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Bacterial infections, including those caused by Pseudomonas aeruginosa, often lead to sepsis, necessitating effective antibiotic treatment like carbapenems. The key pharmacokinetic/pharmacodynamic (PK/PD) index correlated to carbapenem efficacy is the fraction time of unbound plasma concentration above the minimum inhibitory concentration (MIC) of the pathogen (%fT > MIC). While multiple targets exist, determining the most effective one for critically ill patients remains a matter of debate. This study evaluated meropenem\'s bactericidal potency and its ability to combat drug resistance in Pseudomonas aeruginosa under three representative PK/PD targets: 40% fT > MIC, 100% fT > MIC, and 100% fT > 4× MIC. The hollow fiber infection model (HFIM) was constructed, validated, and subsequently inoculated with a substantial Pseudomonas aeruginosa load (1 × 108 CFU/mL). Different meropenem regimens were administered to achieve the specified PK/PD targets. At specified intervals, samples were collected from the HFIM system and subjected to centrifugation. The resulting supernatant was utilized to determine drug concentrations, while the precipitates were used to track changes in both total and drug-resistant bacterial populations over time by the spread plate method. The HFIM accurately reproduced meropenem\'s pharmacokinetics in critically ill patients. All three PK/PD target groups exhibited a rapid bactericidal response within 6 h of the initial treatment. However, the 40% fT > MIC and 100% fT > MIC groups subsequently showed bacterial resurgence and resistance, whereas the 100% fT > 4× MIC group displayed sustained bactericidal activity with no evidence of drug resistance. The HFIM system revealed that maintaining 100% fT > 4× MIC offers a desirable microbiological response for critically ill patients, demonstrating strong bactericidal capacity and effective prevention of drug resistance.

The FDA announced a boxed warning for tigecycline due to progression of infections caused by Gram-negative bacteria and increased risk of mortality during treatment. Plasma exposure of tigecycline might not prevent bacteraemia in these cases from the focuses. Hence, we evaluated intensified dosing regimens and breakpoints that might suppress bloodstream infections, caused by progression of infection by e.g., Gram-negatives. A pharmacometric model was built from tigecycline concentrations (100-600 mg daily doses) against clinical Klebsiella pneumoniae isolates (MIC 0.125-0.5 mg/L). Regrowth occurred at clinically used doses and stasis was only achieved with 100 mg q8h for the strain with the lowest studied MIC of 0.125 mg/L. Stasis at 24 h was related to fAUC/MIC of 38.5. Our study indicates that even intensified dosing regimens might prevent bloodstream infections only for MIC values ≤0.125 mg/L for tigecycline. This indicates an overly optimistic breakpoint of 1 mg/L for Enterobacterales, which are deemed to respond to the tigecycline high dose regimen (EUCAST Guidance Document on Tigecycline Dosing 2022).

OBJECTIVE: Mechanistic understanding of pharmacodynamic interactions is key for the development of rational antibiotic combination therapies to increase efficacy and suppress the development of resistances. Potent tools to provide those insights into pharmacodynamic drug interactions are semi-mechanistic modeling and simulation techniques. This study uses those techniques to provide a detailed understanding with regard to the direction and strength of the synergy of ceftazidime-avibactam and ceftazidime-fosfomycin in a clinical Escherichia coli isolate expressing extended spectrum beta-lactamase (CTX-M-15 and TEM-4) and carbapenemase (OXA-244) genes. Enhanced killing effects in combination were identified as a driver of the synergy and were translated from static time-kill experiments into the dynamic hollow fiber infection model. These findings in combination with a suppression of the emergence of resistance in combination emphasize a potential clinical benefit with regard to increased efficacy or to allow for dose reductions with maintained effect sizes to avoid toxicity.

