Jan M.M. Walboomers

A recent report that 93 per cent of invasive cervical cancers worldwide contain human papillomavirus (HPV) may be an underestimate, due to sample inadequacy or integration events affecting the HPV L1 gene, which is the target of the polymerase chain reaction (PCR)-based test which was used. The formerly HPV-negative cases from this study have therefore been reanalyzed for HPV serum antibodies and HPV DNA. Serology for HPV 16 VLPs, E6, and E7 antibodies was performed on 49 of the 66 cases which were HPV-negative and a sample of 48 of the 866 cases which were HPV-positive in the original study. Moreover, 55 of the 66 formerly HPV-negative biopsies were also reanalyzed by a sandwich procedure in which the outer sections in a series of sections are used for histological review, while the inner sections are assayed by three different HPV PCR assays targeting different open reading frames (ORFs). No significant difference was found in serology for HPV 16 proteins between the cases that were originally HPV PCR-negative and -positive. Type-specific E7 PCR for 14 high-risk HPV types detected HPV DNA in 38 (69 per cent) of the 55 originally HPV-negative and amplifiable specimens. The HPV types detected were 16, 18, 31, 33, 39, 45, 52, and 58. Two (4 per cent) additional cases were only HPV DNA-positive by E1 and/or L1 consensus PCR. Histological analysis of the 55 specimens revealed that 21 were qualitatively inadequate. Only two of the 34 adequate samples were HPV-negative on all PCR tests, as against 13 of the 21 that were inadequate ( p< 0.001). Combining the data from this and the previous study and excluding inadequate specimens, the worldwide HPV prevalence in cervical carcinomas is 99.7 per cent. The presence of HPV in virtually all cervical cancers implies the highest worldwide attributable fraction so far reported for a specific cause of any major human cancer. The extreme rarity of HPV-negative cancers reinforces the rationale for HPV testing in addition to, or even instead of, cervical cytology in routine cervical screening.

Sequence analysis of human papillomavirus (HPV) general primer GP5/6 mediated PCR products revealed the presence of short highly conserved sequences adjacent to the 3' ends of both primers. Part of these sequences was used to elongate GP5 and GP6 at their 3' ends to generate the primers GP5+ and GP6+, respectively. Compared with the GP5/6 PCR, GP5+/6+ specific PCR on 22 cloned mucosotropic HPVs revealed an improved HPV detection, reflected by a 10- to 100-fold higher sensitivity and a markedly increased signal to background ratio, especially at the gel level. As determined on purified DNA, the sensitivity of this GP5+/6+ based assay was at the femtogram level for those HPV genotypes which match strongly with the primers (e.g. HPV-16) and at the picogram level for HPV types (e.g. HPV-39 and -51) having four or more mismatches with one or both primers. Application of both methods on 264 cervical scrapes of a cohort of women participating in a prospective follow-up study revealed an increase of total HPV positivity from 39% (GP5/6 PCR) to 43% (GP5+/6+ PCR) of the scrapes. Additional HPV typing by PCR specific for the HPV-6, -11, -16, -18, -31 and -33 revealed that all GP5+/6+ PCR positive cases which were negative by GP5/6 PCR (n = 12) contained HPV types different from these six types. These data indicate that the GP5+/6+ PCR method provides an increased detection level mainly of uncommon, apparently poorly matched HPV types in cervical scrapes and most likely in the enlargement of the spectrum of HPVs detectable by this assay.

BACKGROUND: The incidence of anal cancer has increased in recent decades, particularly among women. To identify underlying risk factors, we conducted a population-based case-control study in Denmark and Sweden. METHODS: We conducted telephone interviews with 324 women and 93 men in whom invasive or in situ anal cancer was diagnosed between 1991 and 1994, 534 controls with adenocarcinoma of the rectum, and 554 population controls. The interviews covered a wide spectrum of possible risk factors for anal cancer. Odds ratios were calculated by logistic regression. Specimens of anal-cancer tissue and samples of rectal adenocarcinomas were tested for human papillomavirus (HPV) DNA with the polymerase chain reaction. RESULTS: Multivariate analysis revealed consistent and statistically significant associations between measures of sexual promiscuity and the risk of anal cancer in both men and women. There was a significant trend toward an association between higher numbers of partners of the opposite sex in women (P<0.001) and men (P<0.05) and strong associations with a variety of venereal diseases. In women, receptive anal intercourse, particularly before the age of 30 years, and venereal infections in the partner were also associated with an increased risk (odds ratios, 3.4 and 2.4, respectively). Fifteen percent of the men with anal cancer reported having had homosexual contact, as compared with none of the controls (P<0.001). High-risk types of HPV, notably HPV-16, were detected in 84 percent of the anal-cancer specimens examined, whereas all rectal-adenocarcinoma specimens tested were negative for HPV. CONCLUSIONS: Our study provides strong evidence that a sexually transmitted infection causes anal cancer. The presence of high-risk types of HPV, notably HPV-16 (which is known to cause cancer of the cervix), in the majority of anal-cancer tissue specimens suggests that most anal cancers are potentially preventable.