An Investigation Of Theassociation Between HPV Infection And Prognosis Of Oral Andoropharyngeal Squamous Cell Carcinomas

The primary lesions of oropharyngeal squamous cell carcinoma usually occurred in tongue root,soft palate,pharynx,tonsil.Oral squamous cell carcinomas frequently appeared in tongue,buccal mucosa,gums and hard palate.A increasing evidence demonstrated that smoking and alcohol consuming are the main factors that contributing to the development of the cancers mentioned above.However,it had been reported that 20% of patients with oropharyngeal and oral squamous cell carcinomas had no experience of drinking alcohol and smoking,suggestting that existence of other potential risk factors.In 1983,Stina Syrjanen revealed the existence of HPV in head and neck cancers.Additionally,HPV has been detected in the oropharynx,hypopharynx and other areas.Hence,the influence of HPV has attracted more and more attention.Numerious researches focused on the correlations between HPV and oropharyngeal squamous cell carcinomas.By contrast,HPV may also reside in oral squamous cell carcinomas and relevant studies were rare.Moreover,the researches based on Chinese population were also few.Hence,we conducted a systematic investigation concerning the relation between HPV and the clinicopathologic parameters and prognosis of oral and oropharyngeal squamous cell carcinomas.Objectives1.To detect the P16 protein expression in oral and oropharyngeal squamous cell carcinomas and analyze the relation between the the clinicopathologic factors and P16 protein expression and then discuss the clinical significance.2.To probe the independent risk factors affecting postoperative outcome by means of careful postoperative follow-up.3.To investigate the sensitivity to chemotherapy of HPV positive oral and oropharyngeal squamous cell carcinomas.Moreover,culturing HPV positive SCC090 cell and HPV negative Fadu cell to verify the conclusion deriving from the retrospective research.Methods1.The collection of detailed clinical information and specimenWe collected ninety-three oropharyngeal squamous cell carcinomas and ninety-five oral squamous cell carcinomas paraffin specimens and sorted the patients' clinical information containing pathology number,gender,age and other clinicopathological parameters from medical records.2.Detection of HPV infection by P16 immunohistochemistry(IHC)All of the 188 cases of specimens were fixed by 10% formalin and embedded by paraffin and each specimen were recut 5 copies.Consequently,376 slices P16 IHC were conducted and the results were determined by two experts in pathology department.In order to enhance the credibility of the expriments,we used PBS instead of primary antibody as control group.3.Collecting patients' prognostic data including survival time,tumor recurrence time and site,metastasis by means of phone,email,letters,outpatient visit and regular follow-up.All patients were followed up until death or in September 2016 and survival was analyzed by OS and DFS.4.Analyzing the sensitivity of radiotherapy or chemotherapy in P16 protein relevant oral and oropharyngealsquamous cell carcinomas.5.Structuring Nomogram model to bring about individual evaluation.6.Conductting CCK8 and flow cytometry to verify the sensitivity to radio/radiochemotherapy of HPV positive SCC090 cell line.7.Statistical methods: c2 test and Fisher exact probability method;Kaplan-Meier and Log-rank methods;Cox proportional hazard model.Results1.The collections of 188 cases with detailed clinicopathological parameters and archivedspecimens were finished and set up SPSS database.2.For OPSCC patients,age,smoking,histotype,N stage and TNM clinical stage were statistically significant when the P16-positive group was compared to their P16-negative counterparts.Additionally,for OSCC patients,the smoking index,N stage and TNM clinical stage were statistically significant compared to the P16-negative group.However,a higher proportion of patients were male,older than 60 years of age,had a smoking index <20 and were T3+T4 stage patients,although significance was not achieved.3.The univariate analysis revealed that histotype,P16 status,T stage and operative treatment were relevant factors that influenced OPSCC prognosis(P<0.05).The OS of P16 positive groups were higher than their counterparts and Log-Rank tests had statistically significant(P=0.004).Multivariate Cox regression analysis showed that T stage,P16 and treatment were independent prognostic factors.Moreover,The univariate analysis revealed that P16 status,T stage and TNM stage were relevant factors that influenced DFS.The DFS of P16 positive groups were higher than their counterparts and Log-Rank tests had statistically significant(P=0.022).Multivariate Cox regression analysis showed that P16 were independent prognostic factors affecting DFS.4.The univariate analysis showed that T stage,TNM stage,age and smoking index were statistically significant(P<0.05).However,no statistical significance was observed between the two curves within the OSCC group(P=0.343).The results of the Cox regression model showed that the smoking index was an independent factor that affected the prognosis of OSCC patients.In addition,univariate analysis revealed that age,smoking,T stage and P16 statuswere relevant factors that influenced DFS of OSCC patients.P16 negative group's DFS was higher than P16 positive group and the two curves were statistical significant(P=0.042).The results of the Cox regression model revealed that age,smoking and P16 protein expression were independent factors that affected DFS of OSCC patients.5.P16 positive expression OPSCC was more sensitive to radiotherapy(RT)and radiochemotherapy(RCT).6.HPV positive SCC090 cell line displayed more sensitive to ?rays after conducting CCK8 and flow cytometry.ConclusionHPV infection is an important pathogenic factor of oral and oropharyngeal squamous cell carcinoma.P16 IHC could be used to be an ideal substitute for HPV detection and the positive rate of P16 expression was significant higher in oropharyngeal squamous cell carcinoma than oral cancers,which was consistent with current reports.P16 positive patients of OPSCC showed improved OS,DFS and radiochemotherapy sensitivity compared to P16 negative groups.However,the results were reversed in OSCC's survival analysis.Hence,P16 expression could be an important indicator to evaluate the prognosis of patients with OOPSCC.We believed that it will provide more powerful theoretical support for clinical treatment and prognosis evaluation through increasing sample capacity and multicenters' cooperation.