1. Rogozinski TT, Jablonska S, Jarzabek-Chorzelska M: Role of cell-mediated immunity in spontaneous regression of plane warts. Int J Dermatol 27(5):322-326, 1988. Human papillomavirus-induced plane warts most often occur in the second decade of age. Afterward, they either spontaneously regress or are eradicated in the course of various treatments. As proved by in vivo and in vitro tests as well as clinical observations, they most often affect and persist longer in immunocompromised hosts. In this work it was confirmed that specific-ie, anti-HPV-directed, cell-mediated immunologic response plays a role in spontaneous regression of plane warts and that preservation of nonspecific immunity is prerequisite for spontaneous regression of plane warts.

2. Avgerinou G, Georgala S, Theodoridis A, et al.: Reduction of cell mediated immunity in patients with genital warts of long duration. Genitourin Med 62(6):396-398, 1986. Cell mediated immunity was studied by a leucocyte migration inhibition assay and by tuberculin and dinitrochlorobenzene skin tests in 30 patients with recurrent genital warts and in 34 healthy people (with no history of genital warts) who served as controls. Migration inhibition was significantly less in patients suffering from recurrences for more than one year than in controls (p less than 0.001). Dinitrochlorobenzene and tuberculin sensitivity were also found to be impaired in those with infection of long duration (p less than 0.001).

3. Stanley M, Coleman N, Chambers M: The host response to lesions induced by human papillomavirus. Ciba Found Symp 187:21-32; discussion 32-44, 1994. Human papillomaviruses (HPVs) are strictly intraepithelial pathogens: in the natural productive infection they induce benign epithelial proliferations of mucocutaneous surfaces, some of which may progress to malignancy. Benign HPV-induced lesions are chronic persistent growths; high levels of viral antigen are expressed in the apparent absence of a host immune response suggesting that these viruses have evolved efficient mechanisms of immune evasion. Cell-mediated responses are central in the pathogenesis of HPV and regression of both cutaneous and genital warts histologically resembles a delayed-type hypersensitivity response (DTH). The antigen(s) in the wart against which this response is initiated are not known but in an experimental murine model DTH responses to the E6 and E7 proteins of HPV-16 can be elicited when viral antigen is presented via the epithelial route. Priming with low levels of viral antigen in this model induces non-responsiveness and the loss of DTH. In HPV-associated cancers the E6/E7 genes are expressed and an antibody response to the proteins is found in at least 50% of cases indicating that these oncoproteins are potential targets for immunotherapy. [References: 22]

4. van der Steen P, van de Kerkhof P, der Kinderen D, et al.: Clinical and immunohistochemical responses of plantar warts to topical immunotherapy with diphenylcyclopropenone. J Dermatol 18(6):330-333, 1991. A 30-year-old man with bilateral plantar warts of the mosaic type which had been resistant to standard treatment modalities was treated with diphenylcyclopropenone. After 10 weeks, the treated warts had disappeared; the untreated warts, although showing some involution, still persisted. The untreated warts, serving as a control to prove the effectiveness of topical immunotherapy, responded likewise to subsequent treatment with diphenylcyclopropenone. Wart regression was reflected histopathologically by decreases in acanthosis, papillomatosis, granular vacuolation, and hyperkeratosis. Immunohistochemically, Ki-67 expression was markedly reduced, and a reversal of the CD4/CD8 ratio was seen. These findings suggest a major role of a cell-mediated immune response in the spontaneous resolution of warts.

5. Iwatsuki K, Tagami H, Takigawa M, et al.: Plane warts under spontaneous regression. Immunopathologic study on cellular constituents leading to the inflammatory reaction. Arch Dermatol 122(6):655-659, 1986. Immunohistologically, cellular infiltrates in regressing plane warts were mainly composed of lymphocytes and mononuclear phagocytes. There were many infiltrating T lymphocytes. Immunoelectron microscopic observation demonstrated that both helper/inducer and suppressor/cytotoxic phenotypes of T lymphocytes infiltrated in the lesions. OKT6-positive cells were observed in the dermis as well as in the epidermis. Moreover, as noted in allergic contact dermatitis, the apposition of T lymphocytes to Langerhans' cell-like cells could be seen. Lymphocytes and a small number of mononuclear phagocytes were found adjacent to damaged keratinocytes in the epidermis, the picture of which has been described as satellite cell necrosis, a hallmark of cytotoxic reaction by aggressors. These findings suggest that specific cell-mediated immunity against virus-infected keratinocytes takes place in the process of regressing plane warts.

6. Obalek S, Glinski W, Haftek M, et al.: Comparative studies on cell-mediated immunity in patients with different warts. Dermatologica 161(2):73-83, 1980. The distribution of peripheral blood T and B lymphocytes, the in vitro lymphocyte response to PHA, and in vivo experimental DNCB sensitization were studied in patients with different clinical forms of warts (common, 84; flat, 88; plantar, 22; genital, 14) and in 15 cases of epidermodysplasia verruciformis (EV). The percentage of T lymphocytes forming E rosettes was significantly decreased in patients with common (54.8%), flat (47.5%) and plantar (58.3%) warts, and those with EV (47.6%) in comparison with normal controls (68.4%). The DNCB sensitivity developed less frequently and it was less intensive in patients with common and flat warts than in the normal population. 60% of EV cases were anergic to challenging doses of DNCB. The lymphocyte response to PHA was reduced in all groups of patients studied as compared to normals. T cell function was found to be most defective in patients with EV and those with flat warts. Only a slight but statistically significant defect was demonstrated in the common wart group. CMI in patients with both plantar and genital warts was shown to be almost normal; except minor alterations of PHA-induced lymphocyte transformation and E rosetting T lymphocyte counts. These data have shown the divergency of CMI defect in the patients with different clinical forms of warts caused by various HPV types. This could indicate that distinct HPV types varied in their infectiveness and host cell-mediated resistance is a fundamental factor preventing viral infection.

7. Steele K, Shirodaria P, O'Hare M, et al.: Monochloroacetic acid and 60% salicylic acid as a treatment for simple plantar warts: effectiveness and mode of action. Br J Dermatol 118(4):537-543, 1988. Monochloroacetic acid crystals and 60% salicylic acid ointment was found to be more effective than placebo as a treatment for simple plantar warts in a double blind study on 57 patients. Nineteen (66%) patients in the active treatment group compared with five (18%) patients in the placebo group were cured after 6 weeks (P = 0.002). The active treatment was associated with a significantly higher cure rate 6 months after entry (P = 0.04). Treatments were well tolerated. IgG or IgM antibodies or both to human papilloma virus (HPV) types 1 or 2 or both were detected significantly more frequently in the actively treated group 6 weeks after entry (P = 0.0005). Twelve (50%) patients considered to be cured had no detectable secondary immune response. Our results suggest that cure does not depend primarily on the humoral system but rather on mechanical destruction of wart tissue, or occurs as a result of cell mediated immunity.

8. Kilkenny M, Marks R: The descriptive epidemiology of warts in the community. Australas J Dermatol 37(2):80-86, 1996. Warts are common skin infections caused by human papillomavirus (HPV) and affect most people sometime in their life. A number of epidemiological studies on the prevalence of warts have been completed in schools, various occupational groups, general practices and hospitals. All studies have relied on a subjective measure for the diagnosis of warts. Cross-sectional studies completed in schools have shown the prevalence in children to vary from 2 to 20%. Occupational handlers of meat, poultry and fish have a higher prevalence than other workers. Children and young adults are the groups most affected. Future studies are needed to investigate the true frequency of warts in the community and the likelihood of an individual developing these lesions during his/her lifetime. [References: 40]

9. Rudlinger R, Bunney M, Grob R, et al.: Warts in fish handlers. Br J Dermatol 120(3):375-381, 1989. Fish handlers frequently suffer from hand warts. The clinical form and HPV type in these lesions were studied. Eleven individuals (10 fishmongers and one fisherman) with multiple hand warts were examined clinically and samples from their warts examined by Southern blot and reverse blot analysis. Clinically, with one exception, the warts were of the common type. HPV DNA was detected in all but one individual. HPV4 was found in one sample, HPV1 related virus in three, a virus hybridizing with both HPV27 and HPV2 in five (four individuals) and HPV7 in seven (six individuals). More than one type was detected in four individuals. HPV7 infection was related to the greater length of time spent in handling fish. These findings indicate that HPV7 is not, as was previously thought, found exclusively in those handling butcher meat and suggest that environmental conditions may be a factor in the clinical manifestation of HPV7 infection. The exact nature of a virus designated HPV2/27 and the significance of its presence in these fish handlers remains uncertain.

10. Jackson V, Chalkley R: Separation of newly synthesized nucleohistone by equilibrium centrifugation in cesium chloride. Biochemistry 13(19):3952-3956, 1974.

11. Keefe M, al-Ghamdi A, Coggon D, et al.: Cutaneous warts in butchers. Br J Dermatol 130(1):9-14, 1994. Several studies have indicated a high prevalence of hand warts in meat handlers, although the reasons for this are not clear. The high prevalence may be partly due to HPV7, a virus found almost exclusively in meat handlers, but the source of HPV7 is not known. We have carried out a cross-sectional survey of hand warts in male meat workers and controls from other occupational groups, to investigate the reasons for the high prevalence of warts, and particularly of HPV7, in butchers. We studied 240 abattoir workers, 246 retail and wholesale butchers, 308 engineering fitters and 292 office workers. Each subject was interviewed using a standard questionnaire, and his hands were examined by a dermatologist. Scrapings from the warts were tested for HPV1, HPV2 and HPV7 by a polymerase chain reaction method. The prevalence of hand warts was 33.3% in the abattoir workers, 34.1% in the butchers, 19.5% in the engineers and 14.7% in the office workers. Scrapings were taken from 247 of 267 subjects with warts, and HPV DNA was detected in 151 samples. The most common viruses were HPV2 (94 men) and HPV7 (76 men). The excess of warts in meat workers was largely due to HPV7, which was found in only two of the office workers, and was not found in any of the engineers. Logistic regression analysis showed no association between the prevalence of hand warts (or HPV2 and HPV7 specifically) and hand trauma, cold and wet working conditions, smoking, atopy, or handling any particular kind of meat. We suggest that some constituent of animal flesh predisposes to replication of HPV7 in keratinized epithelium.

12. Corley E, Pueyo S, Goc B, et al.: Papillomaviruses in human skin warts and their incidence in an Argentine population. Diagn Microbiol Infect Dis 10(2):93-101, 1988. Human papillomavirus genomic types present in human warts of an Argentine population were studied. HPV DNA from single warts was obtained using an alkaline extraction procedure that resulted in a clean DNA preparation, which could be analyzed with several endonucleases. This method was used to isolate and insert the HPV DNAs of two genomic types into the Bam HI site of the pBR322 plasmid. Restriction maps of both HPV DNAs were constructed. According to these maps, one of the genomic variations was identical to HPV1a and the other to HPV2a. The incidence of HPV2 and of HPV1 in different types of skin warts was studied by a dot blot hybridization assay. Twenty-two out of 28 common warts were positive for HPV2 and negative for HPV1; four were positive for HPV1 and negative for HPV2 and two were negative for both. Five out of six plantar warts were positive for HPV1, and one was negative for both. Three out of seven filiform warts were positive for HPV2, three were positive for both probes, and one was negative for both. Southern blot analysis of HPV2 positive samples indicated that 80% were HPV2a and 20% another subtype not yet characterized. All plantar warts contained HPV1a. Msp I/Hpa II restriction analysis confirms previous results indicating that HPV1a DNA is partially methylated, while no evidence of methylation was found for HPV2a DNA.

