This subprogram's overall goal is discovering novel natural products that are active in oncology screens. The subprogram will maximize the likelihood of discovering novel natural products by focusing on endophytic fungi from unexplored ecological niches and uncultured soil microbes. The subprogram will also utilize DNA-based techniques for identifying productive sources. Active compounds will be isolated using bioassay-guided fractionation and structurally characterized by spectroscopic, crystallographic, and/or chemical techniques. The specific aims are: 1. Provide prefractionated fungal extracts for screening in oncology assays. Endophytic fungi, 500 to 750 isolates/year, will be collected in Costa Rica, prioritized by growth characteristics, fermented using productive media and conditions, and extracted. The crude extracts will be prefractionated to provide fractions for screening at Wyeth and the Institute of Chemistry and Cell Biology in cancer relevant assays. Select endophytic fungal strains will be genotyped by 18S ribosomal-DNA sequencing to provide molecular taxonomy for these difficult to classify fungi. 2. Provide extracts from heterologous expression of environmental DNA for screening in oncology assays. Environmental DNA (eDNA) will be collected from a variety of microbial environments, cleaned up and size selected to prepare cosmid libraries. The libraries will initially be selected in E. coli, and probed with a variety of DNA probes based on known natural products synthetic modules by colony hybridization. Positive clones will be retransformed into appropriate hosts and fermented. Crude extracts[unreadable]the DNA selection serves as a stringent prefractionation[unreadable]will be supplied to oncology relevant assays. 3. Carry out refermentation, bioassay-guided fractionation, and structure determination of screening positives. Selected cultures, either fungal of microbial, will be refermented (if necessary), extracted and subjected to bioassay-guided fractionation. Compounds that continue to elicit interest in oncology screens will be structurally characterized by spectroscopic, crystallographic, and/or chemical techniques.