GMI, a Fungal Immunomodulatory Protein From <i>Ganoderma microsporum</i> , Induces Different Cytotoxicity in Parental and Osimertinib‐Resistant EGFR‐Mutated Lung Cancer Cells via Apoptotic and Autophagic Cell Death

Lung adenocarcinoma often carries driver mutations, such as EGFR mutations, which are effectively targeted by EGFR tyrosine kinase inhibitors (EGFR-TKIs) like osimertinib, a third-generation EGFR-TKI approved for treating T790M and other EGFR-activating mutations. However, the development of adaptive drug resistance remains a major challenge and is associated with poor prognosis in cancer therapy. Ganoderma microsporum immunomodulatory protein (GMI) has demonstrated anticancer properties in various cancers and exhibits synergistic cytotoxic effects when combined with several anticancer drugs. This study investigated the cytotoxic mechanisms of GMI on the lung cancer cell line H1975, which harbors the EGFR L858R/T790M double mutation, as well as on H1975/TR cells with osimertinib resistance. GMI treatment triggered apoptosis in H1975 cells, as indicated by plasma membrane phospholipid translocation and loss of mitochondrial membrane potential. GMI-treated cells displayed increased LC3BII and the development of acidic vesicular organelles, both of which are hallmarks of autophagy induction. Autophagy inhibition by 3-methyladenine and ATG gene silencing effectively decreased the cytotoxic effect of GMI, suggesting that GMI induces autophagic cell death in H1975/TR cells. This study is the first to reveal the novel role of GMI in inducing cytotoxic effects in H1975 cells and H1975/TR cells with osimertinib resistance through two different forms of cell death: apoptosis and autophagy, respectively.