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Keywords:

  • HPV;
  • non-melanoma skin cancer;
  • UV;
  • apoptosis;
  • transformation;
  • stem cell

Abstract

  1. Top of page
  2. Abstract
  3. Introduction
  4. HPV and its association with NMSC
  5. Transforming properties of cutaneous HPV early region proteins
  6. Molecular mechanisms
  7. Anti-apoptotic effects of HPV in response to UV irradiation
  8. HPV and potential cancer stem cells in the epidermis
  9. Genetic abnormalities in NMSC
  10. Summary
  11. Acknowledgements
  12. References

Human papillomaviruses (HPVs) are DNA tumour viruses that induce hyperproliferative lesions in cutaneous and mucosal epithelia. The relationship between HPV and non-melanoma skin cancer (NMSC) is important clinically since NMSC is the most common form of malignancy among fair-skinned populations. It is well established that solar ultraviolet (UV) irradiation is the major risk factor for developing NMSC, but a pathogenic role for HPV in the development of NMSC has also been proposed. Recent molecular studies reveal a likely role for HPV infection in skin carcinogenesis as a co-factor in association with UV. This review summarizes the literature describing these data, highlights some of the important findings derived from these studies, and speculates on future perspectives. Copyright © 2006 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.


Introduction

  1. Top of page
  2. Abstract
  3. Introduction
  4. HPV and its association with NMSC
  5. Transforming properties of cutaneous HPV early region proteins
  6. Molecular mechanisms
  7. Anti-apoptotic effects of HPV in response to UV irradiation
  8. HPV and potential cancer stem cells in the epidermis
  9. Genetic abnormalities in NMSC
  10. Summary
  11. Acknowledgements
  12. References

Human papillomaviruses (HPVs) are small DNA tumour viruses that have a circular double-stranded DNA genome of ∼8 kb in length. The viruses are strictly epitheliotropic, infecting keratinocytes at a wide range of body sites. Over 120 different types of HPV have been identified to date 1. Extensive DNA sequence and phylogenetic analysis has revealed that the diverse spectrum of viruses that infect the skin is clustered in groups, and significant sequence divergence exists between the viral types within the different groups. The most frequent manifestation of cutaneous HPV infection is the development of benign self-limiting epithelial proliferations, namely warts. Most interest in the HPV field is centred round the association of HPV infection with the development of cancers at specific body sites, and this review will focus on the potential role of HPV in the development of skin cancers, highlighting recent advances in the understanding of the molecular mechanism by which cutaneous HPV-encoded proteins may contribute towards tumour formation.

While the causative relationship between HPV infection and squamous cell cancers (SCCs) of the genital tract is well established, the role of HPV in the development of cutaneous malignancy is, as yet, unclear. However, there is increasing evidence showing the involvement of cutaneous HPV types in the process of skin carcinogenesis. Solar radiation, in particular the UV-B component of the spectrum, is one of the most important environmental carcinogens for humans. UV-B is known to induce mutations in genomic 2 and mitochondrial DNA 3, which makes it the most important aetiological agent in the development of non-melanoma skin cancer (NMSC) 4. NMSC is the most common cancer in Caucasian populations, with over one million cases reported in the USA and more than 60 000 cases occurring annually in the UK 5–7. Since HPV-associated NMSCs develop predominantly at sun-exposed body sites, a co-factor role for the virus in cancer development is thought to be the most likely way that the virus contributes towards NMSC development. While cutaneous and anogenital HPVs encode similar early region proteins that can participate in lesional development, it is now apparent that the cellular targets of the virally encoded proteins are different. In addition, while anogenital cancers typically contain at least one copy of HPV genome per cell and viral gene expression is required to maintain the transformed phenotype 8, HPV DNA load estimates in cutaneous cancers suggest that the virus is present in only 1 in 20–5000 cells 9, 10. These observations suggest that a new paradigm for understanding the role of HPV in NMSC must be envisaged.

HPV and its association with NMSC

  1. Top of page
  2. Abstract
  3. Introduction
  4. HPV and its association with NMSC
  5. Transforming properties of cutaneous HPV early region proteins
  6. Molecular mechanisms
  7. Anti-apoptotic effects of HPV in response to UV irradiation
  8. HPV and potential cancer stem cells in the epidermis
  9. Genetic abnormalities in NMSC
  10. Summary
  11. Acknowledgements
  12. References

Epidermodysplasia verruciformis

The association of HPV infection and cutaneous carcinomas dates back many years. Epidermodysplasia verruciformis (EV) was described by Lewandowsky and Lutz in 1922 11 and was the first evidence for the involvement of HPV infection in the development of skin cancer 12, 13. EV is a rare heritable disease, which is characterized by a predisposition to infection with specific types of HPV, named EV-HPV types (now termed β-HPV types), including HPV types 5, 8, 9, 12, 14, 15, 17, and 19–25. In contrast to the presence of multiple types in benign lesions, mostly HPV 5 or 8 and sometimes HPV 14, 17, 20 or 47 are found in SCC and are regarded as high-risk types 9. In early childhood, the disease results in the development of warts and characteristic macular lesions (Figure 1), which occur predominantly on the face, dorsa of the hands, and legs, and which usually persist throughout life without a tendency to regress 14. About 30–60% of the patients develop carcinoma in situ or SCC at sunlight-exposed body sites by the fourth decade 12, 15, 16. A characteristic of this disorder appears to be a specific inability to control HPV infection at the keratinocyte level. The persistence of β-HPV infection in EV has been suggested to be due to the inability of the patient's immune system to reject the EV-HPV-harbouring keratinocytes, by a still unknown immunogenetic defect 17–19. Partial defects in cell-mediated immunity, such as inhibition of natural killer cell activity 20 and of cytotoxic T cells 21, have been demonstrated in EV patients and it is likely that interacting immunogenetic and environmental factors, particularly UV radiation, are important 17.

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Figure 1. Organotypic raft cultures were grown for 14 days at the air–liquid interface, conditions that induce epidermal differentiation. After embedding in paraffin wax, 4 µm sections were stained with haematoxylin and eosin for histological examination. Cells infected with the empty retrovirus (A) generate a normal epithelium with the distinct strata of keratinocyte differentiation on top of the dermis. The epidermis generated by keratinocytes expressing HPV 8 E6–E7 oncoproteins (B) shows parakeratosis; the basal cells look dysplastic and have pleiomorphic nuclei. (C) Normal human skin. (D) The characteristic cytopathic effect of an EV-specific HPV in a patient found to be infected with HPV 8. Histology shows pronounced vacuolation of the keratinocytes in the supraspinous and granular layers. Original magnification: (A–D) ×200

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The first susceptibility locus to EV (EV1) was initially mapped to a region of chromosome 17 where the p53 gene is also located 22. The second susceptibility locus for EV (EV2) was mapped to chromosome 2 23. At the EV1 locus, two EV sensitivity genes (EVER1 and EVER2) have recently been discovered and EV-associated mutations have been identified 24, 25. The long splice isoforms of EVER1 and EVER2 are identical in sequence and genomic localization to the transmembrane channel-like (TMC) proteins TMC6 and TMC8 26. The EVER1 and EVER2 gene products have features of integral membrane proteins and are localized in the endoplasmic reticulum when expressed in a human keratinocyte cell line 24. It is, at present, unclear how the EVER1 and EVER2 genes are involved in the innate or adaptive immune responses to control EV-HPV infection in epidermal keratinocytes. Future work is needed to determine the function of EVER proteins to define their mechanism of action and their role in the control of EV-HPV infection. This should open new perspectives for the study of the susceptibility to oncogenic cutaneous HPVs.

