Bioinformatics analysis of the role of CXC ligands in the microenvironment of head and neck tumor.

Chemokines play a significant role in cancer. CXC-motif chemokine ligands (CXCLs) are associated with the tumorigenesis and progression of head and neck squamous cell carcinoma (HNSC); however, their specific functions in the tumor microenvironment remain unclear. Here, we analyzed the molecular networks and transcriptional data of HNSC patients from the Oncomine, GEPIA, String, cBioPortal, Metascape, TISCH, and TIMER databases. To verify immune functions of CXCLs, their expression was analyzed in different immune cell types. To our knowledge, this is the first report on the correlation between CXCL9-12 and 14 expression and advanced tumor stage. CXCL2, 3, 8, 10, 13, and 16 were remarkably related to tumor immunity. Kaplan-Meier and TIMER survival analyses revealed that high expression of CXCL1, 2, 4, and 6-8 is correlated with low survival in HNSC patients, whereas high expression of CXCL9, 10, 13, 14, and 17 predicts high survival. Only CXCL13 and 14 were associated with overall survival in human papilloma virus (HPV)-negative patients. Single-cell datasets confirmed that CXCLs are associated with HNSC-related immune cells. Thus, CXCL1-6, 8-10, 12-14, and 17 could be prognostic targets for HNSC, and CXCL13 and 14 could be novel biomarkers of HPV-negative HNSC.

Undetected human papillomavirus DNA and uterine cervical carcinoma: Association with cancer recurrence.

BACKGROUND: The time course of human papillomavirus (HPV) DNA clearance was studied in patients with carcinoma of the cervix during follow-up after primary radical radiotherapy (RT). This study investigated the relationship between timing of HPV clearance and RT effectiveness. PATIENTS AND METHODS: A total of 71 consecutive patients who were treated for cervical cancer with primary radical radiotherapy and high-dose rate intracavitary brachytherapy with or without chemotherapy were enrolled in the study. Samples for HPV DNA examination were taken before (1) treatment, (2) every brachytherapy, and (3) every follow-up examination. The times when HPV DNA was undetected were analyzed for association with recurrence-free survival. RESULTS: HPV DNA was not detected in 13 patients (18 %) before RT. Of the 58 patients with HPV DNA detected before treatment, HPV DNA was not detected in 34 % during treatment and in 66 % after the treatment. Within 6 months after RT, HPV DNA was detected in 0 % of all patients. The patients were followed up for a median period of 43 months (range 7-70 months). In all, 20 patients were found to develop recurrence. The 3-year cumulative disease-free survival (DFS) rate was 71 ・} 5.4 % for all 71 patients. In multivariate analysis, DFS was significantly associated with HPV (detected vs. not detected) with a hazard ratio of 0.07 (95 % confidence interval 0.008-0.6, p = 0.009). CONCLUSION: In this study, patients in whom HPV was not detected had the worst prognosis. Six months after RT, HPV DNA was detected in 0 % of the patients. Patients in whom HPV DNA could not be detected before treatment need careful follow-up for recurrence and may be considered for additional, or alternative treatment.

Prevalence of HPV-DNA in Pap smears containing ASC and AGC performed within Population Programme of Prophylaxis and Early Detection of Early Cervical Cancer.

AIM: to assess the incidence of HPV -DNA in women with ASC/AGC compared to patients with normal Pap smears. MATERIAL AND METHODS: The study group consisted of 242 women (207 ASC and 35 AGC cases). The control group counted 200 age-matched women with negative Pap smears. Cervical samples collected from all the participants were tested for the presence of HPV-DNA using the Hybrid Capture-2 test. RESULTS: Total HPV infection was significantly higher in the study than in the control group (43.0% vs.14.0%) (p=0.005). There was no difference in the incidence of HPV -DNA between ASC and AGC groups. Prevalence of HPV-DNA ASC-H was significantly higher in ASC-US group (83.3% vs. 40.5%) (p=0.004). HPV positive endometrial AGC significantly outnumbered HPV positive endocervical AGC (88.9% vs. 26.9%) (p=0.003). Similar trends were observed for the high-risk type of HPV (p<0.001). CONCLUSIONS: The significant difference in HPV -DNA incidence between the study and control groups suggests that HPV plays a role in the development of ASC and AGC. The implementation of HPV testing in all women diagnosed with ASC or AGC can lead to tailored therapeutic management and more careful follow-up care.

