Khamas, A. B. | bioRxiv Science

bioRxiv2024

Mitomycin C Retains Efficacy after Adaptive Laboratory Evolution of Staphylococcus aureus

Antibiotic resistance is one of the greatest threats against human health and the misuse and overuse of antibiotics is a key factor driving resistance development. During prolonged antibiotic treatment of chronic infections, the antimicrobial pressure facilitates selection of antibiotic resistance mutations. It has been suggested that using antibiotics in combinations may reduce the emergence of resistance. Furthermore, antibiotic tolerant persister cells may be a reservoir for resistance development, so targeting persister cells with anti-persister drugs could also reduce the emergence of resistance. In this study, we conducted a 42-day adaptive laboratory evolution experiment using Staphylococcus aureus exposed to common antibiotics and the anti-persister drug mitomycin C, either alone or in combination. We monitored susceptibility daily and assessed phenotypic changes in growth and biofilm formation in evolved strains. Whole-genome sequencing revealed mutations linked to antibiotic resistance and phenotypic shifts. Resistance developed rapidly against rifampicin, while ciprofloxacin and daptomycin showed slower resistance emergence. Treatments with vancomycin or mitomycin C resulted in minimal changes in susceptibility. Combination therapies generally delayed resistance, though resistance was not fully prevented. Notably, mitomycin C combined with rifampicin effectively suppressed rifampicin resistance. Sub-inhibitory antibiotic concentrations were associated with both known and novel mutations, including in the nucleotide excision repair system and azoreductase, following mitomycin C treatment--mutations not previously reported. While combination therapy delayed resistance, mitomycin Cs efficacy and ability to prevent rifampicin resistance highlights its potential in combating antibiotic resistance. Further investigation is needed to evaluate the broader application of anti-persister drugs in resistance prevention.

microbiology↗

bioRxiv2023

Antibody-drug conjugates to treat bacterial biofilms

Implant-associated infections remain a grand unmet medical need because they involve biofilms that protect bacteria from the immune system and harbour antibiotic-tolerant persister cells. There is an urgent need for new biofilm-targeting therapies with antimicrobials, to treat these infections via a non-surgical way. In this work, we address this urgent medical need and engineer antibody-drug conjugates (ADC) that kill bacteria in suspension and in biofilms, in vitro and in vivo. The ADC contains an anti-neoplastic drug mitomycin C, which is also a potent antimicrobial against biofilms. While most ADCs are clinically validated as anti-cancer therapeutics where the drug is released after internalisation of the ADC in the target cell, the ADCs designed herein release the conjugated drug without cell entry. This is achieved with a novel mechanism of drug, which likely involves an interaction of ADC with thiols on the bacterial cell surface. ADC targeted towards bacteria were superior by the afforded antimicrobial effects compared to the non-specific counterpart, in suspension and within biofilms, in vitro and in vivo. An implant-associated murine osteomyelitis model was then used to demonstrate the ability of the antibody to reach the infection, and the superior antimicrobial efficacy compared to standard antibiotic treatment in vivo. Our results illustrate the development of ADCs into a new area of application with a significant translational potential.

microbiology↗