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Aminoglycosides are a group of antibiotics used since the 1940s to primarily treat a broad spectrum of bacterial infections. The primary resistance mechanism against these antibiotics is enzymatic modification by aminoglycoside-modifying enzymes that are divided into acetyl-transferases, phosphotransferases, and nucleotidyltransferases. To overcome this problem, new semisynthetic aminoglycosides were developed in the 70s. The most widely used semisynthetic aminoglycoside is amikacin, which is refractory to most aminoglycoside modifying enzymes. Amikacin was synthesized by acylation with the l-(−)-γ-amino-α-hydroxybutyryl side chain at the C-1 amino group of the deoxystreptamine moiety of kanamycin A. The main amikacin resistance mechanism found in the clinics is acetylation by the aminoglycoside 6′-N-acetyltransferase type Ib [AAC(6′)-Ib], an enzyme coded for by a gene found in integrons, transposons, plasmids, and chromosomes of Gram-negative bacteria. Numerous efforts are focused on finding strategies to neutralize the action of AAC(6′)-Ib and extend the useful life of amikacin. Small molecules as well as complexes ionophore-Zn+2 or Cu+2 were found to inhibit the acetylation reaction and induced phenotypic conversion to susceptibility in bacteria harboring the aac(6′)-Ib gene. A new semisynthetic aminoglycoside, plazomicin, is in advance stage of development and will contribute to renewed interest in this kind of antibiotics.

Aminoglycosides are a class of antibiotics that bind to ribosomal RNA and exert pleiotropic effects on ribosome function. Amikacin, the semisynthetic derivative of kanamycin, is commonly used for treating severe infections with multidrug-resistant, aerobic Gram-negative bacteria. Amikacin carries the 4-amino-2-hydroxy butyrate (AHB) moiety at the N 1 amino group of the central 2-deoxystreptamine (2-DOS) ring, which may confer amikacin a unique ribosome inhibition profile. Here we use in vitro fast kinetics combined with X-ray crystallography and cryo-EM to dissect the mechanisms of ribosome inhibition by amikacin and the parent compound, kanamycin. Amikacin interferes with tRNA translocation, release factor-mediated peptidyl-tRNA hydrolysis, and ribosome recycling, traits attributed to the additional interactions amikacin makes with the decoding center. The binding site in the large ribosomal subunit proximal to the 3’-end of tRNA in the peptidyl (P) site lays the groundwork for rational design of amikacin derivatives with improved antibacterial properties. Here the authors use fast kinetics, X-ray crystallography, and cryo-EM to uncover the mechanism of ribosome inhibition by amikacin and kanamycin. They find that amikacin binds near the P-site tRNA, offering new strategies to fight antibiotic resistance.

Plazomicin is currently the only next-generation aminoglycoside approved for clinical use that has the potential of evading the effects of widespread enzymatic resistance factors. However, plazomicin is still susceptible to the action of the resistance enzyme AAC(2′)-Ia from Providencia stuartii . As the clinical use of plazomicin begins to increase, the spread of resistance factors will undoubtedly accelerate, rendering this aminoglycoside increasingly obsolete. Understanding resistance to plazomicin is an important step to ensure this aminoglycoside remains a viable treatment option for the foreseeable future. Here, we present three crystal structures of AAC(2′)-Ia from P. stuartii , two in complex with acetylated aminoglycosides tobramycin and netilmicin, and one in complex with a non-substrate aminoglycoside, amikacin. Together, with our previously reported AAC(2′)-Ia-acetylated plazomicin complex, these structures outline AAC(2′)-Ia’s specificity for a wide range of aminoglycosides. Additionally, our survey of AAC(2′)-I homologues highlights the conservation of residues predicted to be involved in aminoglycoside binding, and identifies the presence of plasmid-encoded enzymes in environmental strains that confer resistance to the latest next-generation aminoglycoside. These results forecast the likely spread of plazomicin resistance and highlight the urgency for advancements in next-generation aminoglycoside design.

