DDX3X promotes ILF3 stability to accelerate neuroblastoma progression via M2 macrophage polarization

Neuroblastoma (NB) is a common pediatric solid tumor with poor prognosis, in which macrophage polarization is increasingly recognized as a key driver of progression. However, the roles of RNA-binding proteins such as DEAD-box helicase 3 X-linked (DDX3X) and interleukin enhancer-binding factor 3 (ILF3) in this process remain unclear. This study aims to explore the role of the DDX3X-ILF3 axis in modulating macrophage polarization and promoting the malignant progression of NB. ILF3 expression was analyzed by RT-qPCR and Western blotting. Lentiviral-mediated knockdown of ILF3 or DDX3X was performed, followed by evaluation of cell viability, proliferation, invasion, and migration, respectively. In a Transwell co-culture system, the expression of polarization markers was examined by Western blotting, and the secretion of inflammatory cytokines was measured by enzyme-linked immunosorbent assay. Actinomycin D treatment combined with RT-qPCR was used to assess ILF3 mRNA stability. RIP-qPCR was employed to explore the interaction of DDX3X with ILF3 mRNA. NB xenograft and metastasis mouse model (lung, liver, and kidney) was established to evaluate the regulatory effects of the DDX3X inhibitor RK-33 and ILF3 overexpression on NB growth and metastasis. ILF3 expression