Shinya Agena

Viral load, physical status, and E6/E7 mRNA expression of human papillomavirus in head and neck squamous cell carcinoma

The purpose of this study was to determine prospectively both human papillomavirus (HPV) load and physical status in different types of head and neck squamous cell carcinoma (HNSCC).

A comprehensive evaluation of human papillomavirus positive status and p16INK4a overexpression as a prognostic biomarker in head and neck squamous cell carcinoma

Head and neck squamous cell carcinoma (HNSCC) patients with human papillomavirus (HPV) infection have better prognosis than those without HPV infection. Although p16INK4a expression is used as a surrogate marker for HPV infection, there is controversy as to whether p16INK4a reliably indicates HPV infection. Here, to evaluate the accuracy of p16INK4a expression for determining HPV infection and the prognostic value of HPV infection and p16INK4a expression for HNSCC survival, especially oropharyngeal squamous cell carcinoma (OPSCC) survival, 150 fresh-frozen HNSCC samples were analyzed for HPV DNA, E6/E7 mRNA and p16INK4a expression by polymerase chain reaction and immunohistochemistry. p16INK4a expression was scored from 0 to 4 according to the percentage of p16INK4a-positive cells, with overexpression defined as >40% positive cells. Of the 150 tumor samples tested, 10 tumors were nasopharyngeal, 53 oropharyngeal, 39 hypopharyngeal, 24 laryngeal and 24 were located in the oral cavity. HPV DNA was detected in 47 (31.3%) samples, but only 21 also exhibited HPV mRNA expression. Inter-rater agreement was low between p16INK4a expression and HPV DNA presence and between p16INK4a expression and HPV mRNA expression, but was good between the combination of HPV DNA status and p16INK4a overexpression and HPV mRNA expression. Three-year recurrence-free survival was significantly higher for OPSCC patients who were HPV DNA-positive than for OPSCC patients who were HPV DNA-negative (P=0.008) and for OPSCC patients over-expressing p16INK4a than for without overexpressing p16INK4a (P=0.034). Multivariate analysis revealed that T1-3 stage and the combination of HPV DNA positivity and p16INK4a overexpression predicted significantly better recurrence-free survival. This combination is a more accurate marker for active HPV infection in HNSCC than HPV DNA status or general p16INK4a-positive status alone and offers a useful and reliable method for detecting and determining the prognosis of HPV-related HNSCC.

Prognostic value of human papillomavirus and squamous cell carcinoma antigen in head and neck squamous cell carcinoma

To clarify the synergistic influence of human papillomavirus (HPV) status and squamous cell carcinoma antigen (SCCA) mRNA expression on head and neck squamous cell carcinoma (HNSCC) prognosis, HPV DNA presence and SCCA1 and SCCA2 mRNA expression were determined by PCR and quantitative real‐time RT‐PCR, respectively, in 121 patients with primary HNSCC who were receiving curative treatment. HPV DNA was detected in 28.1% (34/121) of HNSCC cases, and only high‐risk types (HPV‐16, HPV‐33, HPV‐35 and HPV‐58) were observed. Positive HPV status showed a significantly better prognosis than negative HPV status (P = 0.022). An elevated SCCA2/SCCA1 mRNA ratio was an independent predictor of disease recurrence (P = 0.004). In addition, HPV‐negative patients with a high SCCA2/SCCA1 ratio (>0.27) had a significantly lower recurrence‐free survival rate than HPV‐negative patients with a low SCCA2/SCCA1 ratio (P < 0.011). Our findings revealed that both HPV status and the SCCA2/SCCA1 mRNA ratio are independently associated with prognosis in HNSCC. Patients with both a HPV‐negative status and a high SCCA2/SCCA1 ratio might need intensified treatment and rigorous follow up after treatment because of the high risk of recurrence.

Human papillomavirus load and physical status in sinonasal inverted papilloma and squamous cell carcinoma.

