Herbert Pfister

Reply: Virological treatment failure of protease inhibitor therapy in an unselected cohort of HIV-infected patients.

Objective: To determine the rate of virological treatment failure with protease inhibitor therapy in unselected patients and to assess underlying risk factors. Design and setting: Retrospective study in two German tertiary care treatment centres. Patients: A total of 198 HIV-infected patients treated with protease inhibitors in 1996. Main outcome measures: Levels of HIV RNA 1-6 months after start of treatment; definition of treatment failure of < 1 log 10 reduction in plasma HIV RNA within 6 months after starting protease inhibitor therapy; multivariate analysis of risk factors for treatment failures. Results: A total of 226 treatment episodes with protease inhibitors were evaluable (saquinavir, 83; ritonavir, 47; indinavir, 96). The rate of virological treatment failure was 44% (saquinavir, 64%; ritonavir, 38%; indinavir, 30%). In a multivariate analysis, the following independent risk factors for virological failure were found: CD4 cell count, pretreatment with antiretroviral drugs (number), and protease inhibitor (compound). The relative risk reduction for each CD4 cell count increase was 0.997 (P = 0.012), 2.64 for pretreatment with one or two drugs versus no drug (P = 0.05), 2.97 for pretreatment with more than two drugs versus no drug (P = 0.05), and 4.62 for treatment with saquinavir versus indinavir (P = 0.001). Conclusion: An unexpectedly high rate of virological treatment failure of protease inhibitor therapy was found in an unselected cohort of HIV-infected patients. Response to antiretroviral combination therapy in normal clinical practice may considerably differ from results of randomized clinical trials. Further studies are warranted to find optimal treatment strategies for both initial and salvage therapy.

Human papilloma viruses (HPV): Characterization of four different isolates

Abstract Out of 50 papilloma virus isolates from individual human warts (verrucae vulgares and verrucae plantares) 36 were analyzed for cleavage pattern of their DNA after restriction enzyme digestion with the endonucleases EcoRl, BamHI, Hind II, Hind III, Hpa II, and Hae III. Furthermore, the electrophoretic mobility of virion proteins from some of the same as well as from the additional isolates was studied by SDS-polyacrylamide gel electrophoresis. Four different cleavage patterns were observed: While three individual isolates (HPV 1, HPV 2, HPV 3) had many cleavage sites in common, and differed only in a few sites, a fourth one (HPV 4) was found to be entirely different. The electrophoretic mobility of proteins of HPV 4 was also observed to differ from those of HPV 1–3. cRNA transcribed either from HPV 1 or HPV 4 did not hybridize with the heterologous isolate. Rabbit antiserum reacting highly against HPV 1 did not react with HPV 4 proteins in complement fixation tests. We thus conclude that several human papilloma viruses exist: While HPV 1–3 are closely related isolates, HPV 4 represents a new human papilloma virus, profoundly different from the previous ones as far as base composition and antigenicity are concerned. Preliminary data suggest that the latter type occurs in approximately 20% of verrucae vulgares with low virus production, whereas HPV 1, representative for about 70% of these papillomas, predominates in warts with high virus yields.

Human papillomaviruses and genital cancer

Publisher Summary Papillomaviruses are clearly proven to be the etiologic agents of anogenital lesions, with the potential to progress to squamous cell carcinomas. Viral particles, DNA, and/or capsid antigens can be demonstrated in benign precursors and the proliferations can be transmitted from person to person by cell-free extracts. The complete nucleotide sequences of Human Papillomaviruses (HPVs)—namely, HPV 6, HPV 11, and HPV 16—revealed the familiar genome organization of papillomaviruses and homologous sequences to those open reading frames that were shown to code for transforming functions in the case of BPV1. Malignant conversion of papillomavirus-induced tumors is well established for a number of animal systems and for the human disease epidennodysplasia verruciformis. HPV DNA persists in genital carcinomas either integrated into the host genome or extrachromosomally and is transcribed at least in some tumors. The worldwide prevalence of HPV 16 is noteworthy and may possibly indicate an increased cancerogenic potential of this virus. A tentative calculation indicates that the cancer risk after HPV infection equals or exceeds the risk attended with the infections by other human tumor viruses such as HTLVI, Epstein-Barr virus, or hepatitis B virus. Tumor progression is certainly subjected to additional factors such as chemical or physical carcinogens, hormones, and systemic or local immune deficiencies. The molecular biology of HPV infection has to be further examined to learn more about the viral role in keratinocyte transformation, which will be essential to define relevant diagnostic probes and possible candidates for therapeutic intervention.

