Abstract: Background: This study deals with the anti-metastasis and anti-biofilm activities using silver nanoparticles (AgNPs) against human breast cancer cells (MCF-7) via in vitro methods. Different studies have been reported that biofilms formed by Klebsiella pneumoniae, Escherichia coli and Staphylococcus aureus are resistance to most of the recently used antibiotics due to MDR effect. Materials and methods: The characterization of biosynthesized particles was done by UV-vis spectroscopy, FTIR, SEM and XRD techniques. Cytotoxicity analysis was done by MTT assay and Neutral Red Uptake (NRU) assay. Anti-biofilm activity was carried out by using the bacterial strains of Klebsiella pneumoniae, Escherichia coli and Staphylococcus aureus respectively. Results: AgNPs exhibit strong absorption peak in the 420–500 nm range due to Surface Plasmon Resonance in UV- Vis spectroscopy. In FTIR analysis, we found phenol and alcohol compounds (at 3447 cm-1), carbonyl groups (at 1631 cm-1), alcoholic groups (at 1384 cm-1) and polysaccharides (at 1093 cm-1). In XRD that AgNPs formed were highly crystalline in nature. In SEM the diameter of AgNPs ranges from 30-90 nm and TEM depicts the size of AgNPs to be 15 ± 2 nm. T. natans-AgNPs exhibits strong cytotoxic activity against MCF-7 breast cancer cell line. Conclusion: The presence of different bioactive molecules in T.natans shows that, green synthesized AgNPs significantly inhibited proliferation of MCF-7 breast cancer cells by inducing apoptosis as well as antibiofilm activity using different bacterial colonies.For the very first time we are reporting the anticancer drugs repositioned as a potential antibiofilm agent in the Klebsiella pneumoniae, Escherichia coli and Staphylococcus aureus organisms. Key words: Trapa natans, Silver Nanoparticles (AgNPs),Human Breast Cancer Cells (MCF-7), Anti-biofilm, Anti-metastasis, Cytotoxicity. Key words: Trapa natans, Silver Nanoparticles (AgNPs),Human Breast Cancer Cells (MCF-7), Anti-biofilm, Anti-metastasis, Cytotoxicity.