Abstract 4693: Characterization of the differential mechanisms of CDKN2A inactivation in oral tongue squamous cell carcinomas and correlation with patient outcome.
Authors: Lim Annette M. , Do Hongdo , Young Richard J. , Wong Stephen Q. , Angel Christopher , Collins Marnie , Takano Elena , Corry June , Fox Stephen B. , Wiesenfeld David , Kleid Stephen , Sigston Elizabeth , Lyon Bernard , Rischin Danny , Dobrovic Alexander , Solomon Benjamin 
Published: August 2014

Abstract
Background. CDKN2A inactivation has been reported to be a frequent event in head and neck squamous cell carcinomas that correlates with worse patient outcome. The differential mechanisms of CDKN2A inactivation in oral tongue squamous cell carcinomas (OTSCC) have been poorly characterised. We sought to determine the frequency of potential mechanisms of CDKN2A inactivation in OTSCC and explore their impact on patient outcome.
Material and methods. We investigated a cohort of 134 OTSCC patients treated between 2002 to 2008 with combinations of surgery, radiotherapy and chemotherapy, who have been comprehensively annotated for patient/tumour characteristics and outcome. 134 formalin fixed paraffin embedded blocks representative of primary tumours prior to treatment were collected. DNA was extracted from 115 patient blocks and a tissue microarray was made representing 123 patient samples. We assessed p16/CDKN2A expression by immunohistochemistry (IHC), promoter methylation status by methylation-sensitive high resolution melting, mutation status by Sanger sequencing, gene copy number variation by fluorescence in-situ hybridisation and correlated each of these with patient outcome.
Results. The majority of OTSCC demonstrated loss of p16 expression (107/118, 90.7%), assessed by IHC. The frequency of CDKN2A inactivating mutations was 20.3% (21/103), homozygous CDKN2A loss was 4.1% (7/98), hemizygous CDKN2A loss 30.6% (30/98), and CDKN2A promoter methylation &gt;10% was 17.7% (20/113). 34/107(31.8%) p16 IHC negative tumours had no evidence of CDKN2A mutation, copy number variation or promoter methylation identified. Of eleven p16 IHC positive patients, only two demonstrated evidence of HPV type 16 and 33 on PCR. No correlation was identified between any of the mechanisms of CDKN2A inactivation and clinicopathological features. Smoking status and age did not influence the frequency of any mechanism of CDKN2A inactivation or patient outcome. There was a non-significant trend for worse outcome for p16 IHC negative patients versus IHC positive patients (HR=0.65, 95% CI=0.23-1.79, p=0.40). No significant relationship between mechanisms of p16 inactivation and patient outcome were found.
Conclusion. Loss of p16 is a frequent event in oral tongue squamous cell carcinomas that can occur through gene copy number variation, mutation, promoter methylation or through other unidentified mechanisms. We identified no correlation between mechanisms of p16 loss and clinicopathological characteristics or patient outcome.
Citation Format: Annette M. Lim, Hongdo Do, Richard J. Young, Stephen Q. Wong, Christopher Angel, Marnie Collins, Elena Takano, June Corry, Stephen B. Fox, David Wiesenfeld, Stephen Kleid, Elizabeth Sigston, Bernard Lyon, Danny Rischin, Alexander Dobrovic, Benjamin Solomon. Characterization of the differential mechanisms of CDKN2A inactivation in oral tongue squamous cell carcinomas and correlation with patient outcome. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4693. doi:10.1158/1538-7445.AM2013-4693