BACKGROUND This study investigated prospectively the role of human papillomavirus (HPV) in paranasal inverted papilloma (IP). METHODS HPV presence and viral load and physical status of HPV-16 were examined by polymerase chain reaction-based methods using fresh frozen samples obtained from 13 patients with IP (IP group), 11 with squamous cell carcinoma in the maxillary sinus (SCC group) and 39 with chronic inflammatory lesions (inflammatory group). RESULTS The presence of the HPV genome was detected in 46.1%, 27.3% and 7.6% of patients in the IP, SCC and inflammatory groups, respectively. The IP group showed significantly higher HPV-positive rates than the inflammatory group. All types of HPV detected were high-risk HPV, especially HPV-16. The relative HPV-16 copy numbers varied from 2.5 to 1524.1 per 50 ng genomic DNA. The viral load was higher in the IP and SCC groups than in the inflammatory group. In the IP group, no significant relationship was found between HPV-16 viral load and clinical characteristics, or between physical status and clinical characteristics. One patient with IP and concomitant squamous cell carcinoma, however, showed high viral load and integration. CONCLUSIONS HPV infection is involved in the pathogenesis of IP, and high viral load and integration of HPV have an important role in malignant lesion in association with IP.

Epstein-Barr Virus and Human Papillomavirus Infections and Genotype Distribution in Head and Neck Cancers

Objective 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). Methods and Materials 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. Results 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). Conclusions 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.

Prediction of concurrent chemoradiotherapy outcome in advanced oropharyngeal cancer

The aim of this study was to investigate human papillomavirus (HPV) infection as a predictor of concurrent chemoradiotherapy (CCRT) response and indicator of planned neck dissection (PND) for patients with advanced oropharyngeal squamous cell carcinoma (OPSCC; stage III/IV). Overall, 39 OPSCC patients (32 men, 7 women; median age 61 years, range 39–79 years) were enrolled. The primary lesion and whole neck were irradiated up to 50.4 Gy, and subsequently the primary site and metastatic lymph nodes were boosted with a further 16.2 Gy. Although several chemotherapy regimens were employed, 82.1% of OPSCC patients received the combination of nedaplatin and 5-fluorouracil. HPV-related OPSCC (16 cases) was defined as both HPV DNA-positive status by polymerase chain reaction and p16INK4a overexpression by immunohistochemistry. Patients with N2 and N3 disease received PND 2–3 months after CCRT completion. Compared to non-responders, CCRT responders showed significantly lower nodal stage (N0 to N2b) and HPV-positive status in univariate analysis. Patients with HPV-related OPSCC had longer time to treatment failure (TTF) than those with HPV-unrelated OPSCC (p=0.040). Three-year TTF was 81.3 and 47.8% in the HPV-related and HPV-unrelated groups, respectively. There were also significant differences in disease-free survival (DFS) between the two OPSCC patient groups (p=0.042). Three-year DFS was 93.8 and 66.7% in patients with HPV-related and HPV-unrelated OPSCC, respectively. Multivariate logistic analysis showed a lower risk of TTF event occurrence in HPV-related OPSCC (p=0.041) than in HPV-unrelated OPSCC. Thus, HPV testing in addition to nodal stage was useful for predicting CCRT response, especially in advanced OPSCC. Because patients who received PND showed moderate locoregional control, PND is an effective surgical procedure for controlling neck lesions in patients with advanced HPV-unrelated disease.

Lymphoepithelial carcinoma in parotid gland related to EBV infection: A case report

Lymphoepithelial carcinoma commonly occurs at the nasopharynx and rarely occurs at other sites in the head and neck region. It is well known to occur at limited patients of local area as Asia or Arctic Circle. Related to this point, it is pointed out that this tumor has strong relation with Epstein-Barr Virus (EBV) infection. In this time, we experienced to treat lymphoepithelial carcinoma with metastatic cervical lymph nodes occurring at parotid gland. The morbidity ratio of this tumor is less than one percent of all parotid gland tumors. Moreover, we proved the infection of EBV to tumor cell by in situ hybridization (ISH). Incidentally, because it is considered that this tumor has well sensitivity against irradiation or anti-tumor drugs, prognosis of this tumor is better than that of other head and neck tumors with different pathological type. Actually, we tried to perform chemotherapy twice in (Nedaplatin (CDGP) 60mg/m2×day 2 and 5-FU 600mg/m2×day 5) and to irradiate about 70Gy dose against parotid gland and cervical lymph nodes. It could not find local recurrence or metastasis as of now after five years from treatment.

