An antibiotic doxorubicin is generated from the bacterium Streptomyces paucities. Since 1960s, it has been used extensively as a chemotherapeutic drug. Doxorubicin belongs to the class of chemotherapy drugs known as anthracyclines. Doxorubicin (Dox) is an effective anticancer medication for solid and hematologic tumors. However, it can poison multiple organs in a variety of people. In this work, the effects of different doxorubicin doses (100,50,25,12.500,6.250 and 3.125 μg/ml) on cell viability were examined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) method in the cell lines HCT116 (colon), PC3 (prostate), Hep-G2 (human hepatocellular carcinoma), and 293T (human embryonic kidney as the control). PI staining and Annexin V/PI staining were used in flow cytometry to measure cell cycle arrest and apoptosis, respectively. The findings demonstrated that cytotoxicity was caused by doxorubicin in HCT116, PC3, Hep-G2, and 293T. IC50 was 24.30, 2.640, 14.72, and 13.43 μg/ml, respectively, using various concentrations. These findings confirmed that doxorubicin has cytotoxic effects on both cancerous and non-cancerous cells by reducing cell viability. Using flow cytometry to measure the cell cycle, and apoptosis, it was discovered that doxorubicin damaged PC3 cells with a considerable increase in apoptosis and cell cycle arrest in the G2/M phase. In addition, when PC3 cell lines were treated with a high dosage of doxorubicin, the mRNA expression levels of p53 and Casp3 dose genes whereas Bcl-2 fell off. At the same time, the effects of doxorubicin on various yeast knockout strains (YKO) were investigated. To determine the three various doxorubicin concentrations, the Comet assay was performed because doxorubicin might lead to DNA damage. The genotypes of YKO were chosen using the Clustal Omega Multiple Sequence Alignment (EMBL-EBI) of saccharomyces and human gene sequence homology. The comet assay showed improved yeast cell sensitivity, which was unquestionably confirmed.