Background: Cutaneous human papillomavirus (HPV) infection may be a risk factor for squamous cell carcinoma (SCC) of the skin.

Methods: To investigate the association between cutaneous HPV and SCC, a case–control study was conducted, including 173 SCC cases from a university dermatology clinic and 300 controls that screened negative for skin cancer. Serum antibodies against cutaneous HPV types in genera alpha, beta, gamma, mu, and nu were measured. Tumor tissue from 159 SCC cases was tested for the presence of DNA for genus-beta HPV types. Using logistic regression ORs and 95% confidence intervals (CI) were estimated for the associations between SCC and cutaneous HPV infection, adjusting for age and sex. The Bonferroni method was used to account for multiple comparisons.

Results: SCC was positively associated with seropositivity to any genus-beta HPV type (OR, 1.93; 95% CI, 1.23–3.02), particularly with types in species-1 (OR, 1.86; 95% CI, 1.22–2.85). Type-specific associations with SCC were observed for HPV 8 (OR, 1.80; 95% CI, 1.14–2.84), 17 (OR, 1.59; 95% CI, 1.02–2.49) and HPV 10 from genus-alpha (OR, 2.24; 95% CI, 1.04–4.85). None of the type-specific associations remained statistically significant after correction for multiple comparisons. When DNA-positive SCC cases were compared with controls, strong serologic associations were observed for HPVs 5 (OR, 3.48; 95% CI, 1.27–9.59), 17 (OR, 3.36; 95% CI, 1.29–8.72), and 24 (OR, 3.79; 95% CI, 1.24–11.5).

Conclusion: Genus-beta HPV infections were associated with SCC in our study population.

Impact: Identifying the role of cutaneous HPV infection in SCC may lead to improved characterization of high-risk individuals and the development of novel prevention strategies. Cancer Epidemiol Biomarkers Prev; 21(8); 1303–13. ©2012 AACR.

Squamous cell carcinoma (SCC) of the skin is the second most frequently occurring cancer among Caucasians in the United States, and the incidence continues to increase each year (1). Established risk factors for SCC include UV radiation exposure, older age, light skin, and immunosuppression (2). Emerging evidence suggests that cutaneous human papillomavirus (HPV) infection may also be a risk factor for SCC (3–8). HPV types that infect cutaneous epithelia have been identified from genera alpha, beta, gamma, mu, and nu (9). Presence of antibodies against one or more of the genus-beta HPV types as a group has been associated with SCC in several case–control studies (3–8) and type-specific associations with SCC have also been observed with HPV 8 (5, 8, 10), 15 (11), 17 (11), and 38 (5, 11). Estimates of HPV DNA prevalence in SCC tissues from immunocompetent individuals range from 20% to 48% (10, 12–16). To our knowledge, only one study has reported findings on the relationship between HPV seroreactivity and SCC in conjunction with detection of HPV DNA in the skin cancer tumor itself, and in that study, DNA-positive SCC cases were more likely to be seropositive for any HPV type than DNA-negative SCC cases (3).

This is the first case–control study in a U.S. population to investigate the association between SCC and seroreactivity to cutaneous HPV types belonging to 5 different genera. In addition, serologic associations between beta HPV types and SCC were further stratified by the presence of DNA of these HPV types in the tumor tissues.

Study design and population

A clinic-based case–control study was conducted in Tampa, FL to investigate the association between cutaneous HPV infection and SCC of the skin. The study design and population have been previously described in detail (17). Briefly, histologically confirmed SCC cases were recruited from the University of South Florida (USF; Tampa, FL) Dermatology clinic (n = 191). Control subjects were recruited from the USF Family Medicine and Moffitt's Lifetime Cancer Screening and Prevention (LCS) clinics. All control participants included in the current study underwent a full-body skin cancer screening exam, were negative for current signs of skin cancer, and had no history of any type of cancer (n = 281). If a patient had a suspicious lesion detected during the skin screening exam that was later determined to be benign based on pathology review, the patient was also included as a control (n = 77). If a patient's screen-detected lesion was histologically confirmed to be an SCC then that patient was included as a case (n = 6). Study participants were ages 18 to 80 and were eligible to participate regardless of immune status. Nine SCC cases (4.7%) reported a history of organ transplantation.

Participants completed a self-administered questionnaire on skin cancer risk factors, and blood samples were obtained from 174 (90.6%) cases and 340 (95.0%) controls. With the exception of 2 non-White controls, the current analysis was restricted to White participants. The final sample size for the analysis of cutaneous HPV seroreactivity was 173 SCC cases and 300 controls. From patients with SCC undergoing surgical excision, a 3-mm punch of the residual SCC tumor was obtained and flash frozen in liquid nitrogen. Analyses were restricted to tumor specimens that tested positive for β-globin, corresponding to 180 SCC tumors from 159 individual patients, including 19 patients who contributed tissues from distinct, concurrent tumors. The final sample size for analyses including cutaneous HPV seroreactivity and DNA status consisted of 146 SCC cases and 300 controls. Written informed consent was provided by all study participants and all study procedures were approved by the Institutional Review Board at the USF.

