Objective: To explore the relationship between the expression of human papillomavirus type16 (HPV-16) infection and p27^Kip1 in non-small cell lung cancer (NSCLC) of hakka. Methods: We collected 43 cases of NSCLC specimens by surgical removal and pathological diagnosis from Meizhou People’s Hospital (Meizhou Hospital Affiliated to Sun Yat-sen University) in March 2004 to December 2008, including 20 cases squamous carcinoma and 23 cases of adenocarcinoma; 27 cases of lung benign lesions during the same period were selected as a control group. The expression of HPV-16 and p27^Kip1 in the above specimens was detected by immunohistochemistry (SABC), and the correlation analysis of two proteins in NSCLC was conducted with Spearman rank correlation test. Results: The cytoplasmic positive rate (39.5%) of HPV-16 in NSCLC tissues was higher than that in lung benign lesions (7.4%), and statistical analysis showed significant difference (P<0.01); p27^Kip1 protein expression was observed in NSCLC tissue cell nucleus or cytoplasm, and the positive rate of cell nuclear expression was 9.3%, which was significantly lower than that of lung benign lesions (44.4%; P<0.05); and cytoplasm expression was abundant and positive rate was 48.8%, which was significantly higher than that of pulmonary benign lesions cytoplasm expression (7.4%, P<0.05); p27^Kip1 protein nuclear expression was negatively correlated with cytoplasmic expression, which was statistically significant. HPV-16 expression and cytoplasm p27^Kip1 protein expression in NSCLC were positively correlated, with statistical significance (P<0.05) and p27^Kip1 protein nucleus expression had a tendency of negative correlation (P=0.094). Conclusion: From the results of HPV-16 and p27^Kip1 expression in NSCLC tissues and pulmonary benign lesions, we speculate that HPV-16 may leakage from nucleus to cytoplasm and stranded in cytoplasm through regulating p27^Kip1 protein, making p27^Kip1 protein not play its role in nuclei of tumor suppression, thus promoting the development of NSCLC.