HPV-associated cancers in immunosuppressed patients and the general population

Historically, the second model of HPV-induced NMSC to receive attention was that of skin cancer occurring in the context of immunosuppression, particularly organ transplantation 27, 28. Renal transplant recipients are highly susceptible to extensive wart infection and have a markedly increased (∼200-fold) incidence of cutaneous SCCs, which arise on sun-exposed body sites 29. Highly sensitive nested PCR has recently been shown to be able to detect EV-associated HPV DNA in a significant proportion of skin tumours 30–33 and using the in situ hybridization technique, gene expression of EV-HPV has also been demonstrated in SCCs 34. The percentage of HPV DNA-positive NMSCs in immunosuppressed patients is about 80%, rising in SCC and premalignancies 32, 35–37, sometimes even reaching 100% 9. EV-HPVs have also been found in normal skin 38–40 and NMSCs 41 in the immunocompetent general population, with detection rates of about 30% for SCC and basal cell carcinoma (BCC) 32, 42, 43. A high prevalence (85%) of EV-HPV DNA in actinic keratoses, which are precursor lesions of SCC in the immunocompetent population 44, could be compatible with a carcinogenic role for HPV in the early stages of skin tumour development. During these stages, the virus stimulates keratinocyte proliferation which may cause enhanced HPV gene expression leading to the first steps in NMSC development.

HPV and other epidermal lesions

In addition to being strongly associated with the development of cervical carcinomas, high-risk mucosal HPVs are also associated with the development of other specific epidermal cancers. HPV 16 DNA is frequently found in vulval disease 45 and expression of surrogate markers of HPV infection, such as p16(INK4a) 46, may be valuable in the diagnosis of vulval premalignant and malignant lesions 47 and in NMSC 48. HPV 16 and other mucosal viruses have also been implicated in the development of periungual lesions, where histology often mimics an association with a wart infection 49, 50. Mechanistic investigations of the role of mucosal HPV in carcinogenesis in these settings may provide valuable insights into skin carcinogenesis in the future. In addition to cancers, cutaneous HPVs have also been detected and proposed to be involved in the development of psoriasis 51, 52. EV-HPV types, in particular HPV types 5 and 36, were detected at high frequency in children with psoriasis 53. In further studies, HPV 5 has also been detected at high frequency in psoriatic plaques 54, and HPV DNA positivity in plucked hairs from psoriatic patients undergoing PUVA treatment has also been noted 55. However, further work will be required to establish whether HPV contributes to the development of psoriasis or is a passenger in these hyperproliferative lesions.

Transforming properties of cutaneous HPV early region proteins

  1. Top of page
  2. Abstract
  3. Introduction
  4. HPV and its association with NMSC
  5. Transforming properties of cutaneous HPV early region proteins
  6. Molecular mechanisms
  7. Anti-apoptotic effects of HPV in response to UV irradiation
  8. HPV and potential cancer stem cells in the epidermis
  9. Genetic abnormalities in NMSC
  10. Summary
  11. Acknowledgements
  12. References

Monolayer cell cultures

In the first experiments using monolayer cultures, the E6 genes of HPVs 5, 8, 14, 21, 25, and 47 were shown to be sufficient to induce anchorage-independent growth and morphological transformation in some established rodent cell lines such as mouse C127, rat 3Y1, and Rat1 56–58. The E7 genes of HPV 8 and HPV 47 failed to induce any detectable transformation in rodent cells 56–59. In co-operation with the activated H-ras gene, however, HPV 5 and HPV 8 E7 gave rise to transformed cell lines 60 and the E7 gene of HPV 1, which is specific for plantar warts, fully transformed the mouse fibroblast line C127 61. In 1994, Schmitt et al61 were the first to study the immortalization capability of cutaneous HPV E6 and E7 genes in primary human foreskin keratinocytes. The E6 and E7 genes of HPV 1, which showed remarkable transforming activities in rodent cell lines, were unable to induce immortalization of human keratinocytes, although weak induction could be observed with HPV 8 E7. In a recent publication, Caldeira et al62 demonstrated that the E6 and E7 proteins of the EV-associated HPV 38, but not HPV 10 and HPV 20, are sufficient to increase the life span of primary keratinocytes.

Cutaneous HPV in organotypic keratinocyte cultures

The dependence of the viral life cycle on cellular differentiation together with the difficulty of generating a fully differentiated stratified epithelium in vitro has hampered the investigation of cutaneous HPV gene function. Organotypic collagen raft cultures using foreskin keratinocytes expressing the E6 and E7 genes of the EV-types HPV 5, 12, 15, 17, 20, and 38 showed no invasive phenotype of the epithelial cells 63. Using a de-epidermalized dermis-based raft culture system (Figure 1), it was demonstrated recently that expression of the E7 gene of the cutaneous HPV 8 in primary human adult keratinocytes induces epithelial hyperproliferation and hypercornification 64. In addition, and most significantly, it promotes a tumourigenic phenotype, as evidenced by the invasive behaviour of the HPV-transduced keratinocytes. Migration of the keratinocytes downward into the dermis was facilitated by the degradation of components of the basement membrane and the extracellular matrix (collagen VII, collagen IV, and laminin V) through induction of the expression of matrix metalloproteinase (MMP) 1, MMP-8, and MT1-MMP. These in vitro findings are in accord with similar observations in skin cancer, thereby supporting a direct role for the E7 protein of HPV 8 in NMSC development. In addition, the invasive phenotype was only observed in cultures generated using the cancer-associated HPV 8, and was not seen in experiments using the E6 and E7 genes of HPV 2 or HPV 3, viral types associated with the development of hyperproliferative and flat warts, respectively (A Storey, unpublished data). In another study, Smola-Hess et al65 confirmed that HPV 8 E7, but not HPV 1 E7, was sufficient to induce MT1-MMP expression in primary human keratinocytes and HaCaT cells in monolayer cultures.

There is preliminary evidence that the E2 protein of HPV 8, in addition to being involved in viral transcription and replication, may also play a role in virus-induced cell transformation. Expression of HPV 8 E2 in rodent fibroblasts led to anchorage-independent cell growth and reduced serum requirement 66 and allowed colony formation in soft agar when transfected into the human skin keratinocyte cell line HaCaT 67. Recent reports have shown that HPV 8 E2 can interact with cellular factors such as C/EBP, p300, and p53 68–70, or regulate the activity of promoters of genes such as p21 or β-integrin, all of which are involved in epithelial differentiation 71, 72. Moreover, when HPV 8 E2 was expressed in de-epidermalized raft cultures, it perturbed epithelial differentiation with coincident expression of MMP-9. Transient reporter gene assays in human adult primary epidermal keratinocytes showed that HPV 8 E2 activated the MMP-9 promoter in a dose-dependent manner (Garcia-Escudero et al, unpublished data).

Transgenic animals

The first transgenic mouse model for skin-associated HPVs was generated by Tinsley et al in 1992 (73) by expressing the early gene region of HPV1, which induces benign plantar warts, under the control of the keratin-6 promoter. Expression of the transgene led to transient epidermal hyperproliferation and abnormal keratinocyte differentiation. Pfister and co-workers have now expressed the complete early region of HPV 8 in murine epidermal basal keratinocytes under the control of the keratin-14 promoter. Over 90% of the transgenic mice spontaneously developed benign skin tumours with significant conversion to SCCs 74. Molecular studies on the skin tumours of these transgenic animals revealed strong gelatinolytic activity, which could be attributed to MMP-9 and the up-regulation and activation of MMP-13 and MT1-MMP 75. The high penetrance of papillomatosis followed by progression to dysplasia is remarkable and in contrast to many other HPV-transgenic mice models 73, 76, 77. Only the high penetrance of papillomatosis and dysplasia in K14-HPV 16 transgenic mice 78 is comparable with HPV 8 transgenics.

Molecular mechanisms

  1. Top of page
  2. Abstract
  3. Introduction
  4. HPV and its association with NMSC
  5. Transforming properties of cutaneous HPV early region proteins
  6. Molecular mechanisms
  7. Anti-apoptotic effects of HPV in response to UV irradiation
  8. HPV and potential cancer stem cells in the epidermis
  9. Genetic abnormalities in NMSC
  10. Summary
  11. Acknowledgements
  12. References

Cutaneous HPV and cell cycle defects

Extensive studies have linked the efficiency of mucosal HPV types in promoting cancer development to the viral E6 and E7 proteins 79. E7 is an oncogene that mediates the initiation of DNA synthesis and transforms cells into continuously growing cell clones 80. It thereby induces centrosome-associated mitotic abnormalities that lead to aneuploidy and enhance the likelihood of malignant progression 81–83.

