Project Summary/Abstract Cancer is one of the leading causes of death in the United States and worldwide. Appropriate activation of the immune system and effective targeting of tumor cells are the primary hurdles to be overcome for successful cancer immunotherapy. Microbes-based therapy was extensively studied recently to fill the critical unmet needs of cancer patients, where the current treatment options have been exhausted. We have developed genetically engineered Salmonella (GMS), which can actively search for cancer cells, induce tumor regression, prolong survival time, and inhibit cancer metastasis in multiple mouse models. They can also eradicate themselves from the host after treatment. Interestingly, Salmonella also processes multiple natural killer (NK) cell stimulators. On the other hand, oncolytic viruses are new cancer immunotherapeutics and have been approved to use in the clinic. Oncolytic Myxoma virus (MYXV) has been tested in multiple preclinical cancer models and has demonstrated immune stimulating properties for cancer treatment. MYXV was able to bring CD45+ leukocytes to the tumor bed. In addition, these large DNA viruses can be armed to express additional immune activating molecules. In this study, we propose to explore a novel cancer treatment approach that combines the targeted cancer cell killing ability of bacteria and immune stimulating abilities of both oncolytic bacteria and viruses. Apart from the direct killing of cancer cells, improved GMS will stimulate NK cells and MYXV will recruit other tumor-infiltrating leukocytes to activate the anti-tumor immune responses. In addition, we will explore the transgene expressing armed MYXVs that will express cytokines and chemokines to recruit more specific tumor-infiltrating leukocytes. The success of our efforts would turn microbes to appropriate onsite attractors and activators of the immune system for effective cancer therapy to be successful and applicable across a wide range of tumor types.