Lubomir P. Turek

Age, sexual behavior and human papillomavirus infection in oral cavity and oropharyngeal cancers.

There are few well‐established patient risk factors associated with human papillomavirus (HPV) infection in cancers of the oral cavity and oropharynx. The purpose of this study was to determine if there were significant different risk factors and tumor characteristics between HPV‐positive and HPV‐negative cancer cases. HPV was evaluated in cancer tissue and exfoliated oral cells of 193 oral cavity/oropharynx cancer patients using PCR and direct DNA sequencing. A patient questionnaire collected information about risk factors, sexual practices and medical history. The prevalence of HPV high‐risk (HR) types was 20% in cancer cases. Three types were identified: HPV‐16 (87%), HPV‐18 (3%) and HPV‐33 (11%). Risk factors for HPV‐HR included younger age (≤ 55 years vs. > 55 years; adjusted OR = 3.4; 95% CI = 1.6–7.3) and younger‐age cases who had more lifetime sex partners (adjusted OR = 3.8; 95% CI = 1.4–10.1), practiced oral‐genital sex (adjusted OR = 4.3; 95% CI = 1.8–10.4) or oral‐anal sex (adjusted OR = 19.5; 95% CI = 3.4–113). Compared to HPV‐negative cancers, HPV‐HR cancers were more likely to have a positive HPV‐HR exfoliated oral cytology test (adjusted OR = 7.8; 95% CI = 3.4–18.4), later stage (adjusted OR = 3.0), nodal involvement (adjusted OR = 4.1) and advanced grade (adjusted OR = 3.0). This study shows new evidence that the prevalence of oncogenic mucosal HPV is higher in younger‐age oral cavity/oropharynx cancer cases whose sexual practices are typically associated with sexual transmission of the virus. HPV detection also appears to be an indicator of advanced disease characteristics that may require different clinical treatment for this subset of patients. An exfoliated oral cytology test for HPV was a significant predictor of HR types in the cancers, suggesting that an oral rinse may provide an early biomarker of infected tumors.

Human papillomavirus infection as a prognostic factor in carcinomas of the oral cavity and oropharynx

Although studies have established human papillomaviruses (HPVs) as a risk factor for oral and oropharyngeal cancer, it is not clear whether viral infection affects survival in head and neck malignancies. This investigation examined the relationship between HPV and survival in carcinomas of the oral cavity and oropharynx. Formalin‐fixed, paraffin‐embedded tumor specimens from 139 newly diagnosed cases were tested for HPVs by PCR and DNA sequencing. Patient and tumor characteristics were obtained from questionnaires, pathology reports and cancer registries. Odds ratios (ORs) and relative risks (RRs) were based on logistic and Cox regression models, respectively. HPVs were detected in 21% of the tumors; 83% were HPV‐16. Greater risk of HPV infection was associated with males (OR = 2.9, 95% CI = 1.0–8.6), a history of oral‐genital sex (OR = 4.2, 95% CI = 1.5–11.7), and oropharyngeal tumors (OR = 10.4, 95% CI = 3.5–31.2). As tobacco usage increased, the odds of HPV detection decreased (OR = 0.97/pack‐year, 95% CI = 0.96–0.99). HPV infected patients had better overall survival (RR = 0.3, 95% CI = 0.1–0.8) than those with HPV‐negative tumors. There was an interaction between gender and HPV for overall (p = 0.05) and disease‐specific (p = 0.03) survival that suggested that HPV infected males had better prognosis than HPV‐negative males, but this was not the case among females. HPV status was identified as an independent prognostic factor in oral and oropharyngeal cancers. This result appeared to be gender‐specific, suggesting the need for further study of the interaction between HPV and gender on survival.

Fundamental Differences in Cell Cycle Deregulation in Human Papillomavirus–Positive and Human Papillomavirus–Negative Head/Neck and Cervical Cancers

Human papillomaviruses (HPV) are associated with nearly all cervical cancers, 20% to 30% of head and neck cancers (HNC), and other cancers. Because HNCs also arise in HPV-negative patients, this type of cancer provides unique opportunities to define similarities and differences of HPV-positive versus HPV-negative cancers arising in the same tissue. Here, we describe genome-wide expression profiling of 84 HNCs, cervical cancers, and site-matched normal epithelial samples in which we used laser capture microdissection to enrich samples for tumor-derived versus normal epithelial cells. This analysis revealed that HPV(+) HNCs and cervical cancers differed in their patterns of gene expression yet shared many changes compared with HPV(-) HNCs. Some of these shared changes were predicted, but many others were not. Notably, HPV(+) HNCs and cervical cancers were found to be up-regulated in their expression of a distinct and larger subset of cell cycle genes than that observed in HPV(-) HNC. Moreover, HPV(+) cancers overexpressed testis-specific genes that are normally expressed only in meiotic cells. Many, although not all, of the hallmark differences between HPV(+) HNC and HPV(-) HNC were a direct consequence of HPV and in particular the viral E6 and E7 oncogenes. This included a novel association of HPV oncogenes with testis-specific gene expression. These findings in primary human tumors provide novel biomarkers for early detection of HPV(+) and HPV(-) cancers, and emphasize the potential value of targeting E6 and E7 function, alone or combined with radiation and/or traditional chemotherapy, in the treatment of HPV(+) cancers.