Merkel Cell Polyomavirus DNA in Persons without Merkel Cell Carcinoma

Merkel cell polyomavirus (MCPyV) DNA was detected in 88% of Merkel cell carcinomas in contrast to 16% of other skin tumors. MCPyV was also found in anogenital and oral samples (31%) and eyebrow hairs (50%) of HIV-positive men and in forehead swabs (62%) of healthy controls. MCPyV thus appears to be widespread.

Oncogenic Human Papillomavirus DNA Loads in Human Immunodeficiency Virus-Positive Women with High-Grade Cervical Lesions Are Strongly Elevated

ABSTRACT Human papillomavirus (HPV) DNA loads of six oncogenic HPV types were measured by real-time PCR in cervical scrapes of human immunodeficiency virus (HIV)-infected and uninfected women. In both groups, HPV loads increased with the grade of cervical disease. HIV infection did not affect HPV loads in low-grade lesions but was associated with significantly higher HPV loads in severe dysplasia; highest loads were found in advanced HIV disease. Our data reflect the aggressive course of HPV infection in HIV-positive women.

The E7 Protein of Cutaneous Human Papillomavirus Type 8 Causes Invasion of Human Keratinocytes into the Dermis in Organotypic Cultures of Skin

Human papillomaviruses (HPV) have been implicated in the development of nonmelanoma skin cancer (NMSC). The molecular mechanisms by which these viruses contribute towards NMSC are poorly understood. We have used an in vitro skin-equivalent model generated by transducing primary adult human epidermal keratinocytes with retroviruses expressing HPV genes to investigate the mechanisms of viral transformation. In this model, keratinocytes expressing HPV genes are seeded onto a mesenchyme composed of deepidermalized human dermis that had been repopulated with primary dermal fibroblasts. Expression of the HPV8 E7 gene caused both an enhancement of terminal differentiation and hyperproliferation, but most strikingly, the acquisition of the ability to migrate and invade through the underlying dermis. The basement membrane integrity was disrupted in a time-dependent manner in areas of invading keratinocytes, as evidenced by immunostaining of its protein components collagen types VII, IV, and laminin 5. This was accompanied by the overexpression of extracellular matrix metalloproteinases MMP-1, MMP-8, and MT-1-MMP. These results suggest that the cutaneous HPV type 8 that is frequently found in NMSC of epidermodysplasia verruciformis patients may actively promote an invasive keratinocyte phenotype. These findings also highlight the importance of epithelial-extracellular matrix-mesenchymal interactions that are required to support cell invasion.

The Papillomavirus E2 Protein Binds to and Synergizes with C/EBP Factors Involved in Keratinocyte Differentiation

ABSTRACT The papillomavirus life cycle is closely linked to the differentiation program of the host keratinocyte. Thus, late gene expression and viral maturation are restricted to terminally differentiated keratinocytes. A variety of cellular transcription factors including those of the C/EBP family are involved in the regulation of keratinocyte differentiation. In this study we show that the papillomavirus transcription factor E2 cooperates with C/EBPα and -β in transcriptional activation. This synergism was independent of an E2 binding site. E2 and C/EBP factors synergistically transactivated a synthetic promoter construct containing classical C/EBPβ sites and the C/EBPα-responsive proximal promoter of the involucrin gene, which is naturally expressed in differentiating keratinocytes. C/EBPα or -β coprecipitated with E2 proteins derived from human papillomavirus type 8 (HPV8), HPV16, HPV18, and bovine papillomavirus type 1 in vitro and in vivo, indicating complex formation by the cellular and viral factors. The interaction domains could be mapped to the C terminus of E2 and amino acids 261 to 302 located within the bZIP motif of C/EBPβ. Our data suggest that E2, via its interaction with C/EBP factors, may contribute to enhancing keratinocyte differentiation, which is suppressed by the viral oncoproteins E6 and E7 in HPV-induced lesions.

