Microbial polysaccharides are a significant group of functional phytochemicals. Numerous studies have shown the advantageous pharmacological impacts of polysaccharides, including their effectiveness against cancer. A halophilic bacterial strain obtained from coastal sediments produced exopolysaccharides (EPS). The strain was morphologically recognized and further confirmed as Bacillus cereus strain MSS1 using 16S rDNA analysis, with accession number OR133726. The heteropolysaccharides were purified and fractionated with a DEAE-cellulose column, and the preliminary chemical analysis of the most potent fraction (EPSMSS1) indicated that the four different monosaccharides were mannuronic acid, xylose, fructose, and glucuronic acid, with a molar ratio of 1:1:2:0.5, respectively. The highest production was 12.76 g/l using a Box-Behnken design. It showed antibacterial activity, antioxidant activity, and antibiofilm activity. The 3D architecture of the EPSMSS1 of Bacillus cereus strain MSS1 is being described, predicted, and aligned against other bacterial species. These studies offer valuable insights into optimizing efficiency. Therefore, the EPSMSS1 fraction was shown to have anticancer activity and significant anticancer activities in a dose-dependent manner, with an IC50 value of 20.1 μg/ml. Subsequently, various apoptotic markers, such as cytochrome c, BAX, BCl2, and the BAX/BCL2 ratio, were assessed. Our findings demonstrate that EPSMSS1 triggers the activation of apoptotic protein BAX, enhances the generation of cytochrome c, reduces the expression of antiapoptotic protein BCl2, and distorts the BAX/BCL2 ratio in EPSMSS1-treated HCT-116 cells relative to untreated cells. The anticancer efficacy of EPSMSS1 was verified through the assessment of cell cycle progression using flow cytometry. It has been found that EPSMSS1 pauses the cell cycle in the G1/S phase, causing apoptosis. The main motivation behind this study was EPSMSS1, an innovative marine polysaccharide with remarkable biological activity, especially anti-cancer properties.