Association between human leukocyte antigen polymorphism and human papillomavirus 16-positive vulval intraepithelial neoplasia in British women. (49/364)

Polymorphisms in human leukocyte antigen (HLA) genes have been implicated in the risk for developing human papillomavirus (HPV)-associated cervical neoplasia. By comparison with local cadaver controls typed for HLA class I (n = 946) and II (n = 144) antigens, HPV-16-positive high grade vulval intraepithelial neoplasia patients (n = 42) showed significantly different frequencies of HLA-A2 [odds ratio (OR), 2.1; confidence interval (CI), 1.4-3.9], HLA-B7 (OR, 2.6; CI, 1.4-4.7), HLA-DRB1*01(01/02/04) (OR, 0.1; CI, 0.03-0.5), HLA-DRB1*11 (OR, 3.3; CI, 1.4-7.1), HLA-DRB1*13 (OR, 0), HLA-DQB1*05 (OR, 0.2; CI, 0.05-0.6), and HLA-DQB1*03032 (OR, 4.6; CI, 1.5-14.0). With the exception of HLA-B7 and HLA-DRB1*11, these significant differences were also seen comparative to local HPV-16-positive cervical carcinoma patients (n = 114), suggesting a specific immunogenetic contribution that is independent of HPV-16 infection in high-grade vulval intraepithelial neoplasia. Such factors are important to the development of HPV vaccines for treatment of cervical and vulval neoplasia.  (+info)

Immunohistochemical analysis of p53 in vulval intraepithelial neoplasia and vulval squamous cell carcinoma. (50/364)

Human papillomavirus (HPV) is thought to cause some vulval squamous cell carcinomas (VSCC) by degrading p53 product. Evidence on whether HPV-negative VSCC results from p53 mutation is conflicting. We performed immunohistochemistry for p53 product on 52 cases of lone vulval intraepithelial neoplasia (VIN), 21 cases of VIN with concurrent VSCC and 67 cases of VSCC. We had previously performed HPV detection and loss of heterozygosity (LOH) analyses on these samples. Abnormal p53 immunoreactivity (p53-positive) rates in HPV-positive VSCC and HPV-negative VSCC were 22% (12/54) and 31% (4/13), respectively (P<0.74). p53 immunoreactivity was associated with LOH at the p53 locus (P<0.004), but neither technique differentiated between HPV-positive and HPV-negative VSCC. p53 immunoreactivity was associated with overall LOH rates (p53-positive VSCC vs p53-negative VSCC mean fractional regional allelic loss 0.41 vs 0.24, respectively, P<0.027). LOH at 3p25 was more frequent in p53-positive VSCC cf p53-negative VSCC (70 vs 21%, respectively, P<0.007). There was a trend in p53 disruption associated with invasive disease; HPV-positive VSCC demonstrated more disruption than VIN associated with VSCC, which had more disruption than lone VIN III (22 vs 10 vs 0%, respectively, P<0.005). In all, three out of 73 cases of VIN were p53-positive. All three were associated with concurrent or previous VSCC. Meta-analysis of previous studies revealed significantly more p53 disruption in HPV-negative VSCC cf HPV-positive VSCC (58 vs 33%, respectively; P<0.0001). p53 immunoreactivity/mutation in VIN only appeared in association with VSCC. These data suggest that HPV-independent vulval carcinogenesis does not exclusively require disruption of p53, p53 disruption may work synergistically with LOH at specific loci and p53-positive VIN should be checked carefully for the presence of occult invasion.  (+info)

Minichromosome maintenance (Mcm) proteins, cyclin B1 and D1, phosphohistone H3 and in situ DNA replication for functional analysis of vulval intraepithelial neoplasia. (51/364)

