DNABII targeting antibodies as vaccines against biofilm diseases

Abstract

Microbial biofilms, i.e. communities of microorganisms embedded in a complex matrix mainly composed by macromolecules of bacterial origin, play a significant role in several chronic and relapsing infections. A major hurdle in treating biofilm-mediated infections, including those formed on the surface of medical devices, is that bacterial cells are physiologically much more resistant to both antibacterials and host innate and adaptive immune mechanisms [[1]Del Pozo J.L Biofilm-related disease.Expert Rev Anti Infect Ther. 2018; 16: 51-65Crossref PubMed Scopus (203) Google Scholar]. Astoundingly, the U.S. National Institutes of Health estimates biofilms account for 80% of human microbial infections [[2]Magana M. Sereti C. Ioannidis A. et al.Options and limitations in clinical investigation of bacterial biofilms.Clin Microbiol Rev. 2018; 31 (e00084-16)Crossref PubMed Scopus (107) Google Scholar]. This translates to roughly 2 million cases per year, with an estimated 268,000 deaths and a cost of $18 billion, which often requires extreme measures, such as the replacement or the removal of implanted devices or debridement of infected wounds [[3]Rumbaugh K.P. Sauer K Biofilm dispersion.Nat Rev Microbiol. 2020; Crossref PubMed Scopus (242) Google Scholar,[4]Omar A. Wright J.B. Schultz G. Burrell R. Nadworny P Microbial biofilms and chronic wounds.Microorganisms. 2017; 5: 9Crossref Scopus (180) Google Scholar]. Therefore, identification of novel strategies to treat or prevent biofilm-mediated infections is an urgent clinical need. In this optic, strategies that disperse bacteria from an established biofilm, or that prevent its formation by active host immunization (e. g. anti-biofilm vaccines), are now considered promising approaches. In previous works, the authors demonstrated that antibodies targeting members of the DNABII family of bacterial DNA-binding proteins, integration host factor (IHF) and the histone-like protein, are able to sequester DNABII proteins from biofilms, resulting in the rapid collapse and subsequent detachment of bacteria from their protective biofilm matrix. This leads to the subsequent pathogen clearance by host immune effectors or antibiotics [5Goodman S.D. Obergfell K.P. Jurcisek J.A. et al.Biofilms can be dispersed by focusing the immune system on a common family of bacterial nucleoid-associated proteins.Mucosal Immunol. 2011; 4: 625-637Crossref PubMed Scopus (146) Google Scholar, 6Brockson M.E. Novotny L.A. Mokrzan E.M. et al.Evaluation of the kinetics and mechanism of action of anti-integration host factor-mediated disruption of bacterial biofilms.Mol Microbiol. 2014; 93: 1246-1258PubMed Google Scholar, 7Novotny L.A. Goodman S.D. Bakaletz L.O Redirecting the immune response towards immunoprotective domains of a DNABII protein resolves experimental otitis media.NPJ Vaccines. 2019; 4: 43Crossref PubMed Scopus (19) Google Scholar, 8Mokrzan E.M. Novotny L.A. Brockman K.L. Bakaletz L.O Antibodies against the majority subunit (PilA) of the type IV pilus of nontypeable Haemophilus influenzae disperse Moraxella catarrhalis from a dual-species biofilm.MBio. 2018; 9 (e02423–18)Crossref Scopus (23) Google Scholar]. Importantly, this approach is species-independent and effective against in vitro biofilms of numerous bacterial species (n = 22), including members of the high priority Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and Enterobacter spp (ESKAPE) pathogens, and also in experimental biofilm models of chronic human diseases, including otitis media (OM) caused by nontypeable Haemophilus influenzae (NTHi) in chinchillas, lung infection by Pseudomonas aeruginosa in mice, and periodontal peri‑implantitis by Aggregatibacter actinomycetemcomitans in rat. In this issue of EBioMedicine, Novotny and colleagues report on significant progress towards the clinical application of this approach, by i) testing the ability of the Fab portion of a monoclonal antibody raised against a DNABII tip-chimeric peptide to resolve OM infection by NTHi, and ii) assessing the potential of this chimeric peptide to promote host's active immunization and thus preventing biofilm formation [[9]Novotny L.A. Goodman S.D. Bakaletz L.O Targeting a bacterial DNABII protein with a chimeric peptide immunogen or humanised monoclonal antibody to prevent or treat recalcitrant biofilm-mediated infections.EBioMedicine. 2020; https://doi.org/10.1016/j.ebiom.2020.102867Summary Full Text Full Text PDF PubMed Scopus (13) Google Scholar]. To this aim, authors firstly demonstrated that Fab fragments obtained from a murine monoclonal antibody raised against the DNA-binding tip region of the β-subunit of NTHi IHF (NTHiIHF), termed β-tip Fabs, were able to significantly disrupt the biofilms formed by all tested bacterial species, including NTHi, Moraxella catarrhalis, P. aeruginosa, Burkholderia cenocepacia and Staphylococcus aureus. More significantly, in a chinchilla model of OM by NTHi, murine β-tip Fabs was capable in reducing the amount of established biofilm, resolving disease-associated inflammation, and eliciting the production of multiple host immune effectors to effectively eradicate biofilm-released bacteria. Subsequently, the authors generated a chimeric peptide composed of the DNA-binding tip domains of both the α-subunit and β-subunit of IHFNTHi to obtain polyclonal rabbit anti-tip chimeric peptide Fabs. Two administrations of anti-tip Fabs were able to resolve NTHi biofilms in chinchillas, and significantly reduce the bacterial load, likely through stimulation of host immune effectors. Excitingly, the observed therapeutic effect was also durable. Furthermore, to assess the potential usage of these novel therapeutic agents in clinical trials, authors obtained a panel of humanised tip chimeric peptide-directed monoclonal antibodies (HuTipsMabs), and evaluated one of these HuTipMabs in vitro. This HuTipMab showed strong affinity to the target peptide, and significantly disrupt in vitro biofilms formed by NTHi, P. aeruginosa and B. cenocepacia. Significantly, this HuTipsMab was able to disrupt preformed NTHi biofilms in chinchillas in a lasting manner, indicating that the humanization process did not diminish its effectiveness. Additional advantage is that HuTipsMab did not induce overt inflammation incurred by Fabs generated from either murine or rabbit chimeric peptide sera. Lastly, the authors evaluated whether active preimmunisation of chinchillas with tip or tail chimeric peptide and adjuvant could prevent the induction of OM in chinchilla by superinfection of adenovirus and NTHi. The vaccine formulation significantly delays the onset of OM and showed an efficacy of 85% when compared to negative-control cohorts. In conclusion, members of the DNABII family of bacterial DNA-binding proteins are critical components found in the biofilm produced by all bacterial species tested to date, and their high-degree of sequence conservation makes these proteins amendable for species-independent novel antibacterials targeting biofilm-mediated infections. The use of DNABII targeting Fabs in place of intact IgGs could prevent the formation of anti-antibodies in cases where repeated treatments are required, thus constituting a significant step forward toward clinical use for the treatment of biofilm-mediated diseases. The authors have no conflicts of interest to declare. The work in the Lau laboratory is funded by the U.S. National Institute of Health grants HL090699 and HL142626A1. D'Andrea laboratory was supported by internal funding.

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