Xanthone Derivatives Enhance the Therapeutic Potential of Neomycin against Polymicrobial Gram-Negative Bacterial Infections.

Gram-negative bacterial infections are difficult to manage as many antibiotics are ineffective owing to the presence of impermeable bacterial membranes. Polymicrobial infections pose a serious threat due to the inadequate efficacy of available antibiotics, thereby necessitating the administration of antibiotics at higher doses. Antibiotic adjuvants have emerged as a boon as they can augment the therapeutic potential of available antibiotics. However, the toxicity profile of antibiotic adjuvants is a major hurdle in clinical translation. Here, we report the design, synthesis, and biological activities of xanthone-derived molecules as potential antibiotic adjuvants. Our SAR studies witnessed that the p-dimethylamino pyridine-derivative of xanthone (X8) enhances the efficacy of neomycin (NEO) against Escherichia coli and Pseudomonas aeruginosa and causes a synergistic antimicrobial effect without any toxicity against mammalian cells. Biochemical studies suggest that the combination of X8 and NEO, apart from inhibiting protein synthesis, enhances the membrane permeability by binding to lipopolysaccharide. Notably, the combination of X8 and NEO can disrupt the monomicrobial and polymicrobial biofilms and show promising therapeutic potential against a murine wound infection model. Collectively, our results unveil the combination of X8 and NEO as a suitable adjuvant therapy for the inhibition of the Gram-negative bacterial infections.

Cholic Acid-Derived Gemini Amphiphile Can Eradicate Interkingdom Polymicrobial Biofilms and Wound Infections.

Biofilm infections are mainly caused by Gram-positive bacteria (GPB) like Staphylococcus aureus, Gram-negative bacteria (GNB) like Pseudomonas aeruginosa, and fungi like Candida albicans. These infections are responsible for antimicrobial tolerance, and commensal interactions of these microbes pose a severe threat to chronic infections. Treatment therapies against biofilm infections are limited to eradicating only 20-30% of infections. Here, we present the synthesis of a series of bile acid-derived molecules using lithocholic acid, deoxycholic acid, and cholic acid where two bile acid molecules are tethered through 3'-hydroxyl or 24'-carboxyl terminals with varying spacer length (trimethylene, pentamethylene, octamethylene, and dodecamethylene). Our structure-activity relationship investigations revealed that G21, a cholic acid-derived gemini amphiphile having trimethylene spacer tethered through the C24 position, is a broad-spectrum antimicrobial agent. Biochemical studies witnessed that G21 interacts with negatively charged lipoteichoic acid, lipopolysaccharide, and phosphatidylcholine moieties of GPB, GNB, and fungi and disrupts the microbial cell membranes. We further demonstrated that G21 can eradicate polymicrobial biofilms and wound infections and prevent bacteria and fungi from developing drug resistance. Therefore, our findings revealed the potential of G21 as a versatile antimicrobial agent capable of effectively targeting polymicrobial biofilms and wound infections, suggesting that it is a promising antimicrobial agent for future applications.

Recent developments in membrane targeting antifungal agents to mitigate antifungal resistance.

Fungal infections cause severe and life-threatening complications especially in immunocompromised individuals. Antifungals targeting cellular machinery and cell membranes including azoles are used in clinical practice to manage topical to systemic fungal infections. However, continuous exposure to clinically used antifungal agents in managing the fungal infections results in the development of multi-drug resistance via adapting different kinds of intrinsic and extrinsic mechanisms. The unique chemical composition of fungal membranes presents attractive targets for antifungal drug discovery as it is difficult for fungal cells to modify the membrane targets for emergence of drug resistance. Here, we discussed available antifungal drugs with their detailed mechanism of action and described different antifungal resistance mechanisms. We further emphasized structure-activity relationship studies of membrane-targeting antifungal agents, and classified membrane-targeting antifungal agents on the basis of their core scaffold with detailed pharmacological properties. This review aims to pique the interest of potential researchers who could explore this interesting and intricate fungal realm.

Targeting Vancomycin-Resistant Enterococci (VRE) Infections and Van Operon-Mediated Drug Resistance Using Dimeric Cholic Acid-Peptide Conjugates.

Emergence of vancomycin resistance in Gram-positive bacteria and the prevalence of vancomycin-resistant Enterococci (VRE) infections are highly alarming as very limited antibiotic options are available against VRE infections. Here, we present the synthesis of cholic acid-derived dimeric amphiphiles where two cholic acid moieties are tethered through carboxyl terminals using different alkylene spacers. Our investigations revealed that dimer 5 possessing a propylene spacer and glycine-valine peptides tethered on hydroxyl groups is the most effective antimicrobial against VRE. Dimer 5 can permeabilize bacterial membranes, generate reactive oxygen species, and clear preformed biofilms. We further demonstrate that dimer 5 downregulates vancomycin-mediated transcriptional activation of the vanHAX gene cluster and does not allow VSE to develop vancomycin resistance until 100 generations. Therefore, this study, for the first time, presents a bacterial membrane-targeting amphiphile that can mitigate VRE infections and inhibit the emergence of vancomycin resistance.

