The National Cancer Institute of the United States (NCI) has initiated a Cancer Moonshot program entitled the NCI Program for Natural Product Discovery.

An automated, high-capacity, and high-throughput procedure for the rapid isolation and identification of biologically active natural products from a prefractionated library is presented. The semi-preparative HPLC method uses one milligram of the primary hit fraction and produces 22 subfractions in an assay ready format. Following screening, all active fractions are analyzed by NMR, LCMS, and FTIR and the active principle structural classes elucidated. In the proof-of-concept study we show the processes involved in generating the subfractions, the throughput of the structural elucidation work, as well as the ability to rapidly isolate and identify new and biologically active natural products. Overall, the rapid second stage purification conserves extract mass, requires much less chemist time and introduces knowledge of structure early in the isolation workflow.

Coibamide A (CbA) is a marine natural product with potent antiproliferative activity against human cancer cells and a unique selectivity profile. Despite promising antitumor activity, the mechanism of cytotoxicity and specific cellular target of CbA remain unknown. Here, we develop an optimized synthetic CbA photoaffinity probe (photo-CbA) and use it to demonstrate that CbA directly targets the Sec61α subunit of the Sec61 protein translocon. CbA binding to Sec61 results in broad substrate-nonselective inhibition of ER protein import and potent cytotoxicity against specific cancer cell lines. CbA targets a lumenal cavity of Sec61 that is partially shared with known Sec61 inhibitors, yet profiling against resistance conferring Sec61α mutations identified from human HCT116 cells suggests a distinct binding mode for CbA. Specifically, despite conferring strong resistance to all previously known Sec61 inhibitors, the Sec61α mutant R66I remains sensitive to CbA. A further unbiased screen for Sec61α resistance mutations identified the CbA-resistant mutation S71P, which confirms non-identical binding sites for CbA and apratoxin A and supports the susceptibility of the Sec61 plug region for channel inhibition. Remarkably, CbA, apratoxin A and ipomoeassin F do not display comparable patterns of potency and selectivity in the NCI60 panel of human cancer cell lines. Our work connecting CbA activity with selective prevention of secretory and membrane protein biogenesis by inhibition of Sec61 opens up possibilities for developing new Sec61 inhibitors with improved drug-like properties that are based on the coibamide pharmacophore.