Combining Activity Profiling with Advanced Annotation to Accelerate the Discovery of Natural Products Targeting Oncogenic Signaling in Melanoma

The discovery of bioactive natural products remains a time-consuming and challenging task. The ability to link high-confidence metabolite annotations in crude extracts with activity would be highly beneficial to the drug discovery process. To address this challenge, HPLC-based activity profiling and advanced UHPLC-HRMS/MS metabolite profiling for annotation were combined to leverage the information obtained from both approaches on a crude extract scaled down to the submilligram level. This strategy was applied to a subset of an extract library screening aiming to identify natural products inhibiting oncogenic signaling in melanoma. Advanced annotation and data organization enabled the identification of compounds that were likely responsible for the activity in the extracts. These compounds belonged to two different natural product scaffolds, namely, brevipolides from a Hyptis brevipes extract and methoxylated flavonoids identified in three different extracts of Hyptis and Artemisia spp. Targeted isolation of these prioritized compounds led to five brevipolides and seven methoxylated flavonoids. Brevipolide A (1) and 6-methoxytricin (9) were the most potent compounds from each chemical class and displayed AKT activity inhibition with an IC50 of 17.6 +/- 1.6 and 4.9 +/- 0.2 mu M, respectively.

Automated summary

The combination of HPLC-based activity profiling and advanced UHPLC-HRMS/MS metabolite profiling allows for the identification of highly active compounds in crude extracts, which can be beneficial to the drug discovery process. This strategy was successfully applied to a subset of extract library screening, leading to the identification and isolation of potent natural products inhibiting oncogenic signaling in melanoma.