Detection of human papillomavirus in branchial cleft cysts

High-risk human papillomavirus (HPV) DNA has been reported to be present in branchial cleft cysts, but further information is required to clarify the role of HPV infection in branchial cleft cysts. The presence of HPV, the viral load and the physical statuses in samples from six patients with branchial cleft cysts were investigated using the polymerase chain reaction (PCR), quantitative PCR, in situ hybridization (ISH) using HPV DNA probes and p16INK4a immunohistochemical analysis. High-risk type HPV-16 DNA was identified in four of the six branchial cleft cysts analyzed. Of the HPV-positive branchial cleft cysts, three exhibited mixed-type integration of HPV. HPV DNA was distributed among the basal-to-granular layers of the cystic wall in ISH analysis, and p16INK4a was weakly expressed in the nuclei and cytoplasm of the same layers in patients with integration. ISH revealed that one patient with episomal-type infection exhibited HPV DNA in the cyst wall and did not express p16INK4a. Two patients without evidence of HPV infection exhibited weak p16INK4a expression in the superficial cyst-lining cells of branchial cleft cysts. These results indicate that infection with high-risk HPV types may be common in branchial cleft cysts. In addition, p16INK4a is not a reliable surrogate marker for HPV infection in branchial cleft cysts.

An Invention on Operative Scar of the Parotid Gland Tumor

Objective: A parotid gland tumor is an important disease for otolaryngologists to be able to treat through surgical technique in Japan. It is necessary to preserve facial nerves completely and to not make a prominent postoperative incisional scar of the skin, particularly in the case of benign tumors. Method: We experienced 42 cases of parotid gland tumors in 2011. We tried to diagnose by CT scan, MRI imaging, and fine needle aspiration at preoperation. Results: Benign tumors made up 37 cases, with the rest malignant tumors. We could identify a benign tumor in almost all cases at preoperation. In case of benign tumor, permanent facial nerve palsy was not found, except the schwwanoma in facial nerve at postoperation. Moreover, when we removed the tumor, the following steps were taken: 1) A skin incision was established with minimum length. 2) Incising the preauricular area, we drew a cutting line on the external auditory canal inside a tragus. 3) The sigmoid curve in the postauricular area was designed as gently as possible. The cervical incision line was drawn along the digastic muscle. Conclusion: It is clear that during operations for the facial nerves, complete extraction of the tumors and minimum skin incision in the parotid gland tumor yield compatible results.

Residual Lymph Node Metastasis in Planned Neck Dissection

Objective: Clarify the effectiveness of planned neck dissection (PND) after concurrent chemoradiotherapy (CCRT), we investigated residual lymph node metastasis; imaging tests such as positron emission tomography (PET), computed tomography (CT), and ultrasonography at pre- and post-CCRT; and the adverse effects of PND. Method: CCRT combined with PND was performed on 13 patients with oropharyngeal carcinoma, 4 with hypopharyngeal carcinoma, and 2 with laryngeal carcinoma. All primary lesions showed complete response to CCRT. Histopathologic examination of neck specimens and imaging tests were employed to evaluate the effect of CCRT. Results: Eight of 19 patients (47%) had viable carcinoma in their lymph nodes. Patients with poorly differentiated carcinoma in the primary lesion tended to have residual lymph nodes. Most of the residual neck specimens belonged to the level III or level IV lymph node group. Major complications of PND were prolonged laryngeal edema, dysphasia, and disturbance of shoulder movement. Tracheotomies were performed in 42% of all cases. Positive and negative predictive values of PET examination before PND were 50% and 73%, respectively. None of the patients with PND experienced local or lymph node recurrence. Conclusion: Residual lymph nodes were frequently observed in head and neck carcinomas. Of the imaging tests, PET was sufficient for detecting residual regions. Although PND is a useful tool, related complications were frequently observed. A more accurate detection system is needed to reduce unnecessary neck dissections.