Call: Antimicrobial Resistance through the JPIAMR

Control of bacterial infections is threatened by the rapid emergence of drug resistance, drug tolerance that mitigates antimicrobial efficacy, and the lack of new antibiotics in recent years. Combination treatments and/or re-purposing of known drugs can provide a cost- and time-efficient solution.

Completed project

We propose a comprehensive and powerful strategy to repurpose or improve FDA-approved drugs including neglected/disused (ND) antibiotics, and identify combinations to re-sensitize resistant/tolerant bacteria. Using high-throughput screening (HTS), we will test thousands of pairwise drug combinations on bacterial growth, viability and bacterial persistence in three clinically relevant pathogens: uropathogenic Escherichia coli (UPEC), Staphylococcus aureus (Sa), and Streptococcus pneumoniae (Sp). Our scope thus spans the Gram-negative/-positive divide, and targets several bacterial states implicated in both acute and chronic infections, including pneumonia, pyelonephritis, bacteremia and endocarditis.

To optimize antimicrobial treatment, we will use state-of-the-art and further advance translational pharmacokinetic (PK)/ pharmacodynamic (PD) modeling. This will prioritize and validate lead combinations, which will be tested in several toxicity and animal models. In vivo PK/PD modeling will be also performed for our top leading combinations. Together, these approaches will provide invaluable pre-clinical data that will inform future drug development. In parallel, we will systematically probe for modes of action of our top 100 synergistic drug combinations using HT reverse genetic screens to provide the framework for subsequent in-depth mechanistic studies targeting processes such as the role of redox and iron homeostasis, membrane permeability/stability, drug efflux/influx and bacterial metabolism and stress responses in antimicrobial activity.

We have assembled a strong team with extensive expertise in the cellular processes directly related to antimicrobial activity. For this project, we will combine HT chemical and genetic screening, large-scale data analysis, mechanistic biology, PK/PD modelling and infection models to discover and exploit antimicrobial combination therapy. The breadth and scope of our ambitious strategy will undoubtedly significantly advance the quest for novel treatment strategies against bacterial infections.

Project partners

• Typas Athanasios, European Molecular Biology Laboratory, Germany (Coordinator)
• Jan-Willem Veening, University of Groningen, Netherlands
• Frédéric Barras, CNRS, Aix‐Marseille Université, France
• Birgitta Henriques Normark, Karolinska Institutet, Sweden
• Charlotte Kloft, Freie Universität Berlin, Germany
• Bernt Eric Uhlin, Umeå University, Sweden

Publications

• Antibiotics, 2020. Meropenem Plasma and Interstitial Soft Tissue Concentrations in Obese and Nonobese Patients—A Controlled Clinical Trial
• bioRxiv, 2020. Dissecting the collateral damage of antibiotics on gut microbes
• Cell Host & Microbe, 2020. Proton Motive Force Disruptors Block Bacterial Competence and Horizontal Gene Transfer
• Clinical Pharmacology & Therapeutics, 2020. Perspectives on Model‐Informed Precision Dosing in the Digital Health Era: Challenges, Opportunities, and Recommendations
• PLOS Genetics, 2020. Oxidative stress antagonizes fluoroquinolone drug sensitivity via the SoxR-SUF Fe-S cluster homeostatic axis
• Genes, 2019. Three new integration vectors and fluorescent proteins for use in the opportunistic human pathogen Streptococcus pneumoniae
• Nature Microbiology, 2019. RocS drives chromosome segregation and nucleoid protection in Streptococcus pneumoniae
• bioRxiv, 2018. RocS drives chromosome segregation and nucleoid occlusion in Streptococcus pneumoniae
• Cell Reports, 2018. Antibiotic-induced cell chaining triggers pneumococcal competence by reshaping quorum sensing to autocrine signaling
• Nucleic Acid Research, 2018. Deep genome annotation of the opportunistic human pathogen Streptococcus pneumoniae D39
• Nature, 2018. Species-specific activity of antibacterial drug combinations
• Nucleic Acids Research, 2018. High-resolution analysis of the pneumococcal transcriptome under a wide range of infection-relevant conditions
• CPT: Pharmacometrics & Systemics Pharmacology, 2017. Translational Pharmacometric Evaluation of Typical Antibiotic Broad‐Spectrum Combination Therapies Against Staphylococcus Aureus Exploiting In Vitro Information
• Molecular Microbiology, 2017. Silver potentiates aminoglycoside toxicity by enhancing their uptake

