Gérard Agius

Human papillomavirus genotype distribution in oropharynx and oral cavity cancer in France—The EDiTH VI study

BACKGROUND The incidence of oropharyngeal cancers has gradually increased over the last decades. Recent studies suggest an association between human papillomavirus (HPV) infection and several head and neck cancers, especially oropharyngeal and oral cavity invasive carcinomas. OBJECTIVES The objective was to assess the overall and type specific HPV prevalence in oropharyngeal and oral cavity carcinomas in France. STUDY DESIGN Paraffin-embedded tumour specimens were retrospectively collected in 12 French centres and centrally tested for HPV detection and genotyping (INNO-LiPA assay). RESULTS A total of 523 cases (77% males) were collected, among which 60% were oropharyngeal and 40% oral cavity carcinomas. The most frequent anatomical sites were tonsil (58.9%) and base of tongue (13.7%) for the oropharynx and floor of mouth (41.1%) and oral tongue (38.3%) for the oral cavity. Overall HPV prevalence was 46.5% in oropharyngeal carcinomas and 10.5% in oral cavity carcinomas and was higher in female than in male cases (63.5% vs 42.2% in oropharynx and 17.2% vs 8.0% in oral cavity). About 95% of HPV-positive cases were infected by a single HPV type. HPV 16 was the most prevalent type and was found in 89.7% and 95.5% of HPV-positive oropharyngeal and oral cavity carcinoma cases, respectively. All other HPV types had prevalence below 5%. CONCLUSIONS Our results indicate that HPV is common among oropharyngeal and oral cavity carcinoma cases in France and emphasize the predominance of HPV 16. The potential benefit of HPV vaccination on the occurrence of head and neck carcinomas should be further evaluated.

Clinical Characteristics Are Similar across Type A and B Influenza Virus Infections

Background Studies that aimed at comparing the clinical presentation of influenza patients across virus types and subtypes/lineages found divergent results, but this was never investigated using data collected over several years in a countrywide, primary care practitioners-based influenza surveillance system. Methods The IBVD (Influenza B in Vircases Database) study collected information on signs and symptoms at disease onset from laboratory-confirmed influenza patients of any age who consulted a sentinel practitioner in France. We compared the clinical presentation of influenza patients across age groups (0–4, 5–14, 15–64 and 65+ years), virus types (A, B) and subtypes/lineages (A(H3N2), pandemic A(H1N1), B Victoria, B Yamagata). Results Overall, 14,423 influenza cases (23.9% of which were influenza B) were included between 2003–2004 and 2012–2013. Influenza A and B accounted for over 50% of total influenza cases during eight and two seasons, respectively. There were minor differences in the distribution of signs and symptoms across influenza virus types and subtypes/lineages. Compared to patients aged 0–4 years, those aged 5–14 years were more likely to have been infected with type B viruses (OR 2.15, 95% CI 1.87–2.47) while those aged 15–64 years were less likely (OR 0.83, 95% CI 0.73–0.96). Males and influenza patients diagnosed during the epidemic period were less likely to be infected with type B viruses. Conclusions Despite differences in age distribution, the clinical illness produced by the different influenza virus types and subtypes is indistinguishable among patients that consult a general practitioner for acute respiratory infections.

HPV and head and neck cancer

Head and neck cancer is frequent worldwide and oropharyngeal locations are presently sharply on the increase, in relation with an increasing incidence of oropharyngeal infection by oncogenic type-16 human papillomavirus (HPV). The clinical and biologic profile of these patients is distinct from that of other oropharyngeal carcinoma patients, with earlier onset, cystic cervical nodes and basaloid carcinoma histopathology. Detection of intratumoral viral DNA is essential to confirm the role of HPV, and E6/E7 mRNA expression is the most relevant indicator for stratification. Several methods can reveal intratumoral oncogenic HPV DNA, but PCR with hybridization is the most sensitive and most widely used. According to several reports, prognosis in terms of survival and locoregional control is better in HPV-positive oropharyngeal carcinoma than in oropharyngeal carcinoma associated with smoking and alcohol consumption. The future lies in vaccination, but further studies will determine whether the rate of oropharyngeal carcinoma falls in women vaccinated against cervical cancer.

Human papillomavirus genotype distribution in tonsil cancers.

BackgroundThe incidence of tonsil cancers has increased in several countries. French data on HPV prevalence in tonsil cancers are scarce. The objective of this study was thus to assess the overall and type specific HPV prevalence in tonsil histological samples.MethodsThis French retrospective multicenter study involved 12 centres located throughout the country. Were included 185 histological samples collected from year 2000 to 2009 with a validated diagnosis of tonsil invasive carcinomas. HPV prevalence was studied according to gender, age and histological type of cancer.ResultsOverall HPV prevalence was 57% in tonsil cancers. Mean age of diagnosis was comparable in HPV positive tonsils cases (60 ± 11.2) and HPV negative tonsil cases (59 ± 9.6). HPV prevalence was significantly higher in female than in male cases (28/35 versus 78/150 in tonsil cases, respectively, P = 0.003). About 53% of tonsil cases were infected by a single HPV type. Only eight (4%) samples were infected by more than one HPV type. Among HPV positive samples, HPV 16 was found in 89% of tonsil cases. All other HPV types had prevalence below 5%.ConclusionsOur results indicate that HPV is common in tonsil carcinomas and emphasize the predominant role of HPV 16.