Bacteriophage (phage) therapy is being explored as a possible response to the antimicrobial resistance public health emergency. Administering a mixture of different phage types as a cocktail is one proposed strategy for therapeutic applications, but the optimal method for formulating phage cocktails remains a major challenge. Each phage strain has complex pharmacokinetic/pharmacodynamic (PK/PD) properties which depend on the nano-scale size, target-mediated, self-dosing nature of each phage strain, and rapid selection of resistant subpopulations. The objective of this study was to explore the pharmacodynamics (PD) of three unique and clinically relevant anti-Pseudomonas phages after simulation of dynamic dosing strategies. The Hollow Fiber Infection Model (HFIM) is an in vitro system that mimics in vivo pharmacokinetics (PK) with high fidelity, providing an opportunity to quantify phage and bacteria concentration profiles over clinical time scales with rich sampling. Exogenous monotherapy-bolus (producing max concentrations of Cmax = 7 log10 PFU/mL) regimens of phages LUZ19, PYO2, and E215 produced Pseudomonas aeruginosa nadirs of 0, 2.14, or 2.99 log10 CFU/mL after 6 h of treatment, respectively. Exogenous combination therapy bolus regimens (LUZ19 + PYO2 or LUZ19 + E215) resulted in bacterial reduction to <2 log10 CFU/mL. In contrast, monotherapy as a continuous infusion (producing a steady-state concentration of Css,avg = 2 log10PFU/mL) was less effective at reducing bacterial densities. Specifically, PYO2 failed to reduce bacterial density. Next, a mechanism-based mathematical model was developed to describe phage pharmacodynamics, phage-phage competition, and phage-dependent adaptive phage resistance. Monte Carlo simulations supported bolus dose regimens, predicting lower bacterial counts with bolus dosing as compared to prolonged phage infusions. Together, in vitro and in silico evaluation of the time course of phage pharmacodynamics will better guide optimal patterns of administration of individual phages as a cocktail.

Spectinamides are a novel series of spectinomycin analogs being developed for the treatment of tuberculosis. The preclinical lead spectinamide 1599 is an antituberculosis drug that possesses robust in vivo efficacy, good pharmacokinetic properties, and excellent safety profiles in rodents. In individuals infected with Mycobacterium tuberculosis or Mycobacterium bovis, causative agents of tuberculosis, the host immune system is capable of restraining these mycobacteria within granulomatous lesions. The harsh microenvironmental conditions of these granuloma lead to phenotypic transformation of mycobacteria. Phenotypically transformed bacteria display suboptimal growth, or complete growth arrest and are frequently associated with drug tolerance. Here we quantified the effect of spectinamide 1599 on log-phase and phenotypically tolerant isoforms of Mycobacterium bovis BCG using various in vitro approaches as a first indicator of spectinamide 1599 activity against various mycobacterial isoforms. We also used the hollow fiber infection model to establish time-kill curves and deployed pharmacokinetic/pharmacodynamic modeling to characterize the activity differences of spectinamide 1599 towards the different phenotypic subpopulations. Our results indicate that spectinamide 1599 is more efficacious against log phase bacteria when compared to its activity against other phenotypically tolerant forms such as acid phase bacteria and hypoxic phase bacteria, a behavior similar to the established antituberculosis drug isoniazid.

OBJECTIVE: Employ the hollow fiber infection model (HFIM) to study sequential antibiotic administration (ampicillin, ciprofloxacin and fosfomycin) using human pharmacokinetic profiles to measure changes in the rate of antibiotic resistance development and compare this to simultaneous combination therapy with the same antibiotic combinations.
METHODS: Escherichia coli CFT073, a clinical uropathogenic strain, was exposed individually to clinically relevant pharmacokinetic concentrations of ampicillin on day 1, ciprofloxacin on day 2 and fosfomycin on day 3. This sequence was continued for 10 days in the HFIM. Bacterial samples were collected at different time points to enumerate total and resistant bacterial populations. The results were compared with the simultaneous combination therapy previously studied.
RESULTS: Sequential antibiotic treatment (ampicillin-ciprofloxacin-fosfomycin sequence) resulted in the early emergence of single and multi-antibiotic-resistant subpopulations, while the simultaneous treatment regimen significantly delayed or prevented the emergence of resistant subpopulations.
CONCLUSIONS: Sequential administration of these antibiotic monotherapies did not significantly delay the emergence of resistant subpopulations compared to simultaneous treatment with combinations of the same antibiotics. Further studies are warranted to evaluate different sequences of the same antibiotics in delaying emergent resistance.