Vulval intraepithelial neoplasia (VIN) is defined histopathologically by distinctive abnormalities of cellular maturation and differentiation. To investigate the functional properties of VIN, the expression of several proteins involved in the regulation of the cell cycle as well as in situ DNA replication competence was analysed by immunohistochemistry. Snap-frozen vulval biopsies were graded as normal squamous epithelium (n=6), undifferentiated HPV positive VIN 1 (n=3), VIN 2 (n=8) and VIN 3 (n=20). Immunohistochemistry was performed using the following markers: cyclin D1 (expressed in middle/late G1), cyclin B1 (expressed in G2/early M), phosphorylated histone H3 (expressed during mitosis) and minichromosome maintenance (Mcm) proteins 2 and 5 (expressed during the cell cycle, but not in differentiated or quiescent cells). In situ DNA replication competence was used to identify S-phase cells. The percentage of positively stained nuclei in three representative microscopic fields was calculated per biopsy. In normal vulva, the expression of all markers was restricted to the proliferative compartment of the basal layer of the epithelium. In contrast in high-grade VIN, the majority of epithelial cells expressed the Mcm proteins from basal to superficial layer. The detection of cyclins B1 and D1, phospho-histone H3 and in situ DNA replication was also found through the full thickness of these lesions but by a lower proportion of the cells. This is consistent with these markers providing a series of 'snapshots' of the cell cycle status of individual cells. The low-grade VIN showed reduced expression of the cell cycle markers in relation to the level of dysplasia. The combination of these analyses establishes that the majority of VIN cells remain in a functional replicative or prereplicative state of the cell cycle. Clinical application of these analyses may provide a basis for improved diagnosis of VIN.  (+info)

Subcellular localization of CrmA: identification of a novel leucine-rich nuclear export signal conserved in anti-apoptotic serpins. (52/364)

The cowpox virus-encoded anti-apoptotic protein cytokine response modifier A (CrmA) is a member of the serpin family that specifically inhibits the cellular proteins caspase 1, caspase 8 and granzyme B. In this study, we have used Flag- and yellow fluorescent protein (YFP)-tagged versions of CrmA to investigate the mechanisms that regulate its subcellular localization. We show that CrmA can actively enter and exit the nucleus and we demonstrate the role of the nuclear export receptor CRM1 in this shuttling process. CrmA contains a novel leucine-rich nuclear export signal (NES) that is functionally conserved in the anti-apoptotic cellular serpin PI-9. Besides this leucine-rich export signal, additional sequences mapping to a 103-amino-acid region flanking the NES contribute to the CRM1-dependent nuclear export of CrmA. Although YFP-tagged CrmA is primarily located in the cytoplasm, shifting its localization to be predominantly nuclear by fusion of a heterologous nuclear localization signal did not impair its ability to prevent Fas-induced apoptosis. We propose that nucleocytoplasmic shuttling would allow CrmA to efficiently target cellular pro-apoptotic proteins not only in the cytoplasm, but also in the nucleus, and thus to carry out its anti-apoptotic function in both compartments.  (+info)

Pitting of agar surface by Pseudomonas stutzeri. (53/364)

A strain of Pseudomonas stutzeri which exhibited pitting of agar surfaces is described.  (+info)

Vulvar syringoma, report of a case and review of the literature. (54/364)

Syringomas are common intraepidermal sweat gland tumors most often found in women around the time of adolescence. Frequent sites of involvement include the lower eyelids and malar areas, however vulvar involvement is relatively rare. These lesions often present as small, multiple, skin-colored-to-yellowish papules and are often associated with increased vulvar discomfort and itching. We present a case of a 29-year old female who presented to her gynecologist complaining of vulvar itching and burning. A small condylomatous-type wart observed on her vulva was biopsied and found to be a syringoma. Because of their clinical presentation and associated symptoms, vulvar syringomas should be considered in the differential diagnosis of any multicentric papular lesion of the vulva, vulvar pain syndrome, and pruritus vulvae.  (+info)

Epidermal growth factor receptor (EGFR)-targeted immunoliposomes mediate specific and efficient drug delivery to EGFR- and EGFRvIII-overexpressing tumor cells. (55/364)

