Hybrid Molecules of Azithromycin with Chloramphenicol and Metronidazole: Synthesis and Study of Antibacterial Properties.

The sustained rise of antimicrobial resistance (AMR) causes a strong need to develop new antibacterial agents. One of the methods for addressing the problem of antibiotic resistance is through the design of hybrid antibiotics. In this work, we proposed a synthetic route for the conjugation of an azithromycin derivative with chloramphenicol and metronidazole hemisuccinates and synthesized two series of new hybrid molecules 4a-g and 5a-g. While a conjugation did not result in tangible synergy for wild-type bacterial strains, new compounds were able to overcome AMR associated with the inducible expression of the ermC gene on a model E. coli strain resistant to macrolide antibiotics. The newly developed hybrids demonstrated a tendency to induce premature ribosome stalling, which might be crucial since they will not induce a macrolide-resistant phenotype in a number of pathogenic bacterial strains. In summary, the designed structures are considered as a promising direction for the further development of hybrid molecules that can effectively circumvent AMR mechanisms to macrolide antibiotics.

Comparative Evaluation of the Antibacterial and Antitumor Activities of 9-Phenylfascaplysin and Its Analogs.

Based on the results of our own preliminary studies, the derivative of the marine alkaloid fascaplysin containing a phenyl substituent at C-9 was selected to evaluate the therapeutic potential in vivo and in vitro. It was shown that this compound has outstandingly high antimicrobial activity against Gram-positive bacteria, including antibiotic-resistant strains in vitro. The presence of a substituent at C-9 of the framework is of fundamental importance, since its replacement to neighboring positions leads to a sharp decrease in the selectivity of the antibacterial action, which indicates the presence of a specific therapeutic target in bacterial cells. On a model of the acute bacterial sepsis in mice, it was shown that the lead compound was more effective than the reference antibiotic vancomycin seven out of nine times. However, ED50 value for 9-phenylfascaplysin (7) was similar for the unsubstituted fascaplysin (1) in vivo, despite the former being significantly more active than the latter in vitro. Similarly, assessments of the anticancer activity of compound 7 against various variants of Ehrlich carcinoma in mice demonstrated its substantial efficacy. To conduct a structure-activity relationship (SAR) analysis and searches of new candidate compounds, we synthesized a series of analogs of 9-phenylfascaplysin with varying aryl substituents. However, these modifications led to the reduced aqueous solubility of fascaplysin derivatives or caused a loss of their antibacterial activity. As a result, further research is required to explore new avenues for enhancing its pharmacokinetic characteristics, the modification of the heterocyclic framework, and optimizing of treatment regimens to harness the remarkable antimicrobial potential of fascaplysin for practical usage.

Antibacterial Bionanocomposites Based on Drug-Templated Bifunctional Mesoporous Silica Nanocontainers.

The creation of antibacterial nanocomposites that provide prolonged release of encapsulated drugs is of great interest for various fields of medicine (dentistry, tissue regeneration, etc.). This article demonstrates the possibility of creating such nanocomposites based on sodium alginate and drug-templated mesoporous silica nanocontainers (MSNs) loaded with two bioactive substances. Herein, we thoroughly study all stages of the process, starting with the synthesis of MSNs using antiseptic micelles containing the hydrophobic drug quercetin and ending with assessing the activity of the resulting composites against various microorganisms. The main emphasis is on studying the quercetin solubilization in antiseptic micelles as well as establishing the relationship between the conditions of MSN synthesis and micelle morphology and capacity. The effect of medium pH on the release rate of encapsulated drugs is also evaluated. It was shown that the MSNs contained large amounts of encapsulated drugs and that the rate of drug unloading depended on the medium pH. The incorporation of such MSNs into the alginate matrix allowed for a prolonged release of the drugs.

Formation Kinetics and Antimicrobial Activity of Silver Nanoparticle Dispersions Based on N-Reacetylated Oligochitosan Solutions for Biomedical Applications.