HPV antibody measurement

Sera were tested for antibodies to the major capsid protein L1 of cutaneous HPV type(s) within genera alpha (2, 3, 7, 10, 27, 57, and 77); beta (5, 8, 9, 15, 17, 20, 23, 24, 36, 38, 49, 75, 76, 92, 96, and 107); gamma (4, 48, 50, 65, 88, 95, 101, and 103); mu (1); and nu (41). Sera were also tested for antibodies to the VP1 capsid protein of 2 human polyomaviruses, JC virus (JCV) and KI virus (KIV), to test the specificity of associations observed between cutaneous HPV and SCC.

The antibody detection method used is based on glutathione S-transferase (GST) capture ELISA (18, 19) in combination with fluorescent bead technology (Luminex; refs. 20, 21), as previously described. Individual cutoff values to define HPV type–specific seropositivity were applied as described previously to allow for the direct comparison of cutaneous HPV seroprevalence across studies that used the same assay (4, 22).

HPV DNA detection

DNA extraction from frozen SCC tumor tissues was carried out using the Qiagen BioRobot EZ1 with the EZ1 DNA tissue kit according to the manufacturer's instructions (Qiagen). Briefly, frozen tissues were incubated in proteinase K and a buffer G2 (Qiagen) at 56°C until the tissue was completely lysed. To monitor the possible occurrence of cross-contamination between the different specimens during DNA extraction, tubes containing buffer only were also included.

HPV DNA was measured in all samples using an assay that detects 25 genus-beta HPV types (5, 8, 9, 12, 14, 15, 17, 19, 20, 21, 22, 23, 24, 25, 36, 37, 38, 47, 49, 75, 76, 80, 92, 93, and 96), based on the same techniques used to measure mucosal HPV types (23, 24). The assay combines 2 different techniques: multiplex PCR using HPV type–specific primer for amplification of viral DNA and array primer extension (APEX) for typing with 2 type-specific probes each.

Statistical analysis

Skin cancer risk factors were compared between SCC cases and controls using the χ2 test, and independent associations between these factors and case–control status were estimated by calculating ORs and 95% confidence intervals (CI) using conditional logistic regression with adjustment for all other factors. The same approach was used to compare factors between controls who were seropositive for at least one cutaneous HPV type and controls who were seronegative for all HPV types. HPV type–specific seroprevalence was calculated as the proportion of patients who tested positive for antibodies to a given HPV type. Genus-specific HPV seroprevalence was calculated as the proportion of patients who tested seropositive for at least one HPV type in a given genus. Cutaneous HPV seropositivity to any type was defined as testing seropositive to at least one of the 33 cutaneous HPV types tested across the 5 genera. Logistic regression was used to calculate ORs and 95% CIs to estimate the associations between SCC and (i) type-specific cutaneous HPV seropositivity, (ii) genus-specific seropositivity, and (iii) the number of types in a given genus for which an individual tested seropositive. Tests for trend in risk for SCC associated with seropositivity for increasing number of cutaneous HPV types within a given genus were conducted by assigning ordinal values to each category and including the ordinal variable in the logistic regression model. The Bonferroni method was used to account for multiple comparisons, reducing the statistical significance level for SCC associated with genus-specific HPV seropositivity to P < 0.01 and type-specific HPV seropositivity to P < 0.002.

Tumor tissues obtained from SCC cases were classified as positive or negative for the presence of HPV DNA for types in genus-beta. The 19 cases who contributed more than one SCC tumor tissue were considered HPV DNA–positive if at least one of the tumor tissues provided tested positive for genus-beta HPV DNA. Logistic regression was then used to calculate the OR and 95% CI for the association between seropositivity to any genus-beta HPV type and SCC, stratified by the presence or absence of DNA to any genus-beta HPV type in the tumor tissue (i.e., genus-beta HPV–positive vs. genus-beta HPV–negative). This analysis was restricted to the genus-beta HPV types included in both the antibody and PCR assays (5, 8, 20, 24, 36, 9, 15, 17, 23, 38, 75, 76, 92, and 96). Type-specific concordance was calculated among the SCC cases as the proportion that tested seropositive for a given HPV type and had DNA in their tumor tissue corresponding to the same HPV type. Using logistic regression, additional stratified analyses were then conducted to compare genus-beta type–specific HPV seropositivity between controls and SCC cases that had DNA in their tumors corresponding to the same HPV type for which antibodies were detected.

