Abstract: Multi-Purpose Medicinal Plants (MMPs) are gaining unprecedented attention apparently because of their potency to contribute to the prevention and treatment of myriads of diseases. As such the toxicity of many herbs including the MMPs has always been a concern particularly relating to internal organs. The present study investigates the local knowledge use and in vitro cytotoxicity against two cancer cell lines; BGC-823 and HeLa cells on three (MMPs); Moringa oleifera (Lam) (Moringaceae) Andrographis paniculata (Burm. f) and Asystasia vogeliana (Benth) (Acanthaceae). The local knowledge was collated through Participatory Rural Appraisal (PRA) approach from selected locations in the Oke-Ogun area of Oyo state, Nigeria. Ethanolic extracts were used for the in vitro cytotoxicity study. The extracts were screened in two cancer cell lines (BGC-823 and HeLa cells) using the Sulpho Rhodamine B (SRB) assay. For the treatment of diseases, the local knowledge shows that A. paniculata recorded higher fidelity level on the treatment of malaria (95%), diabetes (80%), high blood pressure (87.5%), cancer and tumor (65.7%). The infusion of A. vogeliana in combination with the leaves of Cassia alata, Cymbopogon citrutus and fruit juice of Citrus aurantifolia recorded higher fidelity level in the treatment of malaria and chronic fever (74.8%), gonorrhea (65%) and leprosy (40%) suggesting better alternative to M. oleifera and A. paniculata. The medicinal relevance of the species in the treatment of diseases such as malaria, fever, high blood pressure, cancer, diabetes among others in local herbal medicine were revealed. Cytotoxicity assay on the two cell lines, BGC-823 and HeLa cells revealed that only the ethanolic extract of A. paniculata exhibited some level (moderate) of cytotoxic activity with IC50 values of 24.7 50 and 23.1 μg/mL, respectively. M. oleifera and A. vogeliana did not show any significant activity on the cell lines. The study highlights the importance of local knowledge in finding cost effective, potent and safe herbs for people and screening of the plant species for their activities against cancer cell lines.