Infecciones vaginales y lesiones celulares cervicales(I). Programa de cribado de otras infecciones y ETS simultáneas / Vaginal infections and cervical cell abnormalities (I). Simultaneous screening for sexually-transmitted diseases and other infections

Objetivo: Programa de cribado de otras infecciones y ETS simultáneas en un grupo de mujeres afectas de infecciones vaginales y lesiones celulares cervicales diagnosticadas por la citología cervicovaginal, durante 1 año. Material y métodos A 399 mujeres con infección vaginal, por C (95), vaginosis bacteriana (234), Gardnerella vaginalis (14), C + vaginosis bacteriana (9), Tricomonas (9), vaginosis bacteriana + Tricomonas (1), virus del papiloma humano (VPH, 16), antecedentes de VPH (A-VPH, 21); y 32 con lesiones celulares cervicales: 7 con ASCUS, 1 con AGUS, 22 con LSIL, y 2 con HSIL; se les practica analítica general con velocidad de sedimentación globular, proteína C reactiva, leucocitos y linfocitos, serología con HBsAg, Anticuerpos para VHC, VIH y VHS-2, y RPR, reagina rápida en plasma para sífilis, y prueba de ADN-VPH (Digene VPH), para ver VPH de alto y bajo riesgo (AR y BR).Resultados Eran extranjeras 131 casos (30,39%). Las infecciones más frecuentes fueron: vaginosis bacteriana, 234 casos (58,64%) y C, 95 casos (23,80). VPH, A-VPH y lesiones celulares, se dan en las más jóvenes, y tienen menos gestaciones y partos, y hay más nuligestas. Vaginosis bacteriana y C se dan en (..) (AU)

Objective: To evaluate a screening program for simultaneous detection of sexually-transmitted diseases and other infections in women with vaginal infections and cervical cell abnormalities diagnosed by cervicovaginal cytology. Material and methods: There were 399 women with vaginal infections due to the following: Candida (95 cases), bacterial vaginosis (234 cases), Gardnerella vaginalis (14 cases), Candidaplus bacterial vaginosis (9 cases), Trichomonas vaginalis (9 cases), bacterial vaginosis plus Trichomonas(1 case), human papillomavirus (HPV) (16 cases), and a history of HPV infection (21cases). Thirty-two women had cervical cell abnormalities: atypical squamous cells of undetermined significance (ASCUS) (7 cases), atypical glandular cells of undetermined significance (AGUS)(1 case), low-grade squamous intraepithelial lesion (LSIL) (22 cases) and high-grade squamous intraepithelial lesion (HSIL) (2 cases). Analysis included erythrocyte sedimentation rate (ESR),C reactive protein (CRP), leukocyte count and lymphocyte count; HBsAg serology, hepatitis C virus (HCV) antibodies, HIV antibodies, herpes simplex virus (HSV)-2 antibodies, rapid plasmareagin (RPR), and DNA-HPV probe (Digene HPV) for high-risk (HR) and low-risk (LR) HPV. Results: A total of 131 women (30.39%) were foreigners. The most frequent infections were (AU)

Distribution of human papillomavirus genotypes in the patients with cervical carcinoma and its precursors in Zhejiang Province, China.

