The investigation of epigenetic modifiers in cultures of an endophytic fungus to enhance PPT production

Podophyllotoxin (PPT) is a natural plant lignan used for the semi-synthesis of anti-cancer drugs. Due to a variety of factors, PPT is becoming less available. An endophytic fungus, PPE7, from Podophyllum peltatum, can serve as an alternate source of PPT, but production declined over several generations of subculture, perhaps due to gene silencing. Small molecule epigenetic modifiers, such as DNA methyl transferase (DNMT) and histone deacytelase (HDAC) inhibitors, can activate silent fungal pathways and enhance the production of fungal metabolites. Re-activation of the PPT pathway may make it possible to identify the genes involved in PPT synthesis through cDNA analysis. The HDAC inhibitor, suberoylanilide hydroxamic acid (SAHA) and the DNMT inhibitor 5-azacytidine (5-aza) were used to treat cultures of PPE7. Extracts were prepared by solid phase extraction (SPE) and analyzed by HPLC. Cultures treated with SAHA and 5-aza showed gain of peaks, loss of peaks and altered amounts of individual peaks compared to the untreated cultures. Thus, both epigenetic modifiers are capable of affecting the compounds produced in cultures of PPE7. LC-MS analysis is in progress to further confirm these findings and explore the production of PPT and related lignans in the treated cultures.