ASPP2: A molecular switch of cancer initiating cells in Squamous Cell Carcinoma?

Epithelial tissue lines the surface of the body (i.e. the skin), along organs and glands inside it. Around 80% of all human cancers develop in the epithelium, and of the different epithelial cancer types, Squamous Cell Carcinoma (SCC) is the most deadly. How SCC develops is currently poorly understood, however, it is known that as well as expressing normal basal cell markers such as keratin 14, they also express markers normally found in differentiated squamous cells such as keratin 1. While it is possible that SCCs initially develop in basal cells, it is unclear why such basal cells in SCCs begin to express differentiation markers. One explanation is that a selective advantage may be derived from their expression that enables basal cells to gain new properties. Alternatively, it is possible that SCC instead initially develops from the differentiated keratin 1-positive and keratin 14-negative suprabasal layer of the epithelium, which sits above the basal layer. We have recently developed an in vivo mouse model system in which the expression of ASPP2, a previously identified tumour suppressor and regulator of p53, is knocked out. Over 90% of tumour-bearing ASPP2 heterozygous mice developed spontaneous SCC in less than 1 year, indicating that ASPP2 is able to suppress the development of SCC and that this model system is ideal for investigating the origin of SCC within the epithelium. We therefore propose to cross our ASPP2 mutant mice with mice in which either keratin 1 or 14 is expressed. We will also examine the mechanism by which ASPP2's suppression of SCC development is achieved, in particular, by examining the expression levels of genes involved in the p63, Notch and Wnt signalling pathways, all of which are known to be affected by ASPP2's activity.

Around 80% of human cancers are epithelial in origin. Of the different tumour types arising in epithelial tissues, squamous cell carcinoma (SCC) is the most malignant and has the highest metastatic potential. Our preliminary results show that the tumour suppressor ASPP2 is able to repress p63 expression, is highly expressed in keratin 1-positive and keratin 14-negative stratified epithelial cells, and that differentiation induces ASPP2's expression in vitro. Most importantly of all, ASPP2 heterozygous and null mice spontaneously develop SCC of the skin. These observations challenge the current view that SCC is derived from basal cells (p63+, keratin 14+/keratin 1-), as ASPP2 is hardly expressed in this cell population. SCC cells may instead be derived from differentiated keratin 1-positive cells in the suprabasal layer. Due to changes in their epigenetic regulation, differentiated keratin 1-positive squamous epithelial cells may start to re-express keratin 14 and p63 to gain a proliferative potential. To determine the precise origin of SCC, ASPP2 will be deleted in keratin 14-positive basal cells and keratin 1-positive suprabasal cells using keratin 14-Cre or keratin 1-Cre respectively. The ability of these mice to develop spontaneous SCC will be studied. To identify genes that ASPP2 uses to suppress SCC, we will also conduct gene array analysis on SCC samples from ASPP2 exon3-null and heterozygous mice, and compare them with tissue samples from wild type mice. The identified ASPP2-regulated genes will then be analysed further to determine whether or not they are involved in epithelial stratification. In particular, genes involved in epithelial stratification and that are targets of the p63, Notch and Wnt signaling pathways will be examined to determine the molecular mechanisms by which ASPP2 is able to suppress the development of SCC.

The proposed project seeks to address a key gap in the current knowledge regarding the cellular origin of SCC. It is therefore to have significant scientific impact, and it is anticipated that it will lead to future, more directed studies being conducted by the scientific community in order to derive better therapies against SCC. SCC is the most malignant epithelial tumour type, with the highest metastatic potential. As SCC is often curable if detected and treated early, a better understanding of the origin of SCC within the epithelium is also likely to lead to improvements in early detection of the disease and, ultimately, improved patient prognosis and quality of life. The ASPP family of proteins is evolutionarily conserved, and ASPP2 has been found to be a haploinsufficient tumour suppressor. While this proposal encompasses basic research in an in vivo model system, the knowledge gained from this study is likely to have long-term, important implications for early diagnosis and the development of new treatment strategies for SCC. The proposed project therefore has the potential to have a significant impact on treatment practices.