Investigation of Anticancer Activity of Pongamia Pinnata and Stevia Rebaudiana Against Breast Cancer: An In Vitro And Phytochemical Study

Background: Breast cancer remains the most commonly diagnosed malignancy among women globally, with approximately 2.3 million new cases and 670,000 deaths annually. The limitations of conventional therapies, including significant adverse effects and the development of therapeutic resistance, have necessitated the search for alternative and adjunctive treatment strategies. Medicinal plants have emerged as promising sources of bioactive phytochemicals with anticancer potential. Methods: Plant materials were collected from designated geographical locations and authenticated. Phytochemicals were extracted using successive solvent extraction methods, and bioactive compounds karanjin from P. pinnata and stevioside from S. rebaudiana were isolated and characterized using chromatographic and spectroscopic techniques. The in vitro anticancer potential was evaluated through brine shrimp lethality assay, Allium cepa root tip assay for anti-mitotic activity, and MTT assay for anti-proliferative activity against MCF-7 (ER+) and MDA-MB-231 (triple-negative) human breast cancer cell lines. Results: Karanjin was successfully isolated from P. pinnata seed oil with a yield of approximately 20% and purity exceeding 95%. The brine shrimp lethality assay revealed LC₅₀ values of 42.1 ± 3.1 μg/mL for karanjin and 58.7 ± 2.9 μg/mL for stevioside, indicating significant cytotoxicity. The Allium cepa assay demonstrated dose-dependent reduction in mitotic index, with karanjin and stevioside reducing the mitotic index to 8.2 ± 1.4% and 14.1 ± 1.6%, respectively, at 100 μg/mL, accompanied by chromosomal abnormalities including bridges, fragments, and micronuclei. The MTT assay revealed IC₅₀ values of 18.5 ± 2.1 μg/mL (karanjin) and 32.4 ± 3.8 μg/mL (stevioside) against MCF-7 cells, and 24.6 ± 2.8 μg/mL and 38.9 ± 4.2 μg/mL against MDA-MB-231 cells, respectively. Morphological examination and AO/EB staining confirmed apoptosis induction in treated cells. Conclusion: The findings demonstrate that Pongamia pinnata and Stevia rebaudiana possess significant anticancer activity against breast cancer cell lines, with karanjin exhibiting the highest potency. The bioactive compounds induce cytotoxicity, inhibit mitotic division, and suppress cancer cell proliferation through apoptosis-mediated mechanisms. The activity against both hormone receptor-positive and triple-negative breast cancer subtypes, along with the chemosensitizing potential of stevioside, highlights their therapeutic promise. These results provide scientific validation for the traditional medicinal uses of these plants and support their further investigation as potential phytopharmaceuticals for breast cancer management.