Heather L. Lehman

p120-Catenin Down-Regulation and Epidermal Growth Factor Receptor Overexpression Results in a Transformed Epithelium That Mimics Esophageal Squamous Cell Carcinoma

Esophageal squamous cell carcinoma (ESCC) is an aggressive malignancy with a poor prognosis due to its highly invasive and metastatic potential. The molecular pathogenesis underlying the invasive mechanism of ESCC is not well known because of the lack of existing models to study this disease. p120-Catenin (p120ctn) and the epidermal growth factor receptor (EGFR) have each been implicated in several cancers, including ESCC. p120ctn is down-regulated in 60% of ESCC tumors, whereas EGFR is the most commonly overexpressed oncogene in ESCC. For these reasons, we investigated the cooperation between p120ctn and EGFR and its effect on ESCC invasion. We show that p120ctn down-regulation is commonly associated with EGFR overexpression. By using a three-dimensional culture system, we demonstrate that the inverse relationship between p120ctn and EGFR has biological implications. Specifically, p120ctn down-regulation coupled with EGFR overexpression in human esophageal keratinocytes (EPC1-PE) was required to promote invasion. Morphological comparison of EPC1-PE cells grown in three-dimensional culture and human ESCC revealed identical features, including significantly increased cellularity, nuclear grade, and proliferation. Molecular characteristics were measured by keratin expression patterns, which were nearly identical between EPC1-PE cells in three-dimensional culture and ESCC samples. Altogether, our analyses have demonstrated that p120ctn down-regulation and EGFR overexpression are able to mimic human ESCC in a relevant three-dimensional culture model.

p120-Catenin Downregulation and PIK3CA Mutations Cooperate to Induce Invasion through MMP1 in HNSCC

Despite recent improvements in treatment for head and neck squamous cell carcinoma (HNSCC), half of all patients with a regional or advanced disease will die within 5 years from diagnosis. Therefore, identification of mechanisms driving the aggressive behavior of HNSCC is of utmost importance. Because p120-catenin (CTNND1/P120CTN) downregulation and PIK3CA mutations are commonly found in HNSCC, the objective of this study was to identify their impact on fundamental processes of metastasis, specifically, migration and invasion. Furthermore, this study aimed to identify the key effector proteins regulated by P120CTN downregulation and PIK3CA mutations. Studies using oral keratinocytes demonstrated that P120CTN downregulation and PIK3CA mutations increased migration and invasion. In addition, P120CTN downregulation and PIK3CA mutations resulted in elevated matrix metallopeptidase 1 (MMP1) levels. Inhibition of MMP1 resulted in decreased invasion, suggesting that MMP1 plays a critical role in HNSCC invasion. Moreover, analysis of HNSCC patient specimens from The Cancer Genome Atlas confirmed these findings. Tumors with low P120CTN and PI3K pathway mutations have higher levels of MMP1 compared to tumors with high P120CTN and no PI3K pathway mutations. In conclusion, this study demonstrates that P120CTN downregulation and PIK3CA mutations promote MMP1-driven invasion, providing a potential novel target for limiting metastasis in HNSCC. Implications: Because of its role in invasion, MMP1 represents a novel, potential target for limiting metastasis in a subset of HNSCCs with P120CTN downregulation and PIK3CA mutations. Mol Cancer Res; 15(10); 1398–409. ©2017 AACR.

Increasing diagnostic accuracy to grade dysplasia in Barrett's esophagus using an immunohistochemical panel for CDX2, p120ctn, c-Myc and Jagged1.

