PROJECT SUMMARY/ABSTRACT Esophageal squamous cell carcinoma (ESCC) accounts for over 80% of all cases of esophageal cancer and has a poor prognosis because of a lack of symptoms at early stages. ESCC develops from squamous dysplasia as a typical histologic precursor lesion. Given that early diagnosis of ESCC may lead to better outcomes, understanding the mechanisms of esophageal neoplasia is imperative. However, current ESCC models are not practical for the study of ESCC preneoplasia or neoplasia. A three-dimensional organoid system featuring the physiologic and pathologic processes of organs has drawn tremendous attention in studying stem cells and diseases. Because organoids can be generated from fresh tissues from surgery, biopsy specimens, or frozen tissues, organoids have been proposed as a promising tool for translation into personalized medicine for cancer treatment or biomarker identification. According to the genetic profiling of ESCC, we genetically engineered normal esophageal organoids (EOs). Intriguingly, such genetic manipulations induced the EO hyperplasia, dedifferentiation, cell polarity loss, and nuclear polymorphism, which is comparable to the early lesion of ESCC. Furthermore, syngeneic transplantation of these transformed organoids into mice developed tumors, similar to ESCC. Based on the preliminary results, we hypothesize that the genetic manipulation of EOs recapitulates ESCC initiation. This will be tested by the following two Aims: Aim 1. Determine genetic interaction required for ESCC neoplasia, Aim 2. Dissect cellular and transcriptional network of ESCC neoplasia. Together, a new model for murine esophageal neoplasia will lay the foundation for the study of pathophysiologic mechanisms of human ESCC initiation, which may lead to the improvement of early ESCC detection, prevention, and treatment.