Neisseria gonorrhoeae: Gonococcus, a bacterium resistant to most antibiotics causing more than 80 million cases of gonorrhea annually, is a WHO high priority pathogen. Recently, vaccine development prospects were boosted by reports that licensed meningococcus serogroup B (MenB) vaccines provided partial protection against gonococcus infection. Thanks to a collaboration with the University Hospital in Siena (Italy), we enrolled donors immunized with the MenB vaccine 4CMenB (Bexsero) to collect their blood and isolate human peripheral blood mononuclear cells (PBMCs). From these samples over 3,000 MenB outer membrane vesicles (OMVs)-specific MBCs were single cell sorted and almost 400 antibodies showed to cross-bind MenB and gonococcus. To identify mAbs able to kill gonococcus and MenB strains, all cross-binding antibodies were tested through a bactericidal assay. From this screening we identified a panel of 17 gonococcus bactericidal mAbs (b-mAbs). In addition to the bactericidal screening, b-mAbs were also tested through innovative assays based on the high-content confocal microscopy platform Opera Phenix to evaluate their opsonophagocytic activity. All but one b-mAb were able to promote Opsonophagocytosis in vitro. Next, we aimed to identify the antigen targeted by identified b-mAbs. Our data revealed that the majority of b-mAbs targeted the PorB (n=9) antigen followed by antibodies directed against the LOS (n=4). The antigen recognized by the remaining 4 b-mAbs remained unknown. The most potent and broadly reactive b-mAb, named 01K12, was tested in vivo and demonstrated protection against gonococcus infection when administered prophylactically.
Thanks to our study we identified PorB and LOS as key antigens of gonococcal and meningococcal immunity providing a mechanistic explanation of the cross-protection observed in the clinic. In addition, we showed that isolating human monoclonal antibodies from vaccinees can be instrumental for rapid bacterial antigen discovery which can be used to address the challenge posed by AMR.
SARS-CoV-2: The coronavirus disease 2019 (COVID-19) pandemic has had a devastating economic and health impact on our society. Over 7 million deaths worldwide have been attributed to SARS-CoV-2, the causative agent of COVID-19. Since the beginning of 2020, we started isolating MBCs from the blood of COVID-19 survivors to discover extremely potent nAbs and develop new medical countermeasures. We isolated over 4,000 S protein-specific MBCs by single cell sorting from 14 donors and identified 453 nAbs. Only 1.4% of them neutralized the authentic virus in vitro with a potency of 1-10 ng/mL. The lead candidate, named J08 (or MAD0004J08), showed to neutralize SARS-CoV-2 and the initial variants (Alpha, Beta, Gamma and Delta) with a 100% inhibitory concentration (IC100) below 10 ng/ml and to protect and treat SARS-CoV-2 infection in vivo at a concentration as low as 0.25 mg/Kg. MAD0004J08 was developed, manufactured and tested in a Phase II/III clinical trial. Unfortunately, after treating around hundreds of individuals, the phase II/III trial was suspended as MAD0004J08 showed a reduced neutralization activity against the newly emerged and globally spread omicron variant.
From 2021 until the end of the vAMRes project we enrolled donors that received two (seronegative 2 doses; SN2) and three vaccine doses (seronegative 3 doses; SN3), and individuals with hybrid immunity, i.e. infection followed by 2 mRNA vaccine doses. From these cohorts, we isolated over 10,000 S protein-specific MBCs, identifying over 700 nAbs against the live SARS-CoV-2 virus. All antibodies were tested in neutralization against emerging SARS-CoV-2 variants to identify the most potent and broadly reactive nAbs. All nAbs were also characterized to understand their epitope region, evaluate their Fc functions and identify their germlines. Thanks to the work performed through the vAMRes project we were able to identify potent and variant resistant nAbs, define protective epitope regions and shed light on the evolution of the B cell and antibody response during infection and after vaccination. This information will be instrumental to design the next generation of therapeutics and vaccines against COVID-19 and human coronavirus related threats.
From the above projects, seventeen manuscripts were published in top tier journals including PNAS, Cell, Nature, Nature Medicine, Nature communications and Journal of Experimental Medicine.
