Identification of EI-17056 as a novel small-molecule radiosensitizer in parental H460 and radioresistant H460R lung cancer cells.

Abstract

Radiotherapy is a major treatment modality for non-small cell lung cancer (NSCLC); however, acquired radioresistance remains a significant challenge that limits its therapeutic efficacy. Therefore, we aimed to evaluate our in-house chemical library of heterocycle-based compounds spanning diverse chemical scaffolds to identify molecules capable of enhancing radiation responsiveness in lung cancer cells, including those with acquired radioresistance. Briefly, H460 cells were subjected to repeated fractionated irradiation (2 Gy per fraction) at a cumulative dose of 80 Gy to generate a radioresistant H460 subline (H460R). The radioresistant phenotype was confirmed by clonogenic survival, Annexin V apoptosis, and intracellular oxidative signal assays following irradiation. Subsequently, a library of 59 structurally diverse heterocycle-based small molecules was screened in H460 and H460R cells. Among the compounds tested, EI-17056, a benzylpyrazole scaffold, was identified as a lead radiosensitizing candidate and exhibited minimal cytotoxicity when administered alone under the screening conditions. Pretreatment with EI-17056 enhanced radiation-induced cell death, as demonstrated by clonogenic survival and Annexin V apoptosis assays. Furthermore, EI-17056 increased intracellular oxidative signals and γH2AX levels following irradiation in both H460 and H460R cells. Importantly, treatment with N-acetyl-L-cysteine (NAC) attenuated the EI-17056-associated increase in intracellular oxidative signals and reduced the enhancement of radiation-induced apoptosis, providing functional evidence that oxidative stress contributes, at least in part, to EI-17056-enhanced radiation-induced cell death. Collectively, our findings identify EI-17056 as a novel small-molecule radiosensitizing candidate that enhances radiation responsiveness in parental H460 cells and shows encouraging evidence of radiosensitizing activity in acquired radioresistant H460R cells. These results support further investigation of EI-17056 as a candidate therapeutic radiosensitizer, while its broader applicability to NSCLC remains to be established in additional models.

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