The E7 protein can interact with the retinoblastoma protein (pRb), an important negative regulator of entry into the S phase of the cell division cycle. It was thought that only the high-risk mucosal HPVs have the unique ability to induce the proteolytic degradation of pRb 84. Even with the limited number of different cutaneous types that have been investigated so far, there is a large variation in the degree to which the E7 protein can interact with pRb. Relative to HPV 16 E7 (100%), the binding efficiency of HPV 38 E7 to pRb is similar to that of HPV 16 62, whereas HPV 1 E7 binds only 66% and HPV 8 E7 34% as effectively 61, while HPV 10 E7 and HPV 20 E7 bind very weakly 62. Surprisingly, the cutaneous HPV 38 E7 is able to promote the degradation of pRb 62, but HPV 8 E7, in spite of having a weaker binding efficiency to pRb, is also able to inactivate pRb and deregulate G1/S transition control (B Akgül, unpublished data). It seems that the ability to degrade pRb is shared among mucosal and cutaneous high-risk HPV types.

The E6 protein has also been associated with cell cycle effects. Accumulation of the p53 target p21 has been observed in skin cells expressing cutaneous HPV 5 or HPV 77 E6 following UV damage 85. No growth arrest was apparent in these cells, in contrast to the fact that this accumulation would result in p21-induced cell cycle arrest in the normal setting. It appears, therefore, that E6 possesses the intrinsic ability to abrogate the p21 response to cellular stress, and consequently sustain the cell cycle capability of HPV-infected cells, akin to that observed with HPV 18 E6 86.

HPV and UV radiation

The link between UV radiation, HPV, and NMSC was clearly highlighted by EV patients. UV is a component of sunlight and is composed of three regions, subdivided according to wavelength: UV-A (320–400 nm), UV-B (280–320 nm), and UV-C (200–280 nm). UV radiation has been shown to play a significant aetiological role in skin cancer development 87, primarily due to the DNA-damaging properties of UV-B. UV-B-mediated DNA damage can cause mutations in key genes; ultimately, the accumulation of gene mutations in key cellular pathways is the force driving tumourigenesis in UV-induced skin cancer. UV-B irradiation is able to induce lesions in DNA, in the form of mutagenic photoproducts between adjacent pyrimidine residues resulting in the formation of dimers. These dimers consist of two types, cyclobutane pyrimidine dimers (CPDs) and 6-4 photoproducts. Both types of DNA lesion are potentially mutagenic and occur more frequently in tandem pyrimidine ‘hot spot’ residues 88. Efficient repair of CPDs in preventing NMSC is highlighted by the increased incidence of NMSC in xeroderma pigmentosum (XP) patients. These patients have a defect in the nucleotide excision repair (NER) DNA repair pathway 89.

The role that E6 and E7 play in uncoupling keratinocytes from a state of terminal differentiation to allow cell proliferation and thus viral replication has been previously described in this review. The ultimate goal of the virus is to induce a cellular state in which it can replicate efficiently in post-mitotic terminally differentiated cells. An apoptotic response to cellular stress such as hypoxia, excess DNA damage or an intrinsic response to viral infection would result in cell death, and an obvious disadvantage to the virus 90. This highlights the importance of UV irradiation in the cutaneous setting, as skin cells at sun-exposed sites are constantly exposed to this potentially mutagenic radiation. As such, the HPVs have evolved a mechanism to subvert this biological pathway.

Anti-apoptotic effects of HPV in response to UV irradiation

  1. Top of page
  2. Abstract
  3. Introduction
  4. HPV and its association with NMSC
  5. Transforming properties of cutaneous HPV early region proteins
  6. Molecular mechanisms
  7. Anti-apoptotic effects of HPV in response to UV irradiation
  8. HPV and potential cancer stem cells in the epidermis
  9. Genetic abnormalities in NMSC
  10. Summary
  11. Acknowledgements
  12. References

HPV and p53

The tumour suppressor gene p53 acts as a key regulator of apoptosis in many tissues. In the normal setting, p53 is activated and stabilized through key acetylation and phosphorylation events 91 in response to UV-B-induced DNA damage. Regulation of the expression of p53 targets results in either cell cycle arrest, to allow DNA repair before continuation of the cell cycle and DNA replication, or, in the case of large amounts of DNA damage, induction of the apoptotic cascade to allow clearance of damaged cells. To bypass the apoptotic response to cellular stress, HPVs have developed a variety of tissue-specific strategies to eliminate key mediators of the apoptotic cascade, based on the ability of the E6 protein to target these proteins for proteasomal degradation. The E6 proteins of high-risk mucosal HPVs (such as HPV 16 or HPV 18) are able to target p53 for proteasomal degradation by recruitment of the E6AP E3 ubiquitin ligase 92, 93. Thus, elimination of p53 allows HPV-infected cells to escape apoptosis, thus providing a background in which normal growth restraints are absent, allowing viral replication 94. Mouse models have suggested an important role for p53 in skin cancer development. Knockout p53 mice spontaneously develop skin cancer 95, 96 and transgenic mice with concurrent p53 mutations or deletions are more susceptible to UV-induced carcinogenesis 97, 98.

However, unlike mucosal HPVs, in the skin E6 proteins derived from cutaneous HPV types are unable to target p53 for proteasomal degradation 62, 94, 99, 100. E6 proteins can, however, effectively inhibit apoptosis in cells carrying wild-type p53 100. The ability of cutaneous viruses to inhibit UV-induced apoptosis under conditions that induce p53 activation supports the premise that viral proteins may specifically interfere with p53-mediated pathways. Subsequently, the cutaneous virus HPV 77 E6 has been shown to inhibit downstream transcriptional targets of UV-activated p53 selectively. Moreover, E6 inhibits transcription of the pro-apoptotic genes Fas, PUMAβ, Apaf-1, and PIG3, whereas the expression level of p53 cell-cycle and regulatory targets p21 and Hdm2 are unaffected 101.

A weight of data suggesting that p53-independent apoptotic pathways play important roles in the skin now exists. Clonal patches of mutant p53 cells within sun-exposed sites have been shown to have little or no pre-cancerous potential 102, 103. In addition, patients with the Li Fraumeni syndrome, who lack one functional copy of p53, are not pre-disposed to NMSC, but are prone to the development of other cancers 104. In murine systems, UV-B-irradiated epidermal keratinocytes carrying mutant p53 are normally constrained within their epidermal compartments. However, chronic UV-B exposure drives clonal expansion of the mutant keratinocytes by a non-mutational mechanism which involves UV-B-mediated apoptosis that preferentially deletes DNA-damaged unmutated cells, leaving death-resistant p53-mutant cells to expand into the vacant niche 105. It is tempting to speculate that HPV-infected cells, where the pro-apoptotic function of p53 has been ablated, may mirror observations in mice, where cells expressing HPV E6 are permitted to proliferate selectively. This allows a greater pool of apoptosis-resistant cells to develop that can go on to acquire further UV-induced mutations that facilitate tumour progression.

Other p53-independent apoptotic pathways

In addition to inhibiting the UV-induced apoptotic response involving p53 transactivation of target genes, HPV E6 proteins are also able to inhibit apoptosis in a p53 transcription-independent mechanism (reviewed in ref 106). In UV-irradiated cells, p53 rapidly translocates to the mitochondria where it induces Bak oligomerization, thereby permeabilizing the mitochondrial membrane for cytochrome c release 107, 108. The Bak protein is a key pro-apoptotic effector located in the outer mitochondrial membrane and is expressed in human epidermal keratinocytes 109, 110. Following UV irradiation, Bak becomes activated and stabilized in monolayer cells, regenerated epidermal sheets, and skin explants maintained in organ culture. Both mucosal 111, 112 and cutaneous HPV E6 proteins 100 target Bak for proteolytic degradation. Furthermore, studies on NMSC biopsies showed that while HPV-negative lesions had a high rate of both proliferation and apoptosis, little apoptosis was observed in HPV-positive lesions while the proliferation rate remained unchanged. These findings suggested that HPV gene expression had altered the balance between proliferation and apoptosis in these lesions, favouring expansion of the tumour 100, 113 (Figure 2). Hence, by abrogating the activity of Bak, cutaneous E6 proteins also inhibit the transcription-independent mechanism of p53-induced apoptosis even though the p53 protein is not targeted for destruction. It is interesting to note that the E6AP ubiquitin ligase machinery associated with p53 degradation has not been formally identified as being responsible for the proteasomal destruction of Bak mediated by the cutaneous E6 proteins, and it is possible that E6 may recruit a novel ubiquitination complex for Bak degradation.