Call

• Antimicrobial Resistance through the JPIAMR

The CO-ACTION project aims to develop and provide a framework for evaluating and validating the effectiveness of antibiotic- and non-antibiotic combinations (COMs) in the preclinical setting based on pharmacokinetic/pharmacodynamic (PK/PD) principles, with a specific emphasis on Neglected and Disused AntiBiotics (ND-AB) as well as COMs with non-antibiotics (NA) both for human and veterinary medicine.

Completed project

To fulfill this ambitious goal, 6 work-packages with 6 interacting partners were developed involving several steps in the development of useful COMs and are executed partly sequential, partly in parallel: screening for CO-ACTION between ND-AB and NA in a collection of strains with well described resistance mechanisms, selecting potential synergistic COMs, subsequent validation using PK/PD experiments and modelling and finally testing COMs in animal models. A full PK/PD work-up and analyses form an important part of the process. The interaction between ND-AB from at least 6 different classes, including Polymyxin B will be determined using checkerboard experiments in 10 well characterized Gram-negative (e.g. P. aeruginosa, K. pneumoniae) multidrug resistant strains and analysed by surface response modelling. In parallel, a high throughput system (the oCelloscope) will be applied to allow efficient screening for large numbers of COMs.

The CO-ACTION of clearly synergistic COMs will be quantified using kill-curves both in medium as well as intracellulary and PK/PD modelling will be used to predict effective dosing regimens vivo. Effectiveness of the most promising 3-6 COMs will be determined in up to 4 different available animal model systems : a neutropenic mouse thigh and lung model, a rat model and effectiveness in a pig model to evaluate emergence of resistance in the gut of different COMs. A full PK/PD evaluation, including in vivo checkerboards will be performed and assessment of concentrations in ELF and microdialysis.

The potential of the COMs in patients will be evaluated by Monte Carlo Simulations of the COMs both using plasma as well as ELF concentrations and derived PK/PD relationships and PD targets. The development of useful COMs requires a high level of interaction between specialized partners. This international collaboration of six partners in CO-ACTION will lead to synergistic antibiotic COMs useful in patient care by bringing together specialists that each have significant expertise in their own field.

Project partners

• Joseph Meletiadis, Erasmus MC University Medical Center, Netherlands (Coordinator)
• Lena Friberg, Uppsala University, Sweden
• Francoise Van Bambeke, Université Catholique de Louvain, Belgium
• Thomas Tängdén, Uppsala University, Sweden
• William Couet, Université de Poitiers, France
• Alain Bousquet-Melou, National Veterinary School/INRA, France

Popular summary

Increasingly, patients with infections are difficult to treat because the bacteria that cause the infection have become resistant to antibiotics. There is therefore an increasing demand for new antimicrobials to overcome resistance. However, there are few new antibiotics in development. An alternative strategy to overcome resistance that has been insufficiently explored is a more efficient use of existing antibiotics (AB) by combining them, in particular Neglected and Disused AB (ND-AB). Also, the combination of antibiotics and other drugs -non-antibiotics (NA) might be efficacious but has not been well explored.

