[Re(CO)3(phen)]+cation reversibly binds to hen egg white lysozyme (HEWL) while the aspirin ligand holds the ability to inhibit the COX-2 enzyme. The anticancer activity of thefac-[Re(CO)3(phen)(aspirin)] complex could be linked to ROS production, and cell cycle disturbance followed by triggering an apoptotic pathway of cell death.

Objectives: Depression and vital exhaustion are associated with poor cardiovascular prognosis, but there is substantial overlap between these constructs. Factor analytic studies have been inconclusive, and may not be the optimal analytic strategy to assess dimensionality. We assessed whether exhaustion and depression formed a single, hierarchical dimension using a form of nonparametric item response theory.Methods: Patients with acute coronary syndrome (n = 430) completed questionnaires assessing depression and vital exhaustion. Mokken scaling was used to assess dimensionality.Results: Mokken scaling formed a strong unidimensional scale, ordered in a hierarchy reflecting prevalence: fatigue (common), depression (less common) and hopelessness (rare).Conclusions: Depressive symptoms form a clear hierarchy in cardiac patients, from fatigue to hopelessness. Vital exhaustion may be considered a less severe form of depression. Use of hierarchical scales may allow clinicians to better determine clinical significance and target interventions.

Protein- and peptide-based tumor vaccines depend on strong adjuvants to induce potent immune responses. Here, we demonstrated that a recently developed novel adjuvant based on a non-coding, long-chain RNA molecule, termed RNAdjuvant(®) , profoundly increased immunogenicity of both antigen formats. RNAdjuvant(®) induced balanced, long-lasting immune responses that resulted in a strong anti-tumor activity. A direct comparison to Poly(I:C) showed superior efficacy of our adjuvant to enhance antigen-specific multifunctional CD8(+) T-cell responses and mediate anti-tumor responses induced by peptide derived from HPV-16 E7 protein in the syngeneic TC-1 tumor, a murine model of human HPV-induced cervical cancer. Moreover, the adjuvant was able to induce functional memory responses that mediated complete tumor remission. Despite its remarkable immunostimulatory activity, our RNA-based adjuvant exhibited an excellent pre-clinical safety profile. It acted only locally at the injection site where it elicited a transient but strong up-regulation of pro-inflammatory and anti-viral cytokines as well as cytoplasmic RNA sensors without systemic cytokine release. This was followed by the activation of immune cells in the draining lymph nodes. Our data indicate that our RNA-based adjuvant is a safe and potent immunostimulator that may profoundly improve the efficacy of a variety of cancer vaccines.

Neuroblastoma is the most common extra-cranial solid tumor of childhood and it is characterized by the presence of a glycosphingolipid, GD2 ganglioside. Monoclonal antibodies targeting the antigen are currently tested in clinical trials. Additionally, several research groups reported results revealing that ganglioside-specific antibodies can affect cellular signaling and cause direct cytotoxicity against tumor cells. To shed more light on gene expression signatures of tumor cells, we used microarrays to analyze changes of transcriptome in IMR-32 human neuroblastoma cell cultures treated with doxorubicin (DOX) or a mouse monoclonal antibody binding to GD2 ganglioside 14G2a (mAb) for 24 h. The obtained results highlight that disparate cellular pathways are regulated by doxorubicin and 14G2a. Next, we used RT-PCR to verify mRNA levels of selected DOX-responsive genes such as RPS27L, PPM1D, SESN1, CDKN1A, TNFSF10B, and 14G2a-responsive genes such as SVIL, JUN, RASSF6, TLX2, ID1. Then, we applied western blot and analyzed levels of RPS27L, PPM1D, sestrin 1 proteins after DOX-treatment. Additionally, we aimed to measure effects of doxorubicin and topotecan (TPT) and 14G2a on expression of a novel human NDUFAF2 gene encoding for mimitin protein (MYC-induced mitochondrial protein) and correlate it with expression of the MYCN gene. We showed that expression of both genes was concomitantly decreased in the 14G2a-treated IMR-32 cells after 24 h and 48 h. Our results extend knowledge on gene expression profiles after application of DOX and 14G2a in our model and reveal promising candidates for further research aimed at finding novel anti-neuroblastoma targets.

In the present paper, we report the synthesis and evaluation of in vitro antimicrobial activities of aziridine-thiourea derivatives. A series of aziridines in reaction with isocyanates and isothiocyanates to obtain urea and thiourea derivatives were used. The structures of all new products were confirmed based on spectroscopic data (1H-NMR, 13C-NMR, HR-MS). These compounds were screened for their in vitro antimicrobial activity against a panel of Gram-positive and Gram-negative strains of bacteria. Six of the tested compounds appeared to be promising agents against reference strains of Escherichia coli, Staphylococcus aureus and Staphylococcus epidermidis. Subsequently, compounds exhibiting promising antibacterial activity were tested against twelve clinical isolates of S. aureus from three different sources of infection. The most bactericidal compounds (MIC = 16–32 µg/mL) showed better antibacterial activity against MRSA than ampicillin and streptomycin. The in vitro cytotoxicity analysis on L929 murine fibroblast and HeLa human tumor cell line using the MTT assay allowed us to select the least toxic compounds for future investigation.