Papillomavirus DNA in warts of immunosuppressed renal allograft recipients.

SummaryRenal allograft recipients were shown to have an increased incidence of warts and skin cancers. We examined 148 patients for evidence of wart virus infections and tested for papillomavirus types, which are known to be associated with human malignancies. Of the 148, 36 (24.3%) patients were afflicted with warts at the end of our study period, in contrast to 5 of 148 (3.3%) before transplantation. DNA from 16 different biopsies was extracted by phenol treatment for further virological studies. DNA of human papillomavirus (HPV) 2 was detected three times, DNA of HPV 4 and 10 twice, and DNA of HPV 3 and 16 once each by blot hybridization. One probe led to strong signals with HPV types previously found only in epidermodysplasia verruciformis patients. A correlation between histology and virus type exsisted in cases of HPV 2, 3, 4, and 10 infections.

Evolution of raltegravir resistance during therapy.

OBJECTIVES We investigated the prevalence of raltegravir resistance-associated mutations at baseline and their evolution during raltegravir therapy in patients infected with different HIV-1 subtypes. METHODS At pre-treatment screening, the integrase gene from plasma samples from patients infected with subtype B and non-B viruses was analysed. Raltegravir resistance evolution was further evaluated in 10 heavily pre-treated patients. RESULTS Two hundred and nine plasma samples from 94 subtype B and 115 non-B patients were sequenced. No signature/primary raltegravir resistance mutations were detected at baseline. The secondary mutations L74M, T97A, V151I and G163R were observed with a frequency of <4%. The primary mutations N155H, Q148R/H or Q143R were observed during raltegravir therapy. The Q148R/H was detected only in subtype B. A switch of the primary mutation during raltegravir treatment was not restricted to the subtype B viruses. The prevalence of each primary mutation varied depending on the length of the raltegravir therapy. The Q148R/H was mostly detected after short exposure to raltegravir, while the Y143R was observed only after prolonged raltegravir exposure. We detected an association between the presence of the T206S in the baseline genotype and the absence of the primary Q148R/H mutation or any secondary mutation accompanying the N155H following raltegravir failure. CONCLUSIONS A number of secondary and additional mutations were found in baseline genotypes. During therapy, when the virus was not optimally suppressed, resistance mutations developed, which were dependent on subtype and time on raltegravir.

Detection of Adenoviruses and Rotaviruses in Drinking Water Sources Used In Rural Areas of Benin, West Africa

ABSTRACT Diseases associated with viruses also found in environmental samples cause major health problems in developing countries. Little is known about the frequency and pattern of viral contamination of drinking water sources in these resource-poor settings. We established a method to analyze 10 liters of water from drinking water sources in a rural area of Benin for the presence of adenoviruses and rotaviruses. Overall, 541 samples from 287 drinking water sources were tested. A total of 12.9% of the sources were positive for adenoviruses and 2.1% of the sources were positive for rotaviruses at least once. Due to the temporary nature of viral contamination in drinking water sources, the probability of virus detection increased with the number of samples taken at one test site over time. No seasonal pattern for viral contaminations was found after samples obtained during the dry and wet seasons were compared. Overall, 3 of 15 surface water samples (20%) and 35 of 247 wells (14.2%) but also 2 of 25 pumps (8%) tested positive for adenoviruses or rotaviruses. The presence of latrines within a radius of 50 m in the vicinity of pumps or wells was identified as being a risk factor for virus detection. In summary, viral contamination was correlated with the presence of latrines in the vicinity of drinking water sources, indicating the importance of appropriate decision support systems in these socioeconomic prospering regions.