Transcriptional regulation of the human papillomavirus-16 E6−E7 promoter by a keratinocyte-dependent enhancer, and by viral E2 trans-activator and repressor gene products: implications for cervical carcinogenesis

The transcriptional promoter of the candidate E6‐E7 transforming gene region of human papillomavirus (HPV)‐16 (P97) was active in transiently transfected cervical carcinoma cells when linked to the HSV‐1 tk or bacterial cat genes. Sequences 5′ to P97 contain a short enhancer element responding to cellular factor(s) in uninfected human foreskin keratinocytes and in cervical carcinoma cells, but not in human or animal fibroblasts. The E2 trans‐activator products of HPV‐16 or of the related bovine papillomavirus (BPV)‐1 further elevated HPV‐16‐driven transcripts in co‐transfections, and required the presence of E2‐binding ACC(N)6GGT cores in cis. A ‘short E2’ C‐terminal repressor gene product (sE2) of HPV‐16 or the BPV‐1 sE2 repressor not only inhibited viral E2 trans‐activation, but also suppressed enhancer response to keratinocytic factors. Suppression by the sE2 products was abolished by deletion of the E2‐binding cores in cis or by a mutation in the sE2 DNA binding domain. The keratinocyte‐dependent enhancer is likely to contribute to the epithelial cell tropism of HPV‐16, and may direct persistent E6‐E7 gene transcription in response to cellular factors in cervical carcinoma cells in which the viral E2 genes are inactive.

Human papillomavirus and risk of oral cancer

Although human papillomavirus (HPV), a sexually transmitted virus, is established as a necessary cause for more than 95% of cervical carcinomas, the association with oral squamous cell carcinoma is less well delineated. The purpose of this study was to determine the frequency and types of HPV in squamous cells of a group of patients with newly diagnosed oral or pharyngeal cancer (n = 93) compared with an age‐and gender‐frequency‐matched control group of patients with no history of oral cancer (n = 205). HPV was evaluated from a mouth rinse collection of cells in the oral cavity and tested by 32P‐labeled HPV generic probes and DNA sequencing for HPV types. HPV was identified in 15% of the oral cancer cases but in fewer than 5% of the controls (P < .05). The risk of cancer associated with HPV infection was independent of tobacco and alcohol use (adjusted odds ratio [OR] = 3.70; 95% confidence interval [CI]: 1.47–9.32; P < .05). HPV types included similar and other types not identified previously in the genital tract. There was no statistically significant increased risk of cancer among former tobacco users (former vs. never users: adjusted OR = 0.67, 95% CI: 0.31–1.44, P < .05), but the risk was significantly increased for current users (current vs. never: adjusted OR = 2.63; 95% CI: 1.22–5.71; P < .05). Likewise, former alcohol users were not at increased risk of disease (former vs. never: adjusted OR = 1.78; 95% CI: 0.87–3.67), whereas current alcohol users were (current vs. never: adjusted OR = 2.57; 95% CI: 1.22–5.42; P < .05). HPV‐related genital lesions (14.3% vs. 10.6%), oral‐genital sexual behavior (42.4% vs. 45.2%), and number (11 or more) of sexual partners (23% v. 17%) were not significantly different between cases and controls. These data suggest that in addition to tobacco and alcohol, HPV plays a role in the development of oral cancer.

Transcriptional enhancer factor (TEF)-1 and its cell-specific co-activator activate human papillomavirus-16 E6 and E7 oncogene transcription in keratinocytes and cervical carcinoma cells.

The human papillomavirus (HPV)‐16 oncogenes, E6 and E7, are transcribed preferentially in keratinocytes and cervical carcinoma cells due to a 5′ enhancer. An abundant peptide binding to a 37 nt enhancer element was purified from human keratinocytes by sequence‐specific DNA chromatography. This protein was identified as transcriptional enhancer factor (TEF)‐1 by complex mobility, binding to wild‐type and mutant SV40 and HPV‐16 enhansons and antigenic reactivity with two anti‐TEF‐1 antibodies. TEF‐1 is cell‐specific, but its transactivation also depends on a limiting, cell‐specific TEF‐1 ‘co‐activator’. We show that both TEF‐1 and the TEF‐1 co‐activator are active in human keratinocytes and essential for HPV‐16 transcription. TEF‐1 binding in vivo was necessary for HPV‐16 P97 promoter activity. Excess TEF‐1 and chimeric GAL4‐TEF‐1 specifically inhibited the P97 promoter by ‘squelching’, indicating that HPV‐16 transcription also requires a limiting TEF‐1 co‐activator. TEF‐1 and the TEF‐1 co‐activator functions mirrored HPV‐16 transcription by their presence in keratinocytes and cervical carcinoma cells and their absence from lymphoid B‐cells, but also functioned in liver cells where the HPV‐16 promoter is inactive. TEF‐1 and its associated co‐activator are thus part of a complex mechanism which determines the restricted cell range of the HPV‐16 E6 and E7 oncogene promoter.

