Compelling evidence has mounted surrounding the relationship between the gut microbiome and many intestinal and extraintestinal cancers. Few studies exist investigating the relationship between the gut microbiome and sarcoma. We hypothesize that the presence of distant osteosarcoma induces change to the profile of flora within the mouse. Twelve mice were used for this experiment: six were sedated and received an injection of human osteosarcoma cells into the flank, while six served as controls. Baseline stool and weight were collected. Tumor size and mouse weight were recorded weekly, and stool samples were collected and stored. Fecal microbiomes of the mice were profiled by 16S rRNA gene sequencing and analyzed for alpha diversity, relative abundances of microbial taxa, and abundance of specific bacteria at different time points. Alpha diversity was increased in the osteosarcoma group compared with the control group. The family Lachnospiraceae had the second strongest negative net average change in relative abundance over time in the osteosarcoma group whereas it had a positive net average change in the control group. An increased Firmicutes/Bacteroidota (F/B) ratio was observed in the osteosarcoma group relative to the control mice. These differences suggest that there may be an interplay between the gut microbiome and osteosarcoma. Clinical significance: Due to the paucity of literature available, our work can support novel research on this relationship and the development of new, personalized treatments for osteosarcoma.