Identification and characterization of five anti-mitotic sesquiterpene lactones from Arnica cordifolia

Unique ecozones, such as those in Canada, play an important role in the production of distinct natural product chemicals that help plants survive highly variable abiotic conditions and herbivory. Extracts prepared from Arnica cordifolia Hook. (heartleaf arnica), a North American species related to the European medicinal plant Arnica montana L., induce mitotic arrest in human cancer cell lines with a mitotic spindle morphology distinct from other mitotic inhibitors isolated from prairie plant species. This study aimed to identify the anti-mitotic compound(s) of A. cordifolia. The cytotoxic and anti-mitotic activities of A. cordifolia extracts and their active compounds on human cancer cells were characterized by MTT assays, light microscopy, flow cytometry, and immunofluorescence microscopy. The active compounds were isolated by bioassay-guided HPLC fractionation and identified by NMR. Five anti-mitotic sesquiterpene lactones were isolated from A. cordifolia: three previously unidentified structures, and the known compounds aromaticin and pulchellin-2α-O-isovalerate. These compounds induced rounded cells positive for the mitotic marker phospho-histone H3 at concentrations of 5 µM, yet had distinct effects on mitotic spindle morphology. Furthermore, aromaticin treatment induced ubiquitin foci in cells, indicating that it may target the ubiquitin-proteasome pathway. This is the first report of mitotic inhibitors from Arnica cordifolia. Of these five compounds, three have previously undescribed chemical structures, whereas new anti-mitotic activities have been identified for aromaticin and pulchellin-2α-O-isovalerate. Differences in their biological activities suggest that they possess distinct cellular targets. These findings support continued exploration of Canadian botanical species as sources of structurally diverse bioactive compounds.