Furanoheliangolides from <i>Centratherum punctatum</i> and a General Approach for Stereochemical Assignment of Flexible Chiral Side Chains

Human topoisomerase 3β (TOP3B) is a potential molecular therapeutic target for cancer and viral infections. A high-throughput differential cell viability assay using colon cancer cell lines was developed to identify natural product modulators of TOP3B-associated cancer cell viability. The assay identified an organic extract of the plant <i>Centratherum punctatum</i> as having cytotoxic activity. Seven new furanoheliangolides, centratherolides A-G (<b>1</b>-<b>7</b>), along with two known analogues (2,3-epoxybutyryloxy)-goyazensolanolide (<b>8</b>) and goyazensolide (<b>9</b>), were isolated. Compounds <b>1</b>, <b>8</b>, and <b>9</b> exhibited selective cytotoxic activities against the TOP3B-knockout (TOP3B-KO) human colon carcinoma HCT116 cells compared with the wild-type HCT116 cells (TOP3B-WT). The challenging absolute configuration determination of the flexible chiral side chains in selected analogues (<b>1</b>-<b>4</b> and <b>8</b>) was resolved by combined approaches, including synthesis of chemical standards, DFT ECD calculation, and chiral HPLC analysis. Application of this elucidation methodology to a commercial sesquiterpene lactone clarified a contradiction in the stereochemical assignments reported for centaurepensin/chlorohyssopifolin A and 17-<i>epi</i>-chlorohyssopifolin A.