Pre-clinical toxicological investigations are pivotal in the development of safer and more efficacious chemotherapeutic agents. Oleanolic acid (OA), a naturally occurring pentacyclic triterpenoid, has demonstrated anticancer potential but is often limited by the toxic side effects of its derivatives. In the current study, we carried out the facile synthesis of a modified OA analogue, OD2, and studied its cytotoxicity and efficacy analysis across several cell lines. Mechanistic toxicology was explored through fluorescence-based assays. Annexin-V/Propidium Iodide (A-V/PI) staining and TUNEL assays were used to confirm apoptosis. OD2 exhibited dose-dependent cytotoxicity, with a pronounced effect on A549 lung cancer cells compared to other cancerous and non-cancerous cell lines. Apoptosis was found to be the predominant mode of cell death, evidenced by Fluorescence imaging analysis of chromatin condensation and mitochondrial dysfunction. This was further validated by an increase in Annexin-V-positive and TUNEL-positive cells in treated groups. OD2 activated the intrinsic mitochondrial apoptotic pathway as evidenced by increased Bax and decreased Bcl-2 protein abundance levels. While the current study showcases the therapeutic potential of the selective toxicological activity of OD2, future studies will focus on the deconvolution of its potential polypharmacological mode of action and decoding the basis of its selective action, so as to glean important lessons that can be applied in the development of chemotherapeutic agents with favorable toxicological profiles.