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Figure 2. Prevention of apoptosis in HPV-positive SCC. Apoptotic cells were detected using TUNEL, and Bak levels determined by immunohistochemistry, in HPV-positive and HPV-negative SCC (original magnification: ×100). T = tumour; E = epidermis

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In summary, the E6 protein of cutaneous viruses has evolved a mechanism of averting apoptosis in the skin through complex interactions with members of the apoptotic cascade. Ultimately, further investigations are needed to unravel the exact molecular mechanisms underlying this phenomenon and the requirement for E6 anti-apoptotic activity in tumour formation and progression. Inhibition of the anti-apoptotic activity of E6 has potentially important clinical implications, since blockade of the proteasome using inhibitors restores Bak levels in E6-expressing cells that then undergo apoptosis 100.

Interference of DNA repair pathways by HPV

Expression of the E6 protein of high-risk HPV types (β-types) has been shown to play an important role in genomic stability. Specifically, E6 allows skin cells to progress through the G1/S cell-cycle checkpoint. Following UV irradiation, at doses insufficient to induce apoptosis, cells arrest in G0 to carry out DNA repair (reviewed in ref 114). Skin cells expressing the high-risk HPV 5 E6 protein take longer to repair UV-B-induced DNA damage, and enter the cell cycle harbouring DNA damage, specifically bypassing the G1/S checkpoint. This is in contrast to the E6 proteins of HPV 10 (found in plantar warts), HPVs 23, 24, 49 (EV types rarely found in cancers), and HPV 77 (found in NMSCs of renal transplant recipients) 115. Interestingly, E6 proteins of HPV 1 and HPV 8 have been shown to interact with the XRCC1 protein that is involved in the repair of DNA single-strand breaks 116. Thus, E6 appears to help cells to overcome cell cycle checkpoints, potentially resulting in the propagation of UV-induced DNA damage. If cells enter the cell cycle, this could lead to the replication of mutated DNA, which, in concert with the anti-apoptotic activity of E6, could lead to the generation of progeny cells with tumourigenic potential. Therefore, cells expressing E6 may be more genetically unstable and accumulate errors in key genes, which in turn drives tumourigenesis in the setting of NMSC. A working model summarizing the potential role of HPV in NMSC initiation and progression is depicted in Figure 3.

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Figure 3. Model of the roles of UV irradiation and HPV infection in skin cancer development. HPV infection acts as an initiator mechanism for SCC development, in combination with UV, on sun-exposed body sites. Exposure to UV leads to DNA damage in skin cells. In HPV-infected cells, E6 promotes retention of DNA damage and protects cells from UV-mediated apoptosis. Normal cells that are fully competent for apoptosis are preferentially deleted following UV damage, allowing clonal expansion of HPV-containing cells. Accumulation of mutations/genetic damage allows cells to progress to SCC independently of the proliferative and mutagenic effects of HPV infection

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HPV and potential cancer stem cells in the epidermis

  1. Top of page
  2. Abstract
  3. Introduction
  4. HPV and its association with NMSC
  5. Transforming properties of cutaneous HPV early region proteins
  6. Molecular mechanisms
  7. Anti-apoptotic effects of HPV in response to UV irradiation
  8. HPV and potential cancer stem cells in the epidermis
  9. Genetic abnormalities in NMSC
  10. Summary
  11. Acknowledgements
  12. References

In the course of the normal viral life cycle, HPV is presumed to infect cells of the basal layer. Considering the transit time of cells through the differentiation programme of the epidermis from the basal layers to being shed, HPV must reside in cells that are retained for long periods in the epidermis, ie cells in the epidermal stem cell niche, to establish a persistent infection. Whether HPV infects and is maintained in epidermal stem cells, or confers stem-like properties to more differentiated cells, is at present unknown. High-risk cutaneous HPV DNA has been found in plucked eyebrow hairs that contain cells in the hair follicle stem cell niche, although the exact target cell of the HPV was not determined in this case 39, 117. Such HPV-infected persistent cells can then acquire multiple genetic lesions as outlined above, such that they may be considered effectively as cancer stem cells (reviewed in ref 118). In addition, it is possible that HPV may also influence the self-renewal capacity of these cells, leading to the generation of additional cancer stem cells by interfering with mechanisms controlling asymmetric cell division 119 that may prevent differentiation and stratification of progeny cells 120, thereby driving NMSC development. A better understanding of the effects of HPV on keratinocyte stem cells may be important for disease management and treatment in the future.

Genetic abnormalities in NMSC

  1. Top of page
  2. Abstract
  3. Introduction
  4. HPV and its association with NMSC
  5. Transforming properties of cutaneous HPV early region proteins
  6. Molecular mechanisms
  7. Anti-apoptotic effects of HPV in response to UV irradiation
  8. HPV and potential cancer stem cells in the epidermis
  9. Genetic abnormalities in NMSC
  10. Summary
  11. Acknowledgements
  12. References

The molecular genetics of NMSC has been widely studied in both BCC and SCC. In this context, the study of loss of heterozygosity (LOH) patterns has provided important insights into the genetic abnormalities underlying these tumour types. In relation to BCC, the study of the familial naevoid basal cell carcinoma (NBCC) syndrome by LOH analysis revealed consistent loss of chromosome arm 9q 121, 122. This region contains the tumour suppressor gene PTCH, which maps to 9q22–31, and is an important regulator of the sonic hedgehog-signalling pathway. Germline inactivation of PTCH is a common event in NBCC 123, 124 and LOH is commonly observed at 9q22 125–127. Mutations in PTCH can also be detected in sporadic forms of BCC 128. Therefore, PTCH and regulation of the sonic hedgehog pathway play an important role in BCC development, and studies are ongoing into its use as a therapeutic target (reviewed in ref 129). PTCH mutations can be found in SCCs from patients with a history of multiple BCCs 130. However, the role of PTCH, if any, in SCC remains to be elucidated. The molecular genetics of SCC is less clear, however. LOH analysis of SCC tumours where the HPV status was not determined revealed widespread loss of markers from as many as 39 chromosomal arms, with frequent loss observed at 9p, 13q, 17p, 17q, and 3p 131. Therefore, unlike BCC, there is no distinct association between a specific genetic abnormality and predisposition to SCC. It is, however, likely that key genes are involved in the development of SCC, and they may or may not be associated with these regions of LOH. 9p21 loss, containing the p16INK4a and p14ARF tumour suppressor genes, has been shown to be a common event in SCC tumours. Furthermore, p16INK4a and p14ARF are commonly inactivated in SCC, by promoter hypermethylation 132. It is likely that these two genes are important in the development of SCC and that further genetic and epigenetic mechanisms play a role in SCC development. How p16INK4a and p14ARF are inactivated in HPV-positive cells, and whether different patterns of LOH are observed in HPV-positive and -negative cancers, may provide important clues to discovering tumour suppressor genes inactivated in NMSC.

Specific underlying genetic defects may predispose to HPV infection and disease. One example of this is the association between severe combined immune deficiency (SCID) in humans and predisposition to chronic HPV disease. SCID is caused by several known genetic defects 133 and many SCID sufferers have chronic HPV disease. Chronic HPV infections often develop late following corrective haemopoietic stem-cell transplantation and are limited to the skin, and in many cases present with lesions typical of EV patients. The EV HPV types 5, 14, and 36, and the wart-causing types 2, 3, and 57 are commonly isolated from SCID patients 134. The mechanism promoting HPV infection in SCID patients is linked to declining or insufficient immune function, but the model elegantly demonstrates the potential molecular events that may underlie HPV-associated NMSC development.

