Introduction: The p16 gene is a tumor suppressor gene located at chromosome 9p21, that is a cyclin-dependent kinase inhibitor and is essential in regulating the cell cycle. In human papilloma virus (HPV) infection, the HPV oncogenes E6 and E7 can inactivate pRB and thus lead to p16 overexpression. Materials and Methods: This is prospective and descriptive study conducted in the Department of Pathology, IMS, BHU and HIMS over a period of 1 year from 1st January 2016 to 31st December 2016. Archival, formalin fixed tumour specimens from patients were retrieved from the department of pathology for immunohistochemical staining by means of an anti-p16 monoclonal antibody. In total, there were 90 patients. We evaluated p16 expression for its clinicopathological significance. Result: HPV types and status in correlation with clinical parameters and expression of p16. Eighty five out of 90 patients with primary carcinoma of the vagina (PCV) could be evaluated for HPV status. 26 were positive for high-risk HPV and 59 were HPV negative. The majority (17 out of 26, 65%) of HPV-positive patients were positive for HPV16. The others were positive for HPV45 (4 patients, 16.6%), HPV18 (2 patient, 8.3%), HPV35 (1 patient), HPV56 (1 patient), and HPV68 (1 patient). Human papillomavirus positivity was significantly correlated with strong p16 expression (p= 0.045). In all, 7 out of the 59 HPV-negative patients were negative for p16 immunostaining, while the remaining 83% showed varying expression: 39 out of 59 (60.9%) showed moderate or strong p16 expression. Conclusion: The vast majority of HPV positive vaginal cancers showed p16 overexpression, suggesting active involvement of HPV in the malignant transformation process. More in-depth studies are needed to understand the molecular carcinogenesis pathway in these p16-negative tumors and to improve outcomes for this population.