Identification of ISZ-sTRAIL Protein as a Potent Anticancer Agent for EML4-ALK-Positive Non-Small-Cell Lung Cancer

EML4-ALK-positive lung cancer represents an important molecular subtype of non-small-cell lung cancer (NSCLC) that initially responds to ALK inhibitors but invariably develops resistance, highlighting the need for novel targeted therapeutic strategies. Death receptors DR4 and DR5 are frequently upregulated in malignancies and can selectively induce tumor cell apoptosis upon binding TRAIL. ISZ-sTRAIL, a trimer-stabilized soluble TRAIL fusion protein, exhibits potent antitumor effects via DR4/DR5 signaling activation. However, the expression status of DR4/DR5 in EML4-ALK-positive NSCLC cells and the therapeutic potential of targeting this pathway remain poorly defined. In this study, we evaluated DR4/DR5 protein expression in EML4-ALK-positive NSCLC cells and investigated the antitumor effect of ISZ-sTRAIL produced in an Escherichia coli expression system. Our results showed that DR4 and DR5 were abundantly expressed in EML4-ALK-positive NCI-H2228 and NCI-H3122 cells compared with normal human bronchial epithelial 16HBE cells. Furthermore, ISZ-sTRAIL significantly suppressed the proliferation of NCI-H2228 and NCI-H3122 cells, with IC50 values of 4.51 ± 0.22 nM and 14.98 ± 3.34 nM, respectively, while showing low cytotoxicity toward normal 16HBE cells (IC50 > 1 μM). Moreover, ISZ-sTRAIL induced caspase-dependent apoptosis in both cell lines via activation of extrinsic and intrinsic pathway, and these effects were markedly abrogated by the pan-caspase inhibitor Z-VAD. These findings identify DR4/DR5 as a potential therapeutic target and provide preclinical evidence for the development of TRAIL-based strategies in the treatment of EML4-ALK-positive NSCLC.