Engineered Salmonella enterica serovar Typhimurium overcomes limitations of anti-bacterial immunity in bacteria-mediated tumor therapy

Abstract

Cancer is one of the leading causes of death in the industrialized world and represents a tremendous social and economic burden. As conventional therapies fail to provide a sustainable cure for most cancer patients, the emerging unique immune therapeutic approach of bacteria-mediated tumor therapy (BMTT) is marching towards a feasible solution. Although promising results have been obtained with BMTT using various preclinical tumor models, for advancement a major concern is immunity against the bacterial vector itself. Pre-exposure to the therapeutic agent under field conditions is a reasonable expectation and may limit the therapeutic efficacy of BMTT. In the present study, we investigated the therapeutic potential of Salmonella and E. coli vector strains in naïve and immunized tumor bearing mice. Pre-exposure to the therapeutic agent caused a significant aberrant phenotype of the microenvironment of colonized tumors and limited the in vivo efficacy of established BMTT vector strains Salmonella SL7207 and E. coli Symbioflor-2. Using targeted genetic engineering, we generated the optimized auxotrophic Salmonella vector strain SF200 (ΔlpxR9 ΔpagL7 ΔpagP8 ΔaroA ΔydiV ΔfliF) harboring modifications in Lipid A and flagella synthesis. This combination of mutations resulted in an increased immune-stimulatory capacity and as such the strain was able to overcome the efficacy-limiting effects of pre-exposure. Thus, we conclude that any limitations of BMTT concerning anti-bacterial immunity may be countered by strategies that optimize the immune-stimulatory capacity of the attenuated vector strains.

The immunogenic potential of bacterial flagella for Salmonella‐mediated tumor therapy

“…Next, we evaluated whether the strongly induced cytokine pattern and production of OMVs correlated with the therapeutic efficacy of the flagella mutant strains in a murine tumor model. Of note, tumor colonization itself appears not to be affected by the modifications of the bacterial flagella as shown previously and confirmed by plating (data not shown). CT26 tumor‐bearing mice were infected intravenously with 5 × 10 6 bacteria and the bodyweight changes as indicator for the health burden of therapy as well as the therapeutic efficacy was assessed (Fig.…”

“…Flagella represent a major immune‐stimulatory structure. In accordance, introducing flagella of other bacterial species or modulating assembly or expression of flagella has improved the performance of such strains in tumor therapeutic approaches . The strains still showed exquisite safety features in vivo .…”

Part II: Breakthroughs in Tumor-Seeking Bacteria

“…Next, the researchers deployed SF200 in mice with antibacterial immunity, anticipating how the anti-tumor bacteria might behave in humans, pets, or livestock that are sensitized to Salmonella (3). They tested the same concept using Escherichia coli , as this close relative of Salmonella is commonly encountered by humans and has an established safety profile.…”

An obligatory anaerobic Salmonella�typhimurium strain redirects M2 macrophages to the M1 phenotype

“…Isolated macrophages were activated by IL-4. The bacterial YB1 strain, which carried the green fluorescent protein (GFP)-tagged plasmid, was added to the macrophage culture for a final bacterial concentration of 5x10 6 /ml. The cultures were incubated at 37˚C under hypoxic conditions as previously described (8) for 2 h. Then, the bacteria were washed away and the macrophages were harvested for further analysis.…”

“…Bacteria, including Bifidobacterium (1,2), Clostridium (3) and Salmonella have been demonstrated to preferentially target and replicate in the hypoxic and necrotic regions of a tumor, resulting in tumor repression (4)(5)(6)(7). In a previous study, a synthetic biology approach was used to generate the novel Salmonella typhimurium strain YB1 (YB1) (8).…”