Objective: To study the role of peptides in lung adenocarcinoma and its potential molecular mechanisms through peptideomics analysis. Methods: We extracted and LC-MS/MS detected endogenous peptides from 3 pairs of age-matched male lung adenocarcinoma and paracancerous tissues. Using bioinformatics to infer the potential functions of differential peptides. CCK-8 assays, wound-healing assays, and Transwell assays were carried out to verify the effect of key peptides on the malignant phenotype of lung adenocarcinoma cell lines (A549, H1975). Results: A total of 4538 peptides were identified, of which 242 were differentially expressed (fold change≥2.0, FDR<0.05), corresponding to 202 precursor proteins. GO analysis and KEGG pathway enrichment analysis show that these proteins are closely related to a series of important biological activities of tumorigenesis and development. Additionally, we also focused on the novel peptide LACRP2. In vitro experiments confirmed that this peptide can inhibit the proliferation, invasion and migration of lung adenocarcinoma cell lines. Conclusion: The endogenous peptide LACRP2 may be related to the proliferation, invasion and migration of lung adenocarcinoma. Peptidomics can aid in understanding the pathogenesis of lung cancer more comprehensively.