Summary

  1. Top of page
  2. Abstract
  3. Introduction
  4. HPV and its association with NMSC
  5. Transforming properties of cutaneous HPV early region proteins
  6. Molecular mechanisms
  7. Anti-apoptotic effects of HPV in response to UV irradiation
  8. HPV and potential cancer stem cells in the epidermis
  9. Genetic abnormalities in NMSC
  10. Summary
  11. Acknowledgements
  12. References

HPV-encoded proteins abrogate a number of cellular networks, the primary objective of which is to enhance the viral life cycle. However, inappropriate activation or inactivation of these cellular systems generates conditions that favour and enhance tumour initiation and progression. While UV is the primary aetiological agent in NMSC development, it is now becoming clear that HPV infection acts as a co-carcinogen with UV in NMSC development. Studies on the molecular basis of cancer have, over the years, revealed a complex and varied series of epigenetic and genetic events that give rise to a particular cancer with a distinct aetiology. A limited number of critical events may underlie, and be common to, many cancers 135 and, of these, the deregulation of cell proliferation coupled to the inhibition of apoptosis is the most significant. These two events are required to support an expanding population of tumour cell progeny and may provide the minimum basic scaffold upon which further changes necessary for neoplastic progression may be superimposed. Our current model of the role of HPV in NMSC development (Figure 3), in which HPV both stimulates cell proliferation and inhibits apoptosis, creates a cellular environment where additional genetic changes that enhance NMSC progression can occur.

Acknowledgements

  1. Top of page
  2. Abstract
  3. Introduction
  4. HPV and its association with NMSC
  5. Transforming properties of cutaneous HPV early region proteins
  6. Molecular mechanisms
  7. Anti-apoptotic effects of HPV in response to UV irradiation
  8. HPV and potential cancer stem cells in the epidermis
  9. Genetic abnormalities in NMSC
  10. Summary
  11. Acknowledgements
  12. References

Our research programme is supported by funding from Cancer Research UK and the Research Advisory Board of the St Bartholomew's and The Royal London Charitable Foundation, in addition to grant awards from the Dr Mildred Scheel Stiftung für Krebsforschung/Deutsche Krebshilfe (BA) and The British Skin Foundation (JCC).

