Ganoderma Lucidum Polysaccharides Potentiate Low‐Dose Gemcitabine Against 4T1 Breast Tumors, Associated With Immune Activation and Gut Microbiota Remodeling

ABSTRACT Gemcitabine (GEM), a first‐line chemotherapeutic agent, may fail to prevent recurrence in some settings. Traditional Chinese medicine emphasizes Yin–Yang balance and “fuzheng” approaches and may offer complementary strategies. Ganoderma lucidum polysaccharides (GLP) exhibit immune‐enhancing activity, but evidence for direct anti‐tumor efficacy, either alone or in combination with low‐dose GEM, remains limited. Using a murine 4T1 breast cancer model, we integrated flow cytometry, 16S rRNA gene sequencing, and gas chromatography–mass spectrometry to evaluate the anti‐tumor activity of GLP combined with low‐dose GEM. GLP alone enhanced immune parameters but did not significantly suppress tumor growth, whereas low‐dose GEM reduced tumor volume and activated anti‐tumor immunity. Notably, the GLP plus GEM combination produced greater tumor inhibition than either treatment alone and was associated with increased IFN‐γ and TNF‐α levels, enhanced CD8 + T‐cell infiltration, and enrichment of Ligilactobacillus, Duncaniella, and Faecalibaculum in the gut microbiota, and elevated serum levels of short‐chain fatty acids and tryptophan. Notably, the antitumor effect in the combination group was accompanied by reduced tumoral IDO expression and elevated serum tryptophan, which were statistically correlated; however, as kynurenine levels were not quantified in this study, we cannot attribute this correlation to altered IDO enzymatic activity or establish a direct functional link along the IDO–tryptophan metabolic pathway. Thus, the combination of GLP and low‐dose GEM exerts a synergistic anti‐tumor effect.