Abstract 2376: Transitional phenotype of epithelial progenitor cells in human lung adenocarcinoma premalignancy

Abstract Introduction: Lung adenocarcinoma (ADC) is the most common subtype of non-small cell lung cancer with poor 5-year survival. Interventions during the premalignant stage of carcinogenesis can improve outcomes. Epithelial progenitors are critical cells that maintain and repair the epithelium due to their proliferation and multipotent differentiation. Previously, we discovered diminished function of progenitor cells in the premalignant lung containing ADC. In the current study, we tested whether progenitor dysfunction in premalignancy involves changes in progenitor proliferation and/or differentiation. Methods: Lung tissues resected for early-stage ADC (N=14) were used, and progenitor function was determined using a 3D organoid culture. Lung organoids were grown from the uninvolved (UN, 5-15 cm) and tumor-adjacent lungs (TA, 0.2-2 cm) from the tumor. We hypothesized that processes active in these spatially distinct areas from the tumor-bearing lungs represent oncogenic changes in premalignancy. Organoids were cultured under either proliferation or differentiation conditions, and molecular profiles of organoids were examined by single-cell RNA-sequencing (ScRNA-seq). Results: Analysis of proliferation and differentiation showed an opposing outcome for the UN and TA tissues. Under proliferation conditions, greater numbers of organoids were made by the cells from TA, while under differentiation, UN areas gave rise to higher numbers of organoids (p<0.05 both). Furthermore, proliferative organoids from UN tissues expressed higher numbers of KRT5+/P63+ basal stem cells compared to organoids from TA, while higher numbers of Ki67+ proliferative cells were present in TA organoids (p<0.05 both). In contrast, there was no difference in epithelial cell types present in organoids grown from UN and TA tissues under differentiation conditions. ScRNA-seq comparing transcriptomic profiles of organoids from TA and UN tissues showed that under proliferation, TA organoids contained a specific cluster of basal cells that was enriched for genes belonging to EGFR signaling with higher expression of amphiregulin (AREG), a canonical ligand for EGFR. Immunostaining of organoids from TA and UN tissues confirmed significantly higher numbers of AREG+ cells in TA organoids. The addition of exogenous AREG in proliferative media with or without EGFR inhibitor showed that AREG can drive progenitor cell proliferation independent of EGFR. Conclusions: Our results suggest that progenitor dysfunction in ADC premalignancy involves increased proliferation and decreased differentiation. AREG has been described as a marker of transitional cells involved in cancer development. Thus, our findings indicate that progenitors within the TA tissue express transitional cell phenotype. Future analysis will focus on the AREG-mediated signaling in progenitor cell proliferation during premalignancy. Citation Format: Khosbayar Lkhagvadorj, Bridget K Greuel, Alexis S Assante, Amy B Frey, Simran Randhawa, Adrie VanBokheven, York E Miller, Moumita Ghosh. Transitional phenotype of epithelial progenitor cells in human lung adenocarcinoma premalignancy. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 2376.