Antibiotic-resistant Gram-positive Pathogens Research Articles

Establishing the Comprehensive Structure-Activity Relationship of the Natural Antibiotic Kibdelomycin/Amycolamicin.

Kibdelomycin (KBD) and amycolamicin (AMM) are potent natural antibiotics effective against antibiotic-resistant Gram-positive pathogens, including vancomycin-intermediate Staphylococcus aureus (S. aureus, VISA), methicillin-resistant S. aureus (MRSA), and quinolone-resistant S. aureus (QRSA). Their antibacterial activity stems from an unprecedented dual mechanism: the lower binding sites occupy the adenosine triphosphate (ATP) binding pocket of bacterial type II topoisomerases, while the upper binding sites disrupt the enzyme dimer interface. This dual action, combined with their unique chemical structures, positions KBD and AMM as promising scaffolds for developing new antibiotics. However, the structure-activity relationship (SAR) of KBD/AMM remains underexplored due to their highly complex chemical structures. In this study, we utilized total synthesis to produce KBD/AMM analogs with various site modifications and evaluated their antimicrobial activities. Our findings establish the first comprehensive SAR for KBD/AMM, paving the way for the development of novel KBD/AMM-based antibiotics.

Antibiotic Resistance to Molecules Commonly Prescribed for the Treatment of Antibiotic-Resistant Gram-Positive Pathogens: What Is Relevant for the Clinician?

Antibiotic resistance in Gram-positive pathogens is a relevant concern, particularly in the hospital setting. Several antibiotics are now available to treat these drug-resistant pathogens, such as daptomycin, dalbavancin, linezolid, tedizolid, ceftaroline, ceftobiprole, and fosfomycin. However, antibiotic resistance can also affect these newer molecules. Overall, this is not a frequent phenomenon, but it is a growing concern in some settings and can compromise the effectiveness of these molecules, leaving few therapeutic options. We reviewed the available evidence about the epidemiology of antibiotic resistance to these antibiotics and the main molecular mechanisms of resistance, particularly methicillin-resistant Sthaphylococcus aureus, methicillin-resistant coagulase-negative staphylococci, vancomycin-resistant Enterococcus faecium, and penicillin-resistant Streptococcus pneumoniae. We discussed the interpretation of susceptibility tests when minimum inhibitory concentrations are not available. We focused on the risk of the emergence of resistance during treatment, particularly for daptomycin and fosfomycin, and we discussed the strategies that can be implemented to reduce this phenomenon, which can lead to clinical failure despite appropriate antibiotic treatment. The judicious use of antibiotics, epidemiological surveillance, and infection control measures is essential to preserving the efficacy of these drugs.

Total Synthesis of Mannopeptimycin β via β-Hydroxyenduracididine Ligation.

Nonribosomal peptide synthesis in bacteria has endowed cyclic peptides with fascinating structural complexity via incorporating nonproteinogenic amino acids. These bioactive cyclic peptides provide interesting structural motifs for exploring total synthesis and medicinal chemistry studies. Cyclic glycopeptide mannopeptimycins exhibit antibacterial activity against antibiotic-resistant Gram-positive pathogens and act as the lipid II binder to stop bacterial cell wall biosynthesis. Here, we report a strategy streamlining solution phase-solid phase synthesis and chemical ligation-mediated peptide cyclization for the total synthesis of mannopeptimycin β.

Establishing the Structure-Activity Relationship of Daptomycin.

Daptomycin is effective in treating infections caused by antibiotic-resistant Gram-positive pathogens, including methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococci (VRE), and vancomycin-resistant S. aureus (VRSA). Due to its distinct mechanism of action toward multidrug-resistant bacteria, daptomycin provides an attractive structural motif to generate new daptomycin-based antibiotics to combat the problem of bacterial resistance. In this study, we used the total synthesis method to produce daptomycin analogues with a variety in terms of types and sites of modifications. Five classes of daptomycin analogues were synthesized, and the antimicrobial activities of the analogues were analyzed by several biological assays. From this study, we established a comprehensive structure-activity relationship of daptomycin which will lay the foundation for the further development of daptomycin-based antibiotics.

