Background Information

Human papillomavirus (HPV)

Tumor virology is the science that researches the role of viruses in human tumor genesis. Main players in tumor virology are human papilloma viruses that play a role in genital, skin and oropharyngeal cancers. At present more than 100 genotypes have been isolated, 77 of which have been identified in humans. A low mutation rate is suggested by the small differences in base sequences of the different genotypes. Cervical cancer is the second leading cause of cancer in women. Although it has been observed that cervical cancer behaves like a sexual transmitted disease, it was not until the 1970s that HPV was shown to be the causative agent. In cervical cancers, HPV has been detected in up to 95% of the cases. The long latency period between infection and the development of cancer, the variety of progression to cervical cancer and the clonal origin of cervical cancer show that HPV infection alone is not sufficient for the development of cervical cancer, but requires a multi-step event. Epidemiological data on the occurrence of cervical cancer in biologically related women also suggests the involvement of a hereditary factor in the etiology of cervical cancer. The multi-step nature of cervical cancer is illustrated by the distinct pre-neoplastic epithelial changes called intraepithelial neoplasia (CIN) which form a spectrum of atypia, graded CIN I to CIN III. The proteins expressed by the HPV genome interfere with the cell cycle regulation process. Regression or progression in cervical pre-neoplasia thus depends on the genetic instability caused by the interaction of HPV and the cell cycle.

Screening for HPV
Women with an intact immunological response system will not carry the virus their entire live. Infections by HPV subtypes will generate an immune response that protects against subsequent infections by the same type. The derived immune response will temper some infections with other types and result in transient lesions in many women. The overall prevalence of HPV in the target population is 20%. Since HPV exposure is greatest in young sexually active women, detection of the virus will exceed by several-fold the ability to detect abnormalities. In post-menopausal women, the HPV infection index is much lower, but cytological abnormalities are detected more often. It is clear that the PAP screening procedure is effective and it has been demonstrated that a positive HPV test is a powerful independent risk factor for the development of CIN lesions, as 93-100% of invasive carcinomas are associated with HPV infections. However, the relative value of screening for HPV in a population is shown by combining the high prevalence of HPV with the long time frame between infection and development of cancer. In other words, the incidence of cancer in women under 25-30 years old is very low whereas the incidence of HPV infection is about 40%. High throughput population screening without morphological-based diagnosis will result in high numbers of virus positive patients, most of whom will never suffer any consequences of the infection, making such testing redundant and a financial burden for healthcare. It is therefore more advisable to target HPV screening, i.e. screening those women for HPV who have developed early cytological abnormalities in order to monitor possible developments towards more serious abnormalities, thus being able to interfere with the disease at an early stage. The in situ HPV hybridization test enables visual control on the cells and the localization of infection as opposed to testing procedures in liquid formats. The information on histological details in combination with HPV in situ hybridization can therefore exclude false positives and innocent positives (young women). Moreover, in situ hybridization makes it possible to differentiate between episomal or integrated HPV DNA. Studies have shown that episomal HPV induces genomic changes such as tetrasomies and single trisomies, while HPV integration correlates to aneusomies and polysomies, which are predominantly detected in CIN III and micro-invasive carcinoma, demonstrating that integration of HPV DNA is a pivotal step in the transition of CIN to micro-invasive carcinoma. Integration of the HPV genome together with histological information makes it possible to determine between high risk CIN II/III and low risk CIN I/II and it is here that in situ hybridization provides the tools for improved diagnosis: specific HPV type information next to morphological details enabling a distinct diagnosis.
Last but not least:
HPV has also been linked to the etiology of other cancers such as cancer of the vulva, penis, non-melanoma skin cancers (basal and squamous cell carcinoma), cancers of the oral cavity, larynx and esophagus. These observations emphasize the importance of this virus group as proven and suspected human cancer carcinogens.

PanPath's Rembrandt® kit for HPV screening and typing targets the type specific HPV genomes (approx. 8 Kb). The probes show no cross-hybridization with other subtypes, provided that the hybridization conditions as laid out in the protocol are strictly followed.