Individual chemoreceptors of <i>Salmonella typhimurium</i> direct its accumulation in heterogeneous tumor tissue in vitro
The effectiveness of most chemotherapeutics is limited by their inability to penetrate deep into tumor tissue and their ineffectiveness against quiescent cells located far from tumor vasculature. Motile Salmonella typhimurium could overcome these therapeutics barriers. We hypothesize that the accumulation of S. typhimurium in tumors is controlled by two mechanisms: (1) chemotaxis towards compounds produced by quiescent cancer cells and (2) preferential growth within tumor tissue. We also hypothesized that individual chemoreceptors target S. typhimurium to specific tumor microenvironments. Using the tumor cylindroid model, which mimics the microenvironment of in vivo tumors, we elucidated the mechanism by which S. typhimurium is attracted to and accumlates in heterogeneous tumor tissue. Time-lapse fluorescent microscopy was used to quantify the accumulation of wild-type and chemotaxis machinery knockouts, including strains lacking individual cell-surface chemoreceptors, chemotaxis signal transduction pathway enzymes, and the flagella and motor assemblies. To measure the extent of apoptosis induced by individual bacterial strains, caspase-3 activity was measured as a function of time. Spatio-temporal profiles of wild-type bacteria were fit to a mathematical model to calculate two parameters that describe bacterial interaction with tumors: bacterial growth, M, and bacterial chemoattraction, K. It was observed that wild-type S. typhimurium are attracted to cylindroids and accumulate at long time points in the central region of large cylindroids. Both bacterial growth and bacterial chemotaxis were significantly greater in large cylindroids, suggesting that quiescent cells secrete bacterial chemoattractants.
Results also showed how individual chemoreceptors directed bacterial chemotaxis within cylindroids: the aspartate receptor initiated chemotaxis towards cylindroids, the serine receptor initiated penetration, and the ribose/galactose receptor directed S. typhimurium towards necrosis. In addition, strains lacking proper flagella constructs, signal transduction proteins, or active motor function, did not chemotax towards tumor cylindroids, indicating that attraction to specific compounds and not random motility is necessary to promote accumulation in tumors. By deleting the ribose/galactose receptor, bacterial accumulation localized to tumor quiescence and had a greater individual effect on inducing apoptosis than wild-type. This new understanding of the mechanisms of Salmonella migration in tumors will allow for the development of therapies with improved targeting to therapeutically inaccessible regions of tumors.