Hydrophobicity of Cholic Acid-Derived Amphiphiles Dictates the Antimicrobial Specificity.

The unique structural components of cell membranes of Gram-positive bacteria, Gram-negative bacteria, and mycobacteria provide an excellent therapeutic target for developing highly specific antimicrobials. Here, we report the synthesis of nine cholic acid (CA)-derived amphiphiles, where three hydroxyl groups of CA were tethered to dimethylamino pyridine and the C24-carboxyl group was conjugated with different alkyl chains. Structure-activity investigations revealed that amphiphile 1 harboring a methyl group has antimicrobial activity against mycobacterial species. On the other hand, amphiphile 7 containing an octyl chain was selective against Gram-positive and Gram-negative bacilli. Biochemical assays confirmed the selective membrane permeabilization abilities of amphiphiles 1 and 7. Importantly, we demonstrate the selective actions of amphiphiles in clearing biofilms, intracellular bacteria, and wound infections. Therefore, for the first time, we show that the unique structural features of CA-derived amphiphiles dictate selective activity against specific bacterial species.

Correction: Cytocompatible, soft and thick brush-modified scaffolds with prolonged antibacterial effect to mitigate wound infections.

Correction for 'Cytocompatible, soft and thick brush-modified scaffolds with prolonged antibacterial effect to mitigate wound infections' by Shaifali Dhingra et al., Biomater. Sci., 2022, 10, 3856-3877, https://doi.org/10.1039/d2bm00245k.

Polyaspartate-derived synthetic antimicrobial polymer enhances the activity of rifampicin against multidrug-resistant Pseudomonas aeruginosa infections.

Infections caused by multidrug-resistant Pseudomonas aeruginosa (P. aeruginosa) pose major challenges for treatment due to the acquired, adaptive, and intrinsic resistance developed by the bacteria. Accumulation of mutations, the ability to form biofilms, and the presence of lipopolysaccharides in the outer bacterial membranes are the key mechanisms of drug resistance. Here, we show that a polyaspartate-derived synthetic antimicrobial polymer (SAMP) with a hexyl chain (TAC6) is an effective adjuvant for a hydrophobic antibiotic, rifampicin. Our in vitro studies demonstrated that the combination of TAC6 and rifampicin is effective against clinically isolated multidrug-resistant strains of P. aeruginosa. Membrane permeabilization studies showed that TAC6 allows the permeabilization of bacterial membranes, and the accumulation of rifampicin inside the cells, thereby enhancing its activity. The combination of TAC6 and rifampicin can also degrade the P. aeruginosa biofilms, and therefore can mitigate the adaptive resistance developed by bacteria. We further demonstrated that the combination of TAC6 and rifampicin can clear P. aeruginosa-mediated wound infections effectively. Therefore, our study showed polyaspartate-derived SAMP to be an effective antibiotic adjuvant against P. aeruginosa infections.

N-methyl Benzimidazole Tethered Cholic Acid Amphiphiles Can Eradicate S. aureus-Mediated Biofilms and Wound Infections.

Infections associated with Gram-positive bacteria like S. aureus pose a major threat as these bacteria can develop resistance and thereby limit the applications of antibiotics. Therefore, there is a need for new antibacterials to mitigate these infections. Bacterial membranes present an attractive therapeutic target as these membranes are anionic in nature and have a low chance of developing modifications in their physicochemical features. Antimicrobial peptides (AMPs) can disrupt the microbial membranes via electrostatic interactions, but the poor stability of AMPs halts their clinical translation. Here, we present the synthesis of eight N-methyl benzimidazole substituted cholic acid amphiphiles as antibacterial agents. We screened these novel heterocyclic cholic acid amphiphiles against different pathogens. Among the series, CABI-6 outperformed the other amphiphiles in terms of bactericidal activity against S. aureus. The membrane disruptive property of CABI-6 using a fluorescence-based assay has also been investigated, and it was inferred that CABI-6 can enhance the production of reactive oxygen species. We further demonstrated that CABI-6 can clear the pre-formed biofilms and can mitigate wound infection in murine models.

Cytocompatible, soft and thick brush-modified scaffolds with prolonged antibacterial effect to mitigate wound infections.

