Biosynthesis and Biological Profiling of Collinolactone and Semisynthetic Derivatives and MetaboIDent, a Novel Tool for Automated Dereplication

Abstract

The rising incidences of age-related diseases such as Alzheimer and some types of cancer demand novel drugs and therapeutic concepts. Nature has already proven to be an extraordinary rich source of novel structural motifs. They can serve as lead structures for drug development like the prominent antibiotic vancomycin or the anti-cancer drug taxol. In this thesis, a novel natural product named collinolactone was isolated from Streptomyces sp. Gö 40/10. The compound possesses an unprecedented and unique 6-10-7-membered tricyclic system with a cyclodecatriene ring flanked by two lactone rings. Based on stable precursor feeding, a biosynthetic pathway via a type I polyketide synthase was postulated, which was then used for the identification of the encoding gene cluster. Follow-up CRISPR-based genetical engineering, optimization of fermentation parameters and the isolation procedure led to a high-production strain with yields of up to 100 mg/L. A representative and divers set of derivatives of collinolactone was synthesized including the use of Burgess reagent, Strykers reagent and Gilman cuprates. All compounds were profiled in a cell-viability assay on L929 cell line, where some compounds showed cytotoxicity in the micromolar range. The mode of action was studied with fluorescence microscopy on the cell cycle of PtK2 cells and an increased formation of monopolar spindles was observed. The inhibition of putative molecular targets was investigated in a malachite green ATP-based assay. Furthermore, collinolactone was found to exhibit neuroprotective properties in a glutamate induced intramolecular oxidative stress assay on HT22 cells. Only collinolactone showed an effect indicating that the binding is highly specific and even the smallest structural changes will lead to a loss of activity. In addition, the reduction of amyloid beta protein aggregates, one of the hallmarks for the development of Alzheimer disease, was studied. Again, collinolactone showed a reduction of aggregation, making it a very promising lead structure for further development towards a drug for Alzheimer therapy.