13. Rubben A, Krones R, Schwetschenau B, et al.: Common warts from immunocompetent patients show the same distribution of human papillomavirus types as common warts from immunocompromised patients. Br J Dermatol 128(3):264-270, 1993. We studied the papillomaviruses (HPV) found in 131 common warts from 111 immunocompetent patients by amplification of viral DNA sequences with the general-primer-mediated polymerase chain reaction (PCR). The virus types were determined by restriction-enzyme cleavage and reverse-blot analysis. Results were confirmed by using the Southern blot technique. Forty patients harboured HPV 2a, 25 individuals showed HPV 2c and 13 yielded HPV 57. Common warts from 16 patients were induced by a variant of HPV 57. HPV 7 was found in four patients. HPV 1 was identified in two patients, and there was evidence for HPV 4 in only one case. One individual yielded an HPV type which was only weakly related to HPV 2. Three patients were infected by more than one HPV type. PCR did not demonstrate HPV-DNA in warts from six individuals. The distribution and variation of HPV types found in the common warts of immunocompetent patients were similar to the findings in immunocompromised patients reported by other authors.

14. Nuovo GJ, Lastarria DA, Smith S, et al.: Human papillomavirus segregation patterns in genital and nongenital warts in prepubertal children and adults. Am J Clin Pathol 95(4):467-474, 1991. This study compared the segregation patterns of human papillomavirus (HPV) in genital and nongenital warts in prepubertal children and adults. HPV 2 was detected in most nongenital warts in children and adults, whereas neither HPV 6 or 11 was detected at nongenital sites in either group with the use of in situ or Southern blot hybridization analyses. Of nine genital tract lesions in children. HPV 2 was detected in two and HPV 6 or 11 in six. More than 90% of cases of regional tract condylomata in adults contained HPV 6 or 11. HPV 2 was not detected in any of 99 genital tract lesions in adults. It is concluded that HPV 6/11 cannot proliferate at nongenital cutaneous sites and HPV 2 can proliferate in the genital tract of children but not adults. Thus, the detection of HPV 6 or 11 in a genital wart in a child implies, assuming cutaneous transmission, infection from a genital site, whereas the detection of HPV 2 presumes nongenital transmission.

15. Obalek S, Jablonska S, Favre M, et al.: Condylomata acuminata in children: frequent association with human papillomaviruses responsible for cutaneous warts. J Amer Acad Dermatol 23:205-213, 1990.

16. Padayachee A: Human papillomavirus (HPV) types 2 and 57 in oral verrucae demonstrated by in situ hybridization. J Oral Pathol Med 23(9):413-417, 1994. Twenty-one cases of verrucae vulgaris (oral warts) were investigated for human papillomavirus (HPV)-group specific antigen by immunocytochemistry and for HPV types 1, 2, 4, 6, 11, 16, 18 and 57 by DNA in situ hybridization with biotinylated probes. Twelve (57%) cases demonstrated the presence of HPV-group specific antigen. Fifteen (71%) cases showed the presence of HPV DNA, 13 of which (87%) demonstrated both HPV types 2 and 57 in the same cells and 2 of which (13%) demonstrated only HPV 2. Six cases were negative for HPV 2 and 57 and all 21 cases (100%) were negative for HPV types 1, 4, 6, 11, 16 and 18. Results indicate the association of a new and as yet unidentified HPV type, closely related to HPV 2 and 57, with oral warts. The identification of both cutaneous type HPV 2 and another type closely related to HPV 2 and 57 in oral verrucae on keratinized and non-keratinized mucosal surfaces indicates the possibility of a latent infection; three patients had a history of warts on their hands, suggesting autoinoculation. This study indicated that future investigations of oral warts, based on a correlation of clinical and histological features with HPV types by DNA in situ hybridization, are called for.

17. Fierlbeck G, Rassner G, Pfister H: [Condylomata acuminata in children--detection of HPV 6/11 and 2. Local therapy with interferon-beta hydrogel]. Hautarzt 43(3):148-151, 1992. Four cases of genital warts in children (girls) are reported. HPV 6/11-DNA was identified in two cases, and HPV 2-DNA in one. In one case no virus identification was possible. The clinical features of the HPV 2-induced genital warts showed the typical morphology of condylomata acuminata. The mode of transmission of the virus, in absence of sexual contact, could not be explained. The HPV 2-associated genital warts might have been transmitted by autoinoculation from warts on the hands. Topical treatment with IFN-beta-hydrogel was applied over 8 weeks, either as single-agent therapy (1 case) or as adjuvant therapy after removal of the condylomata (3 cases). No remission was seen with the single-agent therapy. In one case the genital warts reappeared after adjuvant therapy, but in the other two cases no recurrence was seen.

18. Obalek S, Misiewicz J, Jablonska S, et al.: Childhood condyloma acuminatum: association with genital and cutaneous human papillomaviruses [see comments]. Pediatr Dermatol 10(2):101-106, 1993. We studied 25 children, age 7 months to 12 years 6 months, with anogenital warts, and their parents. In most children the warts were localized in the anal area, in 3 of 18 girls perianally and on the vulva, and in 4 girls exclusively on the vulva. Southern blot hybridization studies disclosed an association of condylomata with human papillomaviruses (HPV) 6 and 11 in 74% and HPV 2 in 17.4% of patients. The clinical features were similar in warts induced by genital and cutaneous HPVs. Even the HPV 2-associated warts in the vulva of two girls were typical of condyloma acuminatum. In all children with HPV 2-induced condylomata, cutaneous common warts coexisted, also induced by HPV 2. However, three mothers had cutaneous warts, and the children's condylomata were associated with HPV 6. Thus, the mere presence of skin warts in family members does not rule out other sources of infection. Sexual abuse was suspected in four girls and two boys, but was not confirmed in any. Nonsexual transmission could occur by persons with the lesions taking care of children. Perinatal transmission also appears to be an important route of infection in small babies. Infection in utero was probable in one girl in whom anal warts appeared in the first week of life and whose mother had cervical condylomata during pregnancy. This study provides further confirmation of possible nonsexual transmission of genital HPVs and the not infrequent association of childhood condylomata with HPV 2.

19. Naylor M, Neldner K, Yarbrough G, et al.: Contact immunotherapy of resistant warts. J Amer Acad Dermatol 19(4):679-683, 1988. Contact immunotherapy has been proved effective in the treatment of resistant warts. This report chronicles our experience with a new contact immunotherapy agent, diphenylcyclopropenone. We have achieved a cure rate of 62% in 45 patients with resistant warts of all types who came to our general dermatology clinic. Cure rates may be lower in patients who have experienced multiple treatment failures. The majority of cures were obtained within 3 to 4 months. Although it appears somewhat less effective than published reports of dinitrochlorobenzene contact immunotherapy, diphenylcyclopropenone contact immunotherapy is an effective treatment for resistant warts and avoids any potential problems from mutagenicity. [References: 31]

20. Glass AT, Solomon BA: Cimetidine therapy for recalcitrant warts in adults. Arch Dermatol 132(6):680-682, 1996. BACKGROUND: Common warts, or verrucae vulgaris, occur most often in children. However, many adults are plagued by this ubiquitous viral infection. Various modalities have been used to treat warts, but none is uniformly effective or directly antiviral. A recent study showed cimetidine to be effective in the treatment of multiple warts in children. Anecdotal reports have suggested that the administration of high doses of cimetidine, through various proposed immunomodulating mechanisms, can improve recalcitrant warts in adults. There have been no data published to date supporting these claims. OBSERVATIONS: An open-label study was conducted to determine the safety and efficacy of high-dose cimetidine in 20 adult patients with recalcitrant warts. Of the 18 patients who completed the study, 16 patients (84%) had either dramatic clinical improvement or complete resolution of their wart lesions after 3 months of cimetidine therapy without any adverse effects. No patient demonstrated disease progression while receiving the medication and complete responders remained free of lesions at 1-year follow-up. CONCLUSIONS: This study further confirms that high-dose cimetidine therapy appears to be beneficial and safe in the treatment of recalcitrant warts in adults. Further placebo-controlled studies are needed to determine its true efficacy.

21. Bauman C, Francis JS, Vanderhooft S, et al.: Cimetidine therapy for multiple viral warts in children. J Amer Acad Dermatol 35(2 Pt 1):271-272, 1996.

22. Orlow SJ, Paller A: Cimetidine therapy for multiple viral warts in children. J Am Acad Dermatol 28(5 Pt 1):794-6, 1993.

23. Yilmaz E, Alpsoy E, Basaran E: Cimetidine therapy for warts: a placebo-controlled, double-blind study. J Amer Acad Dermatol 34(6):1005-1007, 1996. BACKGROUND: Cimetidine, an H2-receptor antagonist, has been used successfully to treat patients with mucocutaneous candidiasis, common variable immunodeficiency, herpes simplex, and herpes zoster because of its immunomodulatory effects. Recently, some trials have suggested that cimetidine may also be useful for the treatment of warts. OBJECTIVE: The aim of the present study was to determine whether cimetidine is effective in the treatment of warts. METHODS: Seventy patients with multiple warts were included in a placebo-controlled, double-blind study. Patients were randomly allocated to treatment groups equally. The groups received cimetidine, 25 to 40 mg/kg daily, or placebo for 3 months. Patients were examined at monthly intervals. RESULTS: At the end of the therapy, 28 cimetidine-treated and 26 placebo-treated patients were examined to determine the efficacy of treatment. Cure rates obtained were 32% (9 of 28) in the cimetidine-treated group and 30.7% (8 of 26) in the placebo-treated group. No significant difference was found between cimetidine and placebo in effectiveness (p = 0.85). CONCLUSION: Our results show that cimetidine is no more effective than placebo in the treatment of patients with common warts.

24. Messing AM, Epstein WL: Natural history of warts: a two year study. Arch Dermatol 87:301-310, 1963.

25. Esterly NB, Cutaneous viral infections, in Nelson's textbook of pediatrics, R.E. Behrman and V.C. Vaughan, Editor. 1983, WB Saunders: Philadelphia. p. 1721-1722.

26. Urbina Gonzalez F, Cristobal Gil MC, Aguilar Martinez A, et al.: Cutaneous toxicity of intralesional bleomycin administration in the treatment of periungual warts. Arch Dermatol 122(9):974-975, 1986.

27. Epstein E: Intralesional bleomycin and Raynaud's phenomenon. J Amer Acad Dermatol 24(5 Pt 1):785-786, 1991.

28. Jenson AB, Lim LY, Singer E: Comparison of human papillomavirus type 1 serotyping by monoclonal antibodies with genotyping by in situ hybridization of plantar warts. J Cutan Pathol 16(2):54-59, 1989. Thirty plantar warts were analyzed for the presence of HPV-1 type-specific and PV genus-specific capsid antigens by immunofluorescence (IF) using monoclonal and polyclonal antibodies and type-specific HPV-1 DNA employing in situ hybridization methods. Fifteen of 30 plantar warts were positive by IF for PV genus-specific structural viral antigens. Thirteen of the 15 productively infected plantar warts expressed intranuclear HPV-1 type-specific capsid antigens and viral DNA, which were detected in the same distribution in each individual wart. The 2 productively infected plantar warts that did not react with HPV-1 type-specific MoAbs did not react with HPV-1 type-specific DNA by in situ hybridization. Thus, serotyping of HPV-1 capsid antigens by monoclonal antibodies is concordant with genotyping of HPV-1 viral DNA by in situ hybridization in productively infected plantar warts.

29. Fuchs PG, Pfister H: Cloning and characterization of papillomavirus type 2c DNA. Intervirology 22(3):177-180, 1984. Human papillomavirus (HPV) DNA was isolated from a clinically diagnosed flat wart and proved to be related to HPV2. The isolate showed 55% cross-hybridization with HPV2a. A physical map of restriction enzyme cleavage sites differed completely from those of HPV2a and HPV2b. The new HPV2 subtype, which will be classified as HPV2c, was found to be very prevalent in common warts.

