Mycelial biotransformation of selenium-enriched polysaccharides from Pleurotus nebrodensis suppress breast cancer via mitochondrial apoptosis and immune microenvironment remodeling.

Selenium-enriched polysaccharides (Se-Ps) have attracted growing interest as promising antitumor agents; however, the potential of Pleurotus nebrodensis (PN) mycelium as a biotransformation carrier for Se-Ps production remains unexplored. This study aimed to develop a mycelial culture system for Se-Ps production and to elucidate their anti-breast cancer mechanisms. Selenium-enriched PN mycelia were obtained by supplementing the medium with sodium selenite at optimized concentration of 50 mg/L. Two purified Se-Ps fractions, Se-PNP-1 and Se-PNP-2, were isolated via water extraction-alcohol precipitation followed by DEAE-52 chromatography. Structural analyses confirmed selenium incorporation and revealed that Se-PNP-2, with a higher selenium content, exhibited a lower molecular weight and random coil conformation, which correlated with its superior bioactivity. In vitro, Se-PNP-2 dose-dependently suppressed 4 T1 cell proliferation (IC₅₀ = 292.4 μg/mL), migration, and invasion, and induced early apoptosis. In vivo, oral administration of Se-PNP significantly inhibited orthotopic 4 T1 tumor growth without observable toxicity, comparable to cyclophosphamide. Mechanistically, Se-PNP indirectly remodeled the tumor immune microenvironment by increasing TNF-α and IFN-γ while suppressing IL-10 and TGF-β1, and upregulated the Bax/Bcl-2 ratio and cleaved Casp3/Parp levels, indicating activation of the mitochondrial apoptosis pathway. These findings establish mycelial biotransformation as a feasible strategy for producing bioactive Se-Ps and position Se-PNP as a promising candidate for breast cancer therapy, with dual direct cytotoxic and immunomodulatory actions.