Dual Targeting of Cell Wall Precursors by Teixobactin Leads to Cell Lysis

Teixobactin represents the first member of a newly discovered class of antibiotics that act through inhibition of cell wall synthesis. Teixobactin binds multiple bactoprenol-coupled cell wall precursors, inhibiting both peptidoglycan and teichoic acid synthesis. Here, we show that the impressive bactericidal activity of teixobactin is due to the synergistic inhibition of both targets, resulting in cell wall damage, delocalization of autolysins, and subsequent cell lysis. We also find that teixobactin does not bind mature peptidoglycan, further increasing its activity at high cell densities and against vancomycin-intermediate Staphylococcus aureus (VISA) isolates with thickened peptidoglycan layers. These findings add to the attractiveness of teixobactin as a potential therapeutic agent for the treatment of infection caused by antibiotic-resistant Gram-positive pathogens.

From endolysins to Artilysin®s: novel enzyme-based approaches to kill drug-resistant bacteria.

One of the last untapped reservoirs in nature for the identification of new anti-microbials is bacteriophages, the natural killers of bacteria. Lytic bacteriophages encode peptidoglycan (PG) lytic enzymes able to degrade the PG layer in different steps of their infection cycle. Endolysins degrade the bacterial cell wall at the end of the infection cycle, causing lysis of the host to release the viral progeny. Recombinant endolysins have been successfully applied as anti-bacterial agent against antibiotic-resistant Gram-positive pathogens. This has boosted the study of these enzymes as new anti-microbials in different fields (e.g. medical, food technology). A key example is the recent development of endolysin-based anti-bacterials against Gram-negative pathogens in which the exogenous application of endolysins is hindered by the outer membrane (OM). These novel anti-microbials, termed Artilysin®s, are able to pass through the OM and reach the PG where they exert their action. In addition, mycobacteria whose cell wall is structurally different from both Gram-positive and Gram-negative bacteria have also been reported to be inhibited by mycobacteriophage-encoded endolysins. Endolysins and endolysin-based anti-microbials can be considered as ideal candidates for an alternative to antibiotics for several reasons: (1) their unique mode of action and activity against bacterial persisters (independent of an active host metabolism), (2) their selective activity against both Gram-positive and Gram-negative pathogens (including antibiotic resistant strains) and mycobacteria, (3) the limited resistance development reported so far. The present review summarizes and discusses the potential applications of endolysins as new anti-microbials.

EC300: a phage-based, bacteriolysin-like protein with enhanced antibacterial activity against Enterococcus faecalis.

Bacteriophage lytic enzymes, either endolysins or virion-associated lysins, have been receiving considerable attention as potential antibacterial agents, particularly for the combat of antibiotic-resistant Gram-positive pathogens. A conclusion that easily emerges from the careful analysis of a great number of reports on the field is that the activity of phage lytic enzymes is rarely studied in conditions that support robust growth of the target bacteria. Here, we report the construction and study of a chimerical lysin, EC300, which was designed to target and kill Enterococcus faecalis in conditions supporting vigorous bacterial growth. EC300 resulted from the fusion of a predicted M23 endopeptidase domain of a virion-associated lysin to the putative cell wall binding domain of a previously characterized amidase endolysin, both produced by the E. faecalis phage F170/08. This bacteriolysin-like protein exhibited a clear enhanced lytic activity over the parental endolysin when both were assayed in a rich bacterial growth medium. We demonstrate the killing efficacy of EC300 against growing cells of a panel of typed E. faecalis clinical strains with high level of antibiotic resistance. The possible reasons for the marked difference between the lytic performance of EC300 and that of the amidase are discussed.