The paper presents the results of the synthesis, a detailed kinetics study, and an investigation of the biological activity of silver nanoparticles (AgNPs) in aqueous solutions of N-reacetylated oligochitosan hydrochloride. UV-visible spectrophotometry and dynamic light scattering were employed to control silver ion reduction. The process was observed to follow a pseudo-first-order law. Transmission and scanning electron microscopy demonstrated that AgNPs ranging in size from 10 to 25 nm formed aggregates measuring 60 to 90 nm, with the aggregate surface coated by a 2-4 nm chitosan shell. X-ray microanalysis and powder X-ray diffractometry were used to study the phase composition, identifying two crystalline phases, nanocrystalline silver and AgCl, present in the dispersions. The antibacterial effect was assessed using the serial dilution method for dispersions with varying degrees of Ag+ conversion. Nanodispersions exhibited significant activity against Escherichia coli, Pseudomonas aeruginosa, Bacillus cereus, and Staphylococcus aureus. Interestingly, the activity did not appear to be heavily influenced by the presence of the AgCl phase or the concentration of Ag+ ions. These synthesized dispersions hold promise for the development of materials tailored for biomedical applications.

Investigation of the Antifungal and Anticancer Effects of the Novel Synthesized Thiazolidinedione by Ion-Conductance Microscopy.

In connection with the emergence of new pathogenic strains of Candida, the search for more effective antifungal drugs becomes a challenge. Part of the preclinical trials of such drugs can be carried out using the innovative ion-conductance microscopy (ICM) method, whose unique characteristics make it possible to study the biophysical characteristics of biological objects with high accuracy and low invasiveness. We conducted a study of a novel synthesized thiazolidinedione's antimicrobial (for Candida spp.) and anticancer properties (on samples of the human prostate cell line PC3), and its drug toxicity (on a sample of the human kidney cell line HEK293). We used a scanning ion-conductance microscope (SICM) to obtain the topography and mechanical properties of cells and an amperometric method using Pt-nanoelectrodes to register reactive oxygen species (ROS) expression. All data and results are obtained and presented for the first time.

Synthesis of 6″-Modified Kanamycin A Derivatives and Evaluation of Their Antibacterial Properties.

Aminoglycosides are one of the first classes of antibiotics to have been used clinically, and they are still being used today. They have a broad spectrum of antimicrobial activity, making them effective against many different types of bacteria. Despite their long history of use, aminoglycosides are still considered promising scaffolds for the development of new antibacterial agents, particularly as bacteria continue to develop resistances to existing antibiotics. We have synthesized a series of 6″-deoxykanamycin A analogues with additional protonatable groups (amino-, guanidino or pyridinium) and tested their biological activities. For the first time we have demonstrated the ability of the tetra-N-protected-6″-O-(2,4,6-triisopropylbenzenesulfonyl)kanamycin A to interact with a weak nucleophile, pyridine, resulting in the formation of the corresponding pyridinium derivative. Introducing small diamino-substituents at the 6″-position of kanamycin A did not significantly alter the antibacterial activity of the parent antibiotic, but further modification by acylation resulted in a complete loss of the antibacterial activity. However, introducing a guanidine residue led to a compound with improved activity against S. aureus. Moreover, most of the obtained 6″-modified kanamycin A derivatives were less influenced by the resistant mechanism associated with mutations of the elongation factor G than the parent kanamycin A. This suggests that modifying the 6″-position of kanamycin A with protonatable groups is a promising direction for the further development of new antibacterial agents with reduced resistances.

Semisynthetic Amides of Amphotericin B and Nystatin A1: A Comparative Study of In Vitro Activity/Toxicity Ratio in Relation to Selectivity to Ergosterol Membranes.

