Steven P. Tjong-A-Hung

Telomerase activity as a biomarker for (pre)neoplastic cervical disease in scrapings and frozen sections from patients with abnormal cervical smear.

PURPOSE To evaluate the diagnostic value of semi-quantitative telomerase activity assessment in cervical scrapings together with human papillomavirus (HPV) typing for detection of (pre)neoplastic cervical lesions and to compare telomerase activity in cervical scrapings and frozen specimens from the same patients. PATIENTS AND METHODS A cross-sectional study was performed in 161 patients referred for an abnormal cervical cytology report. In cervical scrapings, telomerase activity was determined by modified telomere repeat amplification protocol (TRAP) assay and HPV typing by polymerase chain reaction (PCR) with general and type-specific primers. Final diagnosis was made by pathologic examination of biopsy and/or loop excision specimens. RESULTS Telomerase activity was detectable in assessable scrapings from one of nine (11%) patients without cervical intraepitheleal neoplasia (CIN), in three of 26 (12%) with CIN I, eight of 35 (22%) with CIN II, 18 of 62 (29%) with CIN III, and four of 13 (31%) with cancer. Sensitivity and negative predictive value of the TRAP assay for CIN II/III and cancer lesions were 25% and 28%, respectively, while specificity for no CIN or CIN I was 89%. In representative frozen sections, frequency of detectable telomerase activity was related to grade of CIN/cancer; none of 21 normal cervices, none of two CIN I, two of 12 (17%) CIN II, 10 of 31 (32%) CIN III, and 18 of 21 (86%) cervical cancer lesions were telomerase-positive (P < .0005). Telomerase activity levels in paired scrapings and frozen sections appeared to be only weakly related; telomerase-positive sections with negative scrapings and vice versa (only in CIN III) were observed. In oncogenic HPV-negative scrapings (n = 14), no telomerase activity was detected, but in frozen sections, telomerase activity levels appeared to be unrelated to presence of specific HPV types. CONCLUSION Telomerase activity is more frequent in higher grade CIN/cervical cancer lesions. Telomerase activity assessment in cervical scrapings has a low sensitivity for CIN II/III and/or cervical cancer and does not appear to be useful in primary screening for cervical cancer. However, increased telomerase activity in frozen CIN sections may be a possible marker of progressive disease.

Detection and typing of human papillomaviruses present in fixed and stained archival cervical smears by a consensus polymerase chain reaction and direct sequence analysis allow the identification of a broad spectrum of human papillomavirus types

DNA well suited for polymerase chain reaction (PCR) amplification was purified from archival Papanicolaou smears. The detection of a wide range of human papillomavirus (HPV) types was made possible using a HPV-specific consensus primer pair, and typing was conveniently done by direct sequence analysis of the PCR product. The method could be of unique value in longitudinal and cross-sectional studies aimed at answering a number of fundamental pathological and epidemiological questions regarding HPV infection of the genital tract.

Application of the NASBA nucleic acid amplification method for the detection of human papillomavirus type 16 E6-E7 transcripts

Using a human papillomavirus type 16 (HPV-16) E6-E7 specific primer set in a nucleic acid sequence-based amplification (NASBA) reaction, detection of HPV-16 transcripts was accomplished in a single enzymatic reaction at 41 degrees C. The NASBA reaction product was visualized either by Northern bolt analysis with an HPV-16 E6-E7-specific 32P-labelled oligonucleotide probe or by a non-radioactive enzyme-linked gel assay (ELGA). In combination with a rapid nucleic acid extraction procedure this method appears to be very suitable for the sensitive and specific detection of HPV-16 transcripts on small amounts of HPV-16-expressing cells of various sources, including cervical smears.

Human papillomavirus DNA after treatment of cervical dysplasia: Low prevalence in normal cytologic smears

The presence of human papillomavirus (HPV) DNA in relation to cervical cytology was evaluated after treatment of cervical dysplasia.

Cutaneous squamous cell carcinoma and p53 codon 72 polymorphism: a need for screening?

