Non-small cell lung cancer (NSCLC) is the most common type of lung cancer. We previously identified LHX3 as a new preferentially expressed gene in NSCLC. In the present study, we sought to determine its expression, the clinical relevance and the functional roles in NSCLC. LHX3 expression is sharply increased in carcinoma tissues compared to non-carcinoma tissues. Relational analysis reveals a significant association between LHX3 expression and clinical stage (n=172, p=0.032) or radiotherapy (n=167, p=0.022) of patients. LHX3 expression is much higher in the patients at advanced stages (stage III-IV) than in the patients at early stages (stage I-II, p=0.0304), and LHX3 expression is remarkably increased in the patients with radiotherapy treatment (p=0.0002). Survival analyses indicate that LHX3 is associated with unfavorable survival (n=180, p=0.002) and represents an independent prognostic factor [hazard ratio (HR)=1.834, p=0.004] of the NSCLC patients. Furthermore, LHX3 is associated with unfavorable overall survival (n=866, p=0.004) and represents an independent prognostic factor (HR=2.36, p=0.000) in lung adenocarcinoma (ADC) patients, but is not associated with overall survival of squamous cell carcinoma (SCC) patients (n=524, p=0.27). Further analyses found that LHX3 is an early-stage (n=94, p=0.003) and radiosensitivity (n=45, p=0.002) prognostic factor in ADC patients. The patients without radiotherapy have a significantly prolonged survival compared to those with radiotherapy (p=0.0069). Further functional studies show that forced expression of LHX3 in lung cancer cells obviously promotes cell proliferation and invasion, whereas inhibits cell apoptosis. In summary, LHX3 is an early-stage and radiosensitivity prognostic biomarker, and a novel potential oncogene in ADC.