Extensive assays and instrumentation to assess antibiotic mode of action

CO-ADD has assembled a panel of assays to investigate the mode of action of known and novel antibiotics, including:

• Membrane depolarization
• Critical Micelle Concentration
• Membrane disruption by dye-based assays or electron microscopy
• Membrane interactions by PAMPA, NMR and SPR
• Peptidoglycan synthesis with 14C-labelled precursor incorporation
• Cell morphology by microscopy, including CFM, TEM, SEM
• Thermodynamics of target binding by ITC

Expertise in cell culture, transfection and protein expression, isolation and purification and assay development

CO-ADD helps researchers to identify and validate new antimicrobial targets, via programs that assist in the confirmation of target relevance to the virulence or survival of bacterial pathogens and the examination of selectivity/toxicity profiles.

CO-ADD has the ability to clone and express new targets and develop assays to assess inhibition or activation of the target, while bioinformatics support is available in the form of sequence searching and homology comparison/modelling from bacteria and human databases.

Synthetic and medicinal chemistry expertise

The CO-ADD team has collective medicinal chemistry expertise in small molecule, peptide, carbohydrate, and natural product chemistry for rational hit-to-lead optimisation based on quantitative structure-activity and structure-toxicity relationships, molecular modelling and ligand-target crystal and solution state structures, supported by predictive modelling via chemoinformatics and bioinformatics capabilities.

CO-ADD maintains comprehensive databases of drugs, bioactivities and available compounds with specific focus for microbial and antimicrobial research. We collect bacterial strain information, activity and ADMET data from public databases as well as collating in house data. The databases assist not only the recording of all screening data, but help in developing predictive models, designing novel compounds and optimising hits to leads.

ADMET

Comprehensive capabilities to assess absorption, distribution, metabolism, excretion (ADMET) and toxicity of new antibiotics The development potential of new antimicrobial agents begins with measuring toxicity, haemolysis, gastric, plasma and microsomal stability, and protein binding. These assays can be conducted within CO-ADD or through established collaborations. Additional assays include:

• Cytochrome P450 inhibition and induction
• hERG ion channel inhibition
• Metabolite identification
• Off-target panel testing
• Ames/micronucleus mutagenicity
• Pharmacokinetic (PK) studies have been developed using serial sampling in mice, which minimises both compound and animal use

Efficacy

In vivo efficacy mouse models have been established including the classical thigh infection model with colony forming unit (CFU) readout, and intraperitoneal and thigh infection models using bioluminescent bacteria with scanned whole-body bioluminescence detection. These models include the potential to assess compound exposure within the infected animals, allowing for direct correlation of infection outcome in individual animals with actual compound exposure. Additional models (blood septicaemia or lung infection) can be accessed through collaborating sites.

CO-ADD has access to rapid sequencing technology and bioinformatics pipelines (Illumina HiSeq, MiSeq) for full genome sequencing of clinically-resistant bacterial strains or strains derived from forced induction of resistance. We also have access to data from longitudinal collections of clinical isolates from hospital-acquired infections.