PROJECT SUMMARY Immune checkpoint inhibitors (ICI) have been successful in treating some, but not all, patients with melanoma brain metastases (MBM). Disappointingly, the same immunotherapy has largely been ineffective for the treatment of patients with glioblastoma (GBM). However, recent studies in animal models and humans have suggested that the microbe composition of the gut can positively influence the effectiveness of ICI. Our central hypothesis is that the manipulation of the GBM patient’s microbiota can improve the poor responsiveness to ICI. In the current study, we propose to compare the gut microbe composition of GBM patients to those patients with MBM that have responded favorably to ICI (Aim 1). Using cryopreserved fecal samples from these patients, we will create animals with these human microbiomes to test whether the microbe composition will enhance ICI when the animals are challenged with GBM (Aim 2). The development and use of the humanized animal models in this proposal will establish the role of the human microbiome in the action of immunotherapy. Ultimately, the elucidation as to whether certain microbial communities can enhance the effectiveness of immunotherapy will lead to the development of new approaches that include modification or transplantation of these microbe communities in patients for increased therapeutic impact.