All factors listed in Table 1 were considered as potential confounders. With the exception of age and sex, none of the factors listed in Table 1 altered the ORs and corresponding 95% CIs for the associations between SCC and cutaneous HPV infection (measured by serology or DNA) by more than 10%. Therefore, all associations were adjusted for age (as a continuous variable) and sex only. To rule out the possibility of residual confounding by age, an age-restricted analysis was conducted by restricting both cases and controls to those aged 40 to 69. In addition, sex-stratified ORs and 95% CIs were estimated for the associations between genus-specific HPV seropositivity and SCC, also restricted to cases and controls aged 40 to 49. Analyses were conducted with and without the 9 SCC cases who reported a history of organ transplantation with similar results, thus, all cases were included. Analyses were conducted using the SAS statistical software package (version 9.2; SAS Institute).

The distribution of demographic, lifestyle, and skin cancer risk factors among SCC cases and controls is presented in Table 1. Compared with controls, SCC cases were significantly more likely to be older, male, and less educated, and to have light eye and hair color, past occupational sunlight exposure, history of blistering sunburn, cutaneous sensitivity to sunlight exposure, poor tanning ability, and a history of alcohol consumption and smoking (Table 1). Demographic, lifestyle, and skin cancer risk factors were not associated with cutaneous HPV serostatus for any HPV type among the controls (Table 2).

Associations between genus-specific HPV seroreactivity and SCC are presented in Table 3. SCC was significantly associated with seropositivity to any genus-beta HPV type (OR, 1.93; 95% CI, 1.23–3.02), with increasing risk observed with an increase in the number of genus-beta HPV types for which an individual tested seropositive up to 8 types; seropositivity for >8 types showed no additional risk for SCC. Similar associations with SCC were not observed for seropositivity to HPV type(s) in genus alpha, gamma, mu, or nu or the polyomaviruses JCV and KIV (Table 3).

Analyses restricted to cases and controls ages 40 to 69 years yielded associations between cutaneous HPV seroreactivity and SCC of similar magnitude as those observed for the total study population. Specifically, SCC was significantly associated with seropositivity for at least one HPV type in genus-beta (OR, 2.05; 95% CI, 1.21–3.46) but not with genus-alpha (OR, 1.15; 95% CI, 0.69–1.89), gamma (OR, 1.25; 95% CI, 0.77–2.02), mu (OR, 1.20; 95% CI, 0.74–1.97), or nu (OR, 1.06; 95% CI, 0.49–2.27). In addition, age-restricted, sex-stratified analyses showed similar associations between genus-specific cutaneous HPV seropositivity and SCC among males (genus-alpha: OR, 1.04; 95% CI, 0.54–2.01; beta: OR, 2.15; 95% CI, 1.07–4.29; gamma: OR, 1.02; 95% CI, 0.54–1.91; mu: OR, 1.05; 95% CI, 0.56–1.97; nu: OR, 1.10; 95% CI, 0.43–2.77) and females (genus-alpha: OR, 1.35; 95% CI, 0.62–2.94; beta: OR, 2.17; 95% CI, 0.93–5.07; gamma: OR, 1.69; 95% CI, 0.80–3.60; mu: OR, 1.59; 95% CI, 0.72–3.51; nu: OR, 1.03; 95% CI, 0.27–3.98).

Seroprevalence was greatest for HPV type 4 in genus-gamma for both SCC cases (43.4%) and controls (34.3%); however, seropositivity to HPV 4 was not significantly associated with SCC (Table 4). HPV 10 was the single type from genus-alpha significantly associated with SCC (OR, 2.24; 95% CI, 1.04–4.85; Table 4). Within genus-beta, seroprevalence was greatest for HPV types 49, 8, and 17 among cases and types 49, 17, 15, and 9 among controls with significant associations observed with SCC for types 8 (OR, 1.80; 95% CI, 1.14–2.84) and 17 (OR, 1.59; 95% CI, 1.02–2.49; Table 4). None of the type-specific associations corresponded to a P value of less than 0.002, the significance level adjusted for multiple comparisons using the Bonferroni method. At the species level, SCC was significantly positively associated with seropositivity for HPV types in genus-beta species 1 (OR, 1.86; 95% CI, 1.22–2.85; P = 0.004). No significant associations were observed for genus-beta species 2 to 5 (Table 4) and no significant associations with SCC were observed for individual HPV types in genera gamma, mu, or nu (Table 4).