References

  1. Top of page
  2. Abstract
  3. Introduction
  4. HPV and its association with NMSC
  5. Transforming properties of cutaneous HPV early region proteins
  6. Molecular mechanisms
  7. Anti-apoptotic effects of HPV in response to UV irradiation
  8. HPV and potential cancer stem cells in the epidermis
  9. Genetic abnormalities in NMSC
  10. Summary
  11. Acknowledgements
  12. References
  • 1
    de Villiers EM, Fauquet C, Broker TR, Bernard HU, zur Hausen H. Classification of papillomaviruses. Virology 2004; 324: 1727.
  • 2
    Dahle J, Kvam E, Stokke T. Bystander effects in UV-induced genomic instability: antioxidants inhibit delayed mutagenesis induced by ultraviolet A and B radiation. J Carcinog 2005; 4: 11.
  • 3
    Durham SE, Krishnan KJ, Betts J, Birch-Machin MA. Mitochondrial DNA damage in non-melanoma skin cancer. Br J Cancer 2003; 88: 9095.
  • 4
    Sander CS, Hamm F, Elsner P, Thiele JJ. Oxidative stress in malignant melanoma and non-melanoma skin cancer. Br J Dermatol 2003; 148: 913922.
  • 5
    Stern RS. The mysteries of geographic variability in nonmelanoma skin cancer incidence. Arch Dermatol 1999; 135: 843844.
  • 6
    Jemal A, Clegg LX, Ward E, Ries LA, Wu X, Jamison PM, et al. Annual report to the nation on the status of cancer, 1975–2001, with a special feature regarding survival. Cancer 2004; 101: 327.
  • 7
    Goodwin RG, Holme SA, Roberts DL. Variations in registration of skin cancer in the United Kingdom. Clin Exp Dermatol 2004; 29: 328330.
  • 8
    Munoz N. Human papillomavirus and cancer: the epidemiological evidence. J Clin Virol 2000; 19: 15.
  • 9
    Pfister H. Chapter 8: Human papillomavirus and skin cancer. J Natl Cancer Inst Monogr 2003; 5256.
  • 10
    Weissenborn SJ, Nindl I, Purdie K, Harwood C, Proby C, Breuer J, et al. Human papillomavirus-DNA loads in actinic keratoses exceed those in non-melanoma skin cancers. J Invest Dermatol 2005; 125: 9397.
  • 11
    Lewandowsky F, Lutz W. Ein Fall einer bisher nicht beschriebenen Hauterkrankungn (Epidermodysplasia verruciformis). Arch Dermatol Syphilol 1922; 141: 193203.
  • 12
    Orth G. Epidermodysplasia Verruciformis. Plenum Press: London, 1987.
  • 13
    Jablonska S, Majewski S. Epidermodysplasia verruciformis: immunological and clinical aspects. Curr Top Microbiol Immunol 1994; 186: 157175.
  • 14
    Jablonska S, Dabrowski J, Jakubowicz K. Epidermodysplasia verruciformis as a model in studies on the role of papovaviruses in oncogenesis. Cancer Res 1972; 32: 583589.
  • 15
    Tanigaki T, Kanda R, Yutsudo M, Hakura A. Epidemiologic aspects of epidermodysplasia verruciformis (L-L 1922) in Japan. Jpn J Cancer Res 1986; 77: 896900.
  • 16
    Jablonska S. Epidermodysplasia Verruciformis. WB Saunders: Philadelphia, 1991.
  • 17
    Majewski S, Jablonska S. Epidermodysplasia verruciformis as a model of human papillomavirus-induced genetic cancer of the skin. Arch Dermatol 1995; 131: 13121318.
  • 18
    de Oliveira WR, Festa Neto C, Rady PL, Tyring SK. Clinical aspects of epidermodysplasia verruciformis. J Eur Acad Dermatol Venereol 2003; 17: 394398.
  • 19
    Majewski S, Jablonska S. Do epidermodysplasia verruciformis human papillomaviruses contribute to malignant and benign epidermal proliferations? Arch Dermatol 2002; 138: 649654.
  • 20
    Majewski S, Malejczyk J, Jablonska S, Misiewicz J, Rudnicka L, Obalek S, et al. Natural cell-mediated cytotoxicity against various target cells in patients with epidermodysplasia verruciformis. J Am Acad Dermatol 1990; 22: 423427.
  • 21
    Cooper KD, Androphy EJ, Lowy D, Katz SI. Antigen presentation and T-cell activation in epidermodysplasia verruciformis. J Invest Dermatol 1990; 94: 769776.
  • 22
    Ramoz N, Rueda LA, Bouadjar B, Favre M, Orth G. A susceptibility locus for epidermodysplasia verruciformis, an abnormal predisposition to infection with the oncogenic human papillomavirus type 5, maps to chromosome 17qter in a region containing a psoriasis locus. J Invest Dermatol 1999; 112: 259263.
  • 23
    Ramoz N, Taieb A, Rueda LA, Montoya LS, Bouadjar B, Favre M, et al. Evidence for a nonallelic heterogeneity of epidermodysplasia verruciformis with two susceptibility loci mapped to chromosome regions 2p21–p24 and 17q25. J Invest Dermatol 2000; 114: 11481153.
  • 24
    Ramoz N, Rueda LA, Bouadjar B, Montoya LS, Orth G, Favre M. Mutations in two adjacent novel genes are associated with epidermodysplasia verruciformis. Nature Genet 2002; 32: 579581.
  • 25
    Tate G, Suzuki T, Kishimoto K, Mitsuya T. Novel mutations of EVER1/TMC6 gene in a Japanese patient with epidermodysplasia verruciformis. J Hum Genet 2004; 49: 223225.
  • 26
    Keresztes G, Mutai H, Heller S. TMC and EVER genes belong to a larger novel family, the TMC gene family encoding transmembrane proteins. BMC Genomics 2003; 4: 24.
  • 27
    Walder BK, Robertson MR, Jeremy D. Skin cancer and immunosuppression. Lancet 1971; 2: 12821283.
  • 28
    Boyle J, MacKie RM, Briggs JD, Junor BJ, Aitchison TC. Cancer, warts, and sunshine in renal transplant patients. A case–control study. Lancet 1984; 1: 702705.
  • 29
    Stockfleth E, Ulrich C, Meyer T, Arndt R, Christophers E. Skin diseases following organ transplantation—risk factors and new therapeutic approaches. Transplant Proc 2001; 33: 18481853.
  • 30
    Berkhout RJ, Tieben LM, Smits HL, Bavinck JN, Vermeer BJ, ter Schegget J. Nested PCR approach for detection and typing of epidermodysplasia verruciformis-associated human papillomavirus types in cutaneous cancers from renal transplant recipients. J Clin Microbiol 1995; 33: 690695.
  • 31
    Pfister H, Ter Schegget J. Role of HPV in cutaneous premalignant and malignant tumors. Clin Dermatol 1997; 15: 335347.
  • 32
    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: 289297.
  • 33
    Harwood CA, Proby CM. Human papillomaviruses and non-melanoma skin cancer. Curr Opin Infect Dis 2002; 15: 101114.
  • 34
    Purdie KJ, Surentheran T, Sterling JC, Bell L, McGregor JM, Proby CM, et al. Human papillomavirus gene expression in cutaneous squamous cell carcinomas from immunosuppressed and immunocompetent individuals. J Invest Dermatol 2005; 125: 98107.
  • 35
    de Jong-Tieben LM, Berkhout RJ, Smits HL, Bouwes Bavinck JN, Vermeer BJ, van der Woude FJ, et al. High frequency of detection of epidermodysplasia verruciformis-associated human papillomavirus DNA in biopsies from malignant and premalignant skin lesions from renal transplant recipients. J Invest Dermatol 1995; 105: 367371.
  • 36
    de Villiers EM, Lavergne D, McLaren K, Benton EC. Prevailing papillomavirus types in non-melanoma carcinomas of the skin in renal allograft recipients. Int J Cancer 1997; 73: 356361.
  • 37
    Berkhout RJ, Bouwes Bavinck JN, ter Schegget J. Persistence of human papillomavirus DNA in benign and (pre)malignant skin lesions from renal transplant recipients. J Clin Microbiol 2000; 38: 20872096.
  • 38
    Astori G, Lavergne D, Benton C, Hockmayr B, Egawa K, Garbe C, et al. Human papillomaviruses are commonly found in normal skin of immunocompetent hosts. J Invest Dermatol 1998; 110: 752755.
  • 39
    Boxman IL, Russell A, Mulder LH, Bavinck JN, Schegget JT, Green A. Case–control study in a subtropical Australian population to assess the relation between non-melanoma skin cancer and epidermodysplasia verruciformis human papillomavirus DNA in plucked eyebrow hairs. The Nambour Skin Cancer Prevention Study Group. Int J Cancer 2000; 86: 118121.
  • 40
    Antonsson A, Erfurt C, Hazard K, Holmgren V, Simon M, Kataoka A, et al. Prevalence and type spectrum of human papillomaviruses in healthy skin samples collected in three continents. J Gen Virol 2003; 84: 18811886.
  • 41
    Masini C, Fuchs PG, Gabrielli F, Stark S, Sera F, Ploner M, et al. Evidence for the association of human papillomavirus infection and cutaneous squamous cell carcinoma in immunocompetent individuals. Arch Dermatol 2003; 139: 890894.
  • 42
    de Villiers EM. Human papillomavirus infections in skin cancers. Biomed Pharmacother 1998; 52: 2633.
  • 43
    Wieland U, Ritzkowsky A, Stoltidis M, Weissenborn S, Stark S, Ploner M, et al. Communication: papillomavirus DNA in basal cell carcinomas of immunocompetent patients: an accidental association? J Invest Dermatol 2000; 115: 124128.
  • 44
    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: 273279.
  • 45
    Lerma E, Matias-Guiu X, Lee SJ, Prat J. Squamous cell carcinoma of the vulva: study of ploidy, HPV, p53, and pRb. Int J Gynecol Pathol 1999; 18: 191197.
  • 46
    von Knebel Doeberitz M. New markers for cervical dysplasia to visualise the genomic chaos created by aberrant oncogenic papillomavirus infections. Eur J Cancer 2002; 38: 22292242.
