The gut microbiota is considered a key player modulating the efficacy of immune checkpoint inhibitor therapy. The study investigated the association between the response to anti-PD-1 therapy and the baseline gut microbiome in a Polish cohort of melanoma patients, alongside selected agents modifying the microbiome. Sixty-four melanoma patients enrolled for the anti-PD-1 therapy, and ten healthy subjects were recruited. The response to the treatment was assessed according to the response evaluation criteria in solid tumors, and patients were classified as responders or non-responders. The association between selected extrinsic factors and response was investigated using questionnaire-based analysis and the metataxonomics of the microbiota. In the responders, the Bacteroidota to Firmicutes ratio was higher, and the richness was decreased. The abundance of Prevotella copri and Bacteroides uniformis was related to the response, whereas the non-responders’ gut microbiota was enriched with Faecalibacterium prausnitzii and Desulfovibrio intestinalis and some unclassified Firmicutes. Dietary patterns, including plant, dairy, and fat consumption as well as gastrointestinal tract functioning were significantly associated with the therapeutic effects of the therapy. The specific gut microbiota along with diet were found to be associated with the response to the therapy in the population of melanoma patients.