Ninety-six SCC cases (66%) were DNA positive for at least one genus-beta HPV type tested. Seropositivity to at least one of the genus-beta HPV types tested was associated with overall DNA positivity of genus-beta HPV types in SCC (OR, 2.81; 95% CI, 1.53–5.16; data not shown). Seropositivity to any genus-beta HPV type was not associated with SCC among those cases with tumors negative for HPV DNA (OR, 0.98; 95% CI, 0.53–1.84; data not shown). On the basis of these observations, a decision was made to investigate genus-beta type–specific correlations between seroantibodies and DNA presence in the tumor for the same HPV types. Among the 96 SCC cases with HPV DNA–positive tumors, 41.7% tested seropositive for antibodies to the same HPV type detected in the tumor tissue (data not shown). Concordance between the specific HPV type for which DNA was observed in the tumor and the HPV type for which circulating antibodies were observed ranged from 12.5% for type 9 to 54.5% for type 17 (Table 5). Compared with controls, SCC cases who were DNA positive for the following HPV types had significantly greater seroprevalence for the same HPV types: HPV 5 (OR, 3.48; 95% CI, 1.27–9.59), 24 (OR, 3.79; 95% CI, 1.24–11.5), and 17 (OR, 3.36; 95% CI, 1.29–8.72; Table 5). SCC cases with tumors that tested DNA negative for all genus-beta HPV types were significantly less likely to be seropositive for HPV types 20, 36, 9, 15, 38, and 75 than controls (Table 5).

Observations from the current case–control study support the association between cutaneous HPV seropositivity and SCC. Seropositivity to HPV 10 was the single type from genus-alpha associated with SCC. Seropositivity for cutaneous HPV types in genus-beta overall, and for types 8 and 17 specifically, was significantly associated with SCC in this clinic-based case–control study. In addition, correlations between seropositivity and DNA positivity for the same genus-beta HPV type were observed for HPV types 5, 24, and 17. SCC cases with tumors negative for any genus-beta HPV type were less likely to be seroprevalent for HPV types 20, 36, 9, 15, 38, and 75 than controls.

The positive association observed between SCC and seropositivity to HPV types in genus-beta is consistent with 2 (7, 8) of 5 case–control studies (3, 5, 7, 8, 16) including one study from the United States (7). Type-specific associations observed with SCC for HPV types 8 and 17 in genus-beta agree with findings from case–control studies in the United States (7), the Netherlands (5), and Australia (8) for HPV type 8 and with findings from case–control studies in the United States (7) and Italy (25) for HPV type 17. In contrast to previous case–control studies, SCC was not associated with seropositivity to HPV types 5 (6), 15 (7, 11), and 38 (5, 11), as well as with types 20, 24, 9, 49, 75, 76, 92, and 96 (7). Neither of 2 published cohort studies (4, 26) observed statistically significant increased risks of SCC associated with HPV seropositivity for types in genera alpha (4), beta (4, 26), gamma (4), mu (4), or nu (4).

A positive dose–response between seropositivity to increasing numbers of HPV types in genus-beta and SCC was observed in the current study population, consistent with 3 (7, 8, 27) of 4 (7, 8, 25, 27) previous case–control studies. However, in a recent prospective cohort study (26), an increased risk of SCC was not observed among individuals who were seropositive to at least one genus-beta HPV type or multiple types. Furthermore, neither of the 2 prospective studies reporting increased risks of SCC associated with baseline cutaneous HPV seropositivity observed statistically significant type-specific associations with any individual type from genus-beta (4, 26).

The single type-specific association with SCC observed in this study for a cutaneous HPV type in a genus other than beta was with HPV type 10 in genus-alpha, consistent with the 2 studies in the literature previously reporting on HPV type–specific associations outside of genus-beta (4, 11). HPV 10 has been associated with benign skin lesions (28) but displays weak transforming activities by in vitro experimental models (29), thus, its direct involvement in skin carcinogenesis remains unclear.

Among SCC cases from Sweden and Austria (3), 58% tested DNA positive for at least 1 HPV type in genus-beta compared with 66% in SCC cases from the current study. In addition, a low overall concordance between HPV DNA and seropositivity for the same HPV type (19%) was observed (3) compared with the current study with an overall type-specific concordance of 41.7%. In addition, among participants in the current study, individual type-specific concordance was greater for genus-beta HPV types 5, 8, and 15 but lower for types 9 and 24 as compared with participants from Sweden/Austria (3). The observed difference in overall concordance between HPV types identified from the tumor and circulating antibodies may be explained by a difference in the number of genus-beta HPV types examined, (8 types in the Swedish/Austrian population compared with 13 types in the current study population). Individual type-specific variations in concordance may be explained by differences in the laboratory techniques used to test for the presence of HPV DNA in the tissue specimens. In addition, Andersson and colleagues obtained biopsies from the malignant lesion and adjacent healthy skin and subsequently classified SCC cases as DNA positive if either of the biopsies tested positive for beta HPV DNA (3) in contrast to the current study that defined DNA positivity based on detection in the SCC tumor only.