Biomedical device or implant-associated infections caused by pathogenic bacteria are a major clinical issue, and their prevention and/or treatment remains a challenging task. Infection-resistant antimicrobial coatings with impressive cytocompatibility offer a step towards addressing this problem. Herein, we report a new strategy for constructing highly antibacterial as well as cytocompatible mixed polymer brushes onto the surface of 3D printed scaffold made of biodegradable tartaric acid-based aliphatic polyester blends. The mixed brushes were nothing but a combination of poly(3-dimethyl-(methacryloyloxyethyl) ammonium propane sulfonate) (polyDMAPS) and poly((oligo ethylene glycol) methyl ether methacrylate) (polyPEGMA) with varying chain length (n) of the ethylene glycol unit (n = 1, 6, 11, and 21). Both homo and copolymeric brushes of polyDMAPS with polyPEGMA exhibited antibacterial efficacy against both Gram positive and Gram negative pathogens such as E. coli (Escherichia coli) and S. aureus (Staphylococcus aureus) because of the combined action of bacteriostatic effects originating from strongly hydrated layers present in zwitterionic (polyDMAPS) and hydrophilic (polyPEGMA) copolymer brushes. Interestingly, a mixed polymer brush comprising polyDMAPS and polyPEGMA (ethylene glycol chain unit of 21) at 50/50 ratio provided zero bacterial growth and almost 100% cytocompatibility (tested using L929 mouse fibroblast cells), making the brush-modified biodegradable substrate an excellent choice for an infection-resistant and cytocompatible surface. An attempt was made to understand their extraordinary performance with the help of contact angle, surface charge analysis and nanoindentation study, which revealed the formation of a hydrophilic, almost neutral, very soft surface (99.99% reduction in hardness and modulus) after modification with the mixed brushes. This may completely suppress bacterial adhesion. Animal studies demonstrated that these brush-modified scaffolds are biocompatible and can mitigate wound infections. Overall, this study shows that the fascinating combination of an infection-resistant and cytocompatible surface can be generated on biodegradable polymeric surfaces by modulating the surface hardness, flexibility and hydrophilicity by selecting appropriate functionality of the copolymeric brushes grafted onto them, making them ideal non-leaching, anti-infective, hemocompatible and cytocompatible coatings for biodegradable implants.

Development of a Highly Specific, Selective, and Sensitive Fluorescent Probe for Detection of Mycobacteria in Human Tissues.

Tuberculosis (TB), including extrapulmonary TB, is responsible for more than one million deaths in a year worldwide. Existing methods of mycobacteria detection have poor sensitivity, selectivity, and specificity, especially in human tissues. Herein, the synthesis of a cholic acid-derived fluorescent probe (P4) that can specifically stain the mycobacterium species is presented. It is shown that P4 probe specifically binds with mycobacterial lipids, trehalose monomycolate, and phosphatidylinositol mannoside 6. P4 probe can detect mycobacteria in polymicrobial planktonic cultures and biofilms with high specificity, selectivity, and sensitivity. Moreover, it can detect a single mycobacterium in the presence of 10 000 other bacilli. Unlike the probes that depend on active mycobacterial enzymes, the membrane-specific P4 probe can detect mycobacteria even in formalin-fixed paraffin-embedded mice and human tissue sections. Therefore, the ability of the P4 probe to detect mycobacteria in different biological milieu makes it a potential candidate for diagnostic and prognostic applications in clinical settings.

Unlocking the bacterial membrane as a therapeutic target for next-generation antimicrobial amphiphiles.

Gram-positive bacteria like Enterococcus faecium and Staphylococcus aureus, and Gram-negative bacteria like Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter Spp. are responsible for most of fatal bacterial infections. Bacteria present a handful of targets like ribosome, RNA polymerase, cell wall biosynthesis, and dihydrofolate reductase. Antibiotics targeting the protein synthesis like aminoglycosides and tetracyclines, inhibitors of RNA/DNA synthesis like fluoroquinolones, inhibitors of cell wall biosynthesis like glycopeptides and ß-lactams, and membrane-targeting polymyxins and lipopeptides have shown very good success in combating the bacterial infections. Ability of the bacteria to develop drug resistance is a serious public health challenge as bacteria can develop antimicrobial resistance against newly introduced antibiotics that enhances the challenge for antibiotic drug discovery. Therefore, bacterial membranes present a suitable therapeutic target for development of antimicrobials as bacteria can find it difficult to develop resistance against membrane-targeting antimicrobials. In this review, we present the recent advances in engineering of membrane-targeting antimicrobial amphiphiles that can be effective alternatives to existing antibiotics in combating bacterial infections.

Predicting antimicrobial peptides with improved accuracy by incorporating the compositional, physico-chemical and structural features into Chou's general PseAAC.

Antimicrobial peptides (AMPs) are important components of the innate immune system that have been found to be effective against disease causing pathogens. Identification of AMPs through wet-lab experiment is expensive. Therefore, development of efficient computational tool is essential to identify the best candidate AMP prior to the in vitro experimentation. In this study, we made an attempt to develop a support vector machine (SVM) based computational approach for prediction of AMPs with improved accuracy. Initially, compositional, physico-chemical and structural features of the peptides were generated that were subsequently used as input in SVM for prediction of AMPs. The proposed approach achieved higher accuracy than several existing approaches, while compared using benchmark dataset. Based on the proposed approach, an online prediction server iAMPpred has also been developed to help the scientific community in predicting AMPs, which is freely accessible at http://cabgrid.res.in:8080/amppred/. The proposed approach is believed to supplement the tools and techniques that have been developed in the past for prediction of AMPs.