CO-ACTION project identified several AB+ND-AB combinations with increased activity against multidrug resistant gram negative bacteria Acinetobacter baumannii, Klebsiella pneuoniae and Pseudomonas aeruginosa. Polymyxin B with rifampicin and minocycline was the most promising combinations in in vitro extracellular and intracellular studies verified in several animal models (murine thigh and lung infection models, pig model) where pharmacokinetics, pharmacodynamics and resistance development was studied. Furthermore, in search of effective ND-AB+NA combinations, synergism was found between polymyxin B and some NA with spironolactone identified as a potential candidate for further studies. In addition, a high throughput model for screening multiple ND-AB+AB combinations against large set of isolates was developed whereas a method for testing AB+NA combinations was assessed. Finally, PK-PD models were developed by associating drug concentrations with antimicrobial activity and clinical doses for the combined drugs were proposed.

Publications

• International Journal of Antimicrobial Agents, 2020. Pharmacodynamics of immune response biomarkers of interest for evSaluation of treatment effects in bacterial infections
• Clinical Microbiology and Infection, 2020. Population pharmacokinetics of colistin and the relation to survival in critically ill patients infected with colistin susceptible and carbapenem-resistant bacteria
• Clinical Microbiology and Infection, 2020. Semi-mechanistic PK/PD modelling of combined polymyxin B and minocycline against a polymyxin-resistant strain of Acinetobacter baumannii
• International Journal of Antimicrobial Agents, 2020. Combination of polymyxin B and minocycline against multidrug-resistant Klebsiella pneumoniae: interaction quantified by pharmacokinetic/pharmacodynamic modelling from in vitro data
• Clinical Microbiology and Infection, 2020. Evaluation of polymyxin B in combination with 13 other antibiotics against carbapenemase-producing Klebsiella pneumoniae in time-lapse microscopy and time-kill experiments
• Antimicrobial Agents and Chemotherapy, 2020. Efficacy of Antibiotic Combinations against MultidrugResistant Pseudomonas aeruginosa in Automated Time-Lapse
• Molecular Pharmaceutics, 2020, Model-Informed Drug Development in Pulmonary Delivery: Semimechanistic Pharmacokinetic–Pharmacodynamic Modeling for Evaluation of Treatments against Chronic Pseudomonas aeruginosa Lung Infections
• Antimicrobial Agents and Chemotherapy, 2020, Extension of Pharmacokinetic/Pharmacodynamic Time-Kill Studies To Include Lipopolysaccharide/Endotoxin Release from Escherichia coli Exposed to Cefuroxime
• Journal of Antimicrobial Chemotherapy, 2019. Population pharmacokinetics of piperacillin in febrile children receiving cancer chemotherapy: the impact of body weight and target on an optimal dosing regimen
• International Journal of Antimicrobial Agents, 2019. An alternative strategy for combination therapy: Interactions between polymyxin B and non-antibiotics
• Clinical Microbiology and Infection, 2018. PK/PD characterization of combined antimicrobial agents: a real challenge and an urgent need
• Clinical Microbiology and Infection, 2018. Semi-mechanistic pharmacokinetic–pharmacodynamic modelling of antibiotic drug combinations
• Clinical Microbiology and Infection, 2018. Automated time-lapse microscopy a novel method for screening of antibiotic combination effects against multidrug-resistant Gram-negative bacteria

Call

• Antimicrobial Resistance through the JPIAMR

Bacterial respiratory infections represent a real threat to public health worldwide, especially in hospital-acquired situations where patients frequently present several co-morbidity factors. Although antibiotics are recognized as the most effective therapy, treatments are often associated with failure due to bacteria that are resistant to multiple firstline antibiotics, and patients who are immune compromised.

Completed project

The proposal ABIMMUNE aims to enhance the therapeutic arsenal against respiratory infections. The idea is to combine Neglected and Disused AntiBiotics (ND-AB) with registered immune modulators that impact host innate immunity. ABIMMUNE will select (i) ND-AB that are not used as first-line antibiotics in standalone treatment of respiratory infections, and (ii) marketed immunostimulatory drugs that target distinct immune pathways including macrophage activation, neutrophil potentiation, immuno-metabolism, or pattern-recognition receptors.

