Aims: The attention for new and effective anticancer drugs but less toxic is increasing over time. Streptomyces is the most important and well-known source of their bioactive compound production with useful bioactivities. This work aimed for evaluation of the anticancer potential of methanolic extract of Streptomyces sp. strain KSF 83 against non-cancerous cell lines (CCD-841-CoN), breast (MCF-7, MDA-MB-231) and colon cancer cell lines (HT-29, HCT-116). Methodology and results: The characteristic of the strain KSF 83 was identified by morphology and 16S rRNA sequencing and results confirmed that the strain belonged to the genus of Streptomyces. The crude substance was produced via submerged fermentation from the strain and methanol solvent was used to extract the culture filtrate. Methanolic extract possessed low toxicity against CCD-841-CoN with only 18% of inhibition activity at the 400 µg/mL. Among all tested cancer cells, the methanolic extract was able to inhibit the growth of all cancer cells tested with MCF-7 was the highest anticancer activity recorded. The methanolic extract also exhibited cytotoxicity in a range of EC50 of 65.79 μg/mL to 262.40 μg/mL. This study revealed the anticancer potential of Streptomyces sp. strain KSF 83, which could be sources of prospective anticancer drugs against breast and colon cancer. Conclusion, significance and impact of study: The extract of KSF 83 was non-toxic toward normal cell lines and able to inhibit the growth of breast and cancer cell lines, thus it can be a potential source of the anticancer drug against breast and colon cancer.