Data-independent Acquisition Mass Spectrometry Reveals Exosomal LAMC1 as a Key Determinant of Lung Adenocarcinoma Radiosensitivity, Independent of EGFR Mutation.

Aim: This study is formulated to reveal the variables affecting the radio-sensitivity in lung adenocarcinoma (LUAD).

Background: LUAD patients show varied radiotherapy responses. While epidermal growth factor receptor (EGFR) mutations are often used to predict sensitivity, their reliability is debated, underscoring the need for better biomarkers.

Objective: The aim of this study was to identify key functional proteins that regulate the sensitivity of LUAD to radiotherapy and to assess the potential value of exosomal LAMC1 as a clinical predictive marker.

Method: In this study, 103 LUAD patients receiving concurrent radiotherapy were included to assess the relationship between EGFR mutation and survival. Intrinsic radio-sensitivity and different radio-sensitivities in 14 LUAD cell lines with/out EGFR mutation were examined based on the surviving fraction at 2 Gy (SF2). Dataindependent acquisition (DIA) and Liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomics were used to investigate the proteomics of the LUAD cell lines. Subsequently, GO/KEGG enrichment analysis was combined with protein-protein interaction (PPI) network screening for key proteins. Nano-flow cytometry was employed to validate changes in radiosensitivity-associated protein expression within exosomes, while siRNA-mediated knockdown was performed to assess the functional impact of specific proteins on LUAD cells.

Results: EGFR mutations were not significantly associated with PFS/OS. 14 LUAD cell lines displayed intrinsic variations in SF2, and no difference between the EGFR mutation and wild-type groups was reported. 5425 proteins were identified via DIA in 14 LUAD cell lines. After bio-informatics analysis, LAMC1, ITGB4, ITGA6, and CD44 were the most representative core differential proteins for the radio-sensitivity in LUAD cells. Notably, LAMC1 was confirmed as a radiation-resistant protein. Following radiotherapy, LUAD cells secreted exosomes with reduced LAMC1 levels. Moreover, LAMC1 knockdown significantly affected cellular proliferation and apoptosis post-irradiation.

Conclusion: LAMC1 serves as a critical functional determinant of radiotherapy resistance in LUAD. Its dynamic changes in exosomes demonstrate potential for predicting radiotherapy response, suggesting clinical utility for radiosensitivity assessment and personalized radiotherapy guidance.