  • 47
    Rufforny I, Wilkinson EJ, Liu C, Zhu H, Buteral M, Massoll NA. Human papillomavirus infection and p16(INK4a) protein expression in vulvar intraepithelial neoplasia and invasive squamous cell carcinoma. J Low Genit Tract Dis 2005; 9: 108113.
  • 48
    Nindl I, Meyer T, Schmook T, Ulrich C, Ridder R, Audring H, et al. Human papillomavirus and overexpression of P16INK4a in nonmelanoma skin cancer. Dermatol Surg 2004; 30: 409414.
  • 49
    Moy RL, Eliezri YD, Nuovo GJ, Zitelli JA, Bennett RG, Silverstein S. Human papillomavirus type 16 DNA in periungual squamous cell carcinomas. J Am Med Assoc 1989; 261: 26692673.
  • 50
    Sato T, Morimoto A, Ishida Y, Matsuo I. Human papillomavirus associated with Bowen's disease of the finger. J Dermatol 2004; 31: 927930.
  • 51
    Favre M, Orth G, Majewski S, Baloul S, Pura A, Jablonska S. Psoriasis: a possible reservoir for human papillomavirus type 5, the virus associated with skin carcinomas of epidermodysplasia verruciformis. J Invest Dermatol 1998; 110: 311317.
  • 52
    Weissenborn SJ, Hopfl R, Weber F, Smola H, Pfister HJ, Fuchs PG. High prevalence of a variety of epidermodysplasia verruciformis-associated human papillomaviruses in psoriatic skin of patients treated or not treated with PUVA. J Invest Dermatol 1999; 113: 122126.
  • 53
    Mahe E, Bodemer C, Descamps V, Mahe I, Crickx B, De Prost Y, et al. High frequency of detection of human papillomaviruses associated with epidermodysplasia verruciformis in children with psoriasis. Br J Dermatol 2003; 149: 819825.
  • 54
    Prignano G, Ferraro C, Mussi A, Stivali F, Trento E, Bordignon V, et al. Prevalence of human papilloma virus type 5 DNA in lesional and non-lesional skin scales of Italian plaque-type psoriatic patients: association with disease severity. Clin Microbiol Infect 2005; 11: 4751.
  • 55
    Wolf P, Seidl H, Back B, Binder B, Hofler G, Quehenberger F, et al. Increased prevalence of human papillomavirus in hairs plucked from patients with psoriasis treated with psoralen-UV-A. Arch Dermatol 2004; 140: 317324.
  • 56
    Iftner T, Bierfelder S, Csapo Z, Pfister H. Involvement of human papillomavirus type 8 genes E6 and E7 in transformation and replication. J Virol 1988; 62: 36553661.
  • 57
    Kiyono T, Nagashima K, Ishibashi M. The primary structure of major viral RNA in a rat cell line transfected with type 47 human papillomavirus DNA and the transforming activity of its cDNA and E6 gene. Virology 1989; 173: 551565.
  • 58
    Kiyono T, Hiraiwa A, Ishibashi M. Differences in transforming activity and coded amino acid sequence among E6 genes of several papillomaviruses associated with epidermodysplasia verruciformis. Virology 1992; 186: 628639.
  • 59
    Hiraiwa A, Kiyono T, Segawa K, Utsumi KR, Ohashi M, Ishibashi M. Comparative study on E6 and E7 genes of some cutaneous and genital papillomaviruses of human origin for their ability to transform 3Y1 cells. Virology 1993; 192: 102111.
  • 60
    Yamashita T, Segawa K, Fujinaga Y, Nishikawa T, Fujinaga K. Biological and biochemical activity of E7 genes of the cutaneous human papillomavirus type 5 and 8. Oncogene 1993; 8: 24332441.
  • 61
    Schmitt A, Harry JB, Rapp B, Wettstein FO, Iftner T. Comparison of the properties of the E6 and E7 genes of low- and high-risk cutaneous papillomaviruses reveals strongly transforming and high Rb-binding activity for the E7 protein of the low-risk human papillomavirus type 1. J Virol 1994; 68: 70517059.
  • 62
    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: 21952206.
  • 63
    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: 13971404.
  • 64
    Akgül B, Garcia-Escudero R, Ghali L, Pfister HJ, Fuchs PG, Navsaria H, et al. The E7 protein of cutaneous human papillomavirus type 8 causes invasion of human keratinocytes into the dermis in organotypic cultures of skin. Cancer Res 2005; 65: 22162223.
  • 65
    Smola-Hess S, Pahne J, Mauch C, Zigrino P, Smola H, Pfister HJ. Expression of membrane type 1 matrix metalloproteinase in papillomavirus-positive cells: role of the human papillomavirus (HPV) 16 and HPV8 E7 gene products. J Gen Virol 2005; 86: 12911296.
  • 66
    Iftner T, Fuchs PG, Pfister H. Two independently transforming functions of human papillomavirus 8. Curr Top Microbiol Immunol 1989; 144: 167173.
  • 67
    Fuchs PG, Horn S, Iftner T, May M, Stubenrauch F, Pfister H. Molecular Biology of Epidermodysplasia Verruciformis-Associated Human Papillomaviruses. Verlag Chemie: Weinheim, 1993.
  • 68
    Muller A, Ritzkowsky A, Steger G. Cooperative activation of human papillomavirus type 8 gene expression by the E2 protein and the cellular coactivator p300. J Virol 2002; 76: 11 04211 053.
  • 69
    Akgül B, Karle P, Adam M, Fuchs PG, Pfister HJ. Dual role of tumor suppressor p53 in regulation of DNA replication and oncogene E6-promoter activity of epidermodysplasia verruciformis-associated human papillomavirus type 8. Virology 2003; 308: 279290.
  • 70
    Hadaschik D, Hinterkeuser K, Oldak M, Pfister HJ, Smola-Hess S. The papillomavirus E2 protein binds to and synergizes with C/EBP factors involved in keratinocyte differentiation. J Virol 2003; 77: 52535265.
  • 71
    Steger G, Schnabel C, Schmidt HM. The hinge region of the human papillomavirus type 8 E2 protein activates the human p21(WAF1/CIP1) promoter via interaction with Sp1. J Gen Virol 2002; 83: 503510.
  • 72
    Oldak M, Smola H, Aumailley M, Rivero F, Pfister H, Smola-Hess S. The human papillomavirus type 8 E2 protein suppresses beta4-integrin expression in primary human keratinocytes. J Virol 2004; 78: 10 73810 746.
  • 73
    Tinsley JM, Fisher C, Searle PF. Abnormalities of epidermal differentiation associated with expression of the human papillomavirus type 1 early region in transgenic mice. J Gen Virol 1992; 73: 12511260.
  • 74
    Schaper ID, Marcuzzi GP, Weissenborn SJ, Kasper HU, Dries V, Smyth N, et al. Development of skin tumors in mice transgenic for early genes of human papillomavirus type 8. Cancer Res 2005; 65: 13941400.
  • 75
    Akgül B, Pfefferle R, Marcuzzi GP, Zigrino P, Krieg T, Pfister H, et al. Expression of matrix metalloproteinases-2, -9, -13 and MT1-MMP in skin tumors of human papillomavirus type 8 transgenic mice. Exper Dermatol 2006; 15: 3542.
  • 76
    Greenhalgh DA, Wang XJ, Rothnagel JA, Eckhardt JN, Quintanilla MI, Barber JL, et al. Transgenic mice expressing targeted HPV-18 E6 and E7 oncogenes in the epidermis develop verrucous lesions and spontaneous, rasHa-activated papillomas. Cell Growth Differ 1994; 5: 667675.
  • 77
    Helfrich I, Chen M, Schmidt R, Furstenberger G, Kopp-Schneider A, Trick D, et al. Increased incidence of squamous cell carcinomas in Mastomys natalensis papillomavirus E6 transgenic mice during two-stage skin carcinogenesis. J Virol 2004; 78: 47974805.
  • 78
    Arbeit JM, Munger K, Howley PM, Hanahan D. Progressive squamous epithelial neoplasia in K14-human papillomavirus type 16 transgenic mice. J Virol 1994; 68: 43584368.
  • 79
    zur Hausen H. Papillomaviruses causing cancer: evasion from host-cell control in early events in carcinogenesis. J Natl Cancer Inst 2000; 92: 690698.
  • 80
    Munger K, Basile JR, Duensing S, Eichten A, Gonzalez SL, Grace M, et al. Biological activities and molecular targets of the human papillomavirus E7 oncoprotein. Oncogene 2001; 20: 78887898.
  • 81
    Duensing S, Munger K. Centrosome abnormalities, genomic instability and carcinogenic progression. Biochim Biophys Acta 2001; 1471: M81M88.
  • 82
    Duensing S, Duensing A, Crum CP, Munger K. Human papillomavirus type 16 E7 oncoprotein-induced abnormal centrosome synthesis is an early event in the evolving malignant phenotype. Cancer Res 2001; 61: 23562360.
  • 83
    Southern SA, Lewis MH, Herrington CS. Induction of tetrasomy by human papillomavirus type 16 E7 protein is independent of pRb binding and disruption of differentiation. Br J Cancer 2004; 90: 19491954.
  • 84
    Munger K. The role of human papillomaviruses in human cancers. Front Biosci 2002; 7: d641d649.
  • 85
    Jackson S, Storey A. E6 proteins from diverse cutaneous HPV types inhibit apoptosis in response to UV damage. Oncogene 2000; 19: 592598.
  • 86
    Storey A, Massimi P, Dawson K, Banks L. Conditional immortalization of primary cells by human papillomavirus type 18 E6 and EJ-ras defines an E6 activity in G0/G1 phase which can be substituted for mutations in p53. Oncogene 1995; 11: 653661.
  • 87
    Fisher MS, Kripke ML. Systemic alteration induced in mice by ultraviolet light irradiation and its relationship to ultraviolet carcinogenesis. Proc Natl Acad Sci U S A 1977; 74: 16881692.
