The role of oral microbiota in pre-metastatic niche preparation in the brain

Background: The oral microbiota plays a critical role in maintaining oral health and immune homeostasis. Emerging evidence suggests a link between oral microbiota and the brain, particularly in the context of brain development and neurological diseases. However, its contribution to brain tumors is underexplored. In this study, we investigated the role of oral microbiota dysbiosis (OMD) in brain metastasis (BrM) development. We hypothesized that OMD may alter the brain's physiological state in ways that favor premetastatic niche (PMN) preparation, thereby creating conditions conducive to BrM development. Methods: Mice with subcutaneous BP melanoma tumors, with no intracranial tumors, were used to model the PMN preparation stage in the brain during melanoma development. To induce OMD, 5 days after primary tumor injection, mice were treated with chlorhexidine gluconate mouthwashes every 12 hours for 10 days. Saliva and stool samples were collected longitudinally and analyzed via 16S rRNA amplicon sequencing. Primary melanoma tumors were measured using a caliper. Brain samples were analyzed by mass spectrometry and immunohistochemistry. Digital spatial profiling was used to determine immune composition and transcriptional signatures in both brain and melanoma samples. Results: Chlorhexidine treatment led to OMD, as verified by a reduction in alpha diversity and alterations in bacterial composition of the oral microbiota. The gut microbiota remained unchanged. Mice with OMD demonstrated a significant increase in primary tumor growth compared to controls (p=0.039), associated with lower immune infiltration. In the brain, OMD resulted in upregulation of proteins associated with acute inflammation and a reduction in peri-vascular extracellular matrix and adhesion proteins, suggestive of blood-brain barrier remodeling. Conclusion: Our investigation suggests oral microbial imbalance is associated with alterations of the peri-vascular niche and may contribute to PMN preparation in the brain. Ongoing studies in our group investigate the impact of these OMD-induced alterations on BrM development, providing insights for microbiota-targeted translational applications.