It was unexpected to observe statistically significant inverse associations between seropositivity and SCC among HPV DNA–negative cases. One explanation may simply be that SCC cases with tumors negative for HPV DNA and subsequently low antibody seroprevalence compared with controls may have a different risk profile for disease. In terms of previous findings, these data may explain the low-risk estimates observed in previous studies for the associations between cutaneous HPV seropositivity and SCC when HPV DNA status of the tumor has not been accounted for. In addition, the observations from the current study may underlie the discrepant findings presented across case–control studies. Further research is needed to delineate the role of cutaneous HPV antibodies as either a marker of prevalent and cleared infections or a marker of current active infections and their subsequent association with SCC risk.

The current study provides evidence for the association between genus-beta HPV and SCC, however, the exact mechanism by which cutaneous HPV is associated with SCC remains unclear. It has been hypothesized that the effects of genus-beta HPV on UV-induced DNA damage and apoptosis could lead to the accumulation of mutations which predispose to SCC formation (30). Evidence from in vitro and in vivo studies have shown that E6 and E7 proteins from certain cutaneous HPV types are capable of inhibiting UV-induced cell-cycle check points, apoptosis, and DNA repair machinery (31–36). Observations from epidemiologic studies have shown statistically significant interactions between sun-related factors and genus-beta HPV seropositivity in relation to SCC (27, 37, 38). In addition, HPV 38 E6 and E7 proteins have displayed transforming properties in primary human keratinocyte cells sufficient to deregulate cell-cycle control and growth arrest (39). Taken together, these findings support a role for cutaneous HPV as a cofactor in SCC carcinogenesis, in contrast to the direct role of mucosal HPV infections in the carcinogenesis of cervical cancer.

Clinic based case–control study populations are often not representative of the general population. However, cases and controls were recruited from clinics that serve the same underlying populations, thus preserving internal validity. Because a considerable proportion of the skin cancer screening patients were confirmed to be free of skin cancer only after completion of the follow-up visit with a dermatologist, there was at time lag from study consent to verification of case–control status. As such, matching controls to cases was logistically challenging and significant case–control differences in age and sex resulted. However, all analyses were adjusted for age and sex. Furthermore, the analysis restricted to those aged 40 to 69 yielded similar results as those obtained from the entire study population. In addition, similar associations between genus-specific HPV seropositivity and SCC were observed between males and females in the restricted age range. Therefore, the observed associations between cutaneous HPV seropositivity and SCC are not likely due to residual confounding by age and/or sex. Adjustment for multiple comparisons reduced the statistical significance of most of the observed associations, thus, chance cannot be ruled out as an explanation for these findings. However, multiple comparisons is a methodologic challenge faced by all epidemiologic studies of cutaneous HPV infections, given the sheer number of types potentially implicated in SCC. Benign lesions were not obtained for control subjects that completed dermatologic follow-up, as such, for comparison purposes it was not possible to test the benign lesions for the presence of HPV DNA. If in fact cutaneous HPV is associated with benign skin lesions then the observed results would be attenuated given that a subset of the controls were diagnosed with such lesions.

Several study strengths should also be noted. The current study is the first to present cutaneous HPV genus and type-specific associations outside of genus-alpha and -beta and to incorporate comparisons with genus-beta HPV DNA reactivity in SCC tumor tissues in a U.S. population. Only one study in the published literature has presented similar results among residents of Sweden and Austria (3). This is a major strength of the current study compared with previously published case–control studies of cutaneous HPV serology in SCC, allowing the unique opportunity to investigate specificity of serologic response and the presence of DNA for the same HPV type. In addition, the laboratory used to test for cutaneous HPV seropositivity has been used in most of the seroepidemiologic studies of cutaneous HPV published to date (4, 6, 7, 11, 22, 25, 40, 41), allowing for direct comparison across studies. Finally, the full-body skin examination conducted on all control subjects is another study strength, as it eliminated misclassification of SCC case–control status that can result by inclusion of apparently healthy individuals who in fact have a prevalent, undiagnosed SCC. Of note, 6 of the SCC cases included in the current study were screen-detected.

The current study confirms previous findings supporting the association between genus-beta seropositivity and SCC. In addition, genus-beta type–specific associations with SCC are supported by the observed correlation between HPV seropositivity and tumor DNA positivity for specific types in genus-beta. However, additional longitudinal studies need to be conducted to better understand the direction of the association between cutaneous HPV infection and SCC to rule out the possibility of reverse causality.

T. Waterboer is an employee for Boehringer Ingelheim. No potential conflicts of interest were disclosed by the other authors.