Various evidences reveal that the human papillomavirus (HPV) is the single most important etiologic agent in cervical carcinoma (CC). To investigate the distribution of HPV genotypes in the patients with CC and its precursors in Zhejiang Province, China, a total of 631 eligible samples from patients in Zhejiang Province with CC (N = 181), cervical intraepithelial neoplasia (CIN) II-III (N = 345), and CIN I (N = 105) were detected. Age-matched samples of 217 women without cervical neoplasia were detected as control. An improved polymerase chain reaction (PCR)-restriction fragment length polymorphism assay validated by Hybrid Capture II and PCR sequencing was designed for HPV genotype. The prevalence of HPV was 95.0% in CC, 88.4% in CIN II-III, and 73.3% in CIN I, while only 41.9% in control. High-risk/low-risk HPV ratio showed a significant trend of increase with increased grade of CIN and transformation to carcinoma. A total of 24 HPV genotypes were detected in CC and its precursors. Of those, HPV 16 (65.2%), 18 (9.4%), and 58 (9.4%) were the major HPV genotypes in CC, while HPV 16 (37.6%), 58 (19.1%), 33 (10.6%), and 18 (5.4%) in CIN. In conclusion, the distribution of predominant HPV genotypes in the patients with CC and its precursors in Zhejiang Province is HPV 16, 58, and 18, probably as well as 33, which may be high-risk factors for CC.

Rising incidence of oropharyngeal cancer and the role of oncogenic human papilloma virus.

OBJECTIVES/HYPOTHESIS: To document the increasing incidence of oropharyngeal (OP) cancer and to provide evidence that this increase is caused by oncogenic human papilloma virus (HPV). STUDY DESIGN: Epidemiologic review and retrospective case series analysis. METHODS: We collected data from Colorado and the United States comparing the average annual age-adjusted incidence rates of OP and non-OP head and neck cancer between the periods 1980 to 1990 and 1991 to 2001. We obtained data on 72 patients with OP cancer from a single county in Colorado, from 1980 through 2004. HPV status was determined by DNA-polymerase chain reaction. We assessed disease-specific survival. RESULTS: The average annual age-adjusted incidence of OP cancer in males in Colorado increased from 2.54 per 100,000 to 3.47 (P < .05) or 36.6%, whereas the U.S. rate increased from 4.34 to 4.81 (P < .05) or 10.8%. The rates in females and the rates of non-OP head and neck cancer decreased. Of the 72 cases, 50 (69%) were positive for HPV subtype 16. The ratio of HPV-positive to HPV-negative cases prior to 1995 was 0.72 (8:11) but was 3.81 (42:11) afterward. Survival was positively affected by HPV status (hazard ratio of 0.15, confidence intervals 0.07-0.36, P < .001). Disease-specific survival was 83% in the HPV-positive patients and 15% in the HPV-negative group. CONCLUSIONS: OP cancer incidence is increasing in Colorado males and to a lesser extent in U.S. males. The HPV-positive OP cancer cases were more frequent in the later years of the study. Disease-specific survival was much better in the HPV-positive patients, confirming that HPV testing defines a unique subset of patients. These findings suggest that HPV oncogenesis accounts for the increase in average annual age-adjusted incidence of OP cancer.

The role of HPV DNA in the evaluation and follow-up of asymptomatic male sexual partners of females with CIN3.

PURPOSE: To determine whether and how asymptomatic sexual partners of females with high-grade cervical intraepithelial neoplasia (CIN3) lesions should be examined. METHODS: Sexual partners of females with CIN3 were evaluated for HPV-related lesions by scraping samples for high-risk HPV DNA and androscopy (colposcopic inspection of the penis, scrotum and peri-anal area). Abnormal androscopically detected lesions were sampled for cytology by Pap smears. RESULTS: 74 partners of 87 females were studied and underwent androscopy, and 17 (22.9%) had abnormal findings: 11/74 had clinical genital condyloma acuminata and 6/74 had aceto-white lesions on the penile shaft or scrotum. Cytology of the 17 abnormal androscopies showed that six smears were normal and 11 had atypia and koilocytosis. Positive high-risk HPV DNA indicated that 13/74 (17.5 %) were infected with HPV. Two partners (2/74, 2.8%) had concomitant HPV DNA 16. CONCLUSIONS: Male sexual partners of females with CIN3 should undergo androscopy and cytology of colposcopically detected abnormal areas.

Human papillomavirus detection in self-collected vaginal specimens and matched clinician-collected cervical specimens.

