Mazzilli, SarahDuffy, ElizabethKirkaldy, Cailyn J.2025-10-012025-10-012024https://hdl.handle.net/2144/513862024Lung cancer is the third most diagnosed and deadly cancer in the United States annually, with lung squamous cell carcinoma (LUSC) being heavily associated with smokers. Prognosis declines with the stage of lung cancer at diagnosis, with stage IA patients having a 50% 5-year survival rate and patients diagnosed with stage IIIA lung cancer having only a 19% 5-year survival. While diagnosis and screening protocols have improved survival rates and our understanding of cancer through projects like The Cancer Genome Atlas, focus has shifted toward detecting lung cancer before it becomes established. Premalignant lesions arise before becoming invasive cancer, and can either regress to normal tissue or progress through histological grades before becoming carcinoma in situ and established lung carcinoma. Previous studies focused on LUSC premalignancy within the section of Computational Biomedicine at Boston University’s Chobanian and Avedisian School of Medicine have used genomic and transcriptomic sequencing on patient biopsies to identify common mutations and subsequent transcriptomic changes found in bronchial premalignant lesions. In 41% of the biopsies taken from patients, there are mutations in the NOTCH1 gene. NOTCH is a membrane protein used for cell signaling, and is particularly important for tissue development and injury repair. The NOTCH pathway maintains resident stem cell populations, prompting them to regenerate and differentiate when necessary. The role of NOTCH in tumorigenesis is complex; in some cancers NOTCH mutations are a driver of cancer and in others a tumor suppressor. In this thesis, small molecule inhibitors were used to inhibit NOTCH1 in order to understand the role of NOTCH1 modulation in the bronchial epithelium. Three aims were devised to study the role of NOTCH1 in vitro using bronchial epithelium cell lines. The first aim sought to establish the sensitivity and effectiveness of small molecule NOTCH1 inhibitors in immortalized bronchial epithelial cells to establish the dosages used for future experimentation. Using qRT-PCR, it was observed that 5 uM doses of small molecule inhibitors effectively inhibit NOTCH1 in immortalized bronchial epithelial cells. The second aim focused on the consequences of NOTCH1 modulation on cell growth which was inhibited by the 5 uM dose of small molecule inhibitors NADI-351 and DAPT, but not Crenigacestat. Finally, primary bronchial epithelial cells were cultured at the air-liquid interface to stimulate differentiation and observe change in cell populations when NOTCH1 was inhibited. Primary bronchial epithelial cells grown at the air-liquid interface with 1 uM treatments had noticeable differences in tissue size and cell differentiation in the two γ-secretase inhibitors, Crenigacestat and DAPT.en-USBiologyThesis/Dissertation2025-09-30