Human Papillomavirus (HPV) 16 E6 Variants in Tonsillar Cancer in Comparison to Those in Cervical Cancer in Stockholm, Sweden

Background: Human papillomavirus (HPV), especially HPV16, is associated with the development of both cervical and tonsillar cancer and intratype variants in the amino acid sequence of the HPV16 E6 oncoprotein have been demonstrated to be associated with viral persistence and cancer lesions. For this reason the presence of HPV16 E6 variants in tonsillar squamous cell carcinoma (TSCC) in cervical cancer (CC), as well as in cervical samples (CS), were explored. Methods: HPV16 E6 was sequenced in 108 TSCC and 52 CC samples from patients diagnosed 2000–2008 in the County of Stockholm, and in 51 CS from young women attending a youth health center in Stockholm. Results: The rare E6 variant R10G was relatively frequent (19%) in TSCC, absent in CC and infrequent (4%) in CS, while the well-known L83V variant was common in TSCC (40%), CC (31%), and CS (29%). The difference for R10G was significant between TSCC and CC (p = 0.0003), as well as between TSCC and CS (p = 0.009). The HPV16 European phylogenetic lineage and its derivatives dominated in all samples (.90%). Conclusion: The relatively high frequency of the R10G variant in TSCC, as compared to what has been found in CC both in the present study as well as in several other studies in different countries, may indicate a difference between TSCC and CC with regard to tumor induction and development. Alternatively, there could be differences with regard to the oral an

Characterization and Whole Genome Analysis of Human Papillomavirus Type 16 E1-1374^63nt Variants

Background. The variation of the most common Human papillomavirus (HPV) type found in cervical cancer, the HPV16, has been extensively investigated in almost all viral genes. The E1 gene variation, however, has been rarely studied. The main objective of the present investigation was to analyze the variability of the E6 and E1 genes, focusing on the recently identified E1-1374^63nt variant. Methodology/Principal Findings. Variation within the E6 of 786 HPV16 positive cervical samples was analyzed using high-resolution melting, while the E1-1374^63nt duplication was assayed by PCR. Both techniques were supplemented with sequencing. The E1-1374^63nt duplication was linked with the E-G350 and the E-C109/G350 variants. In comparison to the referent HPV16, the E1-1374^63nt E-G350 variant was significantly associated with lower grade cervical lesions (p=0.029), while the E1-1374^63nt E-C109/G350 variant was equally distributed between high and low grade lesions. The E1-1374^63nt variants were phylogenetically closest to E-G350 variant lineage (A2 sub-lineage based on full genome classification). The major differences between E1-1374^63nt variants were within the LCR and the E6 region. On the other hand, changes within the E1 region were the major differences from the A2 sub-lineage, which has been historically but inconclusively associated with high grade cervical disease. Thus, the shared variations cannot explain the particular association of the E1-1374^63nt variant with lower grade cervical lesions. Conclusions/Significance. The E1 region has been thus far considered to be well conserved among all HPVs and therefore uninteresting for variability studies. However, this study shows that the variations within the E1 region could possibly affect cervical disease, since the E1-1374^63nt E-G350 variant is significantly associated with lower grade cervical lesions, in comparison to the A1 and A2 sub-lineage variants. Furthermore, it appears that the silent variation 109T>C of the E-C109/G350 variant might have a significant role in the viral life cycle and warrants further study

Quercetin elevates p27Kip1 and arrests both primary and HPV16 E6/E7 transformed human keratinocytes in G1

Our previous work with primary bovine fibroblasts demonstrated that quercetin, a potent mutagen found in high levels in bracken fern (Pteridium aquilinum), arrested cells in G1 and G2/M, in correlation with p53 activation. The expression of bovine papillomavirus type 4 (BPV-4) E7 overcame this arrest and lead to the development of tumorigenic cells lines (Beniston et al., 2001). Given the possible link between papillomavirus infection, bracken fern in the diet and cancer of the upper gastrointestinal (GI) tract in humans, we investigated whether a similar situation would occur in human cells transformed by human papillomavirus type 16 (HPV-16) oncoproteins. Quercetin arrested primary human foreskin keratinocytes in G1. Arrest was linked to an elevation of the cyclin-dependent kinase inhibitor (cdki) p27Kip1. Expression of the HPV16 E6 and E7 oncoproteins in transformed cells failed to abrogate cell cycle arrest. G1 arrest in the transformed cells was also linked to an increase of p27Kip1 with a concomitant reduction of cyclin E-associated kinase activity. This elevation of p27Kip1 was due not only to increased protein half-life, but also to increased mRNA transcription

Detection of human papillomavirus DNA and p53 codon 72 polymorphism in prostate carcinomas of patients from Argentina

BACKGROUND: Infections with high-risk human papillomaviruses (HPVs), causatively linked to cervical cancer, might also play a role in the development of prostate cancer. Furthermore, the polymorphism at codon 72 (encoding either arginine or proline) of the p53 tumor-suppressor gene is discussed as a possible determinant for cancer risk. The HPV E6 oncoprotein induces degradation of the p53 protein. The aim of this study was to analyse prostate carcinomas and hyperplasias of patients from Argentina for the presence of HPV DNA and the p53 codon 72 polymorphism genotype. METHODS: HPV DNA detection and typing were done by consensus L1 and type-specific PCR assays, respectively, and Southern blot hybridizations. Genotyping of p53 codon 72 polymorphism was performed both by allele specific primer PCRs and PCR-RFLP (Bsh1236I). Fischer's test with Woolf's approximation was used for statistical analysis. RESULTS: HPV DNA was detected in 17 out of 41 (41.5 %) carcinoma samples, whereas all 30 hyperplasia samples were HPV-negative. Differences in p53 codon 72 allelic frequencies were not observed, neither between carcinomas and hyperplasias nor between HPV-positive and HPV-negative carcinomas. CONCLUSION: These results indicate that the p53 genotype is probably not a risk factor for prostate cancer, and that HPV infections could be associated with at least a subset of prostate carcinomas

