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[Preprint]. 2024 Dec 11:2024.12.10.627638.
doi: 10.1101/2024.12.10.627638.

Monoclonal antibodies targeting the FimH adhesin protect against uropathogenic E. coli UTI

Edward D B Lopatto  1   2 , Jesús M Santiago-Borges  1   2   3 , Denise A Sanick  1   2 , Sameer Kumar Malladi  4 , Philippe N Azimzadeh  1   2 , Morgan W Timm  1   2 , Isabella F Fox  4 , Aaron J Schmitz  4 , Jackson S Turner  4 , Shaza Sayed Ahmed  4 , Lillian Ortinau  1   2   3 , Nathaniel C Gualberto  1   2 , Jerome S Pinkner  1   2 , Karen W Dodson  1   2 , Ali H Ellebedy  1   4   5 , Andrew L Kau  1   2   3 , Scott J Hultgren  1   2   5
Affiliations

Monoclonal antibodies targeting the FimH adhesin protect against uropathogenic E. coli UTI

Edward D B Lopatto et al. bioRxiv. .

Update in

Abstract

As antimicrobial resistance increases, urinary tract infections (UTIs) are expected to pose an increased burden in morbidity and expense on the healthcare system, increasing the need for alternative antibiotic-sparing treatments. Most UTIs are caused by uropathogenic Escherichia coli (UPEC), while Klebsiella pneumoniae causes a significant portion of non-UPEC UTIs. Both bacteria express type 1 pili tipped with the mannose-binding FimH adhesin critical for UTI pathogenesis. We generated and biochemically characterized 33 murine monoclonal antibodies (mAbs) to FimH. Two mAbs protected mice from E. coli UTI. Mechanistically, we show that this protection is Fc-independent and mediated by the ability of these mAbs to sterically block FimH function. Our data reveals that FimH mAbs hold promise as an antibiotic-sparing treatment strategy.

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Conflict of interest statement

S.J.H. is on the advisory board of Sequoia Vaccines, Inc. S.J.H is a co-founder of Fimbrion Therapeutics that is developing FimH targeted therapies and may financially benefit if the company is successful. The Ellebedy laboratory received funding from Emergent BioSolutions, AbbVie, and Moderna that are unrelated to the data presented in the current study. A.H.E. has received consulting and speaking fees from InBios International, Fimbrion Therapeutics, RGAX, Mubadala Investment Company, Moderna, Pfizer, GSK, Danaher, Third Rock Ventures, Goldman Sachs and Morgan Stanley and is the founder of ImmuneBio Consulting. A. J. S., J.S.T., and A.H.E. are recipients of a licensing agreement with Abbvie that is unrelated to the data presented in the current study. All other authors declare that they have no competing interests.

Figures

Fig. 1.
Fig. 1.. FimH mAbs bind to four distinct epitope classes.
Structures of (A) E. coli FimHLD (PDB 1KLF), (B) K. pneumoniae FimHLD (PDB 9AT9), and (C) E. coli FmlHLD (PDB 6AOW). Residue differences from E. coli FimHLD are highlighted in red. (D) ELISA EC50 values for each mAb to the listed protein. White cells with no values indicate EC50 values were above the range measured in the assay. (E) Epitope mapping of mAbs (top labels) to a panel of FimH mutants (right labels). Binding classes were determined by shared residues that abrogated mAb binding which are highlighted in purple on the surface of E. coli FimHLD (above) (PDB 1KLF) and the table (below).
Fig. 2.
Fig. 2.. Structural basis of FimH mAb protection.
(A-E) Cryo-EM density maps of Fabs bound to FimHLD. (A) Kp1 2H04 (teal), (B) Ec1 F7 (cyan), (C) Kp1 2H04 and Ec1 F7 maps superimposed on each other, (D) Kp2 2C07 (sand), and (E) Ec3 B7 (dark green) bound to FimHLD (salmon). (F-I) The binding epitopes of the Fabs on FimHLD colored by the same color scheme as panels A-E. Density map overlaid on model residue interactions of (J) Kp1 2H04 (cyan) with FimH P26 and (K) Ec3 B7 (dark green) with FimH Y64. mAb heavy chain residues are labeled “HC” and light chain residues are labeled “LC”.
Fig. 3.
Fig. 3.. mAbs bind the relaxed conformation of FimH.
(A) FimH mAb binding to UTI89 bacteria and (B) UTI89 overexpressing conformationally shifted FimH variants (n=3, error bars represent SEM). (C) Representative binding curves of Kp1 2H04, Ec1 F7, and Kp2 2C07 Fab binding FimHLD and FimGNTEH. Results are from kinetic measurements of dilution series of one experiment. (D) Observed BLI binding kinetics to Ec FimGNTEH and Ec FimHLD. N.D. means not determined due to the off rate being below the detection limit.
Fig. 4.
Fig. 4.. Relaxed-preferred mAbs can inhibit FimH binding.
(A) Inhibition of FimHLD binding to BSM at a 5:1 molar ratio of mAb to protein (n≥3). (B) Inhibition of UTI89 guinea pig erythrocyte hemagglutination (n≥2). (C) mAb inhibition of FimHLD binding to C3H/HeN mouse bladders. mAb was pre-incubated with FimHLD at a 10:1 molar ratio. Sections were stained with DNA dye Hoechst (blue), Ec or Kp FimHLD (red) and antibody to uroplakin III (green). n=4-5 bladder sections with n=2 technical replicates.
Fig. 5.
Fig. 5.. FimH mAbs protect from E. coli UTI.
(A) 6-7 week old C3H/HeN mice were pretreated with 0.5 mg of mAb 24 h before infection with UTI89. (B) 24 hpi bladder and (C) kidney titers (For 1B03 and 2E08 n=5 with 1 independent replicate, for B7 n=8 with 1 independent replicate, for 2C07 n=10 with 2 independent replicates, for control IgG, F7, 2H04, 1A02, 2E02 n=13-33 with three independent replicates). (D) IBC counts at 6 hpi (n=16 for control IgG, n=14 for Kp1 2H04 with two independent replicates). Representative 5x magnification images of IBCs (green) in splayed mouse bladders for (E) control IgG and (F) Kp1 2H04 treatments. Scale bar represents 200 μm. 24 hpi bladder (G) and kidney titers (H) from the prophylactic model testing control IgG, 2H04, and 2H04LALAPG (n=21 for control group, n=20 for 2H04, and n=19 for 2H04LALAPG with three independent replicates). For D, a Mann-Whitney U test was used to evaluate statistical significance. For B, C, G, and H statistical comparisons were made using Kruskal-Wallis test (nonparametric ANOVA) with Dunn’s comparisons to the control group correcting for multiple comparisons. * P≤0.05, **P≤0.01, ***P≤0.0001., ****P≤0.00001.
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