Ethel Michele De Villiers

Cross-roads in the classification of papillomaviruses

Acceptance of an official classification for the family Papillomaviridae based purely on DNA sequence relatedness, was achieved as late as 2003. The rate of isolation and characterization of new papillomavirus types has greatly depended on and subjected to the development of new laboratory techniques. Introduction of every new technique led to a temporarily burst in the number of new isolates. In the following, the bumpy road towards achieving a classification system combined with the controversies of implementing and accepting new techniques will be summarized. An update of the classification of the 170 human papillomavirus (HPV) types presently known is presented. Arguments towards the implementation of metagenomic sequencing for this rapidly growing family will be presented.

The E6 and E7 Proteins of the Cutaneous Human Papillomavirus Type 38 Display Transforming Properties

ABSTRACT Several studies have suggested the involvement of cutaneous human papillomaviruses (HPVs) in the development of nonmelanoma skin cancers. Here we have characterized the in vitro properties of E7 proteins of three cutaneous HPV types, 10, 20, and 38, which are frequently detected in skin specimens. We show that HPV38 E7 is able to inactivate the tumor suppressor pRb and induces loss of G1/S transition control, a key event in carcinogenesis. In contrast, HPV10 and HPV20 E7 proteins do not display these in vitro transforming activities. We also show that the two early proteins E6 and E7 of HPV38 are sufficient to corrupt the cell cycle and senescence programs in primary cells, inducing active and long-lasting proliferation of primary human keratinocytes, the natural host cells. Our study shows that E6 and E7 of this cutaneous HPV type have transforming activity in primary human cells, suggesting a role for HPV38 infection in skin carcinogenesis. In further support of such a role, we detected HPV38 DNA in approximately 50% of nonmelanoma skin cancers, but only in 10% of healthy skin specimens (P < 0.001).

Cutaneous Human Papillomaviruses Found in Sun-Exposed Skin: Beta-papillomavirus Species 2 Predominates in Squamous Cell Carcinoma

BACKGROUND A spectrum of cutaneous human papillomaviruses (HPVs) is detectable in nonmelanoma skin cancers, as well as in healthy skin, but the significance that the presence of these types of HPV DNA has for the pathogenesis of skin cancer remains unclear. METHODS We studied 349 nonimmunosuppressed patients with skin lesions (82 with squamous cell carcinomas, 126 with basal cell carcinomas, 49 with actinic keratoses, and 92 with benign lesions). After superficial skin had been removed by tape, paired biopsy samples--from the lesion and from healthy skin from the same patient--were tested for HPV DNA. Risk factors for HPV DNA were analyzed in multivariate models. RESULTS Overall, 12% of healthy skin samples were positive for HPV DNA, compared with 26% of benign lesions, 22% of actinic keratoses, 18% of basal cell carcinomas, and 26% of squamous cell carcinomas. HPV DNA was associated with sites extensively exposed to the sun, both for the lesions (odds ratio [OR], 4.45 [95% confidence interval {CI}, 2.44-8.11]) and for the healthy skin samples (OR, 3.65 [95% CI 1.79-7.44]). HPV types of Beta-papillomavirus species 2 predominate in squamous cell carcinomas (OR, 4.40 [95% CI, 1.92-10.06]), whereas HPV types of Beta-papillomavirus species 1 are primarily found in benign lesions (OR, 3.47 [95% CI, 1.72-6.99]). CONCLUSIONS Cutaneous HPV types are primarily detected at sites extensively exposed to the sun. HPV types of Beta-papillomavirus species 2, but not of species 1, are associated with squamous cell carcinoma.

Detection of Human Papillomavirus DNA in Cutaneous Squamous Cell Carcinoma among Immunocompetent Individuals

The presence of certain types of human papillomavirus (HPV) is a known risk factor for the development of anogenital squamous cell carcinomas (SCCs). A similar association has been hypothesized for cutaneous SCCs, although, to our knowledge, no studies to date have combined sensitive HPV DNA detection techniques with epidemiologic data controlling for known risk factors to explore the association. We designed a case-control study examining HPV prevalence using highly sensitive PCR-detection assays in tissue samples from 85 immunocompetent patients with histologically confirmed SCCs and 95 age-matched individuals without a prior history of skin cancer. A standardized interview was administered to all study subjects to collect information pertaining to potential confounding variables. The overall detection rate of HPV DNA was high in case lesions (54%) and perilesions (50%) and in both sun-exposed normal tissue (59%) and non-sun-exposed normal tissue (49%) from controls. In comparing case tissue to control tissue, there was no differential detection of HPV DNA across various HPV species. However, HPV DNA from beta-papillomavirus species 2 was more likely to be identified in tumors than in adjacent healthy tissue among cases (paired analysis, odds ratio=4.0, confidence interval=1.3-12.0). The high prevalence of HPV DNA detected among controls suggests that HPV DNA is widely distributed among the general population. However, the differential detection of HPV beta-papillomavirus species in tumors among cases suggests that certain HPV types may be involved in the progression of cutaneous SCCs.

