Clinical and biological implications of ΔNp63α - an emerging therapeutic oncological target gene
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The p63 gene encodes multiple protein isoforms that function as sequence specific transcription factors, which can regulate both p53 dependent and independant target genes. Of these, ΔNp63α has been identified as one of the most important isoforms. It is the principal isoform that acts as a crucial regulator of embryonic development. It is also the predominant isoform that is upregulated in some human cancers, thus supporting an important role in human neoplasia. p63 protein is widely reported to be overexpressed in more than 80% of primary head and neck squamous cell carcinomas (HNSCC), and this high expression has been reported to have prognostic implications in these patients. It has also been reported that ∆Np63 is expressed in pancreatic cancers. However, expression of ∆Np63 in pancreatic cancers varies from 8% to 68.2%, and it has not yet been related to any clinicopathological features. In addition, little is known about how ∆Np63 affects prognosis and tumour cell biology in pancreatic cancers. In an effort to further understand the role of ΔNp63α in HNSCC and elucidate the contribution of ΔNp63α in human pancreatic ductal adenocarcinoma, this study has sought to identify the functional effects of ΔNp63α on tumour biology and ultimately novel transcriptional targets for ΔNp63α through the use of gene expression profiling. Using immunohistochemistry analysis of primary human HNSCC and PDAC tissue samples, this has confirmed ΔNp63α is overexpressed in a spectrum of neoplasias in both tumour models. Its expression pattern is suggestive of a role in cell differentiation and overall, it correlates with significantly poorer prognosis for primary pancreatic cancers. RNA interference with four independent p63 targeting sequences have been used to downregulate ΔNp63α expression in the p63 positive carcinoma cell lines from the two selected tumour models. Using this approach, several important biological functions have been determined. The cellular effects of ΔNp63α expression on tumourigenicity, cancer cell migration, invasion and its effect on chemosensitivity have been determined. The results have identified an important role for ΔNp63α in cancer initiation and also its part in contributing to tumour progression. Microarrays have been utilised to create a ΔNp63α dependent transcription profile. Statistical comparisons between non-silencing control siRNA and p63 targeting siRNA groups identified a total of 237 gene expression changes (p<0.05). Seven of these gene expression changes, including PRSS23 and BMP2K have been validated using qRT PCR and immunoblotting, where possible. This has confirmed that both PRSS23 and BMP2K are both downstream target genes that are differentially expressed according to ΔNp63α, at transcriptional level. Functional studies have also demonstrated that these genes, in particular PRSS23, are implicated in mediating the effects of ΔNp63α on tumour cell proliferation, migration and chemosensitivity.
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