Nucleotide sequence analysis of human T-cell lymphotropic virus type I pX and LTR regions from patients with Sicca syndrome

Human T‐cell lymphotropic virus type I (HTLV‐I) is associated with adult T‐cell leukemia (ATL) and tropical spastic paraparesis/HTLV‐I‐associated myelopathy (TSP/HAM). Other inflammatory disorders may occur in HTLV‐I‐infected patients, such as sicca syndrome resembling Sjögrens syndrome. The sicca syndrome may be the unique clinical manifestation of HTLV‐I infection, but is associated frequently with TSP/HAM, which could suggest that sicca syndrome might be an early event in disease progression to TSP/HAM in some cases. We investigated whether peculiar pX and LTR mutations could be related to sicca syndrome, or might argue the existence of clinical progression to TSP/HAM. pX, especially pXI, pXII, and pXIV ORFs corresponding to Tax cytotoxic T‐lymphocyte epitopes, and LTR regions from Caribbean patients who have sicca syndrome with or without TSP/HAM, ATL patients, and healthy carriers were sequenced. The sequences were aligned and compared with ATK‐1 prototype and published sequences. LTR sequences exhibited 1.5–2.4% of divergence with ATK‐1. pX‐sequenced regions showed a lower homology within p12I encoding sequences. Only few mutations were found within functionally important regions, but were not associated specifically with the clinical status. Finally, no mutations that could be related to sicca syndrome or argue the existence of clinical progression to TSP/HAM were found. It would be of interest to study the clinical evolution of HTLV‐I–sicca syndrome in patients and to determine HTLV‐I sequences from peripheral blood and salivary glands at different stages. J. Med. Virol. 59:245–255, 1999.

Disseminated varicella with multiorgan failure in an immunocompetent adult.

A case of fulminant disseminated varicella is reported in a 28‐year‐old immunocompetent man. He developed hepatitis, severe pneumonia, rhabdomyolysis and disseminated intravascular coagulation, followed by encephalopathy and multiorgan failure despite acyclovir therapy. He spent a total of 3.5 months in intensive care and rehabilitation units. Real‐time PCR yielded a rapid diagnosis of varicella‐zoster virus (VZV) infection and was used to monitor plasma viral load for 56 days. Plasma viral load peaked at 7.1 log10/ml on day 4 after symptom onset, then gradually declined and became undetectable after between 1 and 2 months; viral load in lung fluid followed a similar pattern. The glycoprotein E variant associated with increased VZV virulence was not detected, and the VZV thymidine kinase gene bore no major mutations associated with acyclovir resistance. This case serves as a reminder that varicella can be life‐threatening in adults and that vaccination of individuals at risk remains essential. J. Med. Virol. 81:747–749, 2009

Immortalized sebocytes can spontaneously differentiate into a sebaceous-like phenotype when cultured as a 3D epithelium

Abstract:  Sebocytes originate from the same lineage as keratinocytes, and both cell types may have similarities in terms of growth and differentiation. We were interested in studying the behaviour of human sebocytes when cultured in conditions validated for epidermal reconstruction. For this purpose, we established a HPV16‐E6/7‐immortalized human sebocyte cell line (SEBO662) growing in keratinocyte defined media. Postconfluent SEBO662 cells in monolayers express the early sebocyte marker, cytokeratin 7 (K7), do not express Epithelia Membrane Antigen (EMA) and do not exhibit strong lipogenic activity. However, when placed at the air–liquid interface, SEBO662 multilayers spontaneously differentiate into a sebaceous‐like structure as shown by the strong polarized expression of the late sebaceous marker EMA, the overexpression of some lipogenic markers and lipid production on the upper side of the epithelium. This work highlights the value of simple 3D models for exhibiting spontaneous differentiation and polarization.

Human papillomavirus 16 oncoprotein E7 stimulates UBF1-mediated rDNA gene transcription, inhibiting a p53-independent activity of p14ARF.

