The targeting protein for Xenopus kinesin-like protein 2 (TPX2) has been demonstrated to be associated with the tumourigenesis of many cancers. In the present study, we investigated the role and preliminary mechanism of TPX2 in the resistance of lung squamous carcinoma to radiation therapy. The results showed that SK-MES-1R and NCI-H226R cells were more resistant to X-ray irradiation than the parental cells (SK-MES-1 and NCI-H226). Moreover, TPX2 was upregulated in the radioresistant cells compared with the parental cells. TPX2 knockdown significantly decreased TPX2 expression in SK-MES-1 cells, while TPX2 overexpression increased TPX2 expression in NCI-H226 cells compared with the corresponding control cells. TPX2 knockdown enhanced the radiosensitivity of SK-MES-1 and promoted cell apoptosis following exposure to irradiation, whereas TPX2 overexpression decreased the radiosensitivity of NCI-H226 and inhibited cell apoptosis. In in vivo studies, the combination of TPX2 knockdown and irradiation significantly inhibited tumour growth, decreased tumour weight, downregulated TPX2 expression in tumour tissue and induced cell apoptosis in nude mice, while TPX2 overexpression exerted an opposite effect. Our results indicated that TPX2 was correlated with cell radioresistance and it might be served as a therapeutic target to enhance cell radiosensitivity in the radiation therapy of lung squamous carcinoma.