GSE116623
The immunogenic potential of bacterial flagella for Salmonella-mediated tumor therapy
Genetically engineered Salmonella Typhimurium are potent vectors for prophylactic and therapeutic measures against pathogens as well as cancer. This is based on the potent adjuvanticity that supports strong immune responses. The physiology of Salmonella is well understood. It simplifies engineering of both enhanced immune-stimulatory properties as well as safety features, thus, resulting in an appropriate balance between attenuation and efficacy for clinical applications. A major virulence factor of Salmonella is the flagellum. It is also a strong pathogen-associated molecular pattern recognized by extra- and intracellular receptors of immune cells of the host. At the same time, it represents a serious metabolic burden. Accordingly, the bacteria evolved tight regulatory mechanisms that control flagella synthesis in vivo. Here, we systematically investigated the immunogenicity and adjuvant properties of various flagella mutants of Salmonella in vitro and in a mouse cancer model in vivo. We found that mutants lacking the flagellum-specific ATPase FliHIJ or the inner membrane ring FliF displayed the greatest stimulatory capacity and strongest anti-tumor effects, while remaining safe in vivo. Scanning electron microscopy revealed the presence of outer membrane vesicles in the ?fliF and ?fliHIJ mutants. Finally, the combination of the ?fliF and ?fliHIJ mutations with our previously described attenuated and immunogenic background strain SF102 displayed strong efficacy against the highly resistant cancer cell line RenCa. We thus conclude that manipulating flagella biosynthesis has great potential for the construction of highly efficacious and versatile Salmonella vector strains. Overall design: The samples of Salmonella Typhimurium strain SF102 (?lpxR9 ?pagL7 ?pagP8 ?aroA) and derivates (EM6331 (SF102 + ?fliHIJ), EM6333 (SF102 + ?fliF)) were analyzed by RNA-seq. The control samples were cultivated in the LB medium until they reached the exponential phase. Four biological replicas were pooled in pairs to get two replicas for RNA extraction, library preparation and sequencing.