Background: Development of effective cancer-chemotherapy is the most challenging field due to toxicity of chemo-agents. Objective: As chalcone has been known to have pharmacological applications, here we aimed to synthesized three chalcone derivatives, 2',4'-dihydroxy-3,4-methylenedioxychalcone (C1), 2'-hydroxy-2,4, 6-trimethoxychalcone (C2) and 2'-hydroxy-4-methylchalcone (C3) and investigate their anti-cancer properties against Ehrlich ascites carcinoma (EAC) cell. Method: Anticancer properties against EAC cells were studied by examining growth inhibition, MTT assays, tumour-bearing mice survival, tumour weight measurement and haematological profiles. Also, apoptosis of EAC cells was investigated by fluorescence microscopy, flow-cytometry and DNA fragmentation assays. Expression of apoptosis related genes were studied by reverse transcriptase-PCR (RT-PCR). Results: Among the compounds, C1 exhibited highest cell growth inhibition at 200 mg/kg/day (81.71%; P < 0.01). C1 treatment also increased life span of EAC-bearing mice (82.60%, P < 0.05) with the reduction of tumour burden (22.2%, P < 0.01) compared to untreated EAC-bearing mice. In vitro study indicated that C1 killed EAC-cells in a dose-dependent manner and induced mitochondria-mediated apoptotic pathways. In addition, C1 treated cells exhibited increased apoptotic features such as membrane-blebbing, chromatin condensation, and nuclear fragmentation after Hoechst 33342 staining. Increased fragmentation of DNA in gel electrophoresis followed by C1 treatment further confirmed apoptosis of EAC cells. EAC cells treated with C1 showed reduced Bcl-2 expression in contrast to notable upregulation of p53 and Bax expression. It implied that C1 could reinstate the expression of pro-apoptotic tumour suppressor and inhibit anti-apoptotic genes. Conclusions: Thus, C1 showed significant growth inhibitory properties and induced apoptosis of EAC cells.