Multiple microbiological studies will need to be carried out to characterize the activity of a drug on target organisms. In reality, it is likely that many of these studies will be carried out in advance of phase 1 studies, as they are necessary to provide confidence that the drug is likely to be efficacious.
Studies are likely to include a combination of in vitro and pre-clinical animal model studies. Key areas of focus include:
- Identification of molecular target and corresponding gene.
- Kinetics of interactions with molecular target.
- Identification of which species possess target gene.
- Establishing the frequency at which resistance arises.
- Identifying the genes involved in resistance.
- Exploring cross-reactivity of resistance to existing antibiotics and classes of antibiotic.
- Investigating whether a drug affects the efficacy of an antibiotic with which it is likely to be co-dosed, using:
- Checkerboard studies (where the growth of bacteria is assessed in the presence of multiple different antibiotic concentrations in combination) or
- Synergy time-kill studies (comparing the effects of an antibiotic on bacterial survival over time in the presence of absence of a second drug).
Susceptibility of different isolates
- Determining the susceptibility (minimum inhibitory concentration, MIC) for 50–100 isolates of each genus and species responsible for the indication being targeted.
- Determining the susceptibility of isolates with known resistance mechanisms to existing antibacterial classes.
Effects of different body fluids
- Assessing activity in the presence of body fluids relevant to the indication being targeted (e.g. urine, whole blood and serum, pulmonary surfactant).
It will also be important to establish methods for in vitro susceptibility testing. Key considerations include:
- Verifying that standard CLSI/EUCAST methodologies can be adopted or whether special culture conditionsare required.
- For drugs forming part of combination products (e.g. beta-lactams/beta-lactamase inhibitors), determining what ratios or drug concentrations to use for each component.
- Establishing appropriate parameters for quality control, following the CLSI M23 protocol.
Disk diffusion testing
Procedures for disk testing – placing antibiotic disks on agar plates of culture pathogens to assess antibacterial activity – must be established before a phase 2 trial if the method is to be used during that trial. Key issues include:
- Determining the mass of drug to be used.
- Obtaining commercial disks from two manufacturers.
Establishing appropriate parameters for quality control, following the CLSI M23 protocol.