FANCI, a member of the Fanconi anemia (FA) complementation group, normally associates with FANCD2 to play an important role in ribosome biogenesis and DNA repair. However, the correlation of FANCI with prognostic value and the molecular mechanism in patients with lung adenocarcinoma (LUAD) remains unclear. In the present study, bioinformatics analysis was performed on LUAD data from TCGA and GEO databases, and further confirmed by in vitro experiments. We found that a high level of FANCI was significantly correlated with a worse survival probability in patients with LUAD. Moreover, the results from in vitro experiments revealed high levels of FANCI in LUAD specimens and cell lines. Knockdown of FANCI expression in A549 and H460 cells significantly inhibited cell viability and clone formation of LUAD cells in vitro and in vivo. Furthermore, high FANCI levels were negatively correlated with a variety of tumor-infiltrating immune cells. Importantly, the overexpression of FANCI significantly inhibited the activation of M1 macrophages. All the data demonstrated that FANCI was a useful prognostic biomarker in patients with LUAD, and knockdown of FANCI inhibited tumor growth of LUAD cells in vitro and in vivo, partly by suppressing the activation of M1 macrophages.