30. Jablonska S, Orth G, Jarzabek-Chorzelska M, et al.: Immunological studies in epidermodysplasia verruciformis. Bull Cancer (Paris) 65(2):183-190, 1978. Immunofluorescence and cell mediated immunity studies have been performed in 14 cases of epidermodysplasia verruciformis (EV), 3 of those abortive or regressing in members of the families of the patients with EV. Two different types of human papillomavirus (HPV)--HPV3 and HPV4--have been found in cases of EV. HPV3 was detected also in flat warts without features of EV. There was no cross-reactivity between these two viruses, neither with HPV1 responsible for plantar warts nor with HPV2 inducing common warts. There was a relationship between the type of HPV and the clinical picture of EV as well as the malignant transformation, namely HPV4 has been found to be more oncogenic. Cell mediated immunity (CMI) seems to be an important factor because it was depressed in a vast majority of active cases and preserved in regressing and abortive cases (in the members of the families of EV patients). However, low CMI has been found in EV cases infected with HPV3 and in persistent flat warts also due to HPV3, which did not undergo malignant transformation. In contrast, in a case of EV due to HPV4 a malignant transformation occured in spite of still preserved, although lowered CMI. Various human papillomaviruses seem to differ in their oncogenic potential. HPV1 responsible for plantar warts, and HPV2 for common warts have no evident oncogenic potential, HPV3 inducing both EV and flat warts has a low oncogenicity, whereas HPV4 inducing some cases of EV seems more oncogenic.

31. Jablonska S, Orth G, Jarzabek-Chorzelska M, et al.: [New developments relating to papillomaviruses]. Hautarzt 30(8):411-7, 1979. Molecular hybridization technique and immunofluorescence studies with use of specific immune sera against the purified virions isolated from various types of warts and wart-like lesions of epidermodysplasia verruciformis (EV) made it possible to detect four different types of human papilloma viruses (HPV). The recognition of the viruses is important because of the different morphology of the lesions induced and their various oncogenic potentials. HPV1 is mainly responsible for plantar warts, HPV2 for common (hand) warts, HPV3 has been found both in flat warts and in the variety of EV in which skin lesions are of flat wart type, the course is relatively more benign, and usually malignant transformation is not to be expected. HPV4 was up to now found exclusively in the cases of EV with prevalent red and red-brownish plaques and hyper- and depigmentations similar to those of pityriasis versicolor. In all cases of this variety of EV malignancies occured invariably. In patients with EV, as also in--to a lesser extent--longstanding flat and/or common warts cell mediated immunity was in general lowered, but humoral specific anti-HPV antibodies were usually present. HPV type seems to be of a decisive significance for potential oncogenesis, because in a vast majority of cases EV due to HPV3 no malignancies occured in spite of anergy, whereas malignant transformation has been found in all cases due to HPV4, even in a patient with still preserved, although lowered, CMI. [References: 47]

32. Majewski S, Jablonska S: Epidermodysplasia verruciformis as a model of human papillomavirus-induced genetic cancer of the skin. Arch Dermatol 131(11):1312-1318, 1995. BACKGROUND: Epidermodysplasia verruciformis is a rare lifelong disease that has raised an enormous interest since it is a model of cutaneous genetic cancer induced by specific human papillomaviruses. OBSERVATIONS: The interacting immunogenetic and environmental factors, especially UV irradiation, result in the inability of the patients' immune system to respond to epidermodysplasia verruciformis-specific human papillomaviruses. The local immunosuppression is an effect, at least in part, of the overproduction of tumor necrosis factor alpha and transforming growth factor beta1 and of the excessive formation of cis-urocanic acid. CONCLUSIONS: Epidermodysplasia verruciformis is a model not only of cutaneous viral oncogenesis but also of local defense mechanisms in the progression of human papillomavirus-associated cancers. [References: 62]

33. de Jong-Tieben LM, Berkhout RJ, Smits HL, 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 105(3):367-371, 1995. Based on immunologic and epidemiologic data, it is plausible that skin cancer in renal transplant recipients is associated with human papillomaviruses (HPV). At present, conflicting evidence exists concerning the presence of HPV DNA in these cancers. We recently described a nested polymerase chain reaction method that enables the detection of all previously isolated epidermodysplasia verruciformis (EV)-associated HPVs. We now describe the detection of EV-associated HPV DNA in 49 (80%) of 61 biopsies from squamous cell carcinomas, in four (50%) of eight basal cell carcinomas, in 14 (93%) of 15 actinic keratoses, in two (40%) of five cases of Bowen's disease, and in four (57%) of seven keratoacanthomas. HPV DNA typing revealed that all detected HPV types belonged to the EV-associated HPV types. A wide spectrum of EV-associated HPVs was found, including six putative new HPV types. In a high percentage of the lesions more than one HPV type was detected. We often found the same HPV types in different skin biopsies from both malignant and premalignant lesions from the same patient. The high frequency of detection of EV-associated HPV types in biopsies from malignant and premalignant lesions is in agreement with the hypothesis that EV-associated HPVs are involved in the pathogenesis of skin cancer in renal transplant recipients.

34. Tieben LM, Berkhout RJ, Smits HL, et al.: Detection of epidermodysplasia verruciformis-like human papillomavirus types in malignant and premalignant skin lesions of renal transplant recipients. Br J Dermatol 131(2):226-230, 1994. To evaluate the putative role of human papillomaviruses (HPV) in the development of skin cancer in renal transplant recipients, a series of skin biopsies from premalignant and malignant skin lesions was analysed using the polymerase chain reaction. Four different consensus primer pairs were used. HPV DNA was detected in five of 24 cases of squamous cell carcinoma, in one of three cases of Bowen's disease, in none of four basal cell carcinomas, in two of seven cases of actinic keratosis and in one of five cases of keratoacanthoma. Typing by direct sequencing of the amplified HPV DNA was possible in seven of nine cases, and revealed epidermodysplasia verruciformis (EV)-associated HPV types, or HPV types related to EV-associated types. Hence, HPV DNA could be detected in a significant proportion of (pre)malignant skin tumours in renal transplant recipients. The finding that some of the detected HPV types were as yet uncharacterized EV-related types, suggests that HPV DNA could be present in a higher percentage of lesions, and might be detected with refinement of the techniques.

35. Rudlinger R, Smith IW, Bunney MH, et al.: Human papillomavirus infections in a group of renal transplant recipients. Br J Dermatol 115(6):681-692, 1986. One hundred and twenty renal transplant recipients were investigated. Fifty-eight (48%) were found to have warts, 13 (11%) keratoses and six (5%) to have, or recently to have had cancers. The longer the time of immunosuppression, the greater the prevalence of warts; of those patients who had had their transplant for at least 5 years, 87% had warts. Those with a graft survival time of 10 years or more are at special risk of warts, keratoses and malignancy. Five (10%) of 50 women had genital warts, four of whom had internal lesions (vaginal, cervical or anal) and one developed a carcinoma of the vulva. These findings indicate the advisability of colposcopy for all female renal transplant recipients, a high risk group. Eighty-eight specimens from 42 patients were examined by DNA restriction enzyme analysis and cross hybridization for the presence and type of human papillomavirus (HPV). HPV DNA was detected in 66% of the warts examined, HPV2 and HPV4 occurring most often and HPV1 and HPV3 only infrequently. In sequential specimens from common hand warts of one individual, an HPV was found which could not be precisely identified but was related to HPV4. HPV16 was detected in a vaginal wart from one patient and an HPV6-related virus in a vulval wart of another. HPV DNA of an unknown type was demonstrated in one of 11 keratoses examined. With the probes used to examine the few samples of skin cancers available, HPV16 was found in a squamous cell carcinoma of the vulva, and faint bands from an unidentified type of HPV were detected in two squamous cell carcinomata from a patient's hand. One woman had plaque lesions morphologically and histologically resembling those found in epidermodysplasia verruciformis (EV). HPV5 was identified in these lesions. This is only the third reported case of HPV5, previously thought to be unique to EV, in a renal transplant recipient.

36. Handley JM, Maw RD, Lawther H, et al.: Human papillomavirus DNA detection in primary anogenital warts and cervical low-grade intraepithelial neoplasias in adults by in situ hybridization. Sex Transm Dis 19(4):225-229, 1992. In this study, 58 consecutive patients with primary anogenital warts were selected from patients attending a genitourinary clinic. Patients were grouped on the basis of clinical lesion site, i.e. patients with genital warts only, patients with perianal or anal canal warts only, and patients with concurrent perianal and genital warts. Of these patients, 38% of the men (12/31) and 33.3% of the women (9/27) had other anogenital infections, such as nonspecific urethritis (NSU) or nonspecific genital infection, which were the most common. Of the patients who had perianal warts, 37% of the men (7/19) and 25% of the women (4/16) also had warts in the anal canal. Of the women who had anogenital warts, 63% (17/27) had concurrent subclinical low-grade cervical intraepithelial neoplasia (CIN) lesions. Human papilloma virus (HPV) DNA (either 6 or 11, 16 or 18, or 31 or 33 or 35) was detected in 53.3% (40/75) of the anogenital wart biopsy samples, and in 35.2% (6/17) of the low-grade CIN lesions. HPV types 6 or 11 were the most common types in anogenital warts (45.3%); and in CIN lesions HPV types 6 or 11 and 16 or 18 were found with equal frequency (17.6% each). There were no significant differences in HPV types between patients with genital warts and patients with perianal and anal canal warts. Anogenital infection with HPV is multicentric; external anogenital warts and subclinical CIN lesions often exist concurrently. The low prevalence of HPV DNA detected in anogenital warts and CIN biopsy samples may be due to insensitivity of the in situ hybridization technique used in this study.

37. Tsao YP, Yang KY, Han CP, et al.: Genital human papillomavirus infections in Taiwan. International Journal of Gynaecology & Obstetrics 44(1):39-45, 1994. OBJECTIVES: Identification and typing of HPV infections in genital condyloma and normal cytological cervix. METHODS: Cervical cells from 289 Pap cases with normal cytological findings were examined for HPV infection by slot blot hybridization. Fifteen condyloma biopsy specimens were studied by Southern blot hybridization. RESULTS: Thirty-six cases (12.5%) with normal cervical cytology were HPV positive. The predominant HPV type in women with normal cytology is HPV-16. Risk factors for HPV infection in women with normal cytology depend on age and history of pregnancies. Twelve cases (80%) of condyloma contained HPV-6 or -11 sequences. The predominant HPV type in genital condyloma is HPV-11. CONCLUSIONS: HPV detection in population-based screening programs for cervical neoplasia can be an important tool in identifying women who are at risk of developing dysplasia and cervical cancer.

38. Czegledy J, Veress G, Konya J, et al.: Genital human papillomavirus (HPV) infection in Hungarian women. Acta Microbiologica Hungarica 40(2):115-122, 1993. The prevalence of genital human papillomavirus (HPV) infection in Hungarian female populations is not essentially different from that found in other countries of Europe and North-America. Using filter in situ hybridization (FISH), we found that, in a group of cytologically normal women some low risk HPV types (such as HPV 6 and 11) and the most important high risk HPV types (HPV 16 and 18) were present in 23% and 8%, respectively. Eighty-eight percent of condyloma acuminatum patients harboured HPV 6 or HPV 11 in their tumours. On the other hand, in precancerous lesions (cervical intraepithelial neoplasia, CIN) HPV 16 was the predominant type, being present in 29-48% of patients, depending on the detection method used (Southern blot hybridization vs. polymerase chain reaction). The detection rate of high risk HPV types was found to rise with the increasing severity of cervical neoplasia. Finally, 48% of invasive cervical carcinoma specimens were positive for HPV 16 DNA in a type-specific polymerase chain reaction. For patients with HPV 16 positive primary tumours, all but one lymph node metastases and about 30% of histologically normal lymph nodes proved positive for HPV 16 DNA. Our results--in accordance with the numerous data found in literature--seem to confirm the hypothesis that certain HPV types are greatly involved in the development of cervical cancer.

