From a virologic perspective, the definitive proof of carcinogenicity of an HPV type is finding transcriptionally active HPV in a tumour. HPV is not a "hit and run" carcinogen, and transcriptional activity is needed for maintenance of the cancer phenotype. In cervical cancer cell lines, blocking transcriptional activity by antisense RNA leads to apoptosis. This level of evidence is simply lacking for virtually all HPV types. The vast body of evidence relates simply to finding HPV DNA at the same time of cervical neoplasia.
However, relying on testing of scrapes or biopsies, by DNA testing, leads to difficulties. The alpha HPV types share a common route of transmission and multiple infections are present in a large minority of women, although they might not be transmitted from the same partner or at the same time. Given the existence of some very powerfully carcinogenic types, notably HPV 16 and HPV 18, determining which weaker and/or less common types are also carcinogenic becomes (for the epidemiologist) an issue of confounding. None of the traditional approaches to control of confounding are entirely successful. Because HPV 16 causes approximately 50% of cases of cervical cancer, logistic regression and similar approaches will parsimoniously attribute cases associated with both HPV16 and a less important type to HPV16. HPV 18 is the second most important cervical carcinogen, responsible for approximately 15–20% of cervical cancer of all histologic types combined (and a higher fraction of adenocarcinomas). If a type occurs with either HPV 16 or HPV 18, its association with cervical cancer might be confounded by either of these powerful carcinogens. For types causing only a very small fraction of cervical cancer, confounding by any of the more important types is possible.
Dealing with confounding by exclusion, i.e. examining the possibility of carcinogenicity of a more minor type among cancer specimens that do not contain a more important type, becomes a problem of misclassification. This main epidemiologic criterion used for classification of an HPV type as a carcinogen, finding the HPV genotype as a single infection in a cervical scrape or biopsy specimen in a woman with cancer, might sometimes be too lax and prone to error. Colposcopic biopsies and cytology specimens can be misdirected and fail to obtain the critical cells, while contamination of scrapes and biopsies from lower-grade lesions that often surround cancers can detect types other than the causal one. Studies relying on testing of microdissected cervical malignancies will address these issues, but large-scale highly accurate data are not yet available.
Difficulty with control selection adds another level of complexity in assessing carcinogenicity. Cervical cancer typically follows age infection by decades. HPV transmitted at young ages usually become undetectable by DNA or RNA assays and no sensitive serologic assay exists to measure HPV exposure. Consequently, odds ratios based on a comparison of HPV prevalence at the time of case diagnosis to age-matched HPV point prevalence in controls do not estimate true relative risks.
There is not much type-specific prospective data on the carcinogenicity of individual HPV genotypes. The available studies have categorically shown the unique carcinogenicity of HPV 16 and, to a lesser extent, HPV 18 [9
]. Khan et al. observed a risk for the remaining women positive by hc2 after excluding those positive for HPV 16 or HPV 18 (including an unknown mix of the types 31, 33, 35, 39, 45, 51, 52, 56, 58, 59 and 68) of only 3.0% (1.9–4.2) compared with 0.8% (0.6–1.1) among women who were HPV negative at baseline. Thus, there is not strong and convincing long-term prospective evidence for individual HPV types other than HPV16 and HPV18.
Finally, the accuracy of detection of HPV genotypes differs between the major PCR-based systems used to generate most of the data [13
]. Each of the systems shows differential sensitivity, and some have exhibited cross-reactivity of detection. These detection issues are not critical for evaluating the most important HPV types, but make it difficult to clarify the role of the most weakly carcinogenic and least common ones.
With these caveats, the cervical carcinogenicity of the HPV types listed above varies in strength in a continuum without clear breakpoint, from extremely strong (i.e. HPV 16 and, to a lesser degree, HPV 18) to weak, but still probably carcinogenic in rare instances (e.g. HPV 68, see below). Evaluators taking one extreme position could claim that there is reasonable evidence for carcinogenicity of virtually all the types in the species listed above, extending further the list established in Monograph 90. Strict interpreters of causal criteria could argue for a return to a much more limited list. But based on current evidence, no cut-point between sufficient, limited, and inadequate epidemiologic evidence is entirely defensible.