Site-specific delivery of chemotherapeutics specifically to neoplastic hepatocytes without affecting normal hepatocytes should be a focus for potential therapeutic management of hepatocellular carcinoma (HCC). The aptamer TLS 9a with phosphorothioate backbone modifications (L5) has not been explored so far for preferential delivery of therapeutics in neoplastic hepatocytes to induce apoptosis. Thus, the objective of the present investigation was to compare the therapeutic potential of L5-functionalized drug nanocarrier (PTX-NPL5) with those of the other experimental drug nanocarriers functionalized by previously reported HCC cell-targeting aptamers and non-aptamer ligands, such as galactosamine and apotransferrin. A myriad of well-defined investigations such as cell cycle analysis, TUNEL (terminal deoxynucleotidyltransferase-mediated deoxyuridine triphosphate nick end labeling) assay, and studies related to apoptosis, histopathology, and immunoblotting substantiated that PTX-NPL5 had the highest potency among the different ligand-attached experimental formulations in inducing selective apoptosis in neoplastic hepatocytes via a mitochondrial-dependent apoptotic pathway. PTX-NPL5 did not produce any notable toxic effects in healthy hepatocytes, thus unveiling a new and a safer option in targeted therapy for HCC. Molecular modeling study identified two cell-surface biomarker proteins (tumor-associated glycoprotein 72 and heat shock protein 70 [HSP70]) responsible for ligand-receptor interaction of L5 and preferential internalization of PTX-NPL5 via clathrin-mediated endocytosis in neoplastic hepatocytes. The potential of PTX-NPL5 has provided enough impetus for its rapid translation from the pre-clinical to clinical domain to establish itself as a targeted therapeutic to significantly prolong survival in HCC.