The emergence and spread of antimicrobial resistance in Neisseria gonorrhoeae is seriously threatening the treatment and control of gonorrhea globally. Novel treatment options are essential, coupled with appropriate methods to pharmacodynamically examine the efficacy and resistance emergence of these novel drugs. Herein, we used our dynamic in vitro hollow fiber infection model (HFIM) to evaluate protein-unbound lefamulin, a semisynthetic pleuromutilin, against N. gonorrhoeae. Dose-range and dose-fractionation experiments with N. gonorrhoeae reference strains: WHO F (susceptible to all relevant antimicrobials), WHO X (extensively drug-resistant, including ceftriaxone resistance), and WHO V (high-level azithromycin resistant, and highest gonococcal MIC of lefamulin (2 mg/l) reported), were performed to examine lefamulin gonococcal killing and resistance development during treatment. The dose-range experiments, simulating a single oral dose of lefamulin based on human plasma concentrations, indicated that ≥1.2 g, ≥2.8 g, and ≥9.6 g of lefamulin were required to eradicate WHO F, X, and V, respectively. Dose-fractionation experiments, based on human lefamulin plasma concentrations, showed that WHO X was eradicated with ≥2.8 g per day when administered as q12 h (1.4 g twice a day) and with ≥3.6 g per day when administered as q8 h (1.2 g thrice a day), both for 7 days. However, when simulating the treatment with 5-10 times higher concentrations of free lefamulin in relevant gonorrhea tissues (based on urogenital tissues in a rat model), 600 mg every 12 h for 5 days (approved oral treatment for community-acquired bacterial pneumonia) eradicated all strains, and no lefamulin resistance emerged in the successful treatment arms. In many arms failing single or multiple dose treatments for WHO X, lefamulin-resistant mutants (MIC = 2 mg/l), containing an A132V amino acid substitution in ribosomal protein L3, were selected. Nevertheless, these lefamulin-resistant mutants demonstrated an impaired biofitness. In conclusion, a clinical study is warranted to elucidate the clinical potential of lefamulin as a treatment option for uncomplicated gonorrhea (as well as several other bacterial STIs).

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease 2019 (COVID-19), has radically altered daily life. Effective antiviral therapies to combat COVID-19, especially severe disease, remain scarce. Molnupiravir is an antiviral that has shown clinical efficacy against mild-to-moderate COVID-19 but failed to provide benefit to hospitalized patients with severe disease. Here, we explained the mechanism behind the failure of molnupiravir in hospitalized patients and identified alternative dosing strategies that would improve therapeutic outcomes in all patients with COVID-19. We showed that delaying therapy initiation markedly decreased the antiviral effect of molnupiravir, and these results were directly related to intracellular drug triphosphate pools and intracellular viral burden at the start of therapy. The adverse influence of therapeutic delay could be overcome by increasing drug exposure, which increased intracellular molnupiravir triphosphate concentrations that inhibited viral replication. These findings illustrated that molnupiravir must be administered as early as possible following COVID-19 symptom onset to maximize therapeutic efficacy. Higher doses may be effective in patients hospitalized with severe disease, but the safety of high-dose molnupiravir regimens is unknown. Our findings could be extended to design effective regimens with nucleoside analogs for other RNA viruses, especially those with pandemic potential. IMPORTANCE In this study, we showed that early intervention with molnupiravir resulted in a greater antiviral effect, and we explained the mechanism behind this phenomenon. Our results predicted and explained the failure of molnupiravir in hospitalized patients and highlighted the utility of preclinical pharmacodynamic studies to design optimal antiviral regimens for the treatment of viral diseases. This contrasts with the procedure that was implemented early in the pandemic in which clinical studies were conducted in the absence of preclinical experimentation. These findings are significant and demonstrated the importance of experimental approaches in antiviral development for treatments against COVID-19 as well as other viral diseases.

Bacteria developing resistance compromise the efficacy of antibiotics or bacteriophages (phages). We tested the association of these two antibacterials to circumvent resistance. With the Hollow Fiber Infection Model (HFIM), we mimicked the concentration profile of ciprofloxacin in the lungs of patients treated orally for Pseudomonas aeruginosa infections and, independently, mimicked a single inhaled administration of phages (one or two phages). Each treatment selects for antibiotic- or phage-resistant clones in less than 30 h. In contrast, no bacteria were recovered from the HFIM at 72 h when ciprofloxacin was started 4 h post phage administration, even when increasing the initial bacterial concentration by 1,000-fold. The combination of phages with antibiotics used according to clinical regimens prevents the growth of resistant clones, providing opportunities to downscale the use of multiple antibiotics. IMPORTANCE In the treatment of bacterial infections, the use of antibiotics or bacteriophages (phages) is limited by the ability of bacteria to develop resistance. The resistance frequency depends on the exposure to antibacterials. Therefore, determination of concentration profiles of antibiotics is key to define optimal regimens during treatments. In the laboratory, the Hollow Fiber Infection Model (HFIM) mimics concentration profiles observed in patients. In this study, we used the HFIM to evaluate the killing efficacy of the combination of phages and ciprofloxacin. We demonstrated that dosing schedule of phages first and the antibiotic second prevent the selection of resistant bacteria. These results demonstrate that combination efficacy relies on a strong initial reduction of the bacterial population by phages followed by antibiotics before any resistant arise.