Beane-Ebel, Jennifer E.Conrad, Regan D.2025-04-162025https://hdl.handle.net/2144/500962025Lung cancer is the leading cause of cancer deaths globally, but early detection significantly improves survival, as evidenced by the success of low-dose computed tomography (LDCT) in reducing mortality rates. While LDCT identifies lung cancer at earlier stages, further improvements to survival may be achieved by detecting and treating premalignant lesions (PMLs), the abnormal airway changes that precede lung cancer. Not all PMLs progress to invasive cancer; some remain stable or regress, emphasizing the need to understand the mechanisms behind these outcomes. Beyond localized lesions, molecular alterations in the airway field reflect processes occurring throughout the lung, providing a potential avenue for noninvasive biomarker opportunities. Studies show gene expression in bronchial and nasal brushings can distinguish between benign and malignant lung nodules, suggesting airway profiling can aid lung cancer risk assessments. This thesis employs single-cell RNA sequencing (scRNA-seq) and bulk transcriptomic analysis to examine immune and epithelial changes linked to PML severity and to identify molecular shifts in the airway field that may serve as early detection biomarkers and inform prevention strategies. Previous studies identified immune changes associated with PML progression but lacked the resolution needed to determine the contributions of specific immune populations. I analyzed immune cells from endobronchial biopsies across histological grades, identifying 14 subpopulations that differed in abundance between low-grade (LGB) and high-grade (HGB) samples. A cluster of CD4+ T regulatory cells and a subset of plasma cells were enriched in HGB, while multiple CD8+ T cell subpopulations and a neutrophil subpopulation were more abundant in LGB. To examine airway-wide molecular alterations, I analyzed scRNA-seq data from bronchial and nasal brushes along with the biopsies. Nasal brush cells were enriched in gene modules associated with detoxification and mucous production, indicating upper airway specialization. Smoking-related transcriptional changes exhibited a gradient, being most pronounced in bronchial biopsies, followed by bronchial brushes, and least evident in nasal brushes. Notably, some cells from bronchial brush sample clustered with high-grade basal cells and an upregulated gene module in those cells correlated with the most severe lesion histology from bronchial brushes analyzed via bulk RNA-seq. Together, these findings highlight how both immune and epithelial cell populations are altered in PMLs and illustrate how molecular changes throughout the airway reflect the biology of nearby lesions. By leveraging profiling of the immune microenvironment in PMLs and samples collected using noninvasive brush techniques, this work offers new insights into the early stages of lung carcinogenesis and the potential for developing biomarkers to improve PML detection and risk stratification.en-USAttribution 4.0 Internationalhttp://creativecommons.org/licenses/by/4.0/BioinformaticsLung cancer premalignancyLung squamous cell carcinomaSingle cell RNA sequencingThesis/Dissertation2025-04-150000-0002-5855-4415