Conception and design: A.R. Giuliano, M. Pawlita, D.E. Rollison

Development of methodology: T. Waterboer, J.L. Messina, K.M. Michael, M. Pawlita, D.E. Rollison

Acquisition of data (provided animals, acquired and managed patients, provided facilities, etc.): M.R. Iannacone, T. Waterboer, J.L. Messina, N.A. Fenske, B.S. Cherpelis, V.K. Sondak, R.G. Roetzheim, K.M. Michael, M. Pawlita, D.E. Rollison

Analysis and interpretation of data (e.g., statistical analysis, biostatistics, computational analysis): M.R. Iannacone, A.R. Giuliano, V.K. Sondak, M. Tommasino, M. Pawlita, D.E. Rollison

Writing, review, and/or revision of the manuscript: M.R. Iannacone, T. Waterboer, A.R. Giuliano, J.L. Messina, N.A. Fenske, B.S. Cherpelis, V.K. Sondak, R.G. Roetzheim, K.M. Michael, M. Pawlita, D.E. Rollison

Administrative, technical, or material support (i.e., reporting or organizing data, constructing databases): M.R. Iannacone, D.E. Rollison

Study supervision: V.K. Sondak, D.E. Rollison

Development of methods for the detection of papillomaviruses (ICB, IARC): T. Gheit

The authors thank the supporting staff at USF and LCS for their assistance with patient recruitment, especially Kristen Jonathan, Jill Weber, and Carolyn Gerow.

This work was supported by a James and Esther King New Investigator grant awarded to DER (06NIR-08) by the Florida Department of Health (FL DOH). This research was also supported in part by the Miles for Moffitt Foundation Funds.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