Plazomicin (formerly ACHN-490) is a next-generation aminoglycoside that was synthetically derived from sisomicin by appending a hydroxy-aminobutyric acid substituent at position 1 and a hydroxyethyl substituent at position 6′. Plazomicin inhibits bacterial protein synthesis and exhibits dose-dependent bactericidal activity. Plazomicin demonstrates activity against both Gram-negative and Gram-positive bacterial pathogens, including isolates harboring any of the clinically relevant aminoglycoside-modifying enzymes. However, like older parenteral aminoglycosides, plazomicin is not active against bacterial isolates expressing ribosomal methyltransferases conferring aminoglycoside resistance. Plazomicin has been reported to demonstrate in vitro synergistic activity when combined with daptomycin or ceftobiprole versus methicillin-resistant Staphylococcus aureus, heteroresistant vancomycin-intermediate S. aureus, vancomycin-intermediate S. aureus, and vancomycin-resistant S. aureus and against Pseudomonas aeruginosa when combined with cefepime, doripenem, imipenem or piperacillin-tazobactam. After intravenous administration of plazomicin to humans at a dose of 15 mg/ kg, the maximum concentraration was 113 μg/ml, the area under the curve0-24 was 239 h·μg/ml, the half-life was 4.0 h and the steady-state volume of distribution was 0.24 L/kg. Results from a Phase II randomized, double-blind study in patients with complicated urinary tract infection and acute pyelonephritis including cases with concurrent bacteremia comparing plazomicin 15 mg/kg intravenously once daily for 5 days with levofloxacin 750 mg intravenously. for 5 days are anticipated in 2012. Human studies to date have not reported nephrotoxicity or ototoxicity, and lack of ototoxicity has been reported in the guinea pig model. Given reported increases in bacterial resistance to current antimicrobial agents and the lack of availability of new agents with novel mechanisms, plazomicin may become a welcomed addition to the antibacterial armamentarium pending positive results from large-scale clinical trials and other required clinical studies.

Background Treatment of patients affected by severe burns is challenging, especially due to the high risk of Pseudomonas infection. In the present work, we have generated a novel model of bioartificial human dermis substitute by tissue engineering to treat infected wounds using fibrin-agarose biomaterials functionalized with nanostructured lipid carriers (NLCs) loaded with two anti- Pseudomonas antibiotics: sodium colistimethate (SCM) and amikacin (AMK). Results Results show that the novel tissue-like substitutes have strong antibacterial effect on Pseudomonas cultures, directly proportional to the NLC concentration. Free DNA quantification, WST-1 and Caspase 7 immunohistochemical assays in the functionalized dermis substitute demonstrated that neither cell viability nor cell proliferation were affected by functionalization in most study groups. Furthermore, immunohistochemistry for PCNA and KI67 and histochemistry for collagen and proteoglycans revealed that cells proliferated and were metabolically active in the functionalized tissue with no differences with controls. When functionalized tissues were biomechanically characterized, we found that NLCs were able to improve some of the major biomechanical properties of these artificial tissues, although this strongly depended on the type and concentration of NLCs. Conclusions These results suggest that functionalization of fibrin-agarose human dermal substitutes with antibiotic-loaded NLCs is able to improve the antibacterial and biomechanical properties of these substitutes with no detectable side effects. This opens the door to future clinical use of functionalized tissues.

Solid lipid nanoparticles (SLNs) of amikacin were designed in this study for pulmonary delivery to reduce the dose or its administration intervals leading to reduction of its toxicities especially in long term treatment. Nanoparticles of amikacin were prepared from cholesterol by solvent diffusion technique and homogenization. The size, zeta potential, loading efficiency, and release profile of the nanoparticles were studied. The conventional broth macrodilution tube method was used to determine the minimum inhibitory concentration (MIC) and minimum bacteriostatic concentration (MBC) of amikacin SLNs with respect to Pseudomonas aeruginosa in vitro. To guarantee the stability of desired SLNs, they were lyophilized using cryoprotectants. Results showed that considering the release profile of amikacin from the studied nanocarrier, MIC and MBC of amikacin could be about two times less in SLNs of amikacin compared to the free drug. Therefore, fewer doses of amikacin in SLNs can clear the infection with less adverse effects and more safety. Particle size enlargement after lyophilization of desired SLNs after two months storage was limited in comparison with non-lyophilized particles, 996 and 194 nm, respectively. Zeta potential of lyophilized particles was increased to +17 mV from +4 mV before lyophilization. Storage of particles in higher temperature caused accelerated drug release.

