This study overcomes the limitations of traditional anticancer effects of Ganoderma lucidum and pioneers an innovative cultivation system for anticancer Reishi based on synergistic bio-enhancement integrating “traditional Chinese medicine (TCM) compound formula — enzymatic pretreatment — gene editing — transgenic metabolism.” The core technologies include: Construction of a cultivation substrate incorporating anticancer TCM compound formulas; Application of targeted enzymatic hydrolysis technology to efficiently cleave macromolecular active components in TCM; Precise modification of membrane transport systems in Ganoderma strains using the CRISPR-Cas9 gene-editing platform; Reconstruction of metabolic pathways through transgenic technology to achieve synergistic enrichment and stable storage of exogenous antitumor active components together with endogenous functional components of Ganoderma. Experimental results demonstrate that this innovatively cultivated Ganoderma lucidum can effectively activate the body’s immune defense, inhibit cancer cell proliferation through multiple targets, and induce apoptosis, while exhibiting safety with no toxic side effects. This research provides a disruptive technological pathway and a highly promising candidate carrier for the development of efficient, safe, and cost-effective cancer biotherapeutic agents.