Interaction between polymorphisms of the Human Leukocyte Antigen and HPV-16 Variants on the risk of invasive cervical cancer

<p>Abstract</p> <p>Background</p> <p>Persistent infection with oncogenic types of human papillomavirus (HPV) is the major risk factor for invasive cervical cancer (ICC), and non-European variants of HPV-16 are associated with an increased risk of persistence and ICC. HLA class II polymorphisms are also associated with genetic susceptibility to ICC. Our aim is to verify if these associations are influenced by HPV-16 variability.</p> <p>Methods</p> <p>We characterized HPV-16 variants by PCR in 107 ICC cases, which were typed for <it>HLA-DQA1</it>, <it>DRB1 </it>and <it>DQB1 </it>genes and compared to 257 controls. We measured the magnitude of associations by logistic regression analysis.</p> <p>Results</p> <p>European (E), Asian-American (AA) and African (Af) variants were identified. Here we show that inverse association between <it>DQB1*05 </it>(adjusted odds ratio [OR] = 0.66; 95% confidence interval [CI]: 0.39–1.12]) and HPV-16 positive ICC in our previous report was mostly attributable to AA variant carriers (OR = 0.27; 95%CI: 0.10–0.75). We observed similar proportions of <it>HLA DRB1*1302 </it>carriers in E-P positive cases and controls, but interestingly, this allele was not found in AA cases (p = 0.03, Fisher exact test). A positive association with <it>DRB1*15 </it>was observed in both groups of women harboring either E (OR = 2.99; 95% CI: 1.13–7.86) or AA variants (OR = 2.34; 95% CI: 1.00–5.46). There was an inverse association between <it>DRB1*04 </it>and ICC among women with HPV-16 carrying the 350T [83L] single nucleotide polymorphism in the <it>E6 </it>gene (OR = 0.27; 95% CI: 0.08–0.96). An inverse association between <it>DQB1*05 </it>and cases carrying 350G (83V) variants was also found (OR = 0.37; 95% CI: 0.15–0.89).</p> <p>Conclusion</p> <p>Our results suggest that the association between HLA polymorphism and risk of ICC might be influenced by the distribution of HPV-16 variants.</p

Going beyond histology. Synchrotron micro-computed tomography as a methodology for biological tissue characterization: from tissue morphology to individual cells

Current light microscopic methods such as serial sectioning, confocal microscopy or multiphoton microscopy are severely limited in their ability to analyse rather opaque biological structures in three dimensions, while electron optical methods offer either a good three-dimensional topographic visualization (scanning electron microscopy) or high-resolution imaging of very thin samples (transmission electron microscopy). However, sample preparation commonly results in a significant alteration and the destruction of the three-dimensional integrity of the specimen. Depending on the selected photon energy, the interaction between X-rays and biological matter provides semi-transparency of the specimen, allowing penetration of even large specimens. Based on the projection-slice theorem, angular projections can be used for tomographic imaging. This method is well developed in medical and materials science for structure sizes down to several micrometres and is considered as being non-destructive. Achieving a spatial and structural resolution that is sufficient for the imaging of cells inside biological tissues is difficult due to several experimental conditions. A major problem that cannot be resolved with conventional X-ray sources are the low differences in density and absorption contrast of cells and the surrounding tissue. Therefore, X-ray monochromatization coupled with a sufficiently high photon flux and coherent beam properties are key requirements and currently only possible with synchrotron-produced X-rays. In this study, we report on the three-dimensional morphological characterization of articular cartilage using synchrotron-generated X-rays demonstrating the spatial distribution of single cells inside the tissue and their quantification, while comparing our findings to conventional histological techniques

Genetic Epidemiology of Glioblastoma Multiforme: Confirmatory and New Findings from Analyses of Human Leukocyte Antigen Alleles and Motifs

Human leukocyte antigen (HLA) class I genes mediate cytotoxic T-lymphocyte responses and natural killer cell function. In a previous study, several HLA-B and HLA-C alleles and haplotypes were positively or negatively associated with the occurrence and prognosis of glioblastoma multiforme (GBM).As an extension of the Upper Midwest Health Study, we have performed HLA genotyping for 149 GBM patients and 149 healthy control subjects from a non-metropolitan population consisting almost exclusively of European Americans. Conditional logistic regression models did not reproduce the association of HLA-B*07 or the B*07-Cw*07 haplotype with GBM. Nonetheless, HLA-A*32, which has previously been shown to predispose GBM patients to a favorable prognosis, was negatively associated with occurrence of GBM (odds ratio=0.41, p=0.04 by univariate analysis). Other alleles (A*29, A*30, A*31 and A*33) within the A19 serology group to which A*32 belongs showed inconsistent trends. Sequencing-based HLA-A genotyping established that A*3201 was the single A*32 allele underlying the observed association. Additional evaluation of HLA-A promoter and exon 1 sequences did not detect any unexpected single nucleotide polymorphisms that could suggest differential allelic expression. Further analyses restricted to female GBM cases and controls revealed a second association with a specific HLA-B sequence motif corresponding to Bw4-80Ile (odds ratio=2.71, p=0.02).HLA-A allelic product encoded by A*3201 is likely to be functionally important to GBM. The novel, sex-specific association will require further confirmation in other representative study populations