Polyene antifungal amphotericin B (AmB) has been used for over 60 years, and remains a valuable clinical treatment for systemic mycoses, due to its broad antifungal activity and low rate of emerging resistance. There is no consensus on how exactly it kills fungal cells but it is certain that AmB and the closely-related nystatin (Nys) can form pores in membranes and have a higher affinity towards ergosterol than cholesterol. Notably, the high nephro- and hemolytic toxicity of polyenes and their low solubility in water have led to efforts to improve their properties. We present the synthesis of new amphotericin and nystatin amides and a comparative study of the effects of identical modifications of AmB and Nys on the relationship between their structure and properties. Generally, increases in the activity/toxicity ratio were in good agreement with increasing ratios of selective permeabilization of ergosterol- vs. cholesterol-containing membranes. We also show that the introduced modifications had an effect on the sensitivity of mutant yeast strains with alterations in ergosterol biosynthesis to the studied polyenes, suggesting a varying affinity towards intermediate ergosterol precursors. Three new water-soluble nystatin derivatives showed a prominent improvement in safety and were selected as promising candidates for drug development.

Scanning ion-conductance microscopy technique for studying the topography and mechanical properties of Candida parapsilosis yeast microorganisms.

Super-resolution microscopy is widely used in the development of novel antimicrobial testing in vitro. In the presented work, a scanning protocol was developed by the method of scanning ion-conducting microscopy (SICM), which makes it possible to study microorganisms without rigid fixation and in saline, obtaining an index map of nanosized structures. The effect of azole and echinocandins drugs on the morphology and mechanical properties of Candida parapsilosis yeast was studied. The findings are consistent with previously proposed drug mechanisms and reports that have examined antifungal agents using AFM, SEM, and TEM. We have shown that the SICM method is capable of scanning and detecting the nanomechanical properties of yeast non-invasively.

Semisynthetic Amides of Polyene Antibiotic Natamycin.

Natamycin is a macrolide polyene antibiotic, characterized by a potent broad spectrum antifungal activity and low toxicity. However, it is not used for the treatment of systemic mycoses due to its low bioavailability and low solubility in aqueous solutions. In order to create new semisynthetic antifungal agents for treatment of mycoses, a series of water-soluble amides of natamycin were synthesized. Antifungal activities of natamycin derivatives were investigated against Candida spp., including a panel of Candida auris clinical isolates and filamentous fungi. Toxicity for mammalian cells was assayed by monitoring antiproliferative activity against human postnatal fibroblasts (HPF) and human embryonic kidney cells (HEK293). By comparing leakage of contents from ergosterol versus cholesterol containing vesicles, a ratio that characterizes the efficacy and safety of natamycin and its derivatives was determined (EI, efficiency index). Ability of all tested semisynthetic natamycines to prevent proliferation of the yeast Candida spp. cells was comparable or even slightly higher to those of parent antibiotic. Interestingly, amide 8 was more potent than natamycin (1) against all tested C. auris strains (MIC values 2 µg/mL vs 8 µg/mL, respectively). Among 7 derivatives, amide 10 with long lipophilic side chains showed the highest EI and strong antifungal activity in vitro but was more toxic against HPF. In vivo experiments with amide 8 showed in vivo efficacy on a mouse candidemia model with a larger LD50/ED50 ratio in comparison to amphotericin B.

Antifungal Thiazolidines: Synthesis and Biological Evaluation of Mycosidine Congeners.

Novel derivatives of Mycosidine (3,5-substituted thiazolidine-2,4-diones) are synthesized by Knoevenagel condensation and reactions of thiazolidines with chloroformates or halo-acetic acid esters. Furthermore, 5-Arylidene-2,4-thiazolidinediones and their 2-thioxo analogs containing halogen and hydroxy groups or di(benzyloxy) substituents in 5-benzylidene moiety are tested for antifungal activity in vitro. Some of the synthesized compounds exhibit high antifungal activity, both fungistatic and fungicidal, and lead to morphological changes in the Candida yeast cell wall. Based on the use of limited proteomic screening and toxicity analysis in mutants, we show that Mycosidine activity is associated with glucose transport. This suggests that this first-in-class antifungal drug has a novel mechanism of action that deserves further study.

