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P. aeruginosa is a major pathogen of nosocomial infections, often multidrug-resistant, and particularly dangerous for patients on mechanical ventilation. It also frequently causes persistent lung infections in patients with cystic fibrosis.
Due to its key role during infection, the Type 3 Secretion System (T3SS), a virulence factor of P. aeruginosa, represents a major therapeutic target. Inhibiting this factor thus reduces the virulence of P. aeruginosa in in vitro and in vivo models.
Following oligomerization of PcrV into a pentamer at the tip of the injectisome, a pore forms in the membrane of the host cell, allowing injection of toxins and ultimately leading to cell death. A single molecule of mAb MEDI3902 binds to the PcrV pentamer and, although not effectively blocking pore formation, strongly inhibits toxin injection.
Conversely, mAb P3D6 can only bind to PcrV monomers, and its mechanism of action appears to involve inhibition of PcrV pentamer assembly. Finally, up to five molecules of mAb 30-B8 can simultaneously bind to the assembled PcrV pentamer, thus very effectively blocking both pore formation and toxin injection.
This latter mechanism of action appears to be the most effective.
© CEA-Irig/IBS/ - Figure created with Biorender (https://BioRender.com)
The project's objective was to isolate human mAbs capable of reducing the pathogenicity of P. aeruginosa by targeting the T3SS. To do this, based on sorting specific B lymphocytes from patients with cystic fibrosis, the researchers succeeded in isolating mAbs targeting T3SS proteins of P. aeruginosa. They thus identified two antibodies that bind to the PcrV protein located at the tip of the injectisome and block toxin injection by the T3SS, reducing the virulence of P. aeruginosa.
By combining approaches from cellular microbiology, genetics, immunology, and structural biology, the consortium also showed that different anti-PcrV mAbs inhibiting T3SS activity, from this study as well as from previous work, act through distinct mechanisms, depending on the epitope recognized.
This work led to the identification of human mAbs capable of blocking T3SS activity, as well as the elucidation of specific modes of action of different T3SS-inhibitory antibodies. The comparative analysis of these modes of action contributes to a better understanding of the differences in efficacy observed among mAbs depending on the epitopes recognized, highlighting that certain mechanisms of action are more effective than others and thus allowing identification of which epitopes should be prioritized for isolating new effective anti-T3SS agents.
Furthermore, these results provide important elements for the structural design of potential anti-Pseudomonas vaccines based on the PcrV protein. Overall, this work opens new perspectives for the development of effective alternative therapies in a context of increasingly marked antibiotic resistance.