RESUMEN
The study of anticancer properties from natural products has regained popularity as natural molecules provide a high diversity of chemical structures with specific biological and medicinal activity. Based on a documented library of the most common medicinal plants used by the indigenous people of North America, we screened and isolated compounds with anti-breast cancer properties from Juniperus communis (common Juniper). Using bioassay-guided fractionation of a crude plant extract, we identified the diterpene isocupressic acid and the aryltetralin lignan deoxypodophyllotoxin (DPT) as potent inducers of caspase-dependent programmed cell death (apoptosis) in malignant MB231 breast cancer cells. Further elucidation revealed that DPT, in contrast to isocupressic acid, also concomitantly inhibited cell survival pathways mediated by the MAPK/ERK and NFκB signaling pathways within hours of treatment. Our findings emphasize the potential and importance of natural product screening for new chemical entities with novel anticancer activities. Natural products research complemented with the wealth of information available through the ethnobotanical and ethnopharmacological knowledge of the indigenous peoples of North America can provide new candidate entities with desirable bioactivities to develop new cancer therapies.
Asunto(s)
Apoptosis/efectos de los fármacos , Neoplasias de la Mama/tratamiento farmacológico , Juniperus/química , Podofilotoxina/análogos & derivados , Antineoplásicos Fitogénicos/química , Antineoplásicos Fitogénicos/farmacología , Productos Biológicos/química , Productos Biológicos/farmacología , Neoplasias de la Mama/metabolismo , Ácidos Carboxílicos/farmacología , Caspasas/metabolismo , Línea Celular Tumoral , Diterpenos/farmacología , Medicamentos Herbarios Chinos , Femenino , Humanos , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , FN-kappa B/metabolismo , Extractos Vegetales/química , Extractos Vegetales/farmacología , Plantas Medicinales/química , Podofilotoxina/química , Podofilotoxina/farmacología , Transducción de Señal/efectos de los fármacos , Tetrahidronaftalenos/farmacologíaRESUMEN
INTRODUCTION: Because of increased resistance to current drugs, there is an urgent need to discover new anti-mycobacterial compounds for the development of novel anti-tuberculosis drugs. The microplate resazurin assay (MRA) is commonly used to evaluate natural products and synthetic compounds for anti-mycobacterial activity. However, the assay can be problematic and unreliable when screening methanolic phytochemical extracts. OBJECTIVE: To optimise the MRA for the screening and bioassay-guided fractionation of phytochemical extracts using Mycobacterium tuberculosis H37Ra. METHODS: The effects of varying assay duration, resazurin solution composition, solvent (dimethyl sulphoxide - DMSO) concentration and type of microtitre plate used on the results and reliability of the MRA were investigated. The optimal bioassay protocol was applied to methanolic extracts of medicinal plants that have been reported to possess anti-mycobacterial activity. RESULTS: The variables investigated were found to have significant effects on the results obtained with the MRA. A standardised procedure that can reliably quantify anti-mycobacterial activity of phytochemical extracts in as little as 48 h was identified. The optimised MRA uses 2% aqueous DMSO, with an indicator solution of 62.5 µg/mL resazurin in 5% aqueous Tween 80 over 96 h incubation. CONCLUSION: The study has identified an optimal procedure for the MRA when used with M. tuberculosis H37Ra that gives rapid, reliable and consistent results. The assay procedure has been used successfully for the screening and bioassay-guided fractionation of anti-mycobacterial compounds from methanol extracts of Canadian medicinal plants.