Expression of P53 and Prognosis in Patients with Head and Neck Squamous Cell Carcinoma (HNSCC)

Introduction: Mutation of TP53 is the most common genetic abnormality in head and neck squamous cell carcinoma (HNSCC) and results in an accumulation and expression of p53 protein in tumor cells. Disruptive TP53 mutations are consistently associated with poor prognosis but correlations of p53 expression with mutation or prognosis have been variable and the usefulness of p53 as a target for immunotherapy is unknown. Favorable prognosis is associated with the accumulation of T lymphocytes (TILs) in the tumor microenvironment and an immune response to p53 has been suggested by demonstration of antibodies to p53 and p53-restricted cytotoxic cells in patients with HNSCC. To investigate if p53 expression is related to accumulation of TILs, p53 expression was measured in a prospective cohort of patients with HNSCC and correlations with TILs and prognosis were determined. Methods: Studied were 534 previously untreated patients (n) with oral cavity (273), oropharynx (158), larynx (81) or hypopharynx (22) cancers. Expression of p53, p21, and p16 and levels of T cell infiltrates (CD4, CD8, FoxP3) were assessed by immunohistology in tissue microarrays from biopsy specimens. HPV testing by routine pathology was available for 401 patients. Associations with clinical variables were tested using Kruskal-Wallis tests (p53, p21) or Chi-square test (p16). Kaplan-Meier and Cox regression methods were used to evaluate univariable and multivariable associations of protein expression with TIL levels and overall survival (OS) and disease specific survival (DSS) after adjusting for known prognostic factors. Median follow up was 44 months. Results: Higher p53 protein expression was associated with worse OS in univariable (HR = 1.05; 95% C.I. = 1.02, 1.09; p = 0.002) but not in multivariable analysis and did not correlate with increased TIL infiltration. Combined TIL weighted score was a significant independent prognostic factor for OS and DSS, (HR = 0.95; 95% C.I. = 0.93, 0.98; p = 0.0003 and HR = 0.96; 95% C.I. = 0.93, 0.99; p = 0.005, respectively). All of the biomarkers (p53, p21, p16, TILs) differed by HPV status and tumor site (p < 0.0001 each). Further analysis by specific tumor site was unremarkable for an association between p53 expression and TILs or prognosis. However, in multivariable analysis for oropharynx cancer, p53 expression was associated with increased risk of death (HR 2.33; 95% C.I. = 1.00, 5.43; p = 0.05) and in particular for the p16 negative oropharyngeal subgroup (HR = 8.62, 95%; C.I. = 1.94, 38.4; p = 0.005). Conclusions: Over-expression of p53 is an unreliable biomarker for prognosis and does not correlate with levels of TILs suggesting that p53 neoantigens are unlikely to be useful targets for future immunotherapy. These findings were confirmed when patients were analyzed by specific tumor site, however in the subset of HPV-negative oropharyngeal cancers, p53 expression was associated with treatment outcomes and could be a useful biomarker. (438 words). ISSN: 2378-3419 DOI: 10.23937/2378-3419/1410122 Lahin et al. Int J Cancer Clin Res 2019, 6:122 • Page 2 of 12 • reliable and reproducible correlations with prognosis or treatment response could be demonstrated in large homogenous groups of patients. Most recently, we have shown that the immune response in the tumor microenvironment as reflected by tumor infiltrating lymphocytes (TILs) is a critical factor in predicting prognosis [28-30]. The role of p53 protein as a neoantigen in the tumor microenvironment is currently unclear and it is Introduction The TP53 gene and its gene products control the cell cycle by inducing arrest in response to DNA damage and apoptosis if the damage is irreparable [1]. Inactivation of TP53 through mutations plays a critical role in early cancer development and progression. TP53 is a key tumor suppressor [2] and TP53 mutation is the most common genetic change in malignancies, involved in 50% of human cancers [3] as well as up to 70% of head and neck squamous cell carcinomas (HNSCC) [4-6]. A spectrum of TP53 mutations have been identified, but disruptive mutations are probably the most important and have consistently been associated with aggressive tumor behavior and poor survival in HNSCC [6-9]. Mutations of the gene most commonly involve a missense mutation involving the p53 DNA-binding domain that leads to a change in the binding properties or conformation of the protein that inactivates its function and frequently results in prolonged protein half-life and accumulation in the cytoplasm, allowing detection by immunohistochemistry in tumor cells [10]. These gene products have been thought to be excellent targets for immunotherapy [11]. However, overexpression of p53 is an unreliable indicator of TP53 mutation because of other loss of function, dominant-negative or gain of function alterations, degradation of p53 by HPV-16 and other causes of post transcriptional modifications [12]. The clinical relevance of p53 protein overexpression as a biomarker and target for immunotherapy in HNSCC has not been fully elucidated and remains controversial. A major goal of the current study was to determine in a large number of patients the potential usefulness of p53 expression as an indicator of immune response and reflection of tumor infiltrating lymphocytes. It is clear that abnormalities in p53 can contribute to aggressive tumor behavior. In laryngeal carcinoma, several investigations showed no association with clinical outcome [13-17]. Some have reported a correlation with reduced survival [18-20], and others with prolonged survival [21]. Similar conflicting data have been reported for other sites in the head and neck region [22,23]. Some investigators have reported a correlation with recurrences but not with overall survival [24,25]. Others have shown an association with improved survival [26]. We previously reported a statistical trend for p53 mutations to predict poor survival [27] and forp53 expression to predict response to chemotherapy [16]. Conclusions from all of these studies have been limited by the small numbers of patients studied and by varying methods of staining and expression scoring. Because of the frequency and important biologic effects of TP53, it is clear that biomarkers such as p53 expression could provide useful clinical information if