Synthesis and Antibacterial Activity Studies of the Conjugates of Curcumin with closo-Dodecaborate and Cobalt Bis(Dicarbollide) Boron Clusters.

A series of novel conjugates of cobalt bis(dicarbollide) and closo-dodecaborate with curcumin were synthesized by copper(I)-catalyzed azide-alkyne cycloaddition. These conjugates were tested for antibacterial activity. It was shown that all derivatives are active when exposed to Bacillus cereus ATCC 10702 and are not active against Gram-negative microorganisms and Candida albicans at the maximum studied concentration of 1000 mg/L. The conjugate of alkynyl-curcumin with azide synthesized from the tetrahydropyran derivative of cobalt bis(dicarbollide) exhibited activity against Gram-positive microorganisms: Staphylococcus aureus ATCC 29213, Enterococcus faecalis ATCC 29212 and the clinical isolate MRSA 17, that surpassed curcumin by 2-4 times.

Synthesis and Characterization of Novel 2-Acyl-3-trifluoromethylquinoxaline 1,4-Dioxides as Potential Antimicrobial Agents.

The emergence of drug resistance in pathogens leads to a loss of effectiveness of antimicrobials and complicates the treatment of bacterial infections. Quinoxaline 1,4-dioxides represent a prospective scaffold for search of new compounds with improved chemotherapeutic characteristics. Novel 2-acyl-3-trifluoromethylquinoxaline 1,4-dioxides with alteration of substituents at position 2 and 6 were synthesized via nucleophilic substitution with piperazine moiety and evaluated against a broad panel of bacteria and fungi by measuring their minimal inhibitory concentrations. Their mode of action was assessed by whole-genomic sequencing of spontaneous drug-resistant Mycobacterium smegmatis mutants, followed by comparative genomic analysis, and on an original pDualrep2 system. Most of the 2-acyl-3-trifluoromethylquinoxaline 1,4-dioxides showed high antibacterial properties against Gram-positive strains, including mycobacteria, and the introduction of a halogen atom in the position 6 of the quinoxaline ring further increased their activity, with 13c being the most active compound. The mode of action studies confirmed the DNA-damaging nature of the obtained quinoxaline 1,4-dioxides, while drug-resistance may be provided by mutations in redox homeostasis genes, encoding enzymes potentially involved in the activation of the compounds. This study extends views about the antimicrobial and antifungal activities of the quinoxaline 1,4-dioxides and can potentially lead to the discovery of new antibacterial drugs.

Stereochemistries and Biological Properties of Oligomycin A Diels-Alder Adducts.

Oligomycin A is a potent antibiotic and antitumor agent. However, its applications are restricted by its high toxicity and low bioavailability. In this study, we obtained Oligomycin A Diels-Alder adducts with benzoquinone and N-benzylmaleimide and determined their absolute configurations by combining 1H and ROESY NMR data with molecular mechanics conformational analysis and quantum chemical reaction modeling. The latter showed that adduct stereochemistry is controlled by hydrogen bonding of the Oligomycin A side-chain isopropanol moiety with the carbonyl group of the dienophile. Biological studies showed that the Diels-Alder modification of the Oligomycin A diene system resulted in a complex antiproliferative potential pattern. The synthesized adducts were determined to be more active against the triple-negative (ERα, PR, and HER2 negative) breast cancer cell line MDA-MB-231 and lung carcinoma cell line A-549 compared to Oligomycin A. Meanwhile, Oligomycin A was more potent against myeloid leukemia cell line K-562 and breast carcinoma cell line MCF-7 than its derivatives. Thus, modification of the diene moiety of Oligomycin A is a promising strategy for developing novel antitumor agents based on its scaffold.

