ALKBH5 promotes the development of lung adenocarcinoma by regulating the polarization of M2 macrophages through CDCA4.

Objective: Lung adenocarcinoma (LUAD) is the most common subtype of lung cancer, with high morbidity and mortality. N6-methyladenosine (m6A) is an important regulator of LUAD progression. Here, we investigated the potential biological functions of ALKBH5 (a m6A demethylated enzyme) and cell division cycle associated protein 4 (CDCA4) in the progression of LUAD.

Methods: The expressions of CDCA4, METTL3, ALKBH5, FTO, YTHDC2 and YTHDC1 mRNA and proteins in LUAD and adjacent tissues, as well as NCI-H1299 and NCI-H157 cells were detected by RT-qPCR and western blot. Meanwhile, the role of ALKBH5 and CDCA4 in macrophage polarization was explored through tumor formation in Lewis lung carcinoma (LLC) mice and the co-culture system of NCI-H1299 and NCI-H157/THP-1 cells. Cell characterization was further analyzed. The expression of Ki-67 in tumor tissue was tested by immunohistochemistry. The scale of M1 and M2 macrophages was determined by flow cytometry.

Results: CDCA4 was significantly overexpressed in NCI-H1299 and NCI-H157 cell lines compared with BEAS-2B cells. The fold enrichment of CDCA4 level in the overexpression (oe)-METTL3 or short hairpin (sh)-ALKBH5 cells was enhanced. Overexpression of CDCA4 promoted the cell viability, proliferation and migration, and inhibited apoptosis, which was reversed by sh-ALKBH5 intervention. Overexpression of YTHDC2 (not YTHDC1) inhibited the effect of CDCA4 on sh-ALKBH5 cells. sh-CDCA4 inhibited tumor growth and weight of LLC cells in mice, and promoted M1/M2 ratio in LLC mice and NCI-H1299/THP-1 and NCI-H157/THP-1 co-culture systems. Oe-CDCA4 promoted the volume and weight of tumor and inhibited the M1/M2 ratio of tumor tissue in LLC mice, but was reversed by sh-ALKBH5 intervention.

Conclusion: m6A demethylase ALKBH5 promotes the development of LUAD through CDCA4 regulation of malignant characterization and M1/M2 macrophage polarization.