The papillomavirus capsid is a nonenveloped icosahedral shell formed by the viral major structural protein, L1. It is known that disulfide bonds between neighboring L1 molecules help to stabilize the capsid. However, the kinetics of inter-L1 disulfide bond formation during particle morphogenesis have not previously been examined. We have recently described a system for producing high-titer papillomavirus-based gene transfer vectors (also known as pseudoviruses) in mammalian cells. Here we show that papillomavirus capsids produced using this system undergo a maturation process in which the formation of inter-L1 disulfide bonds drives condensation and stabilization of the capsid. Fully mature capsids exhibit improved regularity and resistance to proteolytic digestion. Although capsid maturation for other virus types has been reported to occur in seconds or minutes, papillomavirus capsid maturation requires overnight incubation. Maturation of the capsids of human papillomavirus types 16 and 18 proceeds through an ordered accumulation of dimeric and trimeric L1 species, whereas the capsid of bovine papillomavirus type 1 matures into more extensively cross-linked forms. The presence of encapsidated DNA or the minor capsid protein, L2, did not have major effects on the kinetics or extent of capsid maturation. Immature capsids and capsids formed from L1 mutants with impaired disulfide bond formation are infectious but physically fragile. Consequently, capsid maturation is essential for efficient purification of papillomavirus-based gene transfer vectors. Despite their obvious morphological differences, mature and immature capsids are similarly neutralizable by various L1-and L2-specific antibodies.Papillomaviruses, which are etiologically implicated in the development of warts, cervical cancer, and other epithelial tumors, replicate in the stratified squamous epithelium of the skin or mucous membranes (reviewed in reference 18). The papillomavirus life cycle is closely tied to the epithelial differentiation program, which takes many days to complete. The eventual release of the nonenveloped virion is thought to depend primarily on the normal cellular disintegration typically seen near the surface of squamous epithelia.The complexity of mimicking stratified epithelial differentiation in culture has made it challenging to study the papillomaviral life cycle in vitro. As a result, much of what is known about papillomavirus morphogenesis has been learned by studying recombinant versions of the two viral structural proteins, L1 and L2. The major capsid protein, L1, can spontaneously self-assemble into 72-pentamer virus-like particles (VLPs) that closely resemble native papillomavirus virions (20). The minor capsid protein, L2, is important for papillomavirus infectivity (38), but its arrangement in the virion and its role in encapsidation of the 8-kb double-stranded circular viral genomic DNA remain unclear.We have recently described a system for generating hightiter papillomavirus-based gene transfer vectors (also known as...

Multiparametric MR imaging (T2-weighted MR imaging, MR spectroscopy, dynamic contrast-enhanced MR imaging) of the prostate at 3 T enables tumor detection, with reasonable sensitivity and specificity values.

Purpose To determine the prostate cancer detection rate of multi-parametric (MP) MRI at 3T. Precise one to one histopathologic correlation with MRI was possible using prostate MRI based custom-printed specimen molds following radical prostatectomy. Materials and methods This IRB approved prospective study included forty-five patients (mean age 60.2 years, range 49–75 years) with a mean PSA of 6.37ng/mL (range 2.3–23.7ng/mL), who had biopsy proven prostate cancer (mean Gleason score of 6.7; range 6 to 9). Prior to prostatectomy, all patients underwent prostate MRI on a 3T scanner which included tri-plane T2 weighted MRI, apparent diffusion coefficient maps of diffusion weighted MRI, dynamic contrast enhanced MRI, and spectroscopy.. The prostate specimen was whole mount sectioned in the mold allowing geometric alignment to MRI. Tumors were mapped on MRI and histopathology.. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of MRI for cancer detection were calculated. Additionally, the effects of tumor size and Gleason score on sensitivity of MP-MRI were evaluated. Results PPV of MP-MRI to detect prostate cancer was 98%, 98%, and 100% in overall prostate, peripheral zone, and central gland, respectively. Sensitivities of MRI sequences were higher for tumors >5mm in diameter, as well as for tumors with higher Gleason scores (>7) (p<0.05). Conclusion Prostate MRI at 3T allows for the detection of prostate cancer. A multi-parametric approach increases the predictive power of MRI for diagnosis. In this study, accurate correlation between MP-MRI and histopathology was obtained by the patient specific MRI-based mold technique.

Purpose:To investigate whether apparent diffusion coefficients (ADCs) derived from diffusion-weighted (DW) magnetic resonance (MR) imaging at 3 T correlate with the clinical risk of prostate cancer in patients with tumors that are visible on MR images, with MR imaging/transrectal ultrasonography (US) fusion-guided biopsy as a reference. Materials and Methods:Forty-eight consecutive patients (median age, 60 years; median serum prostate-specifi c antigen value, 6.3 ng/mL) who underwent DW imaging during 3-T MR imaging with an endorectal coil were included in this retrospective institutional review board-approved study, and informed consent was obtained from each patient. Patients underwent targeted MR imaging/transrectal US fusion-guided prostate biopsy. Mean ADCs of cancerous target tumors were correlated with Gleason and D'Amico clinical risk scores. The true risk group rate and predictive value of the mean ADC for classifying a tumor by its D'Amico clinical risk score was determined by using linear discriminant and receiver operating characteristic analyses. Results:A signifi cant negative correlation was found between mean ADCs of tumors in the peripheral zone and their Gleason scores ( P = .003; Spearman r = 2 0.60) and D'Amico clinical risk scores ( P , .0001; Spearman r = 2 0.69). ADC was found to distinguish tumors in the peripheral zone with intermediate to high clinical risk from those with low clinical risk with a correct classifi cation rate of 0.73. Conclusion:There

The role of viral structural proteins in the initiation of adaptive immune responses is poorly understood. To address this issue, we focused on the effect of noninfectious papillomavirus-like particles (VLPs) on dendritic cell (DC) activation. We found that murine bone marrow-derived dendritic cells (BMDCs) effectively bound and rapidly internalized bovine papillomavirus VLPs. Exposure to fully assembled VLPs of bovine papillomavirus, human papillomavirus (HPV)16 or HPV18, but not to predominately disordered HPV16 capsomers, induced acute phenotypic maturation of BMDCs. Structurally similar polyomavirus VLPs bound to the DC surface and were internalized, but failed to induce maturation. DCs that had incorporated HPV16 VLPs produced proinflammatory cytokines IL-6 and TNF-α; however, the release of these cytokines was delayed relative to LPS activation. Production of IL-12p70 by VLP-exposed DCs required the addition of syngeneic T cells or rIFN-γ. Finally, BMDCs pulsed with HPV16 VLPs induced Th1-dominated primary T cell responses in vitro. Our data provide evidence that DCs respond to intact papillomavirus capsids and that they play a central role in VLP-induced immunity. These results offer a mechanistic explanation for the striking ability of papillomavirus VLP-based vaccines to induce potent T and B cell responses even in the absence of adjuvant.