Heterologous production of thiostrepton A and biosynthetic engineering of thiostrepton analogs

Thiostrepton A 1, produced by Streptomyces laurentii ATCC 31255 (S. laurentii), is one of the more well-recognized thiopeptide metabolites. Thiostrepton A 1 and other thiopeptides are of great interest due to their potent activities against emerging antibiotic-resistant Gram-positive pathogens. Although numerous lines of evidence have established that the thiopeptides arise from the post-translational modification of ribosomally-synthesized peptides, few details have been revealed concerning this elaborate process. Alteration to the primary amino acid sequence of the precursor peptide provides an avenue to probe the substrate specificity of the thiostrepton post-translational machinery. Due to the difficulties in the genetic manipulation of S. laurentii, the heterologous production of thiostrepton A 1 from an alternate streptomycete host was sought to facilitate the biosynthetic investigations of the peptide metabolite. The production of thiostrepton A 1 from the non-cognate hosts did not lend itself to be as robust as S. laurentii-based production, therefore an alternate strategy was pursued for the production of thiostrepton variants. The introduction of a fosmid used in the heterologous production of thiostrepton A 1, harboring the entire thiostrepton biosynthetic gene cluster, into the tsrA deletion mutant permitted restoration of thiostrepton A 1 production near to that of the wild-type level. The fosmid was then engineered to enable the replacement of wild-type tsrA. Introduction of expression fosmids encoding alternate TsrA sequences into the S. laurentii tsrA deletion mutant led to the production of thiostrepton variants retaining antibacterial activity, demonstrating the utility of this expression platform toward thiopeptide engineering.

Clinical pharmacology of linezolid: an oxazolidinone antimicrobial agent

The oxazolidinones are a new class of antibiotics whose mechanism of action is the inhibition of bacterial protein synthesis by binding to the 50S ribosomal subunit. Linezolid is the first member of this class of antibiotics to be approved by the US FDA and other regulatory bodies for the treatment of infections caused by antibiotic-resistant Gram-positive pathogens, such as methicillin-resistant Staphylococcus aureus, glycopeptide-intermediate S. aureus and vancomycin-resistant Enterococcus, in adults and children.

Antibacterial Activity and Mechanism of Action of a Novel Anilinouracil-Fluoroquinolone Hybrid Compound

The anilinouracils (AUs) such as 6-(3-ethyl-4-methylanilino)uracil (EMAU) are a novel class of gram-positive, selective, bactericidal antibacterials which inhibit pol IIIC, the gram-positive-specific replicative DNA polymerase. We have linked various fluoroquinolones (FQs) to the N-3 position of EMAU to generate a variety of AU-FQ "hybrids" offering the potential for targeting two distinct steps in DNA replication. In this study, the properties of a hybrid, "251D," were compared with those of representative AUs and FQs in a variety of in vitro assays, including pol IIIC and topoisomerase/gyrase enzyme assays, antibacterial, bactericidal, and mammalian cytotoxicity assays. Compound 251D potently inhibited pol IIIC and topoisomerase/gyrase, displayed gram-positive antibacterial potency at least 15 times that of the corresponding AU compound, and as expected, acted selectively on bacterial DNA synthesis. Compound 251D was active against a broad panel of antibiotic-resistant gram-positive pathogens as well as several gram-negative organisms and was also active against both AU- and FQ-resistant gram-positive organisms, demonstrating its capacity for attacking both of its potential targets in the bacterium. 251D also was bactericidal for gram-positive organisms and lacked toxicity in vitro. Although we obtained strains of Staphylococcus aureus resistant to the individual parent compounds, spontaneous resistance to 251D was not observed. We obtained 251D resistance in multiple-passage experiments, but resistance developed at a pace comparable to those for the parent compounds. This class of AU-FQ hybrids provides a promising new pharmacophore with an unusual dual mechanism of action and potent activity against antibiotic-sensitive and -resistant gram-positive pathogens.

Daptomycin (DAP) Use in Patients with Hematologic (HEM) Cancer History; Post-Marketing Experience in a Registry.