Purpose To evaluate the impact of HPV16 load (VL-the number of virus genome copies per cell) and P16 expression on prognosis of patients with squamous cell carcinomas (SCCs) of head and neck (HN). Materials and methods HPV16 presence was assessed in the group of 109 patients with HNSCCs by quantitative polymerase chain reaction (qPCR). VL (assessed by qPCR) and P16 expression (evaluated by immunohistochemistry) were analysed only in the subgroup of HPV16-positive tumours. These features were correlated with 5-year overall survival (OS) and disease-free survival (DFS). Results HPV16 infection was found in 36 tumours (33.0%). Virus-positive patients had better OS and DFS than those without infection (P = 0.041 and 0.005). Among HPV16-positive HNSCCs, 18 (50.0%) had higher VL (median value > 6764.3 copies/cell) and 25 (73.5%) P16 over expression. The significant differences in OS and DFS (P = 0.008 and 0.004) were noticed according to VL, wherein 100% DFS was found for patients with higher VL. According to P16 expression, significant difference was found only for OS (P = 0.020). In multivariate analysis, VL (P = 0.045; HR = 2.795; CI 0.121-1.060) and the level of smoking (P = 0.023, HR = 2.253; CI 1.124-4.514) were independent factors affecting DFS of HPV16-positive patients. Conclusion On the basis of viral load, it is possible to differentiate prognosis of patients with HPV16-positive HNSCCs. In this subgroup, viral load has stronger prognostic potential than P16 expression.

Tumor metastasis and immune evasion present major challenges of cancer treatment. Radiotherapy can overcome immunosuppressive tumor microenvironments. Anecdotal reports suggest abscopal anti-tumor immune responses. This study assesses abscopal effects of radiotherapy in combination with mRNA-based cancer vaccination (RNActive®). C57BL/6 mice were injected with ovalbumin-expressing thymoma cells into the right hind leg (primary tumor) and left flank (secondary tumor) with a delay of 4 days. Primary tumors were irradiated with 3 × 2 Gy, while secondary tumors were shielded. RNA and combined treatment groups received mRNA-based RNActive® vaccination. Radiotherapy and combined radioimmunotherapy significantly delayed primary tumor growth with a tumor control in 15 and 53% of mice, respectively. In small secondary tumors, radioimmunotherapy significantly slowed growth rate compared to vaccination (p = 0.002) and control groups (p = 0.01). Cytokine microarray analysis of secondary tumors showed changes in the cytokine microenvironment, even in the non-irradiated contralateral tumors after combination treatment. Combined irradiation and immunotherapy is able to induce abscopal responses, even with low, normofractionated radiation doses. Thus, the combination of mRNA-based vaccination with irradiation might be an effective regimen to induce systemic anti-tumor immunity.

Developing effective mRNA vaccines poses certain challenges concerning mRNA stability and ability to induce sufficient immune stimulation and requires a specific panel of techniques for production and testing. Here, we describe the production of stabilized mRNA with enhanced immunogenicity, generated using conventional nucleotides only, by introducing changes to the mRNA sequence and by complexation with the nucleotide-binding peptide protamine (RNActive® technology). Methods described here include the synthesis, purification, and protamine complexation of mRNA vaccines as well as a comprehensive panel of in vitro and in vivo methods for evaluation of vaccine quality and immunogenicity.

Developing effective mRNA vaccines poses certain challenges concerning mRNA stability and ability to induce sufficient immune stimulation and requires a specific panel of techniques for production and testing. Here, we describe the production of stabilized mRNA with enhanced immunogenicity, generated using conventional nucleotides only, by introducing changes to the mRNA sequence and by complexation with the nucleotide-binding peptide protamine (RNActive® technology). Methods described here include the synthesis, purification, and protamine complexation of mRNA vaccines as well as a comprehensive panel of in vitro and in vivo methods for evaluation of vaccine quality and immunogenicity.

Trogocytosis is a process which involves the transfer of membrane fragments and cell surface proteins between cells. Various types of T cells have been shown to be able to acquire membrane-bound proteins from antigen-presenting cells and their functions can be modulated following trogocytosis. However, it is not known whether induced regulatory T cells (iTregs) can undergo trogocytosis, and if so, what the functional consequences of this process might entail. In this study, we show that iTregs can be generated from CD80(-/-)CD86(-/-) double knockout (DKO) mice. Using flow cytometry and confocal fluorescence microscopy, we demonstrate that iTregs generated from DKO mice are able to acquire both CD80 and CD86 from mature dendritic cells (mDCs) and that the acquisition of CD86 occurs to a higher extent than that of CD80. Furthermore, we found that after co-incubation with iTregs, dendritic cells (DCs) downregulate their surface expression of CD80 and CD86. The trogocytosis of both CD80 and CD86 occurs in a cytotoxic T lymphocyte-associated antigen-4 (CTLA-4), CD28 and programmed death ligand-1 (PDL1)-independent manner. Importantly, we showed that iTregs that acquired CD86 from mDCs expressed higher activation markers and their ability to suppress naive CD4(+) T-cell proliferation was enhanced, compared to iTregs that did not acquire CD86. These data demonstrate, for the first time, that iTregs can acquire CD80 and CD86 from mDCs, and the acquisition of CD86 may enhance their suppressive function. These findings provide novel understanding of the interaction between iTregs and DCs, suggesting that trogocytosis may play a significant role in iTreg-mediated immune suppression.

Antioxidant enzyme activities and lipid peroxidation in Mytilus galloprovincialis from the French Mediterranean coast In the present work we have studied some of the indicators of oxidative damage of the digestive gland tissue of two populations of mussels Mytilus galloprovincialis: native mussels cultured in an aquaculture farm and contaminated mussels collected from the polluted marine area of the French Mediterranean coast located nearby Tamaris and La Seyne-sur-mer - small towns in close proximity to Toulon.