A novel therapeutic antibody screening method using bacterial high-content imaging reveals functional antibody binding phenotypes of Escherichia coli ST131

Mailis Maes,Josefin Bartholdson Scott,Zoe A Dyson,Stephen Baker,Sarah E Smith,David A Goulding,Stephen T Reece,Catherine Ludden,Gordon Dougan,Paul Kellam

doi:10.1038/s41598-020-69300-8

Mailis Maes, Josefin Bartholdson Scott + Show 8 more

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https://doi.org/10.1038/s41598-020-69300-8

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The increase of antimicrobial resistance (AMR), and lack of new classes of licensed antimicrobials, have made alternative treatment options for AMR pathogens increasingly attractive. Recent studies have demonstrated anti-bacterial efficacy of a humanised monoclonal antibody (mAb) targeting the O25b O-antigen of Escherichia coli ST131. To evaluate the phenotypic effects of antibody binding to diverse clinical E. coli ST131 O25b bacterial isolates in high-throughput, we designed a novel mAb screening method using high-content imaging (HCI) and image-based morphological profiling to screen a mAb targeting the O25b O-antigen. Screening the antibody against a panel of 86 clinical E. coli ST131 O25:H4 isolates revealed 4 binding phenotypes: no binding (18.60%), weak binding (4.65%), strong binding (69.77%) and strong agglutinating binding (6.98%). Impaired antibody binding could be explained by the presence of insertion sequences or mutations in O-antigen or lipopolysaccharide core biosynthesis genes, affecting the amount, structure or chain length of the O-antigen. The agglutinating binding phenotype was linked with lower O-antigen density, enhanced antibody-mediated phagocytosis and increased serum susceptibly. This study highlights the need to screen candidate mAbs against large panels of clinically relevant isolates, and that HCI can be used to evaluate mAb binding affinity and potential functional efficacy against AMR bacteria.

Highlights

Antimicrobial resistance (AMR) is one of the greatest current challenges in global h ealth[1]
We designed a novel monoclonal antibody (mAb) screening method using High-content imaging (HCI) and image-based morphological profiling to measure the antimicrobial potential of a variant of 3E9-11, which targets the O-antigen of E. coli ST131 O25b:H4
To evaluate HCI as a method for screening candidate mAbs against large panels of clinical isolates, whilst simultaneously determining the diagnostic and functional potentials of the antibody, we synthesised KM467, an IgG1 antibody based on the VH and VL sequences of 3E9-11, which targets the O25b O-antigen of E. coli ST131

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Introduction

Antimicrobial resistance (AMR) is one of the greatest current challenges in global h ealth[1]. For an O-antigen antibody to be of clinical utility it is important to demonstrate that these anti-bacterial activities function against a diverse collection of E. coli ST131 O25b associated with disease in healthcare settings. High-content imaging (HCI) is a powerful phenotypic screening approach that combines high-throughput automated microscopy with comprehensive image analysis to quantify multiple morphological and functional cellular features. This type of image-based morphological profiling can be used for high-throughput screening of drugs, simultaneously evaluating potency as well as mode-of-action[17,18]. We designed a novel mAb screening method using HCI and image-based morphological profiling to measure the antimicrobial potential of a variant of 3E9-11, which targets the O-antigen of E. coli ST131 O25b:H4. Our analysis revealed distinct mAb binding phenotypes within the E. coli ST131 O25b:H4 population that were directly associated with mAb function

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