39. Matsukura T, Sugase M: Identification of genital human papillomaviruses in cervical biopsy specimens: segregation of specific virus types in specific clinicopathologic lesions. Int J Cancer 61(1):13-22, 1995. We have established a critical identification method for the full spectrum of genital human papillomaviruses (HPVs) in clinical specimens. It was based on the recognition of PstI, BanI and MspI cleavage patterns of HPV DNA detected by blot hybridization with HPV 58 DNA probe at Tm -40 degrees C. By this method, we identified 24 different types of genital HPV including 5 novel types (HPV 59, 61, 62, 64 and 67) in the specimens collected at one hospital and found almost all the HPVs with the authentic cleavage patterns of their respective prototypes. In 235 cervical biopsy specimens, HPV 6 or 11 was found in exophytic condyloma acuminatum (15/15) but not in any cervical intraepithelial neoplasia (CIN) specimens. In contrast, HPV 18, 30, 43, 54, 56, 59, 62, 66 and 67 were identified in CIN I (28/71) or II (4/56) but not in CIN III, while HPV 16, 31, 33, 35, 39, 51, 52 and 58 were identified in CIN III (83/93) as well as in CIN I (34/71) and II (47/56). The result indicates that heterogeneous genital HPVs prevail all over the world. In addition, HPV 6 and 11 are etiologic agents only of exophytic condyloma, whereas the other HPVs are etiologic agents of CIN with the segregation of specific HPVs in CIN III. We propose a new clinicopathologic grouping of genital HPVs founded on nucleotide homology of the HPV genome.

40. Fisher AA: Severe systemic and local reactions to topical podophyllum resin. Cutis 28(3):233, 236, 242 passim, 1981.

41. Kellokoski JK, Syrjanen SM, Chang F, et al.: Southern blot hybridization and PCR in detection of oral human papillomavirus (HPV) infections in women with genital HPV infections. J Oral Pathol Med 21(10):459-464, 1992. The presence of human papillomavirus (HPV) in biopsies taken from clinically normal buccal mucosa (n = 212) and clinical lesions (n = 60) was examined by Southern blot hybridization (SBH) using 32P-labelled HPV DNA probes. Furthermore, one hundred formalin-fixed, paraffin-embedded biopsies were analyzed by using polymerase chain reaction (PCR), combined with dot blot hybridization and biotinylated HPV DNA probes. With SBH and PCR, 15.4% and 29.4% of the biopsies, respectively, contained HPV DNA. In clinically normal epithelium, 15.6% and 23.1% of the samples were HPV-positive with SBH and PCR, respectively. The HPV types detected in the genital and oral mucosa of index patients differed in all except two cases. Histology could not be relied on distinguishing HPV DNA positive and HPV DNA negative samples. Hand warts were encountered significantly more frequently in patients with a concomitant oral HPV infection. To conclude, oral HPV infections as detected by SBH and PCR are surprisingly common, but similar to the genital tract, the virus seems to exist in a latent form in the vast majority of cases. The frequent concomitant finding of skin warts and oral HPV infection may suggest some kind of HPV-specific immunosuppression.

42. Neville BW, Damm DD, Allen CM, et al., ed. Oral & Maxillofacial Pathology. 1st ed. 1995, W.B. Saunders: Philadelphia.

43. Miller CS, White DK, Royse DD: In situ hybridization analysis of human papillomavirus in orofacial lesions using a consensus biotinylated probe. American Journal of Dermatopathology 15(3):256-259, 1993. The effectiveness of the Viratype Omniprobe in situ human papillomavirus tissue hybridization kit (Digene Diagnostics) was evaluated for the detection of HPV DNA in common orofacial lesions. Seventy mucocutaneous lesions were hybridized with a biotinylated Omniprobe that was specific for HPV types 6, 11, 16, 18, 31, 33, 35, 42, 43, 44, 45, 51, 52, and 56. Eighteen (25.7%) of the specimens analyzed had intranuclear positive signals for HPV. Probing with HPV 6/11, 16/18, and 31/33/35 to delimit the HPV genotype yielded HPV DNA 6/11 in 16 (88.9%) of the Omniprobe-positive specimens. Only squamous papilloma and condyloma acuminatum were found to harbor HPV DNA. Sites most frequently infected were the labial and buccal mucosa (21.7%) and the floor of the mouth (17.4%). These results suggest that hybridization with the Omniprobe provides appropriate sensitivity and specificity for detecting HPV in some benign proliferations of the oral cavity. However, the detection of HPV in oral squamous cell carcinoma, lichen planus, and keratoacanthoma remains problematic until more sensitive and specific molecular techniques are used.

44. Padayachee A, van Wyk CW: Human papillomavirus (HPV) DNA in focal epithelial hyperplasia by in situ hybridization. J Oral Pathol Med 20(5):210-214, 1991. Eighteen cases of focal epithelial hyperplasia (FEH) were investigated for the presence of human papillomavirus (HPV) group specific antigen by immunocytochemistry and HPV types 1, 6, 11, 13, 16, 18 and 32 by DNA in situ hybridization employing biotinylated probes. Seven (39%) specimens demonstrated the presence of HPV group specific antigen. Fifteen (83%) specimens were positive for HPV DNA: 9 (60%) showed HPV 32, of which 6 were on non-keratinized mucosa and 3 on border of keratinized and non-keratinized mucosa; 5 (33%) showed HPV 13, 4 lesions on keratinized mucosa and 1 on non-keratinized mucosa; 1 (7%) specimen on non-keratinized mucosa showed HPV-11 related. Two specimens on different sites from one patient showed the same HPV type and one patient had, in addition to FEH, a squamous papilloma also demonstrating the same HPV type. Results show a specific HPV distribution pattern in the epithelium indicating areas of high viral concentration adjacent to areas of low or no viral concentration. This study also indicates the possibility of tissue-site specificity or a latent infection and the possibility of a yet unidentified HPV type associated with FEH. It is suggested that future monitoring of patients be carried out with special reference to HPV type and anatomical distribution pattern for FEH lesions.

45. Munger K, Yee CL, Phelps WC, et al.: Biochemical and biological differences between E7 oncoproteins of the high- and low-risk human papillomavirus types are determined by amino-terminal sequences. J Virol 65(7):3943-8, 1991. Differences in the biological characteristics of the high-risk human papillomavirus type 16 (HPV-16) and the low-risk HPV-6 E7 proteins were analyzed and shown to correlate with certain biochemical properties. To ascertain which region of E7 conferred these properties, chimeric E7 genes were constructed by the exchange of the amino and carboxyl coding halves of the HPV-6 and HPV-16 E7 genes. The amino-terminal half of E7 determined the affinity for binding to the retinoblastoma protein pRB, the transformation properties, and the ability to abrogate transforming growth factor beta-mediated repression of the c-myc promoter. This region of E7 is therefore responsible for the biological and biochemical differences between the E7 proteins of the low-risk and the high-risk HPVs and consequently is one of the critical determinants distinguishing these two groups of viruses. Transcriptional transactivation of the adenovirus E2 promoter, in contrast, was a property shared by E7 proteins of both low-risk and high-risk HPVs. Author-abstract

46. Sanchez Y, Elledge SJ: Stopped for repairs. Bioessays 17(6):545-548, 1995. The tumor suppressor protein p53 is intimately involved in the cellular response to DNA damage, controlling cell cycle arrest, apoptosis and the transcriptional induction of DNA damage inducible genes. A transcriptional target of p53, Gadd45, was recently found to bind to PCNA, a component of DNA replication/repair complexes, thereby implicating Gadd45 in DNA metabolism. Using biochemical assays, a role for Gadd45 in excision repair in vitro has been demonstrated. Antisense experiments have also indicated an in vivo role for the GADD45 gene in UV-irradiation survival. These discoveries establish a link between p53 and DNA repair through Gadd45. [References: 27]

47. Scheffner M, Werness BA, Huibregtse JM, et al.: The E6 oncoprotein encoded by human papillomavirus types 16 and 18 promoted the degradation of p53. Cell 63:1129-1136, 1990. The E6 protein encoded by the oncogenic human papillomavirus types 16 and 18 is one of two viral products expressed in HPV-associated cancers. E6 is an oncoprotein which cooperates with E7 to immortalize primary human keratinocytes. Insight into the mechanism by which E6 functions in oncogenesis is provided by the observation that the E6 protein encoded by HPV-16 and HPV-18 can complex the wild-type p53 protein in vitro. Wild-type p53 gene has tumor suppressor properties, and is a target for several of the oncoproteins encoded by DNA tumor viruses. In this study we demonstrate that the E6 proteins of the oncogenic HPVs that bind p53 stimulate the degradation of p53. The E6-promoted degradation of p53 is ATP dependent and involves the ubiquitin-dependent protease system. Selective degradation of cellular proteins such as p53 with negative regulatory functions provides a novel mechanism of action for dominant-acting oncoproteins.

48. Das BC, Sharma JK, Gopalkrishna V, et al.: A high frequency of human papillomavirus DNA sequences in cervical carcinomas of Indian women as revealed by Southern blot hybridization and polymerase chain reaction. J Med Virol 36(4):239-245, 1992. Ninety-six colposcopically directed biopsies from squamous epithelial carcinoma of the uterine cervix and 22 age-matched normal control biopsy specimens were examined by both Southern blot hybridization and polymerase chain reaction (PCR) for the presence of different human papillomavirus (HPV) DNA types. Cancer of the uterine cervix, which is the most common malignant disease in Indian women, showed a high frequency (98%) of HPV as compared to those reported from other parts of the world. HPV type 16 was found to be the dominant (64%) type while the frequency of HPV type 18 was very low (3%). On individual typing of HPV, no biopsy was found to contain any other known HPV types under stringent conditions of hybridization except a single case of HPV type 11. Only one case of double infection with HPV types 16 and 18 was recorded. Under low stringency conditions of hybridization with a mixed probe of HPV types 16 and 18, 29 additional biopsies were found to be positive. Southern blot hybridization alone detected HPV DNA in 92% of the cases but none in the controls. By PCR, six (6.25%) more cases and four (18.18%) healthy women were found to be positive for HPVs. Analysis of the physical state of HPV 16 indicated integration in about 70% of carcinoma cases while 30% of them were in episomal form. The findings suggest that infection with HPV is an important etiologic factor for the development of cervical cancer, that a number of such tumours may arise without HPV infection, and that integration of the viral DNA into host genome is not always essential for malignant progression.(ABSTRACT TRUNCATED AT 250 WORDS). Author-abstract.

49. Pirisi L, Creek KE, Doniger J, et al.: Continuous cell lines with altered growth and differentiation properties originate after transfection of human keratinocytes with human papillomavirus type 16 DNA. Carcinogenesis 9(9):1573-1579, 1988. Immortalization of human keratinocytes (HKc) by human papillomavirus type 16 (HPV16) is reproducible at a high frequency, is due directly to the presence of the viral sequences in the cells, and occurs independently from the genetic characteristics of the host cells. Ten human keratinocyte strains, each derived from a different individual, were transfected with pMHPV16d and selected with G418. Eight became established lines. Two strains, which failed to grow shortly after successful G418 selection, were negative for HPV16 DNA. No lines were established following transfection of the same HKc strains with vector sequences only. The immortalized lines maintained a constant number of copies of the viral genome integrated into the cellular DNA. Each line showed a unique integration pattern of HPV16 sequences into the cellular genome, but expressed similar patterns of viral messages. Sublines able to grow in the absence of growth factors (epidermal growth factor and bovine pituitary extract), and others which became resistant to differentiation stimuli (serum and calcium) were obtained by selection in growth factor-free medium and serum-supplemented medium, respectively. The establishment of continuous cell lines is a direct consequence of the presence of viral sequences; however, because none of these lines formed tumors in nude mice, additional events must be necessary for progression of malignancy. HPV16-immortalized human keratinocyte lines can be used to investigate and identify the viral factors involved with the modification of growth and differentiation control by HPV16.

