An mRNA-based platform for the delivery of pathogen-specific IgA into mucosal secretions.

Colonization of the gut and airways by pathogenic bacteria can lead to local tissue destruction and life-threatening systemic infections, especially in immunologically compromised individuals. Here, we describe an mRNA-based platform enabling delivery of pathogen-specific immunoglobulin A (IgA) monoclonal antibodies into mucosal secretions. The platform consists of synthetic mRNA encoding IgA heavy, light, and joining (J) chains, packaged in lipid nanoparticles (LNPs) that express glycosylated, dimeric IgA with functional activity in vitro and in vivo. Importantly, mRNA-derived IgA had a significantly greater serum half-life and a more native glycosylation profile in mice than did a recombinantly produced IgA. Expression of an mRNA encoded Salmonella-specific IgA in mice resulted in intestinal localization and limited Peyer's patch invasion. The same mRNA-LNP technology was used to express a Pseudomonas-specific IgA that protected from a lung challenge. Leveraging the mRNA antibody technology as a means to intercept bacterial pathogens at mucosal surfaces opens up avenues for prophylactic and therapeutic interventions.

An epitope-enriched immunogen expands responses to a conserved viral site.

Pathogens evade host humoral responses by accumulating mutations in surface antigens. While variable, there are conserved regions that cannot mutate without compromising fitness. Antibodies targeting these conserved epitopes are often broadly protective but remain minor components of the repertoire. Rational immunogen design leverages a structural understanding of viral antigens to modulate humoral responses to favor these responses. Here, we report an epitope-enriched immunogen presenting a higher copy number of the influenza hemagglutinin (HA) receptor-binding site (RBS) epitope relative to other B cell epitopes. Immunization in a partially humanized murine model imprinted with an H1 influenza shows H1-specific serum and >99% H1-specific B cells being RBS-directed. Single B cell analyses show a genetically restricted response that structural analysis defines as RBS-directed antibodies engaging the RBS with germline-encoded contacts. These data show how epitope enrichment expands B cell responses toward conserved epitopes and advances immunogen design approaches for next-generation viral vaccines.

Structure-Guided Molecular Grafting of a Complex Broadly Neutralizing Viral Epitope.

Antigenic variation and viral evolution have thwarted traditional influenza vaccination strategies. The broad protection afforded by a "universal" influenza vaccine may come from immunogens that elicit humoral immune responses targeting conserved epitopes on the viral hemagglutinin (HA), such as the receptor-binding site (RBS). Here, we engineered candidate immunogens that use noncirculating, avian influenza HAs as molecular scaffolds to present the broadly neutralizing RBS epitope from historical, circulating H1 influenzas. These "resurfaced" HAs (rsHAs) remove epitopes potentially targeted by strain-specific responses in immune-experienced individuals. Through structure-guided optimization, we improved two antigenically different scaffolds to bind a diverse panel of pan-H1 and H1/H3 cross-reactive bnAbs with high affinity. Subsequent serological and single germinal center B cell analyses from murine prime-boost immunizations show that the rsHAs are both immunogenic and can augment the quality of elicited RBS-directed antibodies. Our structure-guided, RBS grafting approach provides candidate immunogens for selectively presenting a conserved viral epitope.

Influenza Antigen Engineering Focuses Immune Responses to a Subdominant but Broadly Protective Viral Epitope.

Viral glycoproteins are under constant immune surveillance by a host's adaptive immune responses. Antigenic variation including glycan introduction or removal is among the mechanisms viruses have evolved to escape host immunity. Understanding how glycosylation affects immunodominance on complex protein antigens may help decipher underlying B cell biology. To determine how B cell responses can be altered by such modifications, we engineered glycans onto the influenza virus hemagglutinin (HA) and characterized the molecular features of the elicited humoral immunity in mice. We found that glycan addition changed the initially diverse antibody repertoire into an epitope-focused, genetically restricted response. Structural analyses showed that one antibody gene family targeted a previously subdominant, occluded epitope at the head interface. Passive transfer of this antibody conferred Fc-dependent protection to influenza virus-challenged mice. These results have potential implications for next-generation viral vaccines aimed at directing B cell responses to preferred epitope(s).

Mechanosensitivity below Ground: Touch-Sensitive Smell-Producing Roots in the Shy Plant Mimosa pudica.

The roots of the shy plant Mimosa pudica emit a cocktail of small organic and inorganic sulfur compounds and reactive intermediates into the environment, including SO2, methanesulfinic acid, pyruvic acid, lactic acid, ethanesulfinic acid, propanesulfenic acid, 2-aminothiophenol, S-propyl propane 1-thiosulfinate, phenothiazine, and thioformaldehyde, an elusive and highly unstable compound that, to our knowledge, has never before been reported to be emitted by a plant. When soil around the roots is dislodged or when seedling roots are touched, an odor is detected. The perceived odor corresponds to the emission of higher amounts of propanesulfenic acid, 2-aminothiophenol, S-propyl propane 1-thiosulfinate, and phenothiazine. The mechanosensitivity response is selective. Whereas touching the roots with soil or human skin resulted in odor detection, agitating the roots with other materials such as glass did not induce a similar response. Light and electron microscopy studies of the roots revealed the presence of microscopic sac-like root protuberances. Elemental analysis of these projections by energy-dispersive x-ray spectroscopy revealed them to contain higher levels of K(+) and Cl(-) compared with the surrounding tissue. Exposing the protuberances to stimuli that caused odor emission resulted in reductions in the levels of K(+) and Cl(-) in the touched area. The mechanistic implications of the variety of sulfur compounds observed vis-à-vis the pathways for their formation are discussed.