下咽頭癌とアルコールおよびたばこ代謝関連遺伝子多型についての検討

博士(医学)琉球大

Epstein-Barr virus and human papillomavirus infections and genotype distribution in head and neck cancers.

To investigate the prevalence, genotypes, and prognostic values of Epstein-Barr virus (EBV) and human papillomavirus (HPV) infections in Japanese patients with different types of head and neck cancer (HNC).HPV and EBV DNA, EBV genotypes and LMP-1 variants, and HPV mRNA expression were detected by PCR from fresh-frozen HNC samples. HPV genotypes were determined by direct sequencing, and EBV encoded RNA (EBER) was examined by in situ hybridization.Of the 209 HNC patients, 63 (30.1%) had HPV infection, and HPV-16 was the most common subtype (86.9%). HPV E6/E7 mRNA expression was found in 23 of 60 (38.3%) HPV DNA-positive cases detected. The site of highest prevalence of HPV was the oropharynx (45.9%). Among 146 (69.9%) HNCs in which EBV DNA was identified, 107 (73.3%) and 27 (18.5%) contained types A and B, respectively, and 124 (84.9%) showed the existence of del-LMP-1. However, only 13 (6.2%) HNCs were positive for EBER, 12 (92.3%) of which derived from the nasopharynx. Co-infection of HPV and EBER was found in only 1.0% of HNCs and 10.0% of NPCs. Kaplan-Meier survival analysis showed significantly better disease-specific and overall survival in the HPV DNA+/mRNA+ oropharyngeal squamous cell carcinoma (OPC) patients than in the other OPC patients (P = 0.027 and 0.017, respectively). Multivariate analysis showed that stage T1-3 (P = 0.002) and HPV mRNA-positive status (P = 0.061) independently predicted better disease-specific survival. No significant difference in disease-specific survival was found between the EBER-positive and -negative NPC patients (P = 0.155).Our findings indicate that co-infection with HPV and EBV is rare in HNC. Oropharyngeal SCC with active HPV infection was related to a highly favorable outcome, while EBV status was not prognostic in the NPC cohort

High-risk type human papillomavirus infection and p16 expression in laryngeal cancer

Abstract Background Oropharyngeal cancers associated with high-risk type human papillomavirus (HR-HPV) infection have better prognosis than virus negative cancers. Similarly, the HPV status in laryngeal cancer (LC) may be associated with better outcome. Methods Samples from 88 patients with LC were investigated using the polymerase chain reaction (PCR) and p16 immunohistochemistry for HR-HPV analysis. The cut-off point for p16 overexpression was diffuse (≥75%) tumor expression with at least moderate (+ 2/3) staining intensity. Results The 5-year cumulative survival (CS) rate was 80.7% in all patients with LC. According to a combination of HR-HPV DNA status and p16 overexpression, subjects with LC were divided into four groups: HR-HPV DNA-positive/p16 overexpression-positive (n = 5, 5.7%; CS = 100%), HR-HPV DNA-positive/p16 overexpression-negative (n = 11, 12.5%; CS =81.8%), HR-HPV DNA-negative/p16 overexpression-positive (n = 0), and HR-HPV DNA-negative/p16 overexpression-negative (n = 72, 81.8%; CS = 79.5%). HR-HPV DNA-positive/p16-positive cases tended to have integrated HPV infection and high viral load, compared with HR-HPV DNA-positive/p16 overexpression-negative cases. Conclusions LC patients with HPV infection and high levels of p16 expression might have an improved survival outcome; however, it is necessary to recruit additional LC cases with HPV infection to determine the definitive characteristics of HPV-mediated LC and estimate survival outcome. These results may contribute to the development of a useful method for selecting patients with a potentially fair response to treatment and ensure laryngeal preservation

Kaplan-Meier curves of disease-specific survival (A) and overall survival (B) according to HPV status in all OPC patients.

<p>Disease-specific survival and overall survival rates were significantly better in HPV DNA-positive/mRNA-positive OPC patients than in HPV DNA-positive/mRNA-negative and HPV DNA-negative/mRNA-negative OPC patients. However, no significant differences in disease-specific survival or overall survival were found between HPV DNA-positive and HPV DNA-negative OPC patients.</p

PCR results for EBV subtyping from the <i>EBNA-3C</i> gene (upper) and for the 30 bp deletion <i>LMP-1</i> variant (middle), and sequence analysis results for wild-type <i>LMP-1</i> and the 30 bp deletion <i>LMP-1</i> variant (lower Upper panel: the primers generated a specific 153 bp fragment from specimens containing type A EBV (samples 1, 2, 4, 5, and positive control PC-2) and a specific 246 bp fragment for specimens containing type B EBV (samples 3, 6, 7, and PC-1).

<p>Middle panel: a specific 316 bp band showed wild-type <i>LMP-1</i> (samples 3, 4, 6, 7, and positive control PC-1) and a specific 286 bp band indicated the 30 bp deletion <i>LMP-1</i> (samples 1, 2, 5, and PC-2). The products of positive controls were confirmed by direct sequencing. Lower panel: the 30 bp deletion sequence of <i>del-LMP-1</i> from wild-type LMP-1 by sequence analysis.</p

Detection results of HPV DNA, mRNA, <i>EBNA</i>-3C, the 30 bp deletion LMP-1 variant, and <i>EBER</i> in different types of head and neck cancer.

<p>HPV, human papillomavirus; <i>EBNA</i>, Epstein-Barr nuclear antigen; <i>LMP-1</i>, latent membrane protein-1; <i>EBER</i>, Epstein-Barr virus encoded RNA.</p><p>* Positive rate of the variables in different subsites of head and neck cancer.</p>#<p>Positive rate of HPV mRNA in HPV DNA-positive patients; 3 cases with HPV DNA-positive derived from oropharynx, hypopharynx, and larynx were not sufficient to be examined for RNA extraction and <i>E6/E7</i> mRNA expression.</p><p>Detection results of HPV DNA, mRNA, <i>EBNA</i>-3C, the 30 bp deletion LMP-1 variant, and <i>EBER</i> in different types of head and neck cancer.</p

EBV encoded RNA in in situ hybridization.

<p>Micrograph A: most of the neoplasm cells were positive (nasopharyngeal carcinoma, ×100, bar = 100 µm), as indicated by the arrow at high magnification (×200). Micrograph B: although some positive lymphocytes are apparent (arrow), no neoplasm cells were positive. Micrographs C and D: Sections stained with hematoxylin and eosin (HE).</p