50. Woodworth CD, Bowden PE, Doniger J, et al.: Characterization of normal human exocervical epithelial cells immortalized in vitro by papillomavirus types 16 and 18 DNA. Cancer Res 48:4620-4628, 1988. An in vitro system for studying the interaction between human papillomavirus (HPV) 16 and 18 recombinant DNA and normal human exocervical epithelial cells is described. Eight HPV-immortalized human exocervical epithelial cell lines were established; all the lines contained either integrated HPV16 or 18 sequences and expressed HPV mRNAs. Thus, integration and expression appear to be required for immortalization. Immortalized cells (greater than 200 population doublings to date) divided rapidly (doubling time of 30 to 46 h) and morphologically resembled primary cultures of normal human exocervical epithelial cells. They expressed a keratin pattern consistent with their origin from exocervical epithelium. When cultured at high density or in the presence of serum they terminally differentiated. Sublines resistant to terminal differentiation were selected by growth in serum-supplemented medium. Keratin pattern changes suggest they have some properties in common with cervical squamous carcinoma cells. However, HPV-immortalized cell lines were not tumorgenic in nude mice. Thus, HPV16/18 is not carcinogenic by itself. These cell lines represent an appropriate model for studying factors that regulate HPV gene expression in normal cervical epithelial cells and examining the influence of cocarcinogens on neoplastic progression.

51. Woodworth CD, Doniger J, DiPaolo JA: Immortalization of human foreskin keratinocytes by various human papillomavirus DNAs corresponds to their association with cervical carcinoma. J Virol 63(1):159-164, 1989. Normal human foreskin keratinocytes cotransfected with the neomycin resistance gene and recombinant human papillomavirus (HPV) DNAs (types 16, 18, 31, and 33) that have a high or moderate association with cervical malignancy acquired immortality and contained integrated and transcriptionally active viral genomes. Only transcripts from the intact E6 and E7 genes were detected in at least one cell line, suggesting that one or both of these genes are responsible for immortalization. Recombinant HPV DNAs with low or no oncogenic potential for cervical cancer (HPV1a, -5, -6b, and -11) induced small G418-resistant colonies that senesced as did the nontransfected cells. These colonies contained only episomal virus DNA; therefore, integration of HPV sequences is important for immortalization of keratinocytes. This study suggests that the virus-encoded immortalization function contributes to the pathogenesis of cervical carcinoma.

52. Guido MC, Zamorano R, Garrido-Guerrero E, et al.: Early promoters of genital and cutaneous human papillomaviruses are differentially regulated by the bovine papillomavirus type 1 E2 gene product. J Gen Virol 73(Pt 6):1395-1400, 1992. The physical state of the human papillomavirus (HPV) genome is usually different in malignant lesions of the skin, in which it is generally found in episomal form, and genital mucosa, in which it is frequently integrated with disruption of the E2 gene. Using chimeric or natural HPV promoters in the presence of the bovine papillomavirus type 1 E2 gene product, we observed transcription activation or repression, depending on the distance of E2-binding motifs from the start site. We found a clear difference in the positions of E2-binding motifs in cutaneous and genital HPVs that may partly explain the selective pressure for genome integration of genital HPV types in malignant lesions.

53. Swan DC, Vernon SD, Icenogle JP: Cellular proteins involved in papillomavirus-induced transformation. Arch Virol 138(1-2):105-115, 1994. Human papillomaviruses (HPVs) are associated with at least 80% of cervical carcinomas and are classified as high-risk or low-risk based on whether or not they are commonly found in cervical cancers. The high-risk HPVs have early gene products (E6 and E7) that immortalize human keratinocytes and are at least partially responsible for causing cervical carcinoma. E6 and E7 from the high-risk viruses interact strongly with the tumor suppressors p53 and Rb; those from the low-risk HPVs do not. Transformation involves a multi-step process and requires additional factors besides high-risk HPV infection. High-risk HPVs are capable of immortalizing primary human keratinocytes in tissue culture, but such cells become transformed only after certain chromosomal changes take place, possibly having to do with oncogene activation. The DNA of high-risk HPVs is frequently (if not always) integrated into the genome of cancer cells; it is normally episomal in premalignant lesions. Integration disrupts the E2 and E5 genes and viral gene regulation. Cells containing integrated viral DNA show excessively high levels of E6 and E7. While there is some conflicting evidence, it appears that the p53 and Rb tumor-suppressor genes are more frequently mutated in HPV-negative tumors than they are in HPV-positive tumors, suggesting that for tumor formation to proceed the p53 and Rb proteins must be inactivated either by interaction with the viral proteins or by mutation. The presence of an activated oncogene in a cell lacking functional p53 or Rb may then be sufficient to cause tumor progression. [References: 67]

54. Jeon S, Allen-Hoffmann BL, Lambert PF: Integration of human papillomavirus type 16 into the human genome correlates with a selective growth advantage of cells. J Virol 69(5):2989-2997, 1995. Integration of human papillomavirus type 16 (HPV-16) DNA into a host chromosome has been hypothesized to result in altered expression of two viral transforming genes, E6 and E7, in cervical cancers. In order to investigate the role that changes in viral genomic state and gene expression play in cervical carcinogenesis, we have derived clonal populations of human cervical epithelial cells which harbor multiple copies of either extrachromosomal or integrated viral DNA. The clonal populations harboring extrachromosomal HPV-16 DNA stably maintained approximately 1,000 viral copies for at least 15 passages (approximately 100 cell doublings), which contrasted with the unstable HPV-16 replicons in the parental counterpart. In the clonal populations harboring integrated viral DNA, 3 to 60 copies of HPV-16 DNA were found integrated in either of two forms: type 1, in which all the copies of HPV-16 DNA were disrupted in the E2 open reading frame upon integration, and type 2, in which intact viral copies were flanked by disrupted viral copies and cellular sequences. Despite the lower HPV-16 DNA copy number, the clonal populations with integrated viral DNA had levels of E7 protein that were in most cases higher than those found in the clonal populations harboring extrachromosomal viral DNA. Irrespective of viral genomic state, the clonal populations were capable of undergoing terminal differentiation and unable to form colonies in soft agar, which is indicative of the nontumorigenic nature of these cells. Importantly, a cell population with integrated viral DNA was found to outgrow another with extrachromosomal DNA when these cells were cocultured over a period of time. Thus, integration of human papillomaviral DNA correlates with increased viral gene expression and cellular growth advantage. These observations are consistent with the hypothesis that integration provides a selective advantage to cervical epithelial precursors of cervical carcinoma.

55. Chen SL, Tsao YP, Lee JW, et al.: Characterization and analysis of human papillomaviruses of skin warts. Arch Dermatol Res 285(8):460-465, 1993. We analysed human papillomavirus (HPV) infections in 61 tissue specimens of skin warts of Taiwanese patients by DNA hybridization. The prevalence of HPV infection was 69% by Southern blot hybridization. The typing of HPVs was performed by dot blot hybridization under highly stringent conditions with each probe separately. The prevalence of HPV-1, 2/3, 4, 5, 8, 11, 16 and 18 in skin warts was 13, 7, 16, 2, 0, 5, 2 and 8%, respectively. Chi-squared analysis revealed that there was a correlation between HPV type and copy number. Most HPV-4-induced warts were verruca vulgaris. HPV-1 DNA was detected in verruca plantaris and verruca vulgaris. No specific histopathological features were found to be indicative of the presence or absence of HPV, or of the various types of HPV infection.

56. Kienzler JL, Lemoine MT, Orth G, et al.: Humoral and cell-mediated immunity to human papillomavirus type 1 (HPV-1) in human warts. Br J Dermatol 108(6):665-672, 1983. The humoral and cell-mediated immune response to human papillomavirus type 1 (HPV-1) has been studied in 162 patients carrying papillomas of various clinical types: deep plantar wart or myrmecia, common wart, flat wart, and anogenital wart. Circulating antibodies were detected by immunodiffusion and microcomplement fixation, using purified HPV-1 particles as type-specific antigen. A significant association between myrmecia and anti-HPV-1 antibodies was found (39% of the cases). Cell-mediated immunity was evaluated by a study of delayed hypersensitivity (DH). The main capsid components of HPV-1 (HPV-1 CP), consisting mostly of a polypeptide of molecular weight 54,000, were injected intradermally. In addition to the type-specific antigens, HPV-1 CP contain other antigenic determinants shared by various types of human papilloma-viruses and masked in intact viral particles. The DH tests to HPV-1 CP showed no differences between the carriers of different papilloma types, confirming the presence of common antigenic determinants. Moreover, they gave rise to an increase or to new anti-HPV-1 antibody production mostly in myrmecia carriers (78% and 33% of the cases, respectively), and to new DH to HPV-1 CP in all groups of papilloma carriers (33% to 56%, depending on the clinical papilloma type).

57. Jablonska S, Obalek S, Favre M, et al.: The morphology of butchers' warts as related to papillomavirus types. Arch Dermatol Res 279(Suppl):S66-72, 1987. Hand warts were studied in 160 butchers. Clinical and histological studies were performed in 190 warts and virological studies in 165 warts from 104 butchers. Since we found almost perfect correlation between the histological pattern and the type of infecting virus, it was possible to evaluate the virus types in a further 39 of 56 butchers without virological studies, on the basis of the histology of the warts. The most common infection was with HPV-2 (human papilloma virus) and HPV-7. Thirty-three butchers were infected with two types of viruses and three butchers with three HPVs. The morphology of warts varied considerably. The majority were similar to verrucae vulgares or verrucae planae. Some deep warts resembled myrmecia-type verrucae plantares. Often, several types of warts coexisted. Some clinical patterns were shown to be preferentially associated with distinct types of papillomaviruses: common warts with HPV-2, HPV-4, or HPV-7, plane and intermediate warts with HPV-3, HPV-10, HPV-28. HPV-7, previously identified for the first time in these butchers, was found to be associated with common warts or common wart-like, papillomatous lesions.

58. Jacyk WK, De Villiers EM: Epidermodysplasia verruciformis in Africans. Int J Dermatol 32(11):806-810, 1993. BACKGROUND. Epidermodysplasia verruciformis (EV) is a rare cutaneous disorder characterised by persistent, refractory infection with human papillomaviruses (HPV). Although EV does not seem to have racial or geographic preference, there is a scarcity of reports on its occurrence in Africans. METHODS. Twenty Africans with EV were studied, and the literature on this condition in Africans was reviewed. Virologic studies were performed on 10 patients. RESULTS. Three types of lesions were observed: flat warts, pityriasis versicolor-like macules, and seborrheic keratosis-like changes. Malignant transformation occurred in only one patient. HPV-3 was isolated only from flat warts, HPV-5 and HPV-17 were isolated only from PV-like lesions, whereas an HPV-5-related type was found in all three types of changes. HPV-5-related type revealed DNA that was related but not identical to any of the viruses in the HPV-5 group. This particular type was isolated from all five South African patients with EV in whom virologic studies were performed. CONCLUSIONS. The benign nature of EV in dark-skinned Africans has been confirmed. Four HPV types have been isolated, of which HPV-related type was found in all South African patients with EV and in all types of skin changes, regardless of their morphology. African patients with EV frequently present seborrheic keratosis-like changes.