1.
Jemal
A
,
Siegel
R
,
Xu
J
,
Ward
E
. 
Cancer statistics, 2010
.
CA Cancer J Clin
2010
;
60
:
277
300
.
2.
Saladi
RN
,
Persaud
AN
. 
The causes of skin cancer: a comprehensive review
.
Drugs Today(Barc)
2005
;
41
:
37
53
.
3.
Andersson
K
,
Waterboer
T
,
Kirnbauer
R
,
Slupetzky
K
,
Iftner
T
,
de Villiers
EM
, et al
Seroreactivity to cutaneous human papillomaviruses among patients with nonmelanoma skin cancer or benign skin lesions
.
Cancer Epidemiol Biomarkers Prev
2008
;
17
:
189
95
.
4.
Casabonne
D
,
Michael
KM
,
Waterboer
T
,
Pawlita
M
,
Forslund
O
,
Burk
RD
, et al
A prospective pilot study of antibodies against human papillomaviruses and cutaneous squamous cell carcinoma nested in the Oxford component of the European Prospective Investigation into Cancer and Nutrition
.
Int J Cancer
2007
;
121
:
1862
8
.
5.
Feltkamp
MC
,
Broer
R
,
di Summa
FM
,
Struijk
L
,
van der Meijden
E
,
Verlaan
BP
, et al
Seroreactivity to epidermodysplasia verruciformis-related human papillomavirus types is associated with nonmelanoma skin cancer
.
Cancer Res
2003
;
63
:
2695
700
.
6.
Karagas
MR
,
Nelson
HH
,
Sehr
P
,
Waterboer
T
,
Stukel
TA
,
Andrew
A
, et al
Human papillomavirus infection and incidence of squamous cell and basal cell carcinomas of the skin
.
J Natl Cancer Inst
2006
;
98
:
389
95
.
7.
Karagas
MR
,
Waterboer
T
,
Li
Z
,
Nelson
HH
,
Michael
KM
,
Bavinck
JN
, et al
Genus beta human papillomaviruses and incidence of basal cell and squamous cell carcinomas of skin: population based case-control study
.
BMJ
2010
;
341
:
c2986
.
8.
Struijk
L
,
Hall
L
,
van der Meijden
E
,
Wanningen
P
,
Bavinck
JN
,
Neale
R
, et al
Markers of cutaneous human papillomavirus infection in individuals with tumor-free skin, actinic keratoses, and squamous cell carcinoma
.
Cancer Epidemiol Biomarkers Prev
2006
;
15
:
529
35
.
9.
Bernard
HU
,
Burk
RD
,
Chen
Z
,
van Doorslaer
K
,
Hausen
H
,
de Villiers
EM
. 
Classification of papillomaviruses (PVs) based on 189 PV types and proposal of taxonomic amendments
.
Virology
2010
;
401
:
70
9
.
10.
Forslund
O
,
Ly
H
,
Reid
C
,
Higgins
G
. 
A broad spectrum of human papillomavirus types is present in the skin of Australian patients with non-melanoma skin cancers and solar keratosis
.
Br J Dermatol
2003
;
149
:
64
73
.
11.
Waterboer
T
,
Abeni
D
,
Sampogna
F
,
Rother
A
,
Masini
C
,
Sehr
P
, et al
Serological association of beta and gamma human papillomaviruses with squamous cell carcinoma of the skin
.
Br J Dermatol
2008
;
159
:
457
9
.
12.
Forslund
O
,
Iftner
T
,
Andersson
K
,
Lindelof
B
,
Hradil
E
,
Nordin
P
, et al
Cutaneous human papillomaviruses found in sun-exposed skin: Beta-papillomavirus species 2 predominates in squamous cell carcinoma
.
J Infect Dis
2007
;
196
:
876
83
.
13.
Harwood
CA
,
Surentheran
T
,
McGregor
JM
,
Spink
PJ
,
Leigh
IM
,
Breuer
J
, et al
Human papillomavirus infection and non-melanoma skin cancer in immunosuppressed and immunocompetent individuals
.
J Med Virol
2000
;
61
:
289
97
.
14.
Pfister
H
,
Fuchs
PG
,
Majewski
S
,
Jablonska
S
,
Pniewska
I
,
Malejczyk
M
. 
High prevalence of epidermodysplasia verruciformis-associated human papillomavirus DNA in actinic keratoses of the immunocompetent population
.
Arch Dermatol Res
2003
;
295
:
273
9
.
15.
Stockfleth
E
,
Nindl
I
,
Sterry
W
,
Ulrich
C
,
Schmook
T
,
Meyer
T
. 
Human papillomaviruses in transplant-associated skin cancers
.
Dermatol Surg
2004
;
30
:
604
9
.
16.
Termorshuizen
F
,
Feltkamp
MC
,
Struijk
L
,
de Gruijl
FR
,
Bavinck
JN
,
van Loveren
H
. 
Sunlight exposure and (sero)prevalence of epidermodysplasia verruciformis-associated human papillomavirus
.
J Invest Dermatol
2004
;
122
:
1456
62
.
17.
Rollison
DE
,
Guiliano
A
,
Messina
JL
,
Fenske
NA
,
Cherpelis
BS
,
Sondak
VK
, et al
Case-control study of Merkel cell polyomavirus infection and cutaneous squamous cell carcinoma
.
Cancer Epidemiol Biomarkers Prev
2012
;
21
:
74
81
.
18.
Sehr
P
,
Muller
M
,
Hopfl
R
,
Widschwendter
A
,
Pawlita
M
. 
HPV antibody detection by ELISA with capsid protein L1 fused to glutathione S-transferase
.
J Virol Methods
2002
;
106
:
61
70
.
19.
Sehr
P
,
Zumbach
K
,
Pawlita
M
. 
A generic capture ELISA for recombinant proteins fused to glutathione S-transferase: validation for HPV serology
.
J Immunol Methods
2001
;
253
:
153
62
.
20.
Waterboer
T
,
Sehr
P
,
Michael
KM
,
Franceschi
S
,
Nieland
JD
,
Joos
TO
, et al
Multiplex human papillomavirus serology based on in situ-purified glutathione s-transferase fusion proteins
.
Clin Chem
2005
;
51
:
1845
53
.
21.
Waterboer
T
,
Sehr
P
,
Pawlita
M
. 
Suppression of non-specific binding in serological Luminex assays
.
J Immunol Methods
2006
;
309
:
200
4
.
22.
Michael
KM
,