The association between human papillomavirus (HPV)–associated cervical cancer and cutaneous squamous cell carcinoma and codon 72 polymorphism in the p53 gene is not unequivocal. Especially, it is not known whether carriers of the arginine form have an increased risk of cancer that necessitates screening. The alternative is that the polymorphism is a tumor marker instead of a risk factor. We set out a case‐control study to determine the risk of squamous cell carcinoma of the skin in individuals with the p53 codon 72 arginine genotype in order to establish the possible need for screening. The distribution of the different p53 codon 72 genotypes was examined in 86 subjects with a history of cutaneous squamous cell carcinoma and in 168 controls. Additionally, 121 subjects who had had histologically proven basal cell carcinoma and 108 subjects who had had non‐familial malignant melanoma were tested. p53 polymorphism was evaluated by polymerase chain reaction (PCR) using DNA samples from peripheral blood lymphocytes. In a subgroup of patients with squamous cell carcinoma and controls, the presence of epidermodyplasia verruciformis human papillomavirus (EV‐HPV) DNA was determined in plucked eyebrow hair. Differences in the distributions of the genotypes among cases and controls were calculated, and univariate and multivariate analyses were performed to assess the risk to develop cutaneous squamous cell carcinoma in the presence of the p53 codon 72 arginine genotype. Frequency distributions of the three different genotypes (homozygous for the arginine allele, heterozygous for the two alleles, and homozygous for the proline allele) were similar among the squamous cell carcinoma group and the control group: 47.1%, 46.0% and 6.9% versus 47.8%, 45.8% and 6.4%, respectively. Statistical analysis showed no significant differences between these groups. In patients with squamous cell carcinoma and controls who harbored EV‐HPV DNA in their plucked eyebrow hair, similar results were obtained. The distributions of the p53 codon 72 genotypes in the basal cell carcinoma and malignant melanoma group were also not significantly different from the control group. p53 codon 72 arginine homozygosity does not appear to represent a significant risk factor for cutaneous squamous cell carcinoma and screening seems not to be indicated. Mol. Carcinog. 30:56–61, 2001.

Clearance of Cervical Human Papillomavirus Infection by Treatment for Cervical Dysplasia

Objectives: The aim of this study was to investigate the prevalence of human papillomavirus (HPV) after treatment for cervical dysplasia. Study Design: The presence of HPV was investigated in cervical scrapes of 91 patients, before and after treatment, using consensus primers in the polymerase chain reaction. Results: Before treatment for cervical dysplasia, 89 of 91 patients (98%) were HPV‐positive compared with 28 of 91 patients (31%) after treatment. The HPV type present before treatment was found in the scrapes of only 9 of 89 patients (10%). Detection of HPV after treatment was associated with human immunodeficiency virus infection and incomplete conization. Conclusion: The HPV type present before treatment was frequently cleared by treatment for cervical dysplasia.

No evidence of known types of human papillomavirus in squamous cell cancer of the oesophagus in a low-risk area

Controversial results regarding the presence and role of human papillomavirus in the development of oesophageal squamous cell carcinoma have been published. We used multiple broad-spectrum polymerase chain reactions to identify HPV DNA in oesophageal carcinomas from a low-incidence area. Paraffin embedded- and snap-frozen specimens from oesophageal cancer tissues of 63 patients were examined with a PCR technique with several primer pairs, capable of detecting most known HPV types. In none of the oesophagus cancer tissues could HPV DNA be detected. The role of HPV in this type of carcinoma in a low incidence area remains unclear.

RISK FACTORS FOR HPV DETECTION IN ARCHIVAL PAP SMEARS. A POPULATION-BASED STUDY FROM GREENLAND AND DENMARK

The most important risk factor for cervical cancer is genital infection with certain types of human papillomavirus (HPV). The presence of HPV was studied in archival smears from a random sample of women living in Greenland (GW) and Denmark (DW) having, respectively, a high risk and an intermediate risk for cervical cancer. Risk factors were also examined of the original 126 Danish and 129 Greenlandic archived smears collected during October and November 1988. 125 were located from each country including all abnormal smears. HPV DNA was isolated from the smears and detected by means of a consensus polymerase chain reaction (PCR) detecting a broad spectrum of genital HPV types. HPV was detected in all the abnormal smears and in 22 and 33% respectively of the cytological normal smears from DW and GW. Risk of HPV was significantly higher in smears from women who started sexual life relatively recently (respectively, < or = 4 and < or = 6 years ago in DW and GW) compared with > or = 10 years ago (adjusted prevalence-OR: 9.3; 95% CI: 2.2-39.2 in DW and 5.9; 95% CI: 1.4-25.3 in GW). Among other important risk factors were age in both areas, lifetime number of sex partners and current smoking in DW and ever and gonorrhoea in GW. This study confirms the usefulness of the method as all abnormal smears were positive and, furthermore, the predictors for HPV presence in the normal smears corroborate with those found in recent studies of HPV in fresh cervical swabs. Thus, this method can be useful for large-scale epidemiological studies of HPV DNA in already sampled material.