Gausemycins†A,B: Cyclic Lipoglycopeptides from Streptomyces sp.*.

We report a novel family of natural lipoglycopeptides produced by Streptomyces sp. INA-Ac-5812. Two major components of the mixture, named gausemycins A and B, were isolated, and their structures were elucidated. The compounds are cyclic peptides with a unique peptide core and several remarkable structural features, including unusual positions of d-amino acids, lack of the Ca2+ -binding Asp-X-Asp-Gly (DXDG) motif, tyrosine glycosylation with arabinose, presence of 2-amino-4-hydroxy-4-phenylbutyric acid (Ahpb) and chlorinated kynurenine (ClKyn), and N-acylation of the ornithine side chain. Gausemycins have pronounced activity against Gram-positive bacteria. Mechanistic studies highlight significant differences compared to known glyco- and lipopeptides. Gausemycins exhibit only slight Ca2+ -dependence of activity and induce no pore formation at low concentrations. Moreover, there is no detectable accumulation of cell wall biosynthesis precursors under treatment with gausemycins.

Aminoalkylamides of Eremomycin Exhibit an Improved Antibacterial Activity.

After decades, the glycopeptide vancomycin is still the preferred antibiotic against resistant strains of Gram-positive bacteria. Although its clinical use is strictly regulated, the gradual spread of vancomycin-resistant bacteria, such as glycopeptide-resistant and glycopeptide-intermediate Staphylococcus aureus and vancomycin-resistant Enterococcus spp., is a serious health problem. Based on the literature data and previous studies, our main goal was to assess the antimicrobial potential and to study the structure-activity relationship of new eremomycin aminoalkylamides. We designed and synthesized a series of new eremomycin amides in which eremomycin is conjugated with a hydrophobic arylalkyl group via an alkylenediamine spacer, and tested their antibacterial activities on a panel of Gram-positive strains that were sensitive and resistant to a "gold-standard" vancomycin. Based on the data obtained, the structure-activity relationships were investigated, and a lead compound was selected for in-depth testing. Research carried out using an in vivo model of staphylococcus sepsis, acute toxicity studies, and the estimated therapeutic index also showed the advantage of the selected eremomycin amide derivative in particular, as well as the chosen direction in general.

Bacterial Cell Wall Analogue Peptides Control the Oligomeric States and Activity of the Glycopeptide Antibiotic Eremomycin: Solution NMR and Antimicrobial Studies.

For some time, glycopeptide antibiotics have been considered the last line of defense against Methicillin-resistant Staphylococcus aureus (MRSA). However, vancomycin resistance of Gram-positive bacteria is an increasingly emerging worldwide health problem. The mode of action of glycopeptide antibiotics is essentially the binding of peptidoglycan cell-wall fragments terminating in the d-Ala-d-Ala sequence to the carboxylate anion binding pocket of the antibiotic. Dimerization of these antibiotics in aqueous solution was shown to persist and even to enhance the antibacterial effect in a co-operative manner. Some works based on solid state (ss) Nuclear Magnetic Resonance (NMR) studies questioned the presence of dimers under the conditions of ssNMR while in a few cases, higher-order oligomers associated with contiguous back-to-back and face-to-face dimers were observed in the crystal phase. However, it is not proved if such oligomers persist in aqueous solutions. With the aid of 15N-labelled eremomycin using 15N relaxation and diffusion NMR methods, we observed tetramers and octamers when the N-Ac-d-Ala-d-Ala dipeptide was added. To the contrary, the N-Ac-d-Ala or (N-Ac)2-l-Lys-d-Ala-d-Ala tripeptide did not induce higher-order oligomers. These observations are interesting examples of tailored supramolecular self-organization. New antimicrobial tests have also been carried out with these self-assemblies against MRSA and VRE (resistant) strains.

Design, synthesis and biomedical evaluation of mostotrin, a new water soluble tryptanthrin derivative.

