Coumarins and betulinic acid analogues from Mammea americana and their inhibitory activities on oncogenic MAPK/ERK and PI3K/AKT pathways in human melanoma cells

Melanoma, a highly aggressive form of skin cancer, is primarily driven by two key oncogenic signaling pathways: MAPK/ERK and PI3K/AKT. In a discovery program aiming to identify natural products that inhibit one or both pathways, an in-house library of 2'576 plant extracts was screened using a high-content screening assay with melanoma cells expressing ERK/AKT activity biosensors to quantify inhibition at the single-cell level. The ethyl acetate extract from the leaves of Mammea americana was found to inhibit both pathways in the patient-derived cell line MM121224 and was selected for HPLC-based activity profiling. Scale-up isolation of the compounds eluting in the active window of the chromatogram afforded ten previously undescribed betulinic acid analogues (1-10), along with nine known coumarins (11-19). The isolated compounds were individually evaluated for their inhibitory activity on ERK and AKT in MM121224 cells. Interestingly, none of the betulinic acid derivatives 1-10 showed activity in the assay. In contrast, the isolated coumarins were shown to inhibit both pathways, with the most potent theraphin B (11) exhibiting an IC₅₀ value of 37.0 ± 0.4 μM against the AKT pathway, while also demonstrating weaker activity against the ERK pathway.