  • 88
    Pfeifer GP, You YH, Besaratinia A. Mutations induced by ultraviolet light. Mutat Res 2005; 571: 1931.
  • 89
    Cleaver JE, Bootsma D. Xeroderma pigmentosum: biochemical and genetic characteristics. Annu Rev Genet 1975; 9: 1938.
  • 90
    Pan H, Griep AE. Altered cell cycle regulation in the lens of HPV-16 E6 or E7 transgenic mice: implications for tumor suppressor gene function in development. Genes Dev 1994; 8: 12851299.
  • 91
    Wahl GM, Carr AM. The evolution of diverse biological responses to DNA damage: insights from yeast and p53. Nature Cell Biol 2001; 3: E277E286.
  • 92
    Scheffner M, Werness BA, Huibregtse JM, Levine AJ, Howley PM. The E6 oncoprotein encoded by human papillomavirus types 16 and 18 promotes the degradation of p53. Cell 1990; 63: 11291136.
  • 93
    Werness BA, Levine AJ, Howley PM. Association of human papillomavirus types 16 and 18 E6 proteins with p53. Science 1990; 248: 7679.
  • 94
    Elbel M, Carl S, Spaderna S, Iftner T. A comparative analysis of the interactions of the E6 proteins from cutaneous and genital papillomaviruses with p53 and E6AP in correlation to their transforming potential. Virology 1997; 239: 132149.
  • 95
    Donehower LA, Harvey M, Slagle BL, McArthur MJ, Montgomery CA Jr, Butel JS, et al. Mice deficient for p53 are developmentally normal but susceptible to spontaneous tumours. Nature 1992; 356: 215221.
  • 96
    Ziegler A, Jonason AS, Leffell DJ, Simon JA, Sharma HW, Kimmelman J, et al. Sunburn and p53 in the onset of skin cancer. Nature 1994; 372: 773776.
  • 97
    Li G, Ho VC, Berean K, Tron VA. Ultraviolet radiation induction of squamous cell carcinomas in p53 transgenic mice. Cancer Res 1995; 55: 20702074.
  • 98
    Li G, Tron V, Ho V. Induction of squamous cell carcinoma in p53-deficient mice after ultraviolet irradiation. J Invest Dermatol 1998; 110: 7275.
  • 99
    Steger G, Pfister H. In vitro expressed HPV 8 E6 protein does not bind p53. Arch Virol 1992; 125: 355360.
  • 100
    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: 30653073.
  • 101
    Giampieri S, Garcia-Escudero R, Green J, Storey A. Human papillomavirus type 77 E6 protein selectively inhibits p53-dependent transcription of proapoptotic genes following UV-B irradiation. Oncogene 2004; 23: 58645870.
  • 102
    Jonason AS, Kunala S, Price GJ, Restifo RJ, Spinelli HM, Persing JA, et al. Frequent clones of p53-mutated keratinocytes in normal human skin. Proc Natl Acad Sci U S A 1996; 93: 14 02514 029.
  • 103
    Ren ZP, Hedrum A, Ponten F, Nister M, Ahmadian A, Lundeberg J, et al. Human epidermal cancer and accompanying precursors have identical p53 mutations different from p53 mutations in adjacent areas of clonally expanded non-neoplastic keratinocytes. Oncogene 1996; 12: 765773.
  • 104
    Malkin D, Li FP, Strong LC, Fraumeni JF Jr, Nelson CE, Kim DH, et al. Germ line p53 mutations in a familial syndrome of breast cancer, sarcomas, and other neoplasms. Science 1990; 250: 12331238.
  • 105
    Brash DE, Zhang W, Grossman D, Takeuchi S. Colonization of adjacent stem cell compartments by mutant keratinocytes. Semin Cancer Biol 2005; 15: 97102.
  • 106
    Storey A. Papillomaviruses: death-defying acts in skin cancer. Trends Mol Med 2002; 8: 417421.
  • 107
    Erster S, Mihara M, Kim RH, Petrenko O, Moll UM. In vivo mitochondrial p53 translocation triggers a rapid first wave of cell death in response to DNA damage that can precede p53 target gene activation. Mol Cell Biol 2004; 24: 67286741.
  • 108
    Leu JI, Dumont P, Hafey M, Murphy ME, George DL. Mitochondrial p53 activates Bak and causes disruption of a Bak–Mcl1 complex. Nature Cell Biol 2004; 6: 443450.
  • 109
    Mitra RS, Wrone-Smith T, Simonian P, Foreman KE, Nunez G, Nickoloff BJ. Apoptosis in keratinocytes is not dependent on induction of differentiation. Lab Invest 1997; 76: 99107.
  • 110
    Tomkova H, Fujimoto W, Arata J. Expression of bcl-2 antagonist bak in inflammatory and neoplastic skin diseases. Br J Dermatol 1997; 137: 703708.
  • 111
    Thomas M, Banks L. Inhibition of Bak-induced apoptosis by HPV-18 E6. Oncogene 1998; 17: 29432954.
  • 112
    Thomas M, Banks L. Human papillomavirus (HPV) E6 interactions with Bak are conserved amongst E6 proteins from high and low risk HPV types. J Gen Virol 1999; 80: 15131517.
  • 113
    Jackson S, Ghali L, Harwood C, Storey A. Reduced apoptotic levels in squamous but not basal cell carcinomas correlate with detection of cutaneous human papillomavirus. Br J Cancer 2002; 87: 319323.
  • 114
    Sancar A, Lindsey-Boltz LA, Unsal-Kacmaz K, Linn S. Molecular mechanisms of mammalian DNA repair and the DNA damage checkpoints. Annu Rev Biochem 2004; 73: 3985.
  • 115
    Giampieri S, Storey A. Repair of UV-induced thymine dimers is compromised in cells expressing the E6 protein from human papillomaviruses types 5 and 18. Br J Cancer 2004; 90: 22032209.
  • 116
    Iftner T, Elbel M, Schopp B, Hiller T, Loizou JI, Caldecott KW, et al. Interference of papillomavirus E6 protein with single-strand break repair by interaction with XRCC1. EMBO J 2002; 21: 47414748.
  • 117
    Struijk L, Bouwes Bavinck JN, Wanningen P, van der Meijden E, Westendorp RG, Ter Schegget J, et al. Presence of human papillomavirus DNA in plucked eyebrow hairs is associated with a history of cutaneous squamous cell carcinoma. J Invest Dermatol 2003; 121: 15311535.
  • 118
    Tsai RY. A molecular view of stem cell and cancer cell self-renewal. Int J Biochem Cell Biol 2004; 36: 684694.
  • 119
    Lechler T, Fuchs E. Asymmetric cell divisions promote stratification and differentiation of mammalian skin. Nature 2005; 437: 275280.
  • 120
    Benitah SA, Frye M, Glogauer M, Watt FM. Stem cell depletion through epidermal deletion of Rac1. Science 2005; 309: 933935.
  • 121
    Quinn AG, Sikkink S, Rees JL. Delineation of two distinct deleted regions on chromosome 9 in human non-melanoma skin cancers. Genes Chromosomes Cancer 1994; 11: 222225.
  • 122
    Quinn AG, Campbell C, Healy E, Rees JL. Chromosome 9 allele loss occurs in both basal and squamous cell carcinomas of the skin. J Invest Dermatol 1994; 102: 300303.
  • 123
    Hahn H, Wicking C, Zaphiropoulous PG, Gailani MR, Shanley S, Chidambaram A, et al. Mutations of the human homolog of Drosophila patched in the nevoid basal cell carcinoma syndrome. Cell 1996; 85: 841851.
  • 124
    Johnson RL, Rothman AL, Xie J, Goodrich LV, Bare JW, Bonifas JM, et al. Human homolog of patched, a candidate gene for the basal cell nevus syndrome. Science 1996; 272: 16681671.
  • 125
    Shanley SM, Dawkins H, Wainwright BJ, Wicking C, Heenan P, Eldon M, et al. Fine deletion mapping on the long arm of chromosome 9 in sporadic and familial basal cell carcinomas. Hum Mol Genet 1995; 4: 129133.
  • 126
    Holmberg E, Rozell BL, Toftgard R. Differential allele loss on chromosome 9q22.3 in human non-melanoma skin cancer. Br J Cancer 1996; 74: 246250.
  • 127
    Shen T, Park WS, Boni R, Saini N, Pham T, Lash AE, et al. Detection of loss of heterozygosity on chromosome 9q22.3 in microdissected sporadic basal cell carcinoma. Hum Pathol 1999; 30: 284287.
  • 128
    Reifenberger J, Wolter M, Knobbe CB, Kohler B, Schonicke A, Scharwachter C, et al. Somatic mutations in the PTCH, SMOH, SUFUH and TP53 genes in sporadic basal cell carcinomas. Br J Dermatol 2005; 152: 4351.
  • 129
    Daya-Grosjean L, Couve-Privat S. Sonic hedgehog signaling in basal cell carcinomas. Cancer Lett 2005; 225: 181192.
  • 130
    Ping XL, Ratner D, Zhang H, Wu XL, Zhang MJ, Chen FF, et al. PTCH mutations in squamous cell carcinoma of the skin. J Invest Dermatol 2001; 116: 614616.
  • 131
    Quinn AG, Sikkink S, Rees JL. Basal cell carcinomas and squamous cell carcinomas of human skin show distinct patterns of chromosome loss. Cancer Res 1994; 54: 47564759.
  • 132
    Brown VL, Harwood CA, Crook T, Cronin JG, Kelsell DP, Proby CM. p16INK4a and p14ARF tumor suppressor genes are commonly inactivated in cutaneous squamous cell carcinoma. J Invest Dermatol 2004; 122: 12841292.
  • 133
    Fischer A. Primary immunodeficiency diseases: an experimental model for molecular medicine. Lancet 2001; 357: 18631869.
  • 134
    Laffort C, Le Deist F, Favre M, Caillat-Zucman S, Radford-Weiss I, Debre M, et al. Severe cutaneous papillomavirus disease after haemopoietic stem-cell transplantation in patients with severe combined immune deficiency caused by common γc cytokine receptor subunit or JAK-3 deficiency. Lancet 2004; 363: 20512054.
  • 135
    Evan GI, Vousden KH. Proliferation, cell cycle and apoptosis in cancer. Nature 2001; 411: 342348.