Human papillomavirus (HPV) detection is an integral part of cervical cancer screening, and a range of specimen collection procedures are being tested. Preliminary studies have found that the majority of women prefer self-collection of vaginal specimens instead of clinician-collected specimens of the cervix. The purposes of the current study were to explore the social and behavioral predictors of acceptance of self-collection of vaginal specimens among patients and to assess concordance in detection of HPV between clinician-collected cervical specimens and self-collected vaginal specimens. The study was conducted at a university family medicine clinic using a cross-sectional study design, and enrollment of women presenting for routine gynecological examination consecutively in a period of 1 year, self-administered questionnaires, collection of paired vaginal and cervical specimens for HPV DNA using Hybrid Capture 2, and cytologic analysis. Most women (79.8% [398/499]) agreed to collect vaginal specimens. In our study, 76.6% (216/282) African American women (AA), 88.1% (156/176) white non-Hispanic (WNH) women, and 63.4% (26/41) women of other races (P < 0.0001) agreed to self-collect vaginal specimens. HPV was detected in 16.0% (80/499) of clinician-collected cervical specimens and 26.1% (104/398) of self-collected vaginal specimens (P < 0.001). HPV detection was concordant in 13.4% (53/398) women in both cervical and vaginal specimens. Self-collection of vaginal specimens for HPV DNA detection is acceptable to most women presenting for routine gynecological examination. WNH women were more likely to obtain self-collected specimens than AA women. Vaginal specimens were more likely to be positive for HPV than were cervical specimens.

Differentially expressed genes between high-risk human papillomavirus types in human cervical cancer cells.

Cervical carcinoma (CC) is one of the most common cancers among women worldwide and the first cause of death among the Mexican female population. Human papillomavirus (HPV) infection is the most important etiologic factor for CC. Of the oncogenic types, HPV16 and HPV18 are found in 60-70% of invasive CCs worldwide. HPV18 appears to be associated with a more aggressive form of cervical neoplasia than HPV16 infection. At present, there are no studies on differentially expressed cellular genes between transformed cells harboring HPV16 and HPV18 sequences. Based on previous complementary DNA microarray data from our group, 13 genes were found to be differentially overexpressed between HPV16- and HPV18-transformed cells. These genes were as follows: E6BP, UBE4A, C20orf14, ATF7, ABCC8, SLC6A12, WASF3, SUV39H1, SPAG8, CCNC, E2FFE, BIRC5, and DEDD. Differential expression of six selected genes was confirmed by real-time reverse transcription-polymerase chain reaction (RT-PCR). All real-time RT-PCRs confirmed differential expression between HPV18 and HPV(-) samples. The present work identifies genes from signaling pathways triggered by HPV transformation that could be differentially deregulated between HPV16(+) and HPV18(+) samples.

[Analysis of correlation between HPV DNA lymph node presence and pathologic parameters of primary tumour in cervical carcinoma patients treated surgically]. / Ocena korelacji obecnosci DNA HPV w wezlach chlonnych chorych na raka szyjki macicy z parametrami histopatologicznymi w zmianie pierwotnej.

Young-age sexual activity, multiple sexual partners, number of pregnancies, smoking and human papilliomavirus (HPV) infections are factors commonly considered as promoters of cervical tumorogenesis. Cervical carcinoma patients' prognosis correlates significantly with staging (according to FIGO classification). In early stages lymph-node involvement is considered as the most important prognostic factor. Other factors are: initial volume of the lesion, lymphatic and blood vessels involvement, depth of cervical invasion. Also parametrium involvement seems to decrease the rate of 5-year survival. The aim of the research was to estimate correlation between HPV DNA lymph node presence and pathologic parameters of primary tumour in surgically treated cervical carcinoma patients. We analysed DNA isolated from lymph nodes sampled intra-operatively from 134 patients, surgically treated for cervical cancer with PCR. We confirmed the presence of HPV DNA sequences in lymph nodes in 81 (60.5%) cases. Lymph node presence of HPV DNA rate was significantly higher in: advanced tumours, cases with initial lesion volume exceeding 10 cm3, cases with uterus and vaginal involvement and with initial tumour invasion deeper than 10 mm.

