Cancer is one of the most common diseases in the world. Recently, there are many methods developed by researchers to treat this disease. One of these treatments is targeted for chemotherapy. It is preferred by researchers because it is less toxic and has fewer side effects than other cancer treatments. This study emphasizes the anticancer properties of the newly synthesized Isoindole derivatives. Thus, it was hoped to be a significant improvement based on new generation anticancer compounds with high efficacy and fewer side effects. The main objective of this study was to investigate the biological activity of seven newly synthesized Isoindole derivatives. The anticancer activity of these compounds was evaluated against HeLa (cervical carcinoma) and A549 (lung adenocarcinoma) cancer cell lines. This study is divided into three parts. Firstly, the cytotoxic activity of these compounds was determined by measuring the cell viability of each compound on HeLa and A549 cell lines. The main objective of this analysis is to measure the IC50 value of each compound and determine which compound is best to kill at least half of the cells. Secondly, the effects of programmed cell death and cell cycle were investigated for compounds with the best IC50 for each cell line by using Annexin V-FITC in flow cytometry. Finally, a scratch assay was performed to investigate the effect of these new Isoindole derivatives on cell migration.