Presence of papillomavirus sequences in condylomatous lesions of the mamillae and in invasive carcinoma of the breast

BackgroundViruses including Epstein–Barr virus (EBV), a human equivalent of murine mammary tumour virus (MMTV) and human papillomavirus (HPV) have been implicated in the aetiology of human breast cancer. We report the presence of HPV DNA sequences in areolar tissue and tumour tissue samples from female patients with breast carcinoma. The presence of virus in the areolar–nipple complex suggests to us a potential pathogenic mechanism.MethodsPolymerase chain reaction (PCR) was undertaken to amplify HPV types in areolar and tumour tissue from breast cancer cases. In situ hybridisation supported the PCR findings and localised the virus in nipple, areolar and tumour tissue.ResultsPapillomavirus DNA was present in 25 of 29 samples of breast carcinoma and in 20 of 29 samples from the corresponding mamilla. The most prevalent type in both carcinomas and nipples was HPV 11, followed by HPV 6. Other types detected were HPV 16, 23, 27 and 57 (nipples and carcinomas), HPV 20, 21, 32, 37, 38, 66 and GA3-1 (nipples only) and HPV 3, 15, 24, 87 and DL473 (carcinomas only). Multiple types were demonstrated in seven carcinomas and ten nipple samples.ConclusionsThe data demonstrate the occurrence of HPV in nipple and areolar tissues in patients with breast carcinoma. The authors postulate a retrograde ductular pattern of viral spread that may have pathogenic significance.

Papillomavirus in esophageal papillomas and carcinomas

Human papillomavirus (HPV) has been implicated as a possible etiological factor in the development of squamous‐cell carcinoma of the esophagus. Published data demonstrating HPV DNA in these lesions have been conflicting, varying between failure to detect HPV DNA to detection of up to 60% to 70% of the biopsies harboring HPV DNA, mainly HPV 16 and HPV 18. We have analyzed esophageal carcinoma samples from 2 high‐risk areas, China and South Africa, using a degenerate PCR approach. All amplified products were cloned and sequenced. A broad spectrum of HPV types was demonstrated in 10/29 samples from China and 9/34 samples from South Africa. HPV types detected included mucosal types HPV 6, 18, 51, 52 and 57; cutaneous types HPV 9, 20, 24 and 25; and the putative new HPV types DL231, DL428 and DL436, with HPV 6/51, 6/57, 20/9 and 20/DL231 occurring as double infections. HPV 6 predominated (4 samples) in 11 esophageal papillomas originating from patients in Europe and tested for HPV DNA. Other HPV types present included HPV 20, DL284 and DL436. Another putative new HPV type, DL416, was identified in a dysplastic lesion of the esophagus. Int. J. Cancer 80:681–684, 1999.

Papillomavirus infections and human genital cancer

Human papillomas are induced by at least eight distinct types of papillomaviruses. They are listed in Table 1. Condylomata acuminata, human genital warts, represent a group of fibropapillomatous benign tumors with exuberant exophytic growth which are induced by a papillomavirus (reviewed in [l]). They are mainly located at the foreskin of the penis, the glans, the vulva, the introitus vaginae, and intravaginally (see review [l]). They have also been reported on the cervix uteri, perianally, and even within the urethra [2-41. Epidemiological studies demonstrate a venereal mode of transmission [5]. They are prevalent in groups of high sexual promiscuity, such as homosexuals [3, 61 and prostitutes, and are by no means a rare disease. According to British statistics [7] they comprise 6.1% of all reported cases of venereally transmitted diseases (Table 2). Malignant conversion of condylomata acuminata has been reported repeatedly (reviewed in [l]). Anecdotal reports were published mainly from cases of long duration but exceptionally also from young individuals with a rather recent history of genital warts (reviewed in [l]). Malignant tumors developed within genital warts of the vulva, the penis, and also of the vagina. The role of the virus found electron microscopically in a few nuclei of the benign condylomata acuminata in the induction of such malignant tumors has not been established. Our own investigations on a possible role of human genital wart virus in genital cancer were initiated after a number of unsuccessful attempts to demonstrate herpes simplex type II DNA in biopsies of such tumors by nucleic acid hybridization [8]. Since epidemiological features of genital cancer point in a number of parameters (reviewed in [9]), such as dependence on the number of sexual partners, early onset of sexual relations, existence of marital clusters, and correlation in the incidence between cervical and penile cancer [9-151, to an infectious etiology, we started to analyze additional candidate viruses for their possible involvement in this disease. The presence of a virus belonging to a group of clearly oncogenic agents in genital warts and occasional reports on their malignant conversion stimulated the interest in these viruses. Initial experiments were designed to clarify the question of whether the papillomavirus found in genital warts is identical with papillomaviruses observed in other skin papillomas. This led to the identification of distinct types of papil-

Relationship between steroid hormone contraceptives and HPV, cervical intraepithelial neoplasia and cervical carcinoma†

Available data demonstrate an increase in the transcription of high‐risk papillomaviruses by the 16α‐hydroxylation of estrogens, which is in line with the epidemiologic data showing an increased cervical carcinogenesis risk for long‐term contraceptive‐using, HPV‐infected women. No evidence exists for an increase in HPV‐negative contraceptive users.

Isolation of Multiple TT Virus Genotypes from Spleen Biopsy Tissue from a Hodgkin's Disease Patient: Genome Reorganization and Diversity in the Hypervariable Region

ABSTRACT We report the isolation of 24 novel genotypes of TT viruses from a surgically removed spleen of a patient with Hodgkins disease. The sequence analysis of our 24 isolates revealed the remarkable heterogeneity of TT virus isolates not only from the same patient but also from the same biopsy material. These isolates belong to four phylogenetic groups of TT viruses. Nucleotide sequence analyses revealed five distinct genotypes (tth3, tth4, tth5, tth6, and tth7). The limited variation in sequence identity of the other isolates defines the latter as variants of four of these genotypes. A group of 6 isolates (the tth7 group) revealed a reorganization of open reading frame 1 (ORF1) leading to one larger and a varying number of smaller ORFs. The nucleotide difference of the full-length genomes was less than 1%. A variation of 69 to 97% in amino acids of a second group of 8 isolates (the tth3 group) was restricted to the hypervariable region of ORF1, indicating the existence of a quasi-species. These isolates differed by less than 2% in the remainder of their nucleotide sequences. An alignment of these isolates with 79 previously reported TT virus genotypes permits the proposal of TT virus genera and species within the family Anelloviridae in analogy to a previous proposal for the papillomaviruses (family Papillomaviridae).

Seroreactivity to Cutaneous Human Papillomaviruses among Patients with Nonmelanoma Skin Cancer or Benign Skin Lesions.

Cutaneous human papillomaviruses (HPV) are common in nonmelanoma skin cancers, benign skin lesions, and healthy skin. Increased seroprevalences for cutaneous HPV among nonmelanoma skin cancer patients have been described. To determine whether antibodies to cutaneous HPV are related to presence of the virus and/or to skin disease, we collected serum and biopsies from both lesions and healthy skin from 434 nonimmunosuppressed patients (72 squamous cell carcinomas, 160 basal cell carcinomas, 81 actinic keratoses, and 121 benign lesions). Biopsies were analyzed for HPV DNA by PCR, cloning, and sequencing. Serum antibodies to the major capsid protein L1 of HPV 1, 5, 6, 8, 9, 10, 15, 16, 20, 24, 32, 36, 38, and 57 as well as to the oncoproteins E6 and E7 of HPV 8 and 38 were detected using a multiplexed fluorescent bead-based assay. Type-specific seroprevalence among patients with the same type of HPV DNA (sensitivity of serology) varied from 0% to at most 28%. Presence of HPV DNA and antibodies to the same HPV type was not significantly correlated. However, seropositivity to any HPV type was significantly more common among patients positive for HPV DNA of any HPV type (odds ratio, 1.90; 95% confidence interval, 1.55-2.34). Seroprevalences were similar among the different patient groups but was, for most HPV types, somewhat higher among squamous cell carcinoma patients than among basal cell carcinoma patients (P < 0.01). In conclusion, additional studies are required to clarify the biological meaning of seropositivity as a marker of cutaneous HPV infection and skin disease. (Cancer Epidemiol Biomarkers Prev 2008;17(1):189–95)