59. Kanda R, Tanigaki T, Kitano Y, et al.: Types of human papillomavirus isolated from Japanese patients with epidermodysplasia verruciformis. Br J Dermatol 121(4):463-469, 1989. Virological studies were performed on 12 patients with epidermodysplasia verruciformis (EV). Three types of lesions were observed: red plaques, pityriasis versicolor (PV)-like macules and plane warts. Human papillomavirus (HPV) 14, 20 and 21 were isolated from the plaques, HPV 3, 14 and 38 from flat warts and HPV 5, 12, 17, 20 and 38 from PV-like lesions. No clear relationship could be established between the different lesions and the types of HPV. Types 17 and 20 have been isolated most frequently from Japanese EV patients and HPV 5, frequently detected in other countries, is less common, whereas HPV 8 has not been isolated. Skin cancers occurred in six of the cases (50%) and all had benign lesions that were PV-like. At least one type of HPV 5, 17 or 20 could be isolated from these benign lesions and HPV 17 or 20 detected in the cancers. These three types of HPV in EV patients appear to be involved in the malignant transformation. Author-abstract

60. Ostrow RS, Manias D, Mitchell AJ, et al.: Epidermodysplasia verruciformis. A case associated with primary lymphatic dysplasia, depressed cell-mediated immunity, and Bowen's disease containing human papillomavirus 16 DNA. Arch Dermatol 123(11):1511-6, 1987. Epidermodysplasia verruciformis is a rare, often hereditary disease characterized by a generalized cutaneous infection with human papillomavirus (HPV), depressed cell-mediated immunity, and a propensity for transformation of the warty lesions to squamous cell carcinoma on primarily sun-exposed areas of the skin. A 37-year-old man with congenital lymphatic dysplasia and a history of squamous cell carcinoma of the groin and foot was observed by us to have edema of all four extremities, numerous flat warts, and pityriasis versicolor-like papules over the trunk and arms. Condylomatous lesions were noted in the groin and a periungual verrucous nodule on the thumb. Biopsies showed the trunk and arm lesions to be verrucae and the thumb lesion to be Bowen's disease. Results of molecular hybridization studies from four lesions of the arms showed the presence of only HPV 3 DNA; HPV 16-related DNA was detected in the intraepidermal carcinoma on the thumb. Immunologic evaluation revealed anergy to routine skin testing, depressed mitogen-stimulated lymphocyte transformation, decreased B-lymphocyte count, and a severe reversal of the T-lymphocyte helper:suppressor ratio. Author-abstract

61. Kremsdorf D, Jablonska S, Favre M, et al.: Human papillomaviruses associated with epidermodysplasia verruciformis. II. Molecular cloning and biochemical characterization of human papillomavirus 3a, 8, 10, and 12 genomes. J Virol 48(2):340-351, 1983. The DNAs of four human papillomaviruses (HPVs) that were found in the benign lesions of three patients suffering from epidermodysplasia verruciformis have been characterized. The flat wart-like lesions and the macular lesions of patient 1 contained two viruses, HPV-3a and HPV-8, respectively, whose genomes had previously been only partially characterized. The flat wart-like lesions of patient 2 and the macular lesions of patient 3 each contained a virus previously considered as belonging to types 3 and 5, respectively. These viruses are shown in the present study to be different from all of the HPV types so far characterized; they have tentatively been named HPV-10 and HPV-12. The HPV-3a, HPV-8, and HPV-12 DNAs and the two SalI fragments of HPV-10 DNA (94.1 and 5.9% of the genome length) were cloned in Escherichia coli after having been inserted in plasmid pBR322. The cloned HPV genomes have similar sizes (about 7,700 base pairs), but their guanine-plus-cytosine contents differ from 41.8% for HPV-12 DNA to 45.5% for HPV-3a DNA. The study of the sensitivity of the four HPV DNAs to 14 restriction endonucleases permitted the construction of cleavage maps. Evidence for conserved restriction sites was found only for the HPV-3a and HPV-10 genomes since 5 of the 21 restriction sites localized in the HPV-3a DNA seem to be present also in the HPV-10 DNA. Hybridization experiments, performed in liquid phase at saturation, showed a 35% sequence homology between HPV-3a and HPV-10 DNAs, 17 to 29% sequence homology among HPV-5, HPV-8, and HPV-12 DNAs, almost no sequence homology between the HPV-3a or HPV-10 DNA and the other HPV DNAs, and a weak homology between HPV-9 DNA and HPV-8 or HPV-12 DNA. Blot hybridization experiments showed no sequence homology between the HPV-3a, HPV-8, and HPV-12 DNAs and the DNAs of the HPVs associated with skin warts (HPV-1a, HPV-2, HPV-4, and HPV-7) or with mucocutaneous and mucous membrane lesions (HPV-6b and HPV-11a, respectively). One exception was a weak sequence homology between the HPV-2 prototype and HPV-3a or HPV-10 DNA.(ABSTRACT TRUNCATED AT 400 WORDS)

62. Ostrow R, Zachow K, Watts S, et al.: Characterization of two HPV-3 related papillomaviruses from common warts that are distinct clinically from flat warts or epidermodysplasia verruciformis. J Invest Dermatol 80(5):436-440, 1983. We have recently identified two unusual human papillomavirus (HPV) isolates while engaged in an ongoing study of wart disease in meat handlers and veterinarians. The papillomas from which these two viruses were isolated clinically resembled verruca vulgaris rather than either flat warts or epidermodysplasia verruciformis (EV). These two previously uncharacterized HPVs were molecularly cloned and characterized with respect to known HPVs. The genomes of the two viruses exhibited dramatically different restriction endonuclease cleavage patterns but were found to have significant sequence homology to each other, as well as to HPV-3 and a new virus isolated from a patient with EV. Neither of the two new HPV isolates exhibit detectable sequence homology under stringent conditions of hybridization or share similar restriction endonuclease cleavage patterns with previously characterized HPV types 1,2,4,5,6b, or a previously isolated HPV from meat handlers.

63. Pfister H: Human papillomaviruses and skin cancer. Semin Cancer Biol 3(5):263-271, 1992. Human papillomavirus (HPV)-induced skin warts are classically benign lesions. However an association between specific HPV types and skin cancer becomes obvious in epidermodysplasia verruciformis (EV). The analysis of this disease suggests that lesions infected with HPV types 5 and 8 carry a high risk of developing squamous cell carcinomas. The oncogenes of EV-viruses appear to be E6 and E2, rather than E7. The 'high risk' EV-viruses, HPV 5, 8, and 47, differ from related HPV types in the transforming activity of the E6 gene and in the density of positive transcription control elements in the non-coding region (NCR) of the genome. The extrachromosomal viral DNA in cancers may show deletions affecting regulatory sequences. EV-specific lesions occasionally occur in immunosuppressed patients and HPV 5 or 8 persist in some of the skin cancers to which these patients are prone. DNAs of HPV 2, 16, 34, or 41 were identified in few premalignant and malignant skin tumors of the general population. [References: 65]

64. Melchers W, de Mare S, Kuitert E, et al.: Human papillomavirus and cutaneous warts in meat handlers. J Clin Microbiol 31(9):2547-2549, 1993. The association of papillomavirus and hand warts in meat handlers was examined. Human papillomavirus (HPV) DNA was found in 23 (88%) of 26 cutaneous warts, with HPV 7 (27%) and a yet unidentified HPV (HPV X) (42%) being the predominant types. HPV 2 was found in two (7.5%) patients, and HPV 4 was found in three (11.5%) patients. No bovine papillomavirus sequences were detected. In most patients, the warts developed in less than 2 years after they started working with meat. A possible HPV transmission route by protection gloves and professional equipment is suggested.

65. Jablonska S, Obalek S, Golebiowska A, et al.: Epidemiology of butchers' warts. Arch Dermatol Res 280(Suppl):S24-S28, 1988. Studies were carried out in two slaughter-houses in different cities differing in the degree of work automation and, for comparison, in workers of nearby factories of the same two cities. There was a high incidence of warts (49.2%) in a slaughterhouse where the workers had direct contact with animals and meat, while a significantly lower incidence (9%) was observed in a modern slaughterhouse where the work was almost completely automated. The types of human papillomaviruses (HPVs) were similar in warts of butchers from these slaughterhouses and of 63 butchers from various slaughterhouses all over the country. All cutaneous HPVs were present in butchers' warts. The so-called butchers' wart virus HPV-7 was found in about 30% of the butchers from all slaughterhouses. Cell-mediated immunity of the butchers was found to be unimpaired. There was no correlation between the incidence of infection and the frequency of antibodies against HPV-1, HPV-2, or HPV-3.

66. Kremsdorf D, Jablonska S, Favre M, et al.: Biochemical characterization of two types of human papillomaviruses associated with epidermodysplasia verruciformis. J Virol 43(2):436-447, 1982. The DNAs of the human papillomaviruses (HPVs) associated with the benign lesions of two patients suffering from epidermodysplasia verruciformis (patients JD and JK) were analyzed by using 12 restriction endonucleases. None of the restriction endonucleases were one-cut enzymes for the HPV DNA obtained from patient JD, referred to as the prototypical HPV-5, whereas five of them were one-cut enzymes for the DNA of the major virus found in patient JK, referred to as HPV-9. The molecular cloning of the two fragments resulting from the cleavage of HPV-5 DNA by endonuclease HindIII and of the single fragment obtained after treatment of HPV-9 DNA with endonuclease BamHI was performed in Escherichia coli after the fragments were inserted in plasmid pBR322. A cleavage map of the two cloned genomes was constructed. Little sequence homology (4 to 5%) was detected between HPV-5 and HPV-9 DNAs by DNA-DNA hybridization experiments in liquid phase at saturation; this homology was reproducibly higher than that (2 to 3%) detected under the same conditions between these DNAs and HPV-1a DNA. In addition, blot hybridization experiments performed under stringent conditions showed no or little sequence homology between the DNAs of HPV-5 and HPV-9 and those of HPV prototypes of types 1, 2, 3, 4, and 7 associated with skin warts. These results confirm that HPV-5 and HPV-9 are two distinct HPV types.

67. Ficarra G, Adler-Storthz K, Galeotti F, et al.: Focal epithelial hyperplasia (Heck's disease): the first reported case from Italy. Tumori 77(1):83-85, 1991. A case of focal epithelial hyperplasia (Heck's disease) of the oral mucosa observed for the first time in Italy is reported. The patient was of Italian extraction. The lesions, represented by soft nodules, were multiple and located on the vestibular and labial mucosa. Biopsy tissues were studied for the presence of human papilloma virus (HPV) by electron microscopy, in situ hybridization and immunoperoxidase staining to HPV group antigens. No viral particles consistent with HPV were found in the epithelial cells. The tissues were positive for HPV antigen by immunoperoxidase staining and hybridized to the HPV 13 probe. In situ hybridization to the HPV 6, HPV 11, HPV 16, HPV 18 and 32 probes was negative. Our study substantiates that focal epithelial hyperplasia, although rare, may be observed in Whites and that HPV 13 in associated with the disease.

68. Williamson AL, Dennis SJ: The use of the polymerase chain reaction for the detection of human papillomavirus type 13. J Virol Methods 31(1):57-65, 1991. Human papillomavirus type 13 (HPV-13) is associated with oral focal epithelial hyperplasia (FEH). The purpose of this study was to establish conditions for the application of polymerase chain reaction (PCR) to the specific detection and amplification of HPV-13 DNA. To design primers for HPV-13 a part of the HPV-13 genome was sequenced first: the smallest BamHI fragment (597 bp) of HPV-13 was subcloned and sequenced. The sequence was found to be part of a large open reading frame and had significant homology with the L1 gene of other HPVs. HPV-13 specific primers were designed to amplify a 240 bp fragment from the L1 gene by PCR. Conditions for PCR were standardized for this set of primers.

69. Henke RP, Guerin-Reverchon I, Milde-Langosch K, et al.: In situ detection of human papillomavirus types 13 and 32 in focal epithelial hyperplasia of the oral mucosa. J Oral Pathol Med 18(7):419-421, 1989. 17 cases of focal epithelial hyperplasia of the oral mucosa (FEH, Heck's disease) were investigated for the presence of human papillomavirus (HPV) nucleic acid sequences by means of in situ DNA hybridization using biotinylated DNA probes of HPV types 1, 6, 11, 13, 16, 18, and 32. Ten of 17 cases were positive for HPV 13 DNA in contrast to 6 of 17 positive cases obtained after application of the HPV 32 probe, with a double infection in one case. The results of our study suggest, that HPV 13 and HPV 32 are very specifically found in lesions of FEH and can be detected in a high percentage of cases using in situ hybridization.