Waterboer
T
,
Sehr
P
,
Rother
A
,
Reidel
U
,
Boeing
H
, et al
Seroprevalence of 34 human papillomavirus types in the German general population
.
PLoS Pathog
2008
;
4
:
e1000091
.
23.
Gheit
T
,
Billoud
G
,
de Koning
MN
,
Gemignani
F
,
Forslund
O
,
Sylla
BS
, et al
Development of a sensitive and specific multiplex PCR method combined with DNA microarray primer extension to detect Betapapillomavirus types
.
J Clin Microbiol
2007
;
45
:
2537
44
.
24.
Gheit
T
,
Landi
S
,
Gemignani
F
,
Snijders
PJ
,
Vaccarella
S
,
Franceschi
S
, et al
Development of a sensitive and specific assay combining multiplex PCR and DNA microarray primer extension to detect high-risk mucosal human papillomavirus types
.
J Clin Microbiol
2006
;
44
:
2025
31
.
25.
Waterboer
T
,
Neale
R
,
Michael
KM
,
Sehr
P
,
de Koning
MN
,
Weissenborn
SJ
, et al
Antibody responses to 26 skin human papillomavirus types in the Netherlands, Italy and Australia
.
J Gen Virol
2009
;
90
:
1986
98
.
26.
Plasmeijer
EI
,
Pandeya
N
,
O'Rourke
P
,
Pawlita
M
,
Waterboer
T
,
Feltkamp
MC
, et al
The Association between Cutaneous Squamous Cell Carcinoma and Betapapillomavirus Seropositivity: a Cohort Study
.
Cancer Epidemiol Biomarkers Prev
2011
;
20
:
1171
7
.
27.
Bouwes
Bavinck JN
,
Neale
RE
,
Abeni
D
,
Euvrard
S
,
Green
AC
,
Harwood
CA
, et al
Multicenter study of the association between betapapillomavirus infection and cutaneous squamous cell carcinoma
.
Cancer Res
2010
;
70
:
9777
86
.
28.
Kremsdorf
D
,
Jablonska
S
,
Favre
M
,
Orth
G
. 
Human papillomaviruses associated with epidermodysplasia verruciformis. II. Molecular cloning and biochemical characterization of human papillomavirus 3a, 8, 10, and 12 genomes
.
J Virol
1983
;
48
:
340
51
.
29.
Boxman
IL
,
Mulder
LH
,
Noya
F
,
de Waard
V
,
Gibbs
S
,
Broker
TR
, et al
Transduction of the E6 and E7 genes of epidermodysplasia-verruciformis-associated human papillomaviruses alters human keratinocyte growth and differentiation in organotypic cultures
.
J Invest Dermatol
2001
;
117
:
1397
404
.
30.
Tomlins
C
,
Storey
A
. 
Cutaneous HPV5 E6 causes increased expression of Osteoprotegerin and Interleukin 6 which contribute to evasion of UV-induced apoptosis
.
Carcinogenesis
2010
;
31
:
2155
64
.
31.
Dong
W
,
Arpin
C
,
Accardi
R
,
Gissmann
L
,
Sylla
BS
,
Marvel
J
, et al
Loss of p53 or p73 in human papillomavirus type 38 E6 and E7 transgenic mice partially restores the UV-activated cell cycle checkpoints
.
Oncogene
2008
;
27
:
2923
8
.
32.
Dong
W
,
Kloz
U
,
Accardi
R
,
Caldeira
S
,
Tong
WM
,
Wang
ZQ
, et al
Skin hyperproliferation and susceptibility to chemical carcinogenesis in transgenic mice expressing E6 and E7 of human papillomavirus type 38
.
J Virol
2005
;
79
:
14899
908
.
33.
Jackson
S
,
Harwood
C
,
Thomas
M
,
Banks
L
,
Storey
A
. 
Role of Bak in UV-induced apoptosis in skin cancer and abrogation by HPV E6 proteins
.
Genes Dev
2000
;
14
:
3065
73
.
34.
Jackson
S
,
Storey
A
. 
E6 proteins from diverse cutaneous HPV types inhibit apoptosis in response to UV damage
.
Oncogene
2000
;
19
:
592
8
.
35.
Simmonds
M
,
Storey
A
. 
Identification of the regions of the HPV 5 E6 protein involved in Bak degradation and inhibition of apoptosis
.
Int J Cancer
2008
;
123
:
2260
6
.
36.
Viarisio
D
,
Mueller-Decker
K
,
Kloz
U
,
Aengeneyndt
B
,
Kopp-Schneider
A
,
Grone
HJ
, et al
E6 and E7 from beta HPV38 cooperate with ultraviolet light in the development of actinic keratosis-like lesions and squamous cell carcinoma in mice
.
PLoS Pathog
2011
;
7
:
e1002125
.
37.
Hall
L
,
Struijk
L
,
Neale
RE
,
Feltkamp
MC
. 
Re: Human papillomavirus infection and incidence of squamous cell and basal cell carcinomas of the skin
.
J Natl Cancer Inst
2006
;
98
:
1425
6
.
38.
Iannacone
MR
,
Wang
W
,
Stockwell
HG
,
O'Rourke
K
,
Giuliano
AR
,
Sondak
VK
, et al
Sunlight exposure and cutaneous human papillomavirus seroreactivity in basal cell and squamous cell carcinomas of the skin
.
J Infect Dis
. 
2012
Jun 1. [Epub ahead of print]
.
39.
Caldeira
S
,
Zehbe
I
,
Accardi
R
,
Malanchi
I
,
Dong
W
,
Giarre
M
, et al
The E6 and E7 proteins of the cutaneous human papillomavirus type 38 display transforming properties
.
J Virol
2003
;
77
:
2195
206
.
40.
Iannacone
MR
,
Michael
KM
,
Giuliano
AR
,
Waterboer
T
,
Pawlita
M
,
Rollison
DE
. 
Risk factors for cutaneous human papillomavirus seroreactivity among patients undergoing skin cancer screening in Florida
.
J Infect Dis
201
:
760
9
.
41.
Rollison
DE
,
Pawlita
M
,
Giuliano
AR
,
Iannacone
MR
,
Sondak
VK
,
Messina
JL
, et al
Measures of cutaneous human papillomavirus infection in normal tissues as biomarkers of HPV in corresponding nonmelanoma skin cancers
.
Int J Cancer
2008
;
123
:
2337
42
.