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

Influence of hydrocarbon oil structure on adjuvanticity and autoimmunity

Mineral oils are extensively used in our daily life, in food, cosmetics, biomedicine, vaccines and in different industrial applications. However, exposure to these mineral oils has been associated with immune adjuvant effects and the development of autoimmune diseases. Here we investigate the structural impacts of the hydrocarbon oil molecules on their adjuvanticity and autoimmunity. First, we showed that hydrocarbon oil molecules with small atomic differences could result in experimental arthritis in DA rats differing in disease severity, incidence, weight change and serum levels of acute phase proteins. Injection of these hydrocarbon oils resulted in the activation, proliferation and elevated expression of Th1 and especially Th17 cytokines by the T cells, which correlate with the arthritogenicity of the T cells. Furthermore, the more arthritogenic hydrocarbon oils resulted in an increased production of autoantibodies against cartilage joint specific, triple-helical type II collagen epitopes. When injected together with ovalbumin, the more arthritogenic hydrocarbon oils resulted in an increased production of alpha beta T cell-dependent anti-ovalbumin antibodies. This study shows the arthritogenicity of hydrocarbon oils is associated with their adjuvant properties with implications to not only arthritis research but also other diseases and medical applications such as vaccines in which oil adjuvants are involved

Zinc finger arrays binding human papillomavirus types 16 and 18 genomic DNA: precursors of gene-therapeutics for <it>in-situ</it> reversal of associated cervical neoplasia

<p>Abstract</p> <p>Background</p> <p>Human papillomavirus (HPV) types 16 and 18 are the high-risk, sexually transmitted infectious causes of most cervical intraepithelial neoplasias (CIN) or cancers. While efficacious vaccines to reduce the sexual acquisition of these high-risk HPVs have recently been introduced, no virus-targeted therapies exist for those already exposed and infected. Considering the oncogenic role of the transforming (E6 and E7) genes of high-risk HPVs in the slow pathogenesis of cervical cancer, we hypothesize that timely disruption or abolition of HPV genome expression within pre-cancerous lesions identified at screening may reverse neoplasia. We aimed to derive model zinc finger nucleases (ZFNs) for mutagenesis of the genomes of two high-risk HPV (types 16 & 18).</p> <p>Methods and results</p> <p>Using ZiFiT software and the complete genomes of HPV types16 and 18, we computationally generated the consensus amino acid sequences of the DNA-binding domains (F1, F2, & F3) of (i) 296 & 327 contextually unpaired (or single) three zinc-finger arrays (sZFAs) and (ii) 9 & 13 contextually paired (left and right) three- zinc-finger arrays (pZFAs) that bind genomic DNA of HPV-types 16 and 18 respectively, inclusive of the E7 gene (s/pZFA_HpV/E7). In the absence of contextually paired three-zinc-finger arrays (pZFAs) that bind DNA corresponding to the genomic context of the E6 gene of either HPV type, we derived the DNA binding domains of another set of 9 & 14 contextually unpaired E6 gene-binding ZFAs (sZFA_E6) to aid the future quest for paired ZFAs to target E6 gene sequences in both HPV types studied (pZFA_E6). This paper presents models for (i) synthesis of hybrid ZFNs that cleave within the genomic DNA of either HPV type, by linking the gene sequences of the DNA-cleavage domain of the <it>FokI</it> endonuclease F_N to the gene sequences of a member of the paired-HPV-binding ZFAs (pZFA_HpV/E7 + F_N), and (ii) delivery of the same into precancerous lesions using HPV-derived viral plasmids or vectors.</p> <p>Conclusions</p> <p>With further optimization, these model ZFNs offer the opportunity to induce target-mutagenesis and gene-therapeutic reversal of cervical neoplasia associated with HPV types 16 & 18.</p