There are several advantages to this approach: first, antibacterial activity of innate immunity is independent of antibiotic-resistance. Second, it is difficult for the pathogen to develop resistance to innate immunity since this latter involves multiple killer cells and antibacterial molecules, and bacteria would have to develop an entirely new suite of interactions with the host. Third, targeting host innate immunity may reinstate some immune defense in vulnerable patients. Fourth, innate immunity and ND-AB may synergize to kill bacteria thereby allowing for dose reduction of ND-AB and potentially reducing side effects. Fifth, using ND-AB may globally dampen the proportion of bacteria resistant to first-line antibiotics, allowing their maintenance in clinics. These combination therapies will be tested in clinically relevant models of respiratory infections with the leading Grampositive and Gram-negative bacteria causing community- and hospital-acquired pneumonia.

ABIMMUNE will validate in vitro the proof-of-concept of additive or synergistic activity of ND-AB and immunostimulatory drugs with appropriate host target cells and collection of bacterial isolates. The impact on emergence and maintenance of resistance to first-line antibiotics and ND-AB will also be evaluated. Experiments will be accompanied by translational PK/PD modeling analyses to quantify the joint ND-AB/immune modulators interaction for selection of the most promising regimens for pneumonia in the context of immune vulnerability.

Project partners

• Jean-Claud Sirard, INSERM-CIIL, France (Coordinator)
• Martin Rumbo, National Universtity of La Plata, Argentina
• Tom Van der Poll, Academic Medical Center, University of Amsterdam, Netherlands
• Mustapha Si-Tahar, INSERM-CEPR, France
• Charlotte Kloft, Freie Universität Berlin, Germany

Popular summary

Emergence of bacterial resistance has become a major threat in healthcare, in particular in lower respiratory tract infections. To address this important challenge, novel antibacterial approaches are needed. One strategy is the use of immunomodulatory drugs in combination with antibiotics, which might contribute to make better use of the current antibiotics to ultimately minimising both treatment failure and emergence of antibiotic resistance in pathogens.
The beta-lactam amoxicillin, a WHO-listed essential antibiotic, is a common treatment option for such infections and therefore, it is essential to sustain its effectiveness. Here, we aimed to investigate the combination of amoxicillin with monophosphoryl lipid A as an immunostimulatory molecule exploiting which antibiotic concentrations are present in the body and which effects resulted by these concentrations; so called pharmacokinetic (PK) and pharmacodynamic (PD) knowledge. We also addressed the effect of flagellin, pioglitazone, and rapamycin as immunomodulatory drugs to be combined with antibiotics.

Project resources

Coordinator Miguel Camara’s website

Publications

• Mucosal Immunology, 2021. mTOR-driven glycolysis governs induction of innate immune responses by bronchial epithelial cells exposed to the bacterial component flagellin
• bioRxiv, 2020. Boosting Toll-like receptor 4 signaling enhances the therapeutic outcome of antibiotic therapy in pneumococcal pneumonia
• Journal of Clinical Medicine, 2020. Linezolid Concentrations in Plasma and Subcutaneous Tissue are Reduced in Obese Patients, Resulting in a Higher Risk of Underdosing in Critically Ill Patients: A Controlled Clinical Pharmacokinetic Study
• European Journal of Pharmaceutical Sciences, 2019. High voriconazole target-site exposure after approved sequence dosing due to nonlinear pharmacokinetics assessed by long-term microdialysis
• Frontiers in Immunology, 2019. Therapeutic Synergy Between Antibiotics and Pulmonary Toll-Like Receptor 5 Stimulation in Antibiotic-Sensitive or -Resistant Pneumonia
• Drug Metabolism Reviews, 2019. Novel insights into the complex pharmacokinetics of voriconazole: a review of its metabolism
• Talanta, 2019. A rapid, simple and sensitive liquid chromatography tandem mass spectrometry assay to determine amoxicillin concentrations in biological matrix of little volume

Call

• Antimicrobial Resistance through the JPIAMR