Mackintosh, Lorna J.
The role of Human Papillomaviruses and their replication in nonmelanoma skin cancer.
PhD thesis, University of Glasgow.
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Nonmelanoma skin cancer (NMSC) is the commonest cancer worldwide and is a significant and increasing burden on health care resources. NMSC aetiology is fundamentally linked to sun exposure although infection with oncogenic viruses including Human Papillomaviruses (HPV) is thought to be a cofactor. Fair skinned populations residing in geographical areas with high sun exposure have an increased incidence of NMSC and an excess of NMSC is also observed in certain patient groups including immunosuppressed organ transplant recipients.
This thesis describes the epidemiology of NMSC in the immunosuppressed renal transplant population in the West of Scotland. Clinical samples from this population were investigated for the presence of HPV utilising a PCR reverse hybridisation technique. This assay specifically examined for HPV of the genus Beta that have been previously linked to NMSC.
The effect of presence of HPV in these samples was evaluated through an investigation of the expression of cellular biomarkers. A biomarker expression pattern specific to HPV infected lesions would add further support to the link between HPV and NMSC. Samples were probed for Ki67, p16, p53, MCM2 and MCM5 antigens in addition to novel antigen, Topoisomerase II Beta Binding Protein 1 (TopBP1). TopBP1 is a host cellular protein that is involved in the DNA damage response and is an interacting partner for HPV thus making it a likely candidate for involvement in cutaneous carcinogenesis. A biomarker expression pattern specific to HPV infected NMSC was not identified although aberrant expression of TopBP1 was identified in a subset of skin cancers.
To investigate this novel observation, a molecular investigation of the interaction between TopBP1 and the viral replication factor E2 was carried out, firstly in HPV-16 before extending the work to the cutaneous virus HPV-8. A mutant of HPV-16E2 that failed to bind TopBP1 was generated. Failure to bind TopBP1 resulted in a phenotype exhibiting compromised viral replication. The equivalent mutation generated in HPV-8E2 resulted in an even more compromised replicative phenotype. In addition to this work, gene targets of TopBP1 were also identified by a microarray analysis. This was carried out using MCF7 cells untreated or damaged by ultraviolet radiation (UVR) with endogenous TopBP1 depleted by SiRNA treatment. These studies showed that TopBP1 is involved in a number of cellular processes and is likely to be involved in controlling DNA damage targets following UVR induced DNA damage.
The work described in this thesis provides further evidence supporting the role of HPV in the aetiology of nonmelanoma skin cancer. Generation of an E2 mutant that fails to bind TopBP1 and exhibits a compromised replication phenotype provides further evidence to support the hypothesis that the E2-TopBP1 interaction is essential for viral replication and therefore the viral life cycle. This work may facilitate the development of anti-viral therapies for HPV-associated disease by targeting HPV DNA replication through disruption of the E2/TopBP1 interaction.
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