Esophageal adenocarcinoma (EAC) is the most prevalent type of esophageal malignancy in the United States (US) and the rate of occurrence continues to grow rapidly. As the prevalence of risk factors such as obesity and gastroesophageal reflux disease (GERD) rises, the rates of EAC are expected to continue rising as well. Unfortunately, the 5-year survival rate remains low and the lack of targetable, oncogenic drivers presents challenges in developing more effective and less toxic therapeutics. The current standard of care for EAC involves combinations of chemotherapeutics and radiation therapy that can cause severe side effects and often leads to refractory and relapsed disease. According to the cancer stem cell model, a small subset of the tumor cell population is responsible for cancer's ability to replicate, metastasize, and relapse. These cancer stem cells (CSCs) can self-renew and differentiate. Napabucasin, a "stemness" inhibitor, which works by inhibiting STAT3, has shown promising results in pre-clinical and clinical investigations across a variety of solid tumor types. Because a major barrier in treatment of EAC is the likelihood of relapse, targeting the CSC population that results in this phenotype is a therapeutic strategy of great interest. We hypothesize that employment of napabucasin to inhibit stemness through STAT3 represents a viable therapeutic strategy in the EAC setting. In this study, we investigated the efficacy of napabucasin on EAC cells. Napabucasin was shown to reduce phosphorylation of STAT3 as well as levels of MCL1, a cell survival protein downstream of STAT3, and levels of "stemness" markers Nanog, Sox2, and B-catenin via immunoblot analysis. Napabucasin monotherapy showed high efficacy in some EAC settings, with IC50 values in a clinically achievable range. The treatment in combination with cisplatin, a standard of care chemotherapeutic, resulted in reduced cell viability than either treatment alone indicating that a combination strategy could reduce the dosage of each drug needed. The data suggests that STAT3 inhibition in combination with current standard of care treatments could be a viable therapeutic strategy in EAC, and improve the dismal survival for these patients.

Prevalence of esophageal adenocarcinoma (EAC) has increased six-fold in the United States over the past four decades due to increases in associated risk factors, namely obesity and gastroesophageal reflux disease. The most common genomic driver of EAC, tumor protein 53 (TP53) mutation, has previously been therapeutically intractable, affirming the unmet clinical need to deploy novel therapeutic strategies targeting this genomic driver in this tumor type. 72 percent of EAC patients have mutations in TP53, making tumors more reliant on the G2/M checkpoint to repair DNA damage, increasing likelihood of efficacious G2/M abrogation via targeting WEE1 G2 checkpoint kinase (WEE1), a modulator of this checkpoint. We hypothesize that the G2/M checkpoint represents a viable therapeutic avenue against the most common genomic driver of EAC. We investigated the efficacy of the WEE1 inhibitor AZD1775 on EAC cells. WEE1 mRNA expression levels in EAC are elevated compared to normal tissue controls. AZD1775 was shown to induce cyclin dependent kinase 1 (CDK1) mediated cell cycle progression and increased DNA damage markers as exposure increased via immunoblot analysis. SK-GT- 4 EAC cell line viability was significantly reduced by up to 30 percent when treated with AZD1775 and cisplatin when compared to cisplatin alone. AZD1775 monotherapy showed high efficacy in some EAC settings. Simultaneous dual therapy demonstrated the highest overall efficacy, and stepwise sequential treatments offered negligible benefit. Future research will explore the genomic contexts of the EAC celllines used in order to understand the different responses to AZD1775 monotherapy regimens. Ionizing radiotherapy will be employed in order to understand the DNA damage response timeline, providing more information on the mechanisms of G2/M checkpoint inhibitors in this disease setting. This research will provide insight into novel therapeutic targets for EAC leading to therapeutic testing and improved patient outcome.

Esophageal adenocarcinoma is one of the largest growing cancer types in the United States and the whole world. One of the only known precursors to EAC is Barrett’s Esophagus, the changing of the normal squamous cells which line the esophagus into intestinal cells, following repeated exposure to gastric acids via gastroesophageal reflux disease. There is limited knowledge of the mutations and drivers that contribute to EAC’s low 5-year survival rates, demonstrating a need to identify new therapeutic targets. Given the development of EAC from chronic inflammation and acidic microenvironment, elevated expression of tumor necrosis factor receptor super family member 12A (TNFRSF12A, FN14) and its corresponding ligand, TWEAK, is correlated with disease progression. The functional role of the TWEAK/FN14 signaling axis is well documented in other cancer types, contributing to tumor invasion, migration, and survival. However, reports have shown the TWEAK/FN14 signaling axis can contribute “pro-cancer” and “anti-cancer” phenotypes in different tumor microenvironments. In this study, we seek to demonstrate the functional role of TWEAK and FN14 in EAC survival and migration. We hypothesized TWEAK/FN14 signaling would promoted EAC cell survival and migration. In this study, we illustrate increased expression of FN14 with disease progression. Following treatment with TWEAK, human EAC cell lines had increased sensitivity to standard chemotherapy treatment in vitro. Treatment with TWEAK also correlated with increased cellular migration, most likely in correlation with NF-κB activation. Finally, we showed that inhibition of FN14 via siRNA significantly reduced EAC survival and increased efficacy of standard of care treatments. This data suggests a diverse functional role of the TWEAK/FN14 signaling axis in EAC, and may be a potential target for novel therapeutics.

Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related mortality in the USA and throughout the world. Two phenotypes that promote this deadly outcome are the invasive potential of NSCLC and the emergence of therapeutic resistance in this disease. There is an unmet clinical need to understand the mechanisms that govern NSCLC cell invasion and therapeutic resistance, and to target these phenotypes towards abating the dismal five-year survival of NSCLC. The expression of the tumor necrosis factor receptor superfamily, member 12A (TNFRSF12A; Fn14) correlates with poor patient survival and invasiveness in many tumor types including NSCLC. We hypothesize that suppression of Fn14 will inhibit NSCLC cell motility and reduce cell viability. Here we demonstrate that atorvastatin calcium treatment reduces Fn14 expression in NSCLC cell lines. Prior to Fn14 protein suppression, atorvastatin calcium modulated the expression of the Fn14 modulators P-ERK1/2 and P-NF-κβ. Atorvastatin calcium treatment inhibited the migratory capacity in H1975, H2030 and H1993 cells by at least 55%. When chemotactic migration in H2030 cells was induced by the Fn14 ligand TNF-like weak inducer of apoptosis (TWEAK) treatment, atorvastatin calcium successfully negated any stimulatory effects. Inversely, treatment of NSCLC cells with cholesterol resulted in a statistically significant increase in migration. Depletion of Fn14 expression via siRNA suppressed the migratory effect of cholesterol. Finally, atorvastatin calcium treatment sensitized cells to radiation treatment, reducing cell survival. These data suggest that atorvastatin calcium may inhibit NSCLC invasiveness through a mechanism involving Fn14, and may be a novel therapeutic target in NSCLC tumors expressing Fn14.