Mostotrin (MT), a novel compound, at least five orders of magnitude more soluble in water than its mother substance, was designed and synthesised from tryptanthrin (TR). Its structure was established by nuclear magnetic resonance and mass spectrometry data and confirmed by X­ray analysis, revealing that MT is a pentacyclic product with an additional pseudo­cycle formed with the participation of one intramolecular hydrogen bond. Antimicrobial activity and cytotoxic action against tumour cells in vitro, as well as anti­tumour effects, acute toxicity and anti­inflammatory activities in vivo, were evaluated. Antimicrobial properties of MT against Mycobacterium spp and Bacillus cereus ATCC 10702 appeared to be the same as that of TR, but against the other strains used it was weaker. Furthermore, MT exhibited 5­10 times higher cytotoxic activities against tumour cell lines HCT­116, МСF­7 and K­562 than TR, but was less toxic than TR (LD50 of MT was 375 mg/kg, while LD50 for TR was 75 mg/kg). Additionally, compounds MT and TR were studied in DNA binding tests. The quenching of its fluorescence on addition to DNA solution established MT to be capable of binding to DNA. Its anti­tumour action in vivo on mice with the ascitic form of Ehrlich carcinoma was promising, particularly with joint application of MT and the antitumour drug doxorubicin. In this model, the survival and life span for the doxorubicin and 1 co­treatment group were significantly higher compared to doxorubicin treatment alone. The compound MT showed a lower immunosuppressive effect than TR at the early stages of inflammation induced in mice by LPS from E. Ñoli (MT hardly inhibited the release of IL­1, IL­2, or INF­Î³). These results demonstrated that MT is a perspective hit compound for drug development. In our opinion, further evaluation on the biological effects of MT and its synthetic analogues could lead to safer and more effective anti­tumour and anti­tuberculosis agents than TR itself. MT has also the prospect of application in combination with known anti­tumour drugs for the treatment of oncological diseases.

Discovery of Amphamide, a Drug Candidate for the Second Generation of Polyene Antibiotics.

Amphotericin B (AmB, 1) is the drug of choice for treating the most serious systemic fungal or protozoan infections. Nevertheless, its application is limited by low solubility in aqueous media and serious side effects such as infusion-related reactions, hemolytic toxicity, and nephrotoxicity. Owing to these limitations, it is essential to search for the polyene derivatives with better chemotherapeutic properties. With the objective of obtaining AmB derivatives with lower self-aggregation and improved solubility, we synthesized a series of amides of AmB bearing an additional basic group in the introduced residue. The screening of antifungal activity in vitro revealed that N-(2-aminoethyl)amide of AmB (amphamide, 6) had superior antifungal activity compared to that of the paternal AmB. Preclinical studies in mice confirmed that compound 6 had a much lower acute toxicity and higher antifungal efficacy in the model of mice candidosis sepsis compared with that of AmB (1). Thus, the discovered amphamide is a promising drug candidate for the second generation of polyene antibiotics and is also prospective for in-depth preclinical and clinical evaluation.

Crystallomycin revisited after 60 years: aspartocins B and C.

The study of an archived sample of crystallomycin complex using HPLC, ESI HRMS, and 2D NMR showed that two major components of the antibiotic, compounds 1 and 2, are lipopeptides having the same peptide core, Asp1-cyclo(Dab2-Pip3-MeAsp4-Asp5-Gly6-Asp7-Gly8-Dab9-Val10-Pro11-), N-acylated either with Δ3-iso-tetradecenoyl or Δ3-anteiso-pentadecenoyl that are identical to aspartocins C and B, respectively. According to the 2D NMR study, compound 2 in DMSO solution exists as a mixture of four conformers. The producing strain was identified as Streptomyces griseorubens. Compounds 1 and 2 have considerable Ca2+-dependent activity against Gram-positive bacteria including five MRSA strains.