Mannitol, a polyalcohol bacterial metabolite, has been shown to activate dormant persister cells within bacterial biofilm. This study sought to evaluate an injectable blend of mannitol, chitosan, and polyethylene glycol for delivery of antibiotics and mannitol for eradication of Staphylococcal biofilm. Mannitol blends were injectable and had decreased dissociation and degradation in the enzyme lysozyme compared to blends without mannitol. Vancomycin and amikacin eluted in a burst response, with active concentrations extended to seven days compared to five days for blends without mannitol. Mannitol eluted from the paste in a burst the first day and continued through Day 4. Eluates from the mannitol pastes with and without antibiotics decreased viability of established S. aureus biofilm by up to 95.5% compared to blends without mannitol, which only decreased biofilm when loaded with antibiotics. Cytocompatibility tests indicated no adverse effects on viability of fibroblasts. In vivo evaluation of inflammatory response revealed mannitol blends scored within the 2–4 range at Week 1 (2.6 ± 1.1) and at Week 4 (3.0 ± 0.8), indicative of moderate inflammation and comparable to non-mannitol pastes (p = 0.065). Clinically, this paste could be loaded with clinician-selected antibiotics and used as an adjunctive therapy for musculoskeletal infection prevention and treatment.

The aim of the study was to develop self-emulsifying delivery systems (SEDDS) exhibiting improved permeation rate for pulmonary delivery of amikacin for treatment of cystic fibrosis (CF) patients. Solubility of amikacin in lipids was improved by hydrophobic ion pairing with sodium myristyl sulfate. The complex was loaded into SEDDS. Drug-release studies were performed and the permeation properties of SEDDS through human CF mucus were examined. A total of 10% complex could be loaded into SEDDS. SEDDS exhibited sustained release. Up to twofold more amounts of amikacin permeated through the CF mucus compared with reference. The developed SEDDS with amikacin may be a promising tool for the treatment of certain bacterial infections of CF patients.

Abstract Background: Few studies on performance comparison of nebulizer systems coupled with a single-limb circuit bilevel ventilator are available. Most of these data compared the aerosol drug delivery for only two different systems. Using an adult lung bench model of noninvasive ventilation, we compared inhaled and lost doses of three nebulizer systems coupled with a single-limb circuit bilevel ventilator, as well as the influence of the nebulizer position. Method: Three vibrating mesh nebulizers (Aeroneb® Pro, Aeroneb® Solo, and NIVO®), one jet nebulizer (Sidestream®), and one ultrasonic nebulizer (Servo Ultra Nebulizer 145®) coupled with a bilevel ventilator were tested. They were charged with amikacin solution (500 mg/4 mL) and operated at two different positions: before and after the exhalation port (starting from the lung). The inhaled dose, the expiratory wasted dose, and the estimated lost dose were assessed by the residual gravimetric method. Results: The doses varied widely among the nebulizer types and position. When the nebulizer was positioned before the exhalation port, the vibrating mesh nebulizer delivered the highest inhaled dose (p<0.001), the jet nebulizer the highest expiratory wasted dose (p<0.001), and the ultrasonic device the highest total lost dose (p<0.001). When the nebulizer was positioned after the exhalation port, the vibrating mesh nebulizers delivered the highest inhaled (p<0.001) and expiratory wasted doses (p<0.001), and the ultrasonic device the highest total lost dose (p<0.001). The most efficient nebulizers were NIVO and Aeroneb Solo when placed before the exhalation port. Conclusions: In a single-limb circuit bilevel ventilator, vibrating mesh nebulizers positioned between the exhalation port and lung model are more efficient for drug delivery compared with jet or ultrasonic nebulizers. In this position, the improved efficiency of vibrating mesh nebulizers was due to an increase in the inhaled dose and a reduction in the exhaled wasted dose compared with placement between the ventilator and the expiratory port. Because of the high total lost dose, the ultrasonic device should not be recommended. Nebulizer placement before the exhalation port increased the inhaled dose and decreased the expiratory wasted dose, except for the jet nebulizer.

Amikacin (Amk) analysis and quantitation, for pharmacokinetics studies and other types of investigations, is conventionally performed after extraction from plasma. No report exists so far regarding drug extraction from whole blood (WB). This can represent an issue since quantification in plasma does not account for drug partitioning to the blood cell compartment, significantly underrating the drug fraction reaching the blood circulation. In the present work, the optimization of an extraction method of Amk from murine WB has been described. The extraction yield was measured by RP-HPLC-UV after derivatization with 1-fluoro-2,4-dinitrobenzene, which produced an appreciably stable derivative with a favorable UV/vis absorption. Several extraction conditions were tested: spiked Amk disulfate solution/acetonitrile/WB ratio; presence of organic acids and/or ammonium hydroxide and/or ammonium acetate in the extraction mixture; re-dissolution of the supernatant in water after a drying process under vacuum; treatment of the supernatant with a solution of inorganic salts. The use of 5% (by volume) of ammonium hydroxide in a hydro-organic solution with acetonitrile, allowed the almost quantitative (95%) extraction of the drug from WB.