70. Garlick JA, Calderon S, Buchner A, et al.: Detection of human papillomavirus (HPV) DNA in focal epithelial hyperplasia. J Oral Pathol Med 18(3):172-177, 1989. Five focal epithelial hyperplasia (FEH) specimens from four patients were examined by Southern blot hybridization analysis to determine the specific human papillomavirus (HPV) types present. The histomorphologic features of these specimens were also evaluated and a broad variety of changes including koilocytes, mitosoid cells, ballooning cells and cells showing individual cell keratinization were noted. FEH lesions from the three patients sharing a familial relationship demonstrated HPV DNA sequences that were either the prototype HPV-13 or a very closely related HPV-13 subtype. These patients also showed similar clinical features. Lesional tissue from the other patient was found to harbor HPV DNA sequences similar to HPV-32. In view of these findings it is suggested that these specific HPV types are associated with the characteristic FEH histomorphology described.

71. Henke RP, Milde-Langosch K, Loning T, et al.: Human papillomavirus type 13 and focal epithelial hyperplasia of the oral mucosa: DNA hybridization on paraffin-embedded specimens. Virchows Arch A Pathol Anat Histopathol 411(2):193-198, 1987. 16 cases of focal epithelial hyperplasia (Heck's disease) were studied for the presence of human papillomavirus DNA by means of nucleic acid hybridization. Hybridization was carried out in situ with biotin-labelled probes of HPV 1, 6, 11, 13, 16, and 18 DNA under stringent and non-stringent conditions. Under non-stringent conditions, 6 of 16 cases (38%) hybridized to a mixture of HPV 1, 6, 11, 16, and 18 DNA. When these probes were applied under stringent conditions, only one case could be shown to be weakly positive for HPV 6/11 DNA. Further stringent hybridizations, which were conducted with a HPV 13 probe on 12 of our 16 cases, revealed a positive result in 9 of 12 cases (75%). The results of our study strongly substantiate the concept that HPV 13 or a closely related HPV type is associated with lesions morphologically presenting as focal epithelial hyperplasia.

72. Hernandez-Jauregui P, Eriksson A, Tamayo Perez R, et al.: Human papillomavirus type 13 DNA in focal epithelial hyperplasia among Mexicans. Brief report. Arch Virol 93(1-2):131-137, 1987. Human papillomavirus (HPV) type 13 DNA was detected in focal epithelial hyperplasia lesions of the oral mucosa in seven half-caste mexicans. The lesions contained intracellular papillomavirus-like particles with a diameter of about 50 nm. DNA extracted from biopsies contained unintegrated HPV type 13 DNA genomes as revealed by Southern blot hybridization. The HPV 13 DNA that was isolated in the present study had the same restriction enzyme cleavage map as HPV 13 DNA, previously described by others. It was moreover confirmed that HPV type 13 genome is related to the genomes of HPV types 6 and 11.

73. Lutzner MA, Blanchet-Bardon C, Orth G: Clinical observations, virologic studies, and treatment trials in patients with epidermodysplasia verruciformis, a disease induced by specific human papillomaviruses. J Invest Dermatol 83(1 Suppl):18s-25s, 1984. We have studied 11 patients with the papillomavirus-induced disease epidermodysplasia verruciformis (EV). Clinical diagnostic features are widespread, long-lasting, pityriasis versicolor-like macules and flat, wart-like papules, both usually occurring in early childhood, with the subsequent development in the third decade of multiple skin cancers of the Bowenoid in situ and squamous cell types, primarily in sun-exposed skin. Virologic studies using the methods of immunofluorescence microscopy, restriction endonuclease analysis, and DNA blot hybridization have shown benign lesions to be associated with one or several types of the human papillomaviruses (HPVs) specifically associated with EV (at least 15 types recognized on the basis of sequence homology studies of molecularly cloned genomes). Skin cancers in these patients were associated with the genomes of either HPV-5, HPV-8 or HPV-14, suggesting that these three viruses are potentially oncogenic. A genetic factor appears to play a role in the pathogenesis of EV, since 5 of our patients were children of consanguineous parents and 2 had siblings also suffering with EV, suggesting a recessive inheritance pattern. Treatment of 4 EV patients with an oral retinoid resulted in partial temporary improvement of benign lesions, and the treatment of 2 patients with intralesional interferon injections into multiple Bowenoid cancers in situ has resulted in the disappearance of these lesions. Finally, EV serves as a model for studying the interplay of oncogenic viruses, genetic and immunologic factors, and sunlight in the production of skin cancer in humans.

74. Lutzner MA: Papillomaviruses and skin cancer in Africa. IARC Sci Publ (63):607-623, 1984. Although it was suggested long ago that certain epithelial cancers preceded by papillomas might be caused by viruses, the first proof that papillomaviruses were associated with cancer dates from the work on rabbits in 1934 by Shope and Rose. In the 1970s, the introduction of the blot hybridization technique enabled Orth and his co-workers at the Institut Pasteur, Paris, to demonstrate the presence in man of the DNA of human papillomavirus type 5 (HPV-5) in cancers following Lutz-Lewandovsky epidermodysplasia verruciformis. Some years later, it was possible to demonstrate the presence of the same HPV-5 DNA in the skin cancer of an immunosuppressed recipient of a renal transplant. The number of potentially oncogenic papillomaviruses has recently been increased by the demonstration at the Institut Pasteur of the presence of the DNAs of HPV-8 and HPV-14 in the skin cancers of patients with epidermodysplasia verruciformis. In recent years, an association has been demonstrated between HPV-6, -10, -11, -16, and -18 with verrucous cancers of the skin and mucous membranes, Bowenoid papules, Bowenoid skin diseases and cervical cancer. Such cancers of the skin and mucous membranes are usually treated by surgery, but it has been shown that oral administration of retinoids (synthetic derivatives of vitamin A) or the use of leucocyte interferon intralesionally are effective in cancers in situ following epidermodysplasia verruciformis.

75. Bergeron C, Barrasso R, Beaudenon S, et al.: Human papillomaviruses associated with cervical intraepithelial neoplasia. Great diversity and distinct distribution in low- and high-grade lesions. Am J Surg Pathol 16(7):641-649, 1992. All together, 30 genital human papillomavirus (HPV) types have been characterized so far. To evaluate the importance of HPV diversity in associated cervical diseases, we analyzed 188 biopsy specimens obtained from patients with a recent diagnosis of cervical HPV infection or intraepithelial neoplasia (CIN). Of these 188 specimens, 116 were classified as low-grade CIN (48 cases), high-grade CIN (53 cases), condylomata acuminata (10 cases), flat condylomas (five cases). Seventy-two specimens were considered nondiagnostic. Using probes specific for 18 genital HPV types, HPV DNA sequences were detected by Southern blot hybridization in 100 lesions and 21 nondiagnostic specimens. When further analyzed by the polymerase chain reaction, eight HPV-negative biopsy specimens, four CIN, and four nondiagnostic specimens were positive. Of the 129 positive biopsy specimens, 92 contained at least one of 18 known HPV types and 37 HPV that have not yet been identified. Nine specimens had more than one type. Thirteen HPV types were identified in CIN. The detection rate of HPV 16 increased from 21% in low-grade CIN to 57% in high-grade CIN. HPV 18 was detected in only 3% of CIN; HPV 31, 33, and 35 were found in 8%. HPV 30, 39, 45, 51, 52, 56, 58, and 61 were detected in 44% of low-grade CIN but in only 8% of high-grade CIN. Unidentified HPV were detected in about 25% of low-grade and high-grade CIN. Fifty-seven CIN positive for at least one HPV type were further analyzed by in situ hybridization. Thirty-five (65%) biopsy specimens were positive, including 21 of 24 low-grade CIN and 14 of 33 high-grade CIN. Ten of the 13 previously identified HPV types were detected. Thus, CIN represents an heterogeneous disease from a virologic viewpoint. This fact could explain their variable clinical evolution.

76. Gassenmaier A, Pfister H, Hornstein OP: Human papillomavirus 25-related DNA in solitary keratoacanthoma. Arch Dermatol Res 279(2):73-76, 1986. Solitary keratoacanthomas of 32 patients were screened for the presence of human papillomavirus (HPV) 25 DNA, which was originally isolated and molecularly cloned from warts of an epidermodysplasia verruciformis (Ev) patient. Biotinylated virus DNA was hybridized in situ to thin sections obtained from paraffin-embedded material. HPV DNA was detected in 12 of 32 tumors under stringent conditions, and in 2 additional tumors under relaxed conditions.

77. Payne D, Chan TS, Wagner R, et al.: Cloning of mucosal and cutaneous HPV sequences in a metastatic squamous cell carcinoma from an epidermodysplasia verruciformis patient. Anticancer Res 16(3A):1165-1166, 1996. Human papillomavirus (HPV) is a DNA tumor virus strongly associated with cervical neoplasias. There are over 80 different types of HPVs which can infect either mucosal or cutaneous tissue. Cutaneous squamous cell carcinomas (SCC) associated with HPV are often seen in patients with epidermodysplasia verruciformis (EV). EV is characterized by cutaneous lesions that progress to SCC upon UV exposure. In characterizing the HPV types associated with an unusually aggressive form of EV, we have cloned an HPV with homology to the moderately oncogenic genital type HPV 34, the oncogenic EV type HPV 5 and from benign oral mucosal type HPV 32. The presence of sequences from these highly divergent types is a novel finding. These three viral types are from different phylogenetic branches of the HPV family believed to have evolved independently from each other.

78. Kawashima M, Favre M, Obalek S, et al.: Premalignant lesions and cancers of the skin in the general population: evaluation of the role of human papillomaviruses. J Invest Dermatol 95(5):537-542, 1990. To evaluate the role of human papillomaviruses (HPV) in the development of premalignant lesions and cancers of the skin in the general population, 314 biopsies obtained from 227 patients with benign neoplasms, premalignant lesions, and cancers of the skin and from 25 patients with squamous cell carcinoma of the lip were analyzed by Southern blot hybridization. DNA probes specific for various cutaneous and genital HPV types were used in hybridizations conducted under nonstringent or stringent conditions. HPV DNA sequences were only detected in eight specimens obtained from six patients: HPV 34 in one case of periungual Bowen's disease, HPV 36 and an as yet uncharacterized HPV in two cases of actinic keratosis, HPV 20 in one case of basal cell carcinoma, an as yet unrecognized HPV in one case of squamous cell carcinoma, and HPV 16 in one case of squamous cell carcinoma of the lip. None of the specimens of cutaneous horn and keratoacanthoma contained detectable HPV DNA. In contrast, HPV DNA sequences, mostly HPV 16, were detected in 13 of 23 cases of anogenital Bowen's disease and invasive Bowen's carcinoma. HPV DNA sequences were not detected in 90 cutaneous samples further analyzed by the polymerase chain-reaction technique, using amplification primers that contain conserved sequences among the genomes of HPV. These results strongly suggest that the known HPV types play only a minor role, if any, in skin carcinogenesis in the general population. Author-abstract

79. Scheurlen W, Gissmann L, Gross G, et al.: Molecular cloning of two new HPV types (HPV 37 and HPV 38) from a keratoacanthoma and a malignant melanoma. Int J Cancer 37(4):505-510, 1986. Several benign and malignant skin tumors were analyzed for the presence of human papillomavirus (HPV) DNA. By hybridization with different HPV DNA probes under non-stringent conditions (Tm -40 degrees C), two tumors were found to contain HPV-specific DNA sequences in high copy numbers: (1) a keratoacanthoma from a patient who also suffered from a basalioma; (2) a superficial spreading malignant melanoma of an immunosuppressed patient. For further analysis of these DNA sequences genomic libraries from both tumor DNAs were constructed and, out of these, 4 different HPV DNA types have been cloned. By cross-hybridization experiments and restriction map analysis HPV 9 DNA was identified in the keratoacanthoma whereas HPV 17a DNA could be cloned from the malignant melanoma. From each tumor one additional HPV-type not identical to other known HPV-types was cloned. These isolates are closely related to HPV 9, 15, 17, 22 and 23. A physical map of both HPV DNAs was constructed. Size (7.8 kbp), co-linear alignment to HPV 16, cross-hybridization with other HPV-types under conditions of low stringency and monomeric episomal state of the HPV molecules indicate that these two DNA probes represent new HPV types that have been tentatively designated as HPV 37 (keratoacanthoma) and HPV 38 (malignant melanoma). None of these two HPV types could be found in any other of 231 tumor DNAs originating from different tissues. Author-abstract

80. Yutsudo M, Kanda R, Tanigaki T, et al.: Human papillomavirus type 38 isolated from patients with epidermodysplasia verruciformis. Intervirology 26(1-2):104-108, 1986. A new type of human papillomavirus (HPV), HPV-38b, was isolated from patients with epidermodysplasia verruciformis and was molecularly cloned. This HPV was shown by hybridization experiments to have almost no sequence homology with other types of reported HPVs, but did show homology with HPV-38, which was recently isolated from a melanoma. A physical map and a rough genetic map of the organization of this HPV are presented.