Implications regarding atypical squamous cells of undetermined significance among women residing in a US-Mexico border city.

We conducted a study of Mexican American women living in a US-Mexico border city who attended a gynecology clinic for Papanicolaou (Pap) smear. The objective of this study was to describe the cytologic outcomes of women who had atypical squamous cells of undetermined significance (ASCUS) diagnosis after a Pap smear and to observe any changes during follow-up colposcopy. A total of 852 abnormal Pap smear were identified through a computer search for a 6-month period. Histology data were available for 317 cases. Benign findings were observed in 45.4% of cervical biopsies. A clinically significant diagnosis was reported in the remaining tissue sample. The diagnosis report was either single or combined and recorded as follows: human papilloma virus 46.3%, cervical intraepithelial neoplasia (CIN) 1, 23.6%; CIN 2, 5.6%; and CIN 3, 1.5%. There was one case of invasive cervical cancer. Overall, the incidence rate of ASCUS was 5%. However, we found that a significant proportion of this population had CIN 1 through CIN 3. Furthermore, this population has traditionally been noncompliant and routinely failed to attend follow-up appointments. Based on these results, the clinician should not ignore an initial abnormal Pap smear. Therefore, it is not unreasonable to perform colposcopy in Mexican American patients with a first time diagnosis of ASCUS on routine Pap smear.

Human papillomavirus genotype prevalence in cervical biopsies from women diagnosed with cervical intraepithelial neoplasia or cervical cancer in Melbourne, Australia.

Multicenter international phase III clinical trials using multivalent human papillomavirus (HPV) vaccines for cervical cancer (CC) prevention are underway. As HPV immunity is type specific, defining HPV genotype prevalence in different regions to ascertain whether predominant types differ geographically is considerably important prior to vaccine implementation. This study aimed to define HPV genotypes present in CC and high-grade dysplasia among women in Melbourne, Australia. HPV genotype analysis of a cross section of women in Melbourne with cervical dysplasia/cancer was performed. A total of 493 cervical biopsies from patients being treated for moderate (n= 122) or severe (n= 180) cervical intraepithelial neoplasia (CIN II/III) or CC (n= 191) were tested for HPV genotypes using the PGMY09/11 primer system and line blot assay. HPV detection rates were 63.9%, 72.8%, and 86.9% in CIN II, CIN III, and CC biopsies, respectively. The most prevalent HPV genotypes among CC biopsies were HPV-16 (52.9%), HPV-18 (18.3%), HPV-45 (6.3%), HPV-39 (3.1%), and HPV-73 (2.6%). Multiple HPV infections, comprising two to five types, were identified in 14.4% of biopsies, being significantly fewer (5.2%) among CC biopsies (P < 0.0001). These results indicate that the two most prevalent CC-associated HPV genotypes in Australia parallel those described internationally, with type variations thereafter.

Serologic response to human papillomavirus 16 among Australian women with high-grade cervical intraepithelial neoplasia.

This study evaluated the detection of human papillomavirus (HPV) 16 antibody in HPV 16-associated cervical intraepithelial neoplasia (CIN) in Australian women. Seroreactivity to HPV 16 L1 virus-like particles was assessed in patients with CIN 2 (n= 169) and CIN 3 (n= 229) lesions previously tested for the presence of HPV DNA. Seropositivity was significantly commoner in women with HPV 16 DNA-positive lesions (98/184) than in women with no HPV DNA in the lesion (15/47) or with HPV of types other than 16 in the lesion (43/167) (P= 0.0004). In addition, seropositivity was observed in 33% (55/169) of women with CIN 2 and 46% (106/229) of women with CIN 3, in keeping with the lower fraction of CIN 2 (57/169) than CIN 3 (127/229) biopsies positive for HPV 16 DNA. HPV 16 seropositivity is most common in women with HPV 16-associated CIN, but many patients with HPV-associated CIN 3 are seronegative, and HPV 16 seropositivity is common in women with CIN associated with other HPV types. Overall, HPV 16 serology is a poor predictor of presence of HPV 16-associated CIN 3 in patient population studied.

