Conformation and Relative Configuration of a Very Potent Glycosylphosphatidylinositol‐Anchoring Inhibitor with an Unusual Tricarbocyclic Sesterterpenoid δ ‐Lactone Skeleton from the Fungus Paecilomyces inflatus

In the course of our screening for glycosylphosphatidylinositol (GPI) inhibitors, we found a fungal strain, Paecilomyces inflatus , which inhibited GPI anchoring in yeast. Bioassay‐guided fractionation with gel filtration, MPLC on normal phase and prep. HPLC on reversed‐phase yielded a minor secondary metabolite, the substituted hexadecahydroindeno[5′,6′:4,5]cycloocta[1,2‐ c ]pyranyl heptanoate 1 , that inhibited GPI synthesis in vitro by yeast microsomes with a MIC of 3.4 n m. Ester 1 specifically inhibited GPI synthesis in eukaryotic, including mammalian cells, but had no significant activity in protozoa. Based on spectroscopic evidence, including UV, FT‐IR, FAB‐MS, ESI‐HR‐MS, 1 H‐NMR, 13 C‐NMR, DQ‐COSY, ROESY, HSQC, and HMBC data, the metabolite 1 was shown to have an unusual tricarbocyclic sesterterpenoid δ ‐lactone skeleton. Its solution conformation and relative configuration was elucidated with 1 H, 1 H coupling constants and detailed analysis of its ROESY data, coupled with inspection of Dreiding models. In the preliminary investigations on structure‐activity relationships, the three derivatives 2  –  4 of ester 1 were prepared by acetylation, catalytic hydrogenation, and intramolecular alkene addition of a hydroxy group, respectively. The test results revealed that the modifications of substituents at the δ ‐lactone ring or of the C=C bonds of 1 caused a loss of activity by a factor of ca . 500 to 5000. Therefore, it seems that the δ ‐lactone ring and the C=C bonds in 1 are essential for the potent GPI‐inhibitory activity.