Fusobacterium nucleatum is an oral anaerobe recently found to be prevalent in human colorectal cancer (CRC) where it is associated with poor treatment outcome. In mice, hematogenous F. nucleatum can colonize CRC tissue using its lectin Fap2, which attaches to tumordisplayed Gal-GalNAc. Here, we show that Gal-GalNAc levels increase as human breast cancer progresses, and that occurrence of F. nucleatum gDNA in breast cancer samples correlates with high Gal-GalNAc levels. We demonstrate Fap2-dependent binding of the bacterium to breast cancer samples, which is inhibited by GalNAc. Intravascularly inoculated Fap2-expressing F. nucleatum ATCC 23726 specifically colonize mice mammary tumors, whereas Fap2-deficient bacteria are impaired in tumor colonization. Inoculation with F. nucleatum suppresses accumulation of tumor infiltrating T cells and promotes tumor growth and metastatic progression, the latter two of which can be counteracted by antibiotic treatment. Thus, targeting F. nucleatum or Fap2 might be beneficial during treatment of breast cancer.

Neutrophils play a critical role in cancer, with both protumor and antitumor neutrophil subpopulations reported. The antitumor neutrophil subpopulation has the capacity to kill tumor cells and limit metastatic spread, yet not all tumor cells are equally susceptible to neutrophil cytotoxicity. Because cells that evade neutrophils have greater chances of forming metastases, we explored the mechanism neutrophils use to kill tumor cells. Neutrophil cytotoxicity was previously shown to be mediated by secretion of HO We report here that neutrophil cytotoxicity is Ca dependent and is mediated by TRPM2, a ubiquitously expressed HO-dependent Ca channel. Perturbing TRPM2 expression limited tumor cell proliferation, leading to attenuated tumor growth. Concomitantly, cells expressing reduced levels of TRPM2 were protected from neutrophil cytotoxicity and seeded more efficiently in the premetastatic lung. These findings identify the mechanism utilized by neutrophils to kill disseminated tumor cells and to limit metastatic spread. .

Neutrophils are a heterogeneous population of myeloid cells which may either promote or hinder tumor growth and progression. Anti-tumor neutrophils have the capacity to kill tumor cells in a contactdependent manner. However, the molecular mechanisms underlying tumor cell recognition by neutrophils remained unexplored. Tumor cells were shown to express aberrant glycosylation patterns and neutrophils are equipped with receptors capable of recognizing such glycosylations. Accordingly, we hypothesized that the receptor for advanced glycation end products (RAGE) may facilitate neutrophil recognition of tumor cells. Indeed, RAGE decoy receptors and RAGE-specific blocking antibodies dramatically reduce tumor cell susceptibility to neutrophil cytotoxicity. Unexpectedly, we found that tumor cell RAGE rather than neutrophil RAGE is important for the killing process. We further identified neutrophil Cathepsin G as the neutrophil component interacting with tumor cell RAGE. Cathepsin G-deficient neutrophils show impaired ability to kill tumor cells, suggesting that RAGE-Cathepsin G interaction is required for neutrophil cytotoxicity. These data unravel new aspects of neutrophil antitumor activity and identify a novel role for RAGE and Cathepsin G in neutrophil-mediated cytotoxicity. ARTICLE HISTORY

Preexisting diabetes is a risk factor for the development of multiple types of cancer. Additionally, diabetic patients face a poorer prognosis when diagnosed with cancer. To gain insight into the effects of hyperglycemia, a hallmark of diabetes, on tumor growth and metastatic progression, we combined mouse models of cancer and hyperglycemia. We show that while hyperglycemia attenuates primary tumor growth, it concomitantly increases metastatic seeding in a distant organ. We further show that the increase in metastatic seeding is due to impaired secretion of granulocyte colony-stimulating factor (G-CSF) and impaired neutrophil mobilization. Normalizing blood glucose levels using insulin rescues neutrophil recruitment and tumor growth and concomitantly reduces metastatic seeding. These results provide links among hyperglycemia-induced changes in neutrophil mobilization, primary tumor growth, and metastatic progression. Furthermore, our observations highlight the importance of normalizing blood glucose levels in hyperglycemic cancer patients.

We have recently shown that neutrophil antitumor cytotoxicity is Ca dependent and is mediated by TRPM2, an HO-dependent Ca channel. However, neutrophil antitumor activity is dependent on context and is manifested in the premetastatic niche, but not at the primary site. We therefore hypothesized that expression of TRPM2 and the consequent susceptibility to neutrophil cytotoxicity may be associated with the epithelial/mesenchymal cellular state. We found that TRPM2 expression was upregulated during epithelial-to-mesenchymal transition (EMT), and mesenchymal cells were more susceptible to neutrophil cytotoxicity. Conversely, cells undergoing mesenchymal-to-epithelial transition (MET) expressed reduced levels of TRPM2, rendering them resistant to neutrophil cytotoxicity. Cells expressing reduced levels of TRPM2 were protected from neutrophil cytotoxicity and seeded more efficiently in the premetastatic lung. These data identify TRPM2 as the link between environmental cues at the primary tumor site, tumor cell susceptibility to neutrophil cytotoxicity, and disease progression. Furthermore, these data identify EMT as a process enhancing tumor-cell immune susceptibility and, by contrast, MET as a novel mode of immune evasion. EMT is required for metastatic spread and concomitantly enhances tumor cell susceptibility to neutrophil cytotoxicity. .