Esophageal squamous cell carcinoma (ESCC) is a highly aggressive gastrointestinal cancer characterized by a high rate of metastasis, limited therapeutic options, and a poor prognosis. However, there is limited information regarding the molecular mechanisms underlying the metastatic properties of ESCC. Several genetic alterations are known to be involved in the development of ESCC including overexpression of EGFR and cyclin D1, mislocalization/loss of p120 catenin, and mutations in the tumor suppressor gene p53. P53 is one of the most commonly mutated genes in ESCC, and our group has shown that esophageal cells lines expressing a mutation in human p53, specifically R175H, shows signs of malignancy and increased invasion in 3D organotypic culture. Interestingly, preliminary data indicate that these cells exhibit a downregulation of endocytic recycling genes, suggesting that p53 may be mediating invasion through a specific recycling gene set. However, a mouse model of mutant p53 in ESCC is lacking in the field. To elucidate the role of mutant p53 in ESCC we developed a novel mouse model utilizing a combinatorial genetic and carcinogenic approach that results in the development of ESCC following treatment. We hypothesize mutant p53 enhances ESCC invasion and metastasis via dysregulation of endocytic recycling. We will test this hypothesis through two interrelated Specific Aims.
In Aim 1 will we determine the tumorigenic role of mutant p53 in our novel mouse model using in vivo and in vitro approaches. Specifically, we will compare esophageal tumor incidence between mutant, null, and wt p53 mice, and will analyze tumor grade, degree of differentiation, inflammation, and metastasis through histological examination. We will also develop tumor cell lines to perform migration and invasion assays, 3D organoid culture, and transplantation experiments to evaluate mutant p53's metastatic potential and to test its effect on clinically relevant therapeutics.
In Aim 2 we propose to elucidate the role of p53 in mediating endocytic recycling during ESCC invasion and metastasis. This will be achieved through RNA-seq analysis of ESCC cell lines expressing mutant, null, and WT p53 to identify dysregulated genes involved in recycling. Genes will be genetically deleted in vitro using Crispr/Cas9 technology to determine their functional role, and EGFR recycling will be examined as a potential target of p53 mediated recycling during ESCC invasion. Together, these studies will provide new insights into the role of mutant p53 in ESCC tumorigenesis and will advance our understanding of a potential mechanisms of action, potentially leading to the identification of new therapeutic targets, including in other SCCs (head/neck, lung) that share common genomic and genetic properties.
Esophageal squamous cell carcinoma (ESCC) is a highly aggressive disease with limited therapeutic response and a poor survival rate. The proposed study will investigate the role of mutant p53 and recycling in ESCC invasion and metastasis. Results will yield mechanistic insights and new therapeutic targets for ESCC.
| Giroux, Véronique; Lento, Ashley A; Islam, Mirazul et al. (2017) Long-lived keratin 15+ esophageal progenitor cells contribute to homeostasis and regeneration. J Clin Invest 127:2378-2391 |