Markers of cutaneous human papillomavirus infection in individuals with tumor-free skin, actinic keratoses, and squamous cell carcinoma.

Separately, actinic keratosis (AK) and cutaneous squamous cell carcinoma (SCC) have been associated with cutaneous human papillomavirus (HPV) infections. To further explore the association between HPV infection and SCC development, we determined markers of cutaneous HPV infection within a single population in persons with precursor lesions (AK), cancerous lesions (SCC), and without. Serum and plucked eyebrow hairs were collected from 57 tumor-free controls, 126 AK, and 64 SCC cases. Presence of HPV L1 and E6 seroreactivity and viral DNA were determined for HPV types 5, 8, 15, 16, 20, 24, and 38. Significant positive associations with increasing severity of the lesions (controls, AK, and SCC, respectively) were observed for overall HPV L1 seropositivity (13%, 26%, and 37%) and for HPV8 (4%, 17%, and 30%). In parallel, the proportion of L1 seropositive individuals against multiple HPV types increased from 14% to 39% and 45%. The overall E6 seroreactivity, however, tended to decline with AK and SCC, especially for HPV8 (21%, 11%, and 2%). HPV DNA positivity was most prevalent in the AK cases (54%) compared with the SCC cases (44%) and the tumor-free controls (40%). Among all participants, there was a positive trend between overall HPV DNA positivity and L1 seropositivity, but not E6 seropositivity. Taken together, our data suggest that cutaneous HPV infections accompanied by detectable HPV DNA in eyebrow hairs and HPV L1 seropositivity, but not E6 seropositivity, are associated with an increased risk of AK and SCC.

Probe classification of on-off type DNA microarray images with a nonlinear matching measure.

We propose a nonlinear matching measure, called counting measure, as a signal detection measure that is defined as the number of on pixels in the spot area. It is applied to classify probes for an on-off type DNA microarray, where each probe spot is classified as hybridized or not. The counting measure also incorporates the maximum response search method, where the expected signal is obtained by taking the maximum among the measured responses of the various positions and sizes of the spot template. The counting measure was compared to existing signal detection measures such as the normalized covariance and the median for 2390 patient samples tested on the human papillomavirus (HPV) DNA chip. The counting measure performed the best regardless of whether or not the maximum response search method was used. The experimental results showed that the counting measure combined with the positional search was the most preferable.

Use of interferon-alpha in recurrent respiratory papillomatosis: 20-year follow-up.

OBJECTIVES: The aim of this study was analysis of the results of use of interferon-alpha (IFN-alpha) in patients with recurrent respiratory papillomatosis (RRP) and correlation of the results with human papillomavirus (HPV) type. METHODS: A multicenter prospective series (42 patients from 22 hospitals) yielded 20 years of follow-up of patients with RRP and HPV typing who were treated with IFN-alpha in doses of 3 MU/m2 3 times per week. RESULTS: During long-term follow-up (mean +/- SD, 172 +/- 36.8 months), the rate of event-free survival evaluated by Kaplan-Meier analysis was 42.8%, and the overall survival rate was 82.6%. The HPV typing revealed an association of HPV 11 with a more aggressive disease course (64% of HPV 11 patients versus 24% of HPV 6 patients), a lower incidence of long-term response to IFN-alpha therapy (14% of HPV 11 patients versus 64% of HPV 6 patients), and a higher incidence of malignant transformation and mortality during follow-up (36% and 24%, respectively, of HPV 11 patients versus 0% of HPV 6 patients). CONCLUSIONS: The obtained results revealed maximal effectiveness of IFN-alpha therapy in RRP patients with HPV 6 as compared with HPV 11. The association of HPV 11 with a worse long-term response to IFN-alpha therapy and a higher incidence of malignant transformation and mortality is clinically important and indicates the necessity of HPV typing in RRP patients after the first biopsy.

