Molecular Basis of Esophageal Cancer Development and Progression

Over the years, the morbidity and mortality due to cancer have continued to increase worldwide. Plants are well utilized in Nigeria ethno-medicine for cancer treatment. However, scientific evidence, particularly at the molecular level, has been lacking, and biodiversity loss is a threat.

The cytotoxic activity of the leaves of six plants used in Nigeria ethno-medicine was investigated and the molecular pathway of cytotoxic action of two active extracts on four cancer cell lines (A549, RD, MCF-7, and HeLa) was also assessed.

Cytotoxic activities of the plant extracts were assessed on breast (MCF-7), lung (A549), cervical (HeLa), adenocarcinoma and rhabdomyosarcoma (RD) cells using MTT cell viability assay in a time-dependent manner. The sandwich ELISA method was used to assess the protein expressions (Bcl-2, BAX, and executioner caspase 3) involved in the apoptotic pathway in cancer cells after treatment with cytotoxic extracts. UHPLC-MS/MS approach was used to analyze the possible bioactive phytochemicals of the cytotoxic plant extract.

Uvaria chamae and Dicliptera paniculata extracts displayed good cytotoxicity across all cell lines in a time- and concentration-dependent manner (CC₅₀ <30 µg/mL). Increased pro-apoptotic BAX and lowered expression of anti-apoptotic Bcl-2 show the cytotoxic action of the two extracts followed the intrinsic apoptotic pathway compared to untreated cells. The increased expression of executioner caspase 3 indicated that the intrinsic apoptotic pathway was caspase-dependent.

The anticancer potential of U. chamae and D. paniculata in this study validates the use of these plants in Nigerian ethno-medicine, highlighting apoptosis as a molecular pathway of their cytotoxicity.

Esophageal adenocarcinoma (EAC) is one of the most aggressive gastroesophageal cancers. PTGS2, EGFR, ERBB2 and TP53 are the traditional EAC prognostic biomarkers, but they are still limited in their ability to effectively predict the overall survival.

To identify an improved biomarker for predicting the prognosis of EAC by using the expression profile.

Differential co-expression analysis and differential expression analysis were performed to identify the related genes of EAC. The 5-fold cross-validation was used to select a prognostic biomarker from the 532 EAC related genes.

CRNKL1 was identified as a prognostic biomarker to predict the survival of EAC patients. It could significantly stratify EAC patients into high-risk and low-risk groups and was much better than the traditional biomarkers. Furthermore, ROC curve also verified that CRNKL1 with the highest area under the curve (AUC), reaching a sensitivity of 83.33% and a specificity of 78.57%.

Our research proposed that CRNKL1 might be a novel prognostic biomarker with better predictive ability by comparing with the traditional biomarkers, which provided a preferable opportunity in the clinical applications of EAC.

Esophageal squamous cell carcinoma (ESCC) is an aggressive tumor with rapid progression, high recurrence and metastasis rate. Besides, the 5-year survival rate of ESCC remains dismal despite improvements in treatments having developed a lot. That is because the cellular basis of ESCC has not yet been fully understood. Forkhead box family membranes possess various kinds of properties as they mediate apoptosis, cell cycle arrest, autophagy, senescence and so on. Foxhead box Q1 (FOXQ1), which has a major impact on several kinds of tumor forming and metastasis, while whether it triggers ESCC remains largely obscure.

To determine whether aberrant FOXQ1 expression in esophageal cancer, protein level and mRNA level of specimens of cancerous tissues and adjacent non-cancerous tissues were determined by western blot and real-time PCR. Then we overexpressed or knockdowned FOXQ1 in EC9706 cell; cell growth curve and colony formation were analyzed. Cell invasion ability was analyzed by migration chambers. Reporter gene assay was used to study the transcriptional regulation activity.

FOXQ1 was highly expressed in esophageal cancerous tissues compared with adjacent non-cancerous tissues. FOXQ1 overexpression promotes ESCC tumor cell proliferation, whereas FOXQ1 silencing prevents ESCC tumor cell proliferation. FOXQ1 promotes ESCC metastasis via negatively modulation CDH1. CDH1 silencing could rescue the migratory ability which was blemished by FOXQ1 silencing. FOXQ1 could act as transcriptional repressor which binds to the promoter of CDH1 and blocks its transcription.

In this study, we identified FOXQ1 as an oncogene to promote ESCC tumor cell proliferation and metastasis by negatively regulating CDH1 in EC9706 cell. Besides, we deciphered a previously unidentified mechanism of ESCC progression and metastasis.

CRNN gene expression is downregulated in esophageal squamous cell carcinoma (ESCC), although its clinical implications in esophageal cancer remain unclear.

We performed integrated microarray analysis and identified the CRNN gene as 1 of the major downregulated genes in ESCC. CRNN downregulation was validated at the nucleic acid level in 16 ESCC cases using complementary DNA microarray and at the protein level by immunohistochemical stains in an additional 220 ESCC cases. The clinicopathologic relevance and prognostic significance of CRNN expression in ESCC were explored.

Downregulated CRNN expression was noted at the messenger RNA and protein levels. Immunohistochemical staining revealed negative and positive CRNN expression in 171 (77.7%) and 49 (22.3%) patients with ESCC, respectively. Patients with negative CRNN protein expression had an advanced tumor invasion depth (P = .002), advanced nodal involvement (P = .014), and longer tumor length (P = .037). Patients with negative CRNN expression (median survival, 14.0 months; 5-year survival rate, 20.5%) had poorer overall survival than those with positive expression (30.0 months and 40.3%, respectively; P = .006). On multivariate analysis, negative CRNN expression, nodal involvement, and distant metastasis remained significant prognostic factors for poor overall survival (negative vs positive CRNN, hazard ratio, 1.464; P = .047).

Our analysis has highlighted the clinical implications of CRNN expression in ESCC. Loss of CRNN expression correlated with advanced tumor length, greater tumor invasion depth, lymph node metastasis, and poor survival in patients with ESCC.