Prevalence of human papillomavirus in the oral cavity/oropharynx in a large population of children and adolescents.

Objective: Human papillomavirus (HPV) in the oral cavity or oropharynx is associated with an increased risk of laryngeal papillomatosis, head and neck cancer, and cervical and other genital cancers. We evaluated the prevalence of HPV DNA in the oral cavity/oropharynx in a cross section of children aged 2 weeks to 20 years. Methods: A risk factor questionnaire and oral exfoliated cells were collected from children (N = 1235). HPV DNA was detected using PCR, dot blot hybridization, and DNA sequencing. Results: The HPV prevalence was 1.9% in the oral cavity/oropharynx of children. A bimodal age distribution was observed with the highest HPV prevalence in the youngest and oldest groups: 2.5% aged <1 year, 0.8% aged 1 to 4 years, 1.2% aged 5 to 11 years, 1.5% in aged 12 to 15 years, and 3.3% in aged 16 to 20 years. The prevalence of the HPV quadrivalent vaccine types (HPV-6, 11, 16, 18) reached 0.9% in the 16- to 20-year age group. In this age group, female gender [odds ratio (OR): 6.9, P = 0.04], genital warts (OR: 19.3, P < 0.01), and current smoker (OR: 6.5, P = 0.01) were associated with a higher risk of being detected with an oral HPV infection. No risk factors in parents were identified with transmission of HPV to infants. Conclusions: The age-specific prevalence rates of HPV in this large cross section of children and adolescents demonstrate that HPV infection is acquired gradually in childhood. These data support a target age for HPV vaccination before puberty to prevent serious HPV-related genital and oral diseases.

Human papillomavirus prevalence and types in newborns and parents: concordance and modes of transmission.

Background and Objectives The purpose of this investigation was to determine the risk of vertical and early contact transmission of human papillomavirus (HPV) in newborn infants based on concordance and sequence match to HPV types in parents. Study Design The genitals of pregnant women and newborns and oral cavity of parents and newborns were analyzed using polymerase chain reaction and DNA sequencing. Data were collected about reproductive health and risk factors for HPV. Results Only one mother/newborn and no father/newborn pair was concordant for an HPV type. All other infected newborns had uninfected or discordant type infected parents. Conclusion The risk of vertical transmission to the oral or genital region of newborns is rare, and transmission between parents and the hospitalized newborn does not appear to occur. Lack of parent/child concordance suggests that newborns detected with HPV in their oral cavity or genitals could have become infected by their mother at untested intervals during pregnancy or in newborns with infection in the oral cavity by other contacts after birth.

Complex Etiology Underlies Risk and Survival in Head and Neck Cancer Human Papillomavirus, Tobacco, and Alcohol: A Case for Multifactor Disease

Findings are inconsistent about whether tobacco, alcohol, and human papillomavirus (HPV) are two independent HNC risk factor groups that distinguish an infection-associated cancer from a tobacco/alcohol-associated HNC. We found that cancer in the oral cavity risk was greater in HPV-E6/E7 seropositive/heavy tobacco users (adjusted OR = 3.5) than in HPV-seronegative/heavy tobacco users (adjusted OR = 1.4); and HPV-seropositive/heavy alcohol users (adjusted OR = 9.8) had greater risk than HPV-seronegative/heavy alcohol users (adjusted OR = 3.1). In contrast, the risk of oropharyngeal cancer was greater in the HPV-seronegative/heavy tobacco (adjusted OR = 11.0) than in HPV-seropositive/heavy tobacco (adjusted OR = 4.7) users and greater in HPV-seronegative/heavy alcohol users (adjusted OR = 24.3) compared to HPV-seropositive/heavy alcohol users (adjusted OR = 8.5). Disease-specific and recurrence-free adjusted survival were significantly worse in oropharyngeal HPV-seronegative cases with no survival differences by HPV status seen in oral cavity cases. The association between tobacco/alcohol, HPV, and tumor site is complex. There appear to be distinct tumor site differences in the combined exposure risks, suggesting that different molecular pathways are involved.

Perinatal Vertical Transmission of Human Papillomavirus and Subsequent Development of Respiratory Tract Papillomatosis

This study prospectively examined the potential for human papillomavirus (HPV) to be transmitted vertically to newborns during delivery. Exfoliated cervical cells were extracted from 72 pregnant women during the third trimester and again during labor prior to delivery, and tested for the presence of HPV DNA. These results were compared with HPV DNA specimens from their newborns, who were sampled by exfoliated cells from the oral-pharyngeal cavity and vulva or tissue from the foreskin 24 to 72 hours after delivery. Among the mothers, 18.1% (13 of 72) typed HPV-positive by the ViraPap/ViraType DNA hybridization technique. Two neonates (2.8% or 2 of 72) tested positive from oral-pharyngeal specimens. This finding supports the hypothesis that respiratory tract papillomatosis may develop as a result of perinatal vertical transmission of HPV. Furthermore, this study suggests that neither cesarean section nor prepartum treatment of HPV lesions will always protect against neonatal acquisition of HPV.