Structural Diversity of Glycosylated Cytochalasins and Congeners from the Endophytic Fungus Trichoderma harzianum MLJ-4 with Anti-NSCLC Activity

The endophytic fungus Trichoderma harzianum MLJ-4, a proven agricultural biocontrol agent with untapped biosynthetic potential, yielded 11 new (1-11) and 22 known (12-33) macrocyclic cytochalasins (CYTs) via rice solid culture. Structural elucidation, including absolute configurations, employed HRESIMS, NMR, NMR calculations with DP4+ probability analysis, chemical methods, ECD calculations, and X-ray crystallography. Trichoharzchalasins A-C (1-3) represent the first reported glycosylated CYTs, and trichoharzchalasin D (4) is a rare example of 12-nor-cytochalasin. Cytotoxicity screening against non-small-cell lung cancer (NSCLC) H1975 cells identified five active compounds (IC₅₀ = 4.04-14.87 μM), with trichoharzchalasin H (8) showing superior potency (IC₅₀ = 4.04 μM) to gefitinib (IC₅₀ = 11.33 μM) and comparable efficacy to cisplatin (IC₅₀ = 4.34 μM). Mechanistically, 8 induced G2/M phase arrest and mitochondrial-mediated apoptosis. These findings underscore the biosynthetic potential of T. harzianum and highlight CYTs as promising NSCLC drug leads.