[Correlation of hybrid II capture cytologic exam in diagnosis of cervical lesions related to HPV]. / Importância da correlação do exame citológico com a captura híbrida II no diagnóstico de lesões intraepiteliais cervicais relacionadas ao HPV1.

PURPOSE: The aim of the present study was confront the results of the cytological examination with hybrid capture II in the diagnosis of induced cervical intraepithelial lesion-HPV, correlating the cytological findings with biomoleculares. METHODS: The research was carried through in a group of 160 sexually active women who had espontaneamente looked its gynecologists for consultation of routine, having been submitted to the collection of cervicovaginal material for cytology and for examination of hybrid capture II in the Centro de Patologia Clínica and the Hospital e Maternidade Promater, in the city of the Natal-RN. RESULTS: The results had shown to relatively high numbers of positive cases for HPV using hybrid capture II (41.87%) and the cytology (23.75%). The agreement between the two studied methods relatively was raised (59.38%). It was evident also that the viruses with high oncogênico potential had presented found in the compatible cytology with Lesion of low risk (11.88%), followed of Lesion of high risk (NIC II and III); already the viruses with low oncogênico potential were more associates the Lesion of low risk (6.25%), followed of Lesion of high risk. CONCLUSION: The cytology, exactly with its limitations, is an important method in the detention of attributable patologias to the HPV, emphasizing that the molecular method comes to complement it and to consolidate the cytological findings.

Human papillomavirus, Epstein-Barr virus and p53 mutation in vulvar intraepithelial neoplasia.

OBJECTIVE: To clarify the role of human papillomavirus (HPV) and Epstein-Barr virus (EBV) infection in vulvar carcinogenesis in relation to the mutated p53 gene. STUDY DESIGN: Polymerase chain reaction (PCR) was used to amplify DNA sequences of the viruses and PCR-single-strand conformation polymorphism analysis to screen for p53 gene mutations in exons 5-8 from formalin-fixed, paraffin-embedded blocks including 10 undifferentiated vulvar intraepithelial neoplasia (VIN) specimens. RESULTS: HPV and EBV DNA was found in 75% (6/8) and 0% (0/10) of VIN tissues, respectively. Oncogenic HPV 16 was the predominant type. HPV DNA extraction was not possible in 2 VIN specimens. p53 Gene mutation was shown in 20% (2/10) of VIN lesions. No correlation was found between p53 gene mutation the presence of viral HPV or EBV DNA. Mutated p53 was equally distributed between HPV-positive and -negative VIN cases. CONCLUSION: Our results suggest that although most undifferentiated VIN lesions are associated with HPV infection, p53 mutations may occur independent of viral infection even in the presence of oncogenic HPV. HPV, but not EBV or p53 gene mutation, can play a role in the pathogenesis of undifferentiated VIN.

Antisense activity detection by inhibition of fluorescence resonance energy transfer.

Use of antisense nucleic acids to modulate expression of particular genes is a promising approach to the therapy of human papillomavirus type 16 (HPV-16)-associated cervical cancer. Understandably, evaluation of the in vivo performance of synthetic antisense oligodeoxynucleotides (AS-ODNs) or ribozymes is of ultimate importance to development of effective antisense tools. Here we report the use of a bacterial reporter system based on the inhibition of fluorescence resonance energy transfer (FRET) to measure the interaction of AS-ODNs with HPV-16 target nt 410-445, using variants of the green fluorescent protein (GFP). An optimal FRET-producing pair was selected with GFP as the donor and yellow fluorescent protein (YFP) as the acceptor molecule. Hybridization of AS-ODNs with a chimaeric mRNA containing the antisense target site flanked by GFP variants resulted in the inhibition of the FRET effect. Use of different linkers suggested that the amino acid content of the linker has no significant effect on FRET effect. Antisense accessibility, tested by RNaseH assays with phosphorothioated target-specific and mutant AS-ODNs, suggested a specific effect on the chimaeric mRNA. FRET inhibition measurements correlated with the presence of truncated proteins confirming true antisense activity over the target. Therefore, FRET inhibition may be used for the direct measurement of AS-ODNs activity in vivo.