Pterospermum acerifolium (L.) Willd., commonly called Kanak Champa, is a folk medicine for inflammation, wounds, infections, ulcers, and tumours. The present study primarily evaluated the cytotoxic potential of theethanolic leaf extract of P. acerifolium on human cancer cell lines, along with antioxidant and antimicrobial activities. The ethanolic extract was obtained using the Soxhlet extraction technique. The antimicrobial performancewas tested against the bacteria Bacillus subtilis and Escherichia coli and the fungi Aspergillus niger and Candidaalbicans, employing the agar well diffusion technique. The extract demonstrated moderate activity against Escherichia coli and limited inhibitory activity towards B. subtilis. Only low antifungal activity was observed. DPPH assay was employed for assessment of radical scavenging potential, and the extract exhibited maximuminhibition of 57.40%. The MTT and brine shrimp lethality assays were used to measure the cytotoxic activity. Brineshrimp mortality reached 50.0% at 1000 µg/mL, indicating preliminary general cytotoxic potential. The extract produced a concentration-related reduction in cell viability, with IC50 values of 67.50, 78.20, 67.84 and 68.80 µg/mLfor MCF-7, Colo-205, HepG2 and A549 cells, respectively. These findings provide a basis for further phytochemical isolation, mechanistic investigation and evaluation of selectivity toward normal cells. The present work aligns withGood Health and Well-being (SDG-3) by supporting scientific investigation of plant-derived materials relevant tohealth-related research. It also aligns with the Special Issue theme by linking traditional medicinal knowledge withcontemporary biological evaluation