BackgroundPatients with non-dysplastic Barrett’s esophagus (ND-BE) and low-grade dysplasia (LGD) are typically monitored by periodic endoscopic surveillance, while those with high-grade dysplasia (HGD) and esophageal adenocarcinoma (EAC) are usually treated by more aggressive interventions like endoscopic mucosal resection, ablation or surgery. Therefore, the accurate grading of dysplasia in Barrett’s esophagus (BE) is essential for proper patient care. However, there is significant interobserver and intraobserver variability in the histologic grading of BE dysplasia. The objective of this study was to create an immunohistochemical (IHC) panel that facilitates the grading of BE dysplasia and can be used as an adjunct to histology in challenging cases.Methods100 BE biopsies were re-graded for dysplasia independently by 3 subspecialized gastrointestinal pathologists. IHC staining for CDX2, p120ctn, c-Myc and Jagged1 proteins was then performed and assessed by two separate methods of semi-quantitative scoring. Scores were integrated using a principal component analysis (PCA) and receiver operating characteristic (ROC) curve.ResultsPrincipal component analysis demonstrated the ability of this panel of proteins to segregate ND-BE/LGD and HGD/EAC, as the expression of the four proteins is significantly altered between the two subsets. Analysis of the receiver operating characteristic curve showed that this panel has the potential to aid in the grading of dysplasia in these two subcategories with both high sensitivity and specificity. While not able to discriminate between ND-BE and LGD, this panel of four proteins may be used as an adjunct to help discriminate subsets of ND-BE/LGD from HGD/EAC.ConclusionsWe propose that the maximum utility of this IHC panel of CDX2, p120ctn, c-Myc, and Jagged1 proteins would be to distinguish between LGD and HGD in histologically challenging cases, given the aggressive interventions still used for HGD in many institutions, and hence may aid in the optimal patient management. The results of this initial study are promising, though further validation is needed before this panel can be used clinically, including future randomized prospective studies with larger patient cohorts from diverse locations.

Single and Multiple Gene Manipulations in Mouse Models of Human Cancer.

Mouse models of human cancer play a critical role in understanding the molecular and cellular mechanisms of tumorigenesis. Advances continue to be made in modeling human disease in a mouse, though the relevance of a mouse model often relies on how closely it is able to mimic the histologic, molecular, and physiologic characteristics of the respective human cancer. A classic use of a genetically engineered mouse in studying cancer is through the overexpression or deletion of a gene. However, the manipulation of a single gene often falls short of mimicking all the characteristics of the carcinoma in humans; thus a multiple gene approach is needed. Here we review genetic mouse models of cancers and their abilities to recapitulate human carcinoma with single versus combinatorial approaches with genes commonly involved in cancer.

NFkB hyperactivation causes invasion of esophageal squamous cell carcinoma with EGFR overexpression and p120-catenin down-regulation

Four out of five patients diagnosed with esophageal squamous cell carcinoma (ESCC) will die within five years. This is primarily a result of the aggressive invasive potential of the disease. Our research is focused on the interplay between tumor suppressors and oncogenes in the invasive process. Specifically, EGFR and p120-catenin (p120ctn) are commonly dysregulated genes that are indicative of poor prognosis in ESCC. In a previous study we demonstrated that in our 3D organotypic culture model, only when EGFR overexpression is combined with p120ctn inactivation do the cells transform and invade – as opposed to either event alone. The purpose of this present study was to identify the components of the molecular pathways downstream of p120ctn and EGFR that lead to invasion. Using both human esophageal keratinocytes and human ESCC cells, we have identified NFkB as a central regulator of the invasive process downstream of p120ctn down-regulation and EGFR overexpression. Interestingly, we found that NFkB is hyperactivated in cells with EGFR overexpression and p120ctn inactivation than with either EGFR or p120ctn alone. Inhibition of this NFkB hyperactivation results in complete loss of invasion, suggesting that NFkB signaling is necessary for invasion in this aggressive cell type. Furthermore, we have identified RhoA and Rho-kinase as upstream regulators of NFkB in this process. We believe the cooperation of p120ctn down-regulation and EGFR overexpression is not only important in the aggressive mechanisms of ESCC but could be broadly applicable to many other cancer types in which p120ctn and EGFR are involved.

Additional file 1: Figure S1. of Increasing diagnostic accuracy to grade dysplasia in Barrett’s esophagus using an immunohistochemical panel for CDX2, p120ctn, c-Myc and Jagged1

Expression levels of four-marker protein panel in Barrett’s esophagus disease progression. (A) CDX2 expression with the quartile scoring method and (B) IRS scoring, decreased from ND-BE to EAC

Signaling Pathways Supporting Tumor Invasion in Head and Neck Squamous Cell Carcinoma

Head and neck squamous cell carcinoma (HNSCC) is a highly invasive cancer. A number of signaling pathways like PI3K, Rho and TGFβ/SMAD drive the invasive nature of HNSCC. The PI3K pathway is the most altered pathway in HNSCC. Both upstream and downstream members of this pathway have been found to be mutated or overexpressed leading to an increase in cell invasion. The Rho pathway is also commonly activated; however, only overexpression or downregulation of Rho’s upstream regulators are found in HNSCC. Finally, TGFβ/SMAD pathway activation leads to epithelial mesenchymal transition in HNSCC and subsequently invasion, though loss of TGFβ/ SMAD signaling has also been shown to increase cell invasion.