A new cyclic dipeptide penicimutide: the activated production of cyclic dipeptides by introduction of neomycin-resistance in the marine-derived fungus Penicillium purpurogenum G59

A novel cyclic dipeptide, named penicimutide ( 1 ), and four known cyclic dipeptides, cyclo( l -Val- l -Pro) ( 2 ), cyclo( l -Ile- l -Pro) ( 3 ), cyclo( l -Leu- l -Pro) ( 4 ) and cyclo( l -Phe- l -Pro) ( 5 ), were isolated from a neomycin-resistant mutant of the marine-derived fungus Penicillium purpurogenum G59. The structure of 1 , including the absolute configuration, was determined by spectroscopic and chemical methods, especially NMR and Marfey’s analysis. An unusual amino acid in 1 , 4,5-didehydro- l -leucine, was found for the first time occurring in nature. HPLC–ESI–MS analysis evidenced that 1 – 3 were produced only in the mutant strain, but 4 and 5 were produced in both the mutant and parental strains, indicating that the introduction of neomycin-resistance in the mutant activated pathways of 1 – 3 biosynthesis that were silent in the parental strain. Compound 1 selectively inhibited HeLa cells (among five tested human cancer cell lines) with an inhibition rate (IR %) of 39.4 % at 100 µg/mL, a similar inhibition intensity to that of the positive control 5-fluorouracil (IR % of 41.4 % at 100 µg/mL against HeLa cells). The present work exemplifies the effectiveness of our previous DMSO-mediated method for introducing drug-resistance in fungi to activate silent biosynthetic pathways to obtain new bioactive compounds.