Lawsone (LS) a colored napthoquinone compound obtained from the plant Lawsonia inermis L. (Lythraceae) is known for its usefulness of being a precursor for synthesis of some anticancer compounds. Literatures support potent anticancer activity of napthoquinone derivatives in human colon cancer, present study evaluates the effect and mechanism of LS on chemical induced colon cancerous rats and human colon cancer DLD-1 cells, the study was supported by endoscopy, histological and immunohistochemistry analysis. KAD rats were subjected to colon cancer mediated by Azoxymethane (AOM) injections followed Dextran sodium sulfate (DSS) orally in drinking water. After endoscopic confirmation the rats were given LS (200mg/ml) orally for 8 weeks. Presence of aberrant foci, types of tumors and the proliferative effect on tumor lesions was studied by macroscopic, histological and immunohistochemical analysis. To establish the mechanism, human colon DLD-1 cancer cells were exposed to LS and its effect on proliferation were studied. LS reduced aberrant crypt without affecting tumor pathology. Histological study of colon suggested decrease in numbers of adenomas and lesions. Immunohistochemistry confirmed the antiproliferative activity in adenocarcinomas without affecting the cells of normal colon mucosa. Results on human DLD-1 cells showed LS delayed progression of cell cycle by decreasing expression of cyclin B1 as well as cdk1 by inactivating NF-κB without inducing apoptosis. The study concluded role of LS in suppressing cell proliferation of colon tumors. The suppressive activity on DLD-1 cells was not by apoptosis but by decreased NF-κB activity resulting in suppression of expression levels of cyclin B1 and cdk1.