Background: DAP is a novel lipopeptide antibiotic, approved for complicated skin and skin structure infections, with potent in vitro activity against many resistant Gram-positive organisms. Registration studies often limit or exclude patients with severe illness. Antibiotic resistant Gram-positive pathogens are frequently encountered in compromised patient populations; however, minimal information is available pre-approval in these patients.Methods: Cubicin® Outcomes Registry and Experience (CORE) is a Phase IV retrospective observational chart review (45 institutions) to quantitate characteristics and clinical outcomes of patients (pts) receiving DAP. A standard case report form including demographic, disease state, clinical, and microbiologic data was completed by each investigator. Cure, failure, and improvement were assessed using standard definitions. Nonevaluable pts had insufficient data available to determine clinical outcome. Of the 1160 pts in CORE, 56 were reported with a history of HEM cancer. Eighteen (32%) were nonevaluable; 6 of those expired. This report describes the 38 pts with an assigned outcome.Results: Fifty-five percent of pts were female, 63% were 31 to 65 years of age, and 76% of infections developed in a hospital setting. Common comorbidities included cardiovascular disease (24%), immunosuppression (18%), and transplant (11%). The most common infections were; bacteremia catheter-related (53%), skin and skin structure (34%), and bacteremia noncatheter-related (18%). Seventy-one percent (27/38) of pts were culture positive before DAP with the most common pathogens being Enterococcus sp. (52%, of which 71% were VRE), coagulase-negative staphylococci (41%), and S. aureus (19%, of which 80% were MRSA). A high percentage of pts (92%) received antibiotics (abx) prior to DAP. The most frequent initial dosing regimens were 4mg/kg QD (50%), and 6mg/kg QD (29%). The mean duration of DAP therapy was 13.6 days (range 2–44). Dose (P=0.3) and duration (P=0.58) did not vary by primary pathogen. Twenty-nine (76%) pts received concomitant abx therapy, most commonly with fluoroquinolones (31%), metronidazole (28%) or cephalosporins (24%). The mean time to clinical response was 2.4 days (n=28) and unaffected by primary pathogen (P=0.43) or the influence of concomitant abx therapy (P=0.55). All pts were reported as cured (66%) or improved (34%).Conclusion: The data from the CORE registry demonstrates that DAP shows early clinical response and clinical cure for HEM patients where catheter associated infections and VRE are prevalent. These data provide preliminary data on the use of DAP in this compromised patient population. This information should lead to further prospective studies in cancer patients with DAP.

Ramoplanin: A Lipoglycodepsipeptide Antibiotic

To review the pharmacology, antimicrobial activity, pharmacokinetics, clinical applications, and safety of ramoplanin, a lipoglycodepsipeptide antibiotic. Information was obtained from MEDLINE and BIOSIS databases (1984-August 2004) and Oscient Pharmaceuticals using the key words ramoplanin, A 16686, A 16686A, and MDL 62198. Available English-based articles and abstracts were reviewed, along with information from Oscient Pharmaceuticals. Ramoplanin exerts its bactericidal activity against gram-positive aerobic and anaerobic bacteria by blocking peptidoglycan synthesis via lipid II. In vitro susceptibility reports have demonstrated efficacy against antibiotic-resistant gram-positive pathogens. Cross-resistance has not been documented with vancomyin and other glycopeptides. Clinical trials are investigating ramoplanin's oral administration for treatment of Clostridium difficile-associated diarrhea. Previous clinical trials had evaluated the suppression of colonization of vancomycin-resistant Enterococcus with ramoplanin. Adverse effects are minimal, and drug-drug interactions have not been documented. The completion of clinical trials will determine whether ramoplanin has a promising role as a treatment option for diarrhea due to C. difficile.

Enzymatic deacylation of teicoplanin followed by reductive alkylation: synthesis and antibacterial activity of new glycopeptides.

Novel glycopeptides derived from teicoplanin were prepared and evaluated for activity against antibiotic-resistant gram-positive pathogens. Removal of the fatty acid sidechains of teicoplanin was accomplished by enzymatic deacylation. The resulting deacylated teicoplanin was subjected to reductive alkylation resulting in mono- and di-alkylated compounds at the 2 possible primary amines. Deacylated teicoplanin was less active than teicoplanin against enterococci and staphylococci (MIC > or =32 microg/ml). All mono- and di-alkylated products regained some activity, and some had potent activity against both staphylococci and glycopeptide-resistant enterococci. MICs of the most potent di-alkylated compounds ranged from 0.25 approximately 2 microg/ml against glycopeptide-resistant enterococci.