High-risk human papillomavirus oncoproteins E6 and E7 play a major role in HPV-related cancers. One of the main functions of E7 is the degradation of pRb, while E6 promotes the degradation of p53, inactivating the p14ARF-p53 pathway. pRb and p14ARF can repress ribosomal DNA (rDNA) transcription in part by targeting the Upstream Binding Factor 1 (UBF1), a key factor in the activation of RNA polymerase I machinery. We showed, through ectopic expression and siRNA silencing of p14ARF and/or E7, that E7 stimulates UBF1-mediated rDNA gene transcription, partly because of increased levels of phosphorylated UBF1, preventing the inhibitory function of p14ARF. Unexpectedly, activation of rDNA gene transcription was higher in cells co-expressing p14ARF and E7, compared to cells expressing E7 alone. We did not find a difference in P-UBF1 levels that could explain this data. However, p14ARF expression induced E7 to accumulate into the nucleolus, where rDNA transcription takes place, providing an opportunity for E7 to interact with nucleolar proteins involved in this process. GST-pull down and co-immunoprecipitation assays showed interactions between p14ARF, UBF1 and E7, although p14ARF and E7 are not able to directly interact. Co-expression of a pRb-binding-deficient mutant (E7C24G) and p14ARF resulted in EC24G nucleolar accumulation, but not in a significant higher activation of rDNA transcription, suggesting that the inactivation of pRb is involved in this phenomenon. Thus, p14ARF fails to prevent E7-mediated UBF1 phosphorylation, but could facilitate nucleolar pRb inactivation by targeting E7 to the nucleolus. While others have reported that p19ARF, the mouse homologue of p14ARF, inhibits some functions of E7, we showed that E7 inhibits a p53-independent function of p14ARF. These results point to a mutually functional interaction between p14ARF and E7 that might partly explain why the sustained p14ARF expression observed in most cervical pre-malignant lesions and malignancies may be ineffective.

Infections à papillomavirus humains (HPV) des voies aéro-digestives supérieures (VADS)

Resume En dehors de la sphere genitale, les papillomavirus humains (HPV) sont responsables d’infections des voies aero-digestives superieures (VADS) dont l’histoire naturelle est mal comprise. En marge des manifestations benignes (papillomes, verrues, condylomes), connues depuis longtemps, les HPV a haut risque (HR) sont egalement impliques dans 25% des cancers de l’oropharynx toutes localisations confondues et dans au moins 50% des cancers de l’amygdale. HPV16 est le genotype responsable dans pres de 90% des cas. Concernant les autres localisations des cancers des VADS, l’existence d’un lien reste incertaine. L’incidence des cancers des VADS positifs pour la detection d’HPV a augmente dans les pays occidentaux alors que celle des cancers negatifs pour HPV reste stable ou diminue. Les cancers de l’oropharynx positifs pour HPV surviennent preferentiellement chez des patients plus jeunes, ne presentant generalement pas les facteurs de risque classiques que sont le tabac et l’alcool, et sont associes a des facteurs de risque d’infection sexuellement transmissible, notamment les rapports sexuels oro-genitaux. Ils repondent mieux a la radio- et chimiotherapie et sont de meilleur pronostic que les cancers negatifs pour HPV. Ils presentent des profils moleculaires et genetiques distincts de ceux des tumeurs negatives pour HPV refletant les consequences de l’expression des oncoproteines virales E6 et E7. Connaitre le statut HPV des tumeurs deviendra necessaire pour optimiser les choix therapeutiques, et est d’ores et deja indispensable dans les essais therapeutiques. Le debat de la vaccination des garcons, deja pratiquee dans certains pays, est relance mais les vaccins devront faire la preuve de leur efficacite pour prevenir les infections orales et a terme induire une diminution de l’incidence des cancers oropharynges.

Epithelial to mesenchymal transition and HPV infection in squamous cell oropharyngeal carcinomas: the papillophar study.

Background:Human Papillomavirus (HPV) infection is recognised as aetiological factor of carcinogenesis in oropharyngeal squamous cell carcinomas (OPC). HPV-related OPC respond better to treatments and have a significantly favourable outcome. Epithelial to mesenchymal transition (EMT) implicated in tumour invasion, is a hallmark of a poor prognosis in carcinomas.Methods:We have studied the relationship of EMT markers (E-cadherin, β-catenin and vimentin) with HPV infection (DNA and E6/E7 mRNA detection), p16INK4a expression and survival outcomes in a cohort of 296 patients with OPC.Results:Among the 296 OPSSC, 26% were HPV positive, 20.3% had overt EMT (>25% of vimentin positive tumour cells). Lower E-cadherin expression was associated with a higher risk of distant metastasis in univariate (P=0.0110) and multivariate analyses (hazard ratios (HR)=6.86 (1.98; 23.84)). Vimentin expression tends towards worse metastasis-free survival (MFS; HR=2.53 (1.00; 6.41)) and was an independent prognostic factor of progression-free survival (HR=1.55 (1.03; 2.34)).Conclusions:There was a non significant association of EMT with HPV status. This may be explained by a mixed subpopulation of patients HPV positive with associated risk factors (HPV, tobacco and alcohol). Thus, the detection of EMT in OPC represents another reliable approach in the prognosis and the management of OPC whatever their HPV status.