We hypothesized that immunoliposomes (ILs) that target epidermal growth factor receptor (EGFR) and/or its truncated variant EGFRvIII can be constructed to provide efficient intracellular drug delivery in tumor cells overexpressing these receptors. Monoclonal antibody fragments included Fab' fragments derived from C225, which binds both EGFR and EGFRvIII, or novel anti-EGFR scFv C10, which binds EGFR only. Monoclonal antibody fragments were covalently linked to liposomes containing various reporters or drugs. ILs were evaluated for specific binding, internalization, and cytotoxicity in EGFR/EGFRvIII-overexpressing cell lines in vitro. Flow cytometry and fluorescence microscopy showed that EGFR-targeted ILs, but not nontargeted liposomes or irrelevant ILs, were efficiently bound and internalized by EGFR-overexpressing cells, including glioma cells (U-87), carcinoma cells (A-431 and MDA-MB-468), and EGFRvIII stable transfectants (NR-6M). Furthermore, EGFR-targeted ILs did not bind to non-EGFR-overexpressing cells (MCF-7 and parental NR-6). ILs showed 3 orders of magnitude greater accumulation in NR-6-EGFRvIII stable transfectants versus parental NR-6 cells. Quantitative internalization studies indicated binding of EGFR-targeted ILs to target cells within 5 min, followed by intracellular accumulation beginning at 15 min; total uptake reached approximately 13,000 ILs/cell. ILs were used to deliver cytotoxic drugs doxorubicin, vinorelbine, or methotrexate to EGFR/EGFRvIII-overexpressing target cells in vitro. In each case, the IL agent was significantly more cytotoxic than the corresponding nontargeted liposomal drug in target cells, whereas it was equivalent in cells lacking EGFR/EGFRvIII overexpression. We conclude that EGFR-targeted ILs provide efficient and targeted delivery of anticancer drugs in cells overexpressing EGFR or EGFRvIII.  (+info)

Selective delivery of CB300638, a cyclopenta[g]quinazoline-based thymidylate synthase inhibitor into human tumor cell lines overexpressing the alpha-isoform of the folate receptor. (56/364)

The alpha-isoform of the glycosylphosphatidylinositol cell membrane tethered folate receptor (alpha-FR) is overexpressed in some carcinomas (notably ovarian carcinomas) relative to normal tissues. The nonpolyglutamatable folate-based thymidylate synthase (TS) inhibitor, CB300638 (TS K(i) = 0.24 nM) displayed an IC(50) of 0.0028 microM for the inhibition of the growth of human A431-FBP cells transfected with the alpha-FR. In contrast, the IC(50) for the neotransfected A431 cells was 0.81 microM (300-fold higher). Similarly, this compound inhibited the growth of human KB cells that constitutively overexpress the alpha-FR with an IC(50) of 0.0036 microM. These data were derived from cells grown in a physiological concentration of folate (20 nM R,S-leucovorin). Incubation of KB cells with a 1 microM excess of folic acid (FA), to selectively block uptake via the alpha-FR, increased the CB300638 IC(50) to 0.39 microM. The relatively low potency of CB300638 under these conditions, or in cell lines not expressing the alpha-FR, is ascribed to its low affinity for the ubiquitously expressed folate transporter, the reduced-folate carrier (K(i) for inhibition of [(3)H]methotrexate transport >100 microM). The high potency of CB300638 in alpha-FR-overexpressing cell lines is attributable to high affinity of the alpha-FR (53% of FA) and efficient endosomal trafficking mediated by the alpha-FR. Sixteen-h exposure to CB300638 inhibited the rate of (3)H(2)O release from 5-[(3)H]dUrd (in situ TS assay) in A431, A431-FBP, and KB cells with IC(50) values of 0.1 microM, 0.005 microM, and 0.002 microM, respectively. The coaddition of 1 micro M FA increased the IC(50)s for A431-FBP and KB cells to approximately 0.1 microM consistent with alpha-FR-mediated transport of CB300638. In conclusion, alpha-FR-mediated uptake of CB300638 leads to TS and growth inhibition that is highly selective for alpha-FR overexpressing tumor cell lines. The low expression of the alpha-FR in normal tissues, particularly those sensitive to TS inhibitors, together with the low affinity of CB300638 for the reduced-folate carrier, suggests that the compound may have potential as an antitumor agent with a high therapeutic index.  (+info)