François Chable de la Héronnière

A key aspect of cystic fibrosis pathophysiology is the significant role played by neutrophils, central to the inflammatory response in cystic fibrosis airways. Neutrophil-dominated inflammation greatly influences clinical outcomes, including airway damage and lung function decline. People with cystic fibrosis have an increase in circulating neutrophils that exhibit numerous functional and phenotypic abnormalities. We used spectral flow cytometry to delineate the expression of 24 cell surface markers on neutrophils from 24 healthy donors and 45 adults with cystic fibrosis, before and after elexacaftor-tezacaftor-ivacaftor treatment. This comprehensive analysis examined phenotypic markers associated with fundamental neutrophil functions, including differentiation, maturity, activation, antimicrobial activities (degranulation, pathogen sensing and chemotaxis), metabolism, and immunomodulation. Prior to treatment, neutrophils from adults with cystic fibrosis displayed a phenotype suggestive of immune activation and metabolic aberration. Treatment with elexacaftor-tezacaftor-ivacaftor normalized a FPR1high/CXCR2high subset, an observation consistent with enhanced pathogen-sensing capacity. The treatment, however, failed to restore fully normal phenotypes in cystic fibrosis neutrophils but rather induced the modulation of a PD-L1high/CD114high/GLUT-1high subset of mature neutrophils with an increased expression of PD-L1 and adhesion molecules such as CD11c and CD11b. Taken together, our data reveal a specific immunophenotypic signature of neutrophils in cystic fibrosis that is further modified by treatment with elexacaftor-tezacaftor-ivacaftor, suggesting the opportunity for developing adjunctive therapies to enhance beneficial antimicrobial subsets while simultaneously mitigating immunosuppressive phenotypes, thereby opening new perspectives for immune modulation in cystic fibrosis treatment.

Neutrophils are central mediators of the hyperinflammatory response in severe SARS-CoV-2 infection. We report elevated cytosolic levels of proliferating cell nuclear antigen (PCNA) in neutrophils from patients with severe and critical COVID-19, correlating with enhanced NADPH oxidase–dependent reactive oxygen species (ROS) generation and neutrophil extracellular trap (NET) formation. Using T2AA, a small-molecule inhibitor of the PCNA scaffold, we demonstrate potent suppression of NADPH oxidase activation and NET release, particularly in response to SARS-CoV-2 RNA. Mechanistically, we identify a previously unrecognized interaction between PCNA and the heterodimeric S100A8/S100A9 (calprotectin), predominantly enriched in CD16 ^high CD62L ^low neutrophils expanded during COVID-19. PCNA binds the dimeric S100A8/S100A9 complex mediated via S100A8 subunit with micromolar affinity, and this interaction is abrogated by tetramerization, suggesting regulation by intracellular calcium. Disruption of this complex by T2AA inhibited ROS production in an S100A8/S100A9-dependent manner, implicating calprotectin as a functional regulator of neutrophil activation. In a betacoronavirus mouse model, T2AA treatment attenuated lung inflammation, reduced NET and calprotectin levels, and shifted pulmonary neutrophils away from hyperactivated and immunosuppressive phenotypes, consistent with immune reprogramming toward resolution. These findings establish cytosolic PCNA as a central scaffold in neutrophil hyperactivation during COVID-19 and highlight its pharmacological disruption as a promising host-directed strategy to limit inflammation and prevent organ damage.