81. Joste NE, Rushing L, Granados R, et al.: Bethesda classification of cervicovaginal smears: reproducibility and viral correlates. Hum Pathol 27(6):581-585, 1996. Fifty-five cervicovaginal smears from women with squamous intraepithelial lesions (SILs) were independently evaluated on two separate occasions by four cytopathologists using a binary classification system (the Bethesda system). Smears were categorized as low-grade (LSIL) or high-grade (HSIL) using previously published criteria. All women had subsequent cervical biopsies containing human papillomavirus (HPV) DNA amplified with the polymerase chain reaction and typed by restriction fragment polymorphism analysis. Three or more observers agreed on classification in 49 of 55 cases (87%); unanimous diagnoses were rendered in 31 cases (56%). Interobserver and intraobserver reproducibility ranged from fair to near-excellent (kappa values 0.40 to 0.63; 0.63 to 0.74, respectively). HPV types included HPV 16 (27%), 18 (7%), 31 (9%), 35 (4%), 39 (4%), 6 (10%), 11 (2%), novel types (30%), and multiple types (4%). High-risk HPV types (16, 18, 31, 35, and 39) were significantly associated (P = .03) with consensus HSIL diagnoses (agreement of three or more observers). This was primarily because of the strong association of HPV 16 with HSIL (P = .001). After excluding HPV 16, the other high-risk HPV types (18, 31, 35, and 39) were no longer significantly associated with consensus HSIL diagnoses (P > .5). Conversely, LSIL diagnoses were significantly associated with non-high-risk HPV types (all HPV types except 16, 18, 31, 35, and 39; P = .006). Binary cytological classification of cervicovaginal SILs is reproducible among cytopathologists. Such classification correlates well with most low-risk HPV types and with the prototypic high-risk HPV 16 but not with other high-risk HPV types.

82. de Villiers EM, Hirsch-Behnam A, von Knebel-Doeberitz C, et al.: Two newly identified human papillomavirus types (HPV 40 and 57) isolated from mucosal lesions. Virology 171(1):248-253, 1989. Two new papillomaviruses, HPV 40 and HPV 57, were isolated from a PIN lesion and an inverted papilloma of the maxillary sinus, respectively. HPV 40 showed a 13% homology to HPV 7 by reassociation kinetics and HPV 57 showed a 17% homology to HPV 2 and 25% homology to HPV 27. Hybridization of the DNA of these papillomaviruses to a wide variety of different tumor biopsies revealed that HPV 40 was present in a few genital condylomata acuminata as well as in bowenoid lesions. HPV 57 DNA was present in an oral wart, a genital condyloma acuminatum, and verrucae vulgares lesions from two immunosuppressed patients.

83. Hirt L, Hirsch-Behnam A, de Villiers EM: Nucleotide sequence of human papillomavirus (HPV) type 41: an unusual HPV type without a typical E2 binding site consensus sequence. Virus Res 18(2-3):179-189, 1991. The complete nucleotide sequence of human papillomavirus type 41 (HPV-41) has been determined. HPV-41 was originally isolated from a facial wart, but its DNA has subsequently been detected in some skin carcinomas and premalignant keratoses (Grimmel et al., Int. J. Cancer, 1988, 41, 5-9; de Villiers, Grimmel and Neumann, unpublished results). The analysis of the cloned HPV-41 nucleic acid reveals that its genome organisation is characteristic as for other papillomavirus types. Yet, the analysis indicates at the same time that this virus is most distantly related to all other types of human-pathogenic papillomaviruses sequenced thus far and appears to identify HPV-41 as the first member of a new subgroup of HPV. The overall nucleotide homology to other sequenced HPV types is below 50%. The closest other HPV type is represented by HPV-18, sharing 49% identical nucleotides. The typical E2 binding sequence ACCN6GGT, found in all papillomaviruses analyzed to date, does not occur in the URR of the HPV-41 genome. Modified E2 binding sequences, as described for BPV 1 (Li et al., Genes Dev. 1989, 3, 510-526), are located in the domain proximal to the E6 ORF. These are ACCN6GTT, AACN6GGT and the two perfect palindromic sequences AACGAATTCGTT.

84. Grimmel M, de Villiers E-M, Neumann C, et al.: Characterization of a new human papillomavirus (HPV 41) from disseminated warts and detection of its DNA in some skin carcinomas. Int J Cancer 41(1):5-9, 1988. A new human papillomavirus type (HPV 41), distantly related to known HPV prototypes, was isolated from a patient with disseminated facial, peri-anal and foot warts (epidermodysplasia verruciformis was not diagnosed). The viral DNA was molecularly cloned in 2 BamH1 restriction fragments with sizes of 6.5 kb and 0.98 kb, respectively. The classification of this cloned DNA as a new type is based on the degree of cross-hybridization with 40 HPV types under conditions of varying stringency. A total of 106 benign and malignant skin lesions, as well as 71 malignant tumours of different origins, were screened for the presence of HPV-related sequences. In 2 out of 10 squamous-cell carcinomas and in 1 of 3 arsenic keratoses HPV 41 DNA sequences could be detected. Author-abstract

85. Lorincz AT, Quinn AP, Goldsborough MD, et al.: Cloning and partial DNA sequencing of two new human papillomavirus types associated with condylomas and low-grade cervical neoplasia. J Virol 63(6):2829-2834, 1989. Using low-stringency Southern blot analysis and cloning in lambda bacteriophage, two new human papillomavirus types (HPV-43 and HPV-44) were identified and their DNAs were cloned from vulvar tissues. The isolates were characterized by restriction endonuclease mapping and shown to be new HPV types on the basis of their minimal hybridization with all other known HPV types at high stringency. Both HPVs are most closely related to types 6, 11, and 13. HPV-43 did not exhibit any cross-reactivity with these HPV types at high stringency. HPV-44 showed minimal cross-reactivity to HPV-13, which was in the range of 20 to 25% according to liquid hybridization analysis. The deduced genomic organization of each of the two new HPVs was colinear with HPV-6b. Prevalence studies revealed that HPV-43 and HPV-44 together were found in 6 of 439 normal cervical tissues, in 8 of 195 cervical intraepithelial neoplasms, but in none of 56 cervical cancers tested thus far.

86. Kiyono T, Adachi A, Ishibashi M: Genome organization and taxonomic position of human papillomavirus type 47 inferred from its DNA sequence. Virology 177(1):401-405, 1990. The complete nucleotide sequence of human papillomavirus type 47 (HPV-47) DNA isolated from the lesion of epidermodysplasia verruciformis (EV) was determined. The computer-aided comparison of HPV-47 with other EV-associated viruses using the available sequence data on them revealed that HPV-47 resembles both HPV-5 and HPV-8 as much as HPV-5 and HPV-8 resemble each other, and it led us to regard these three viruses as one cluster and HPV-19 and HPV-25 as another. The conclusion implies that HPV-47 as well as HPV-5 and HPV-8 is associated with the cancer occurrence in EV. Two sets of splicing donor and acceptor sequences in HPV-47, which were previously shown to work in vivo, are also conserved in HPV-5 and HPV-8. One of them allows formation of an ORF predicted to encode an E1/E4 fused protein.

87. Favre M, Obalek S, Jablonska S, et al.: Human papillomavirus type 49, a type isolated from flat warts of renal transplant patients. J Virol 63(11):4909, 1989. The cloning and characterization of the genome of human papillomavirus type 49 (HPV-49) is described. The viral DNA, which is most closely related to the DNAs of HPVs seen in patients with epidermodysplasia verruciformis, was aligned to the HPV-5 genome by electron microscopic analysis of heteroduplexes between the cloned viral DNAs.

88. Nuovo GJ, Crum CP, De Villiers EM, et al.: Isolation of a novel human papillomavirus (type 51) from a cervical condyloma. J Virol 62(4):1452-1455, 1988. We cloned the DNA from a novel human papillomavirus (HPV) present in a cervical condyloma. When DNA from this isolate was hybridized at high stringency with HPV types 1 through 50 (HPV-1 through HPV-50), it showed weak homology with HPV-6 and -16 and stronger homology with HPV-26. A detailed restriction endonuclease map was prepared which showed marked differences from the maps for other HPVs that have been isolated from the female genital tract. Reassociation kinetic analysis revealed that HPV-26 and this new isolate were less than 10% homologous; hence, the new isolate is a novel strain of HPV. The approximate positions of the open reading frames of the new strain were surmised by hybridization with probes derived from individual open reading frames of HPV-16. In an analysis of 175 genital biopsies from patients with abnormal Papanicolaou smears, sequences hybridizing under highly stringent conditions to probes from this novel HPV type were found in 4.2, 6.1, and 2.4% of biopsies containing normal squamous epithelium, condylomata, and intraepithelial neoplasia, respectively. In addition, sequences homologous to probes from this novel isolate were detected in one of five cervical carcinomas examined.

89. Frattini MG, Lim HB, Laimins LA: In vitro synthesis of oncogenic human papillomaviruses requires episomal genomes for differentiation-dependent late expression. Proc Natl Acad Sci 93(7):3062-3067, 1996. Human papillomavirus (HPV) types 16, 18, 31, and 51 are the etiologic agents of many anogenital cancers including those of the cervix. These "high risk" HPVs specifically target genital squamous epithelia, and their lytic life cycle is closely linked to epithelial differentiation. We have developed a genetic assay for HPV functions during pathogenesis using recircularized cloned HPV 31 genomes that were transfected together with a drug resistance marker into monolayer cultures of normal human foreskin keratinocytes, the natural host cell. After drug selection, cell lines were isolated that stably maintained HPV 31 DNA as episomes and underwent terminal differentiation when grown in organotypic raft cultures. In differentiated rafts, the expression of late viral genes, amplification of viral DNA, and production of viral particles were detected in suprabasal cells. This demonstrated the ability to synthesize HPV 31 virions from transfected DNA templates and allowed an examination of HPV functions during the vegetative viral life cycle. We then used this system to investigate whether an episomal genome was required for the induction of late viral gene expression. When an HPV 31 genome (31E1*) containing a missense mutation in the E1 open reading frame was transfected into normal human keratinocytes, the mutant viral sequences were found to integrate into the host cell chromosomal DNA with both early and late regions intact. While high levels of early viral gene transcription were observed, no late gene expression was detected in rafts of cell lines containing the mutant viral genome despite evidence of terminal differentiation. Therefore, the induction of late viral gene expression required that the viral genomes be maintained as extrachromosomal elements, and terminal differentiation alone was not sufficient. These studies provide the basis for a detailed examination of HPV functions during viral pathogenesis.

90. Katase K, Teshima H, Hirai Y, et al.: Natural history of cervical human papillomavirus lesions. Intervirology 38(3-4):192-194, 1995. A total of 87 HPV-positive patients with grade I and II cervical intraepithelial neoplasia (CIN I and II) were followed up by cytology and colposcopy every 3 months for more than 5 years following the first biopsy. These patients were classified into three groups (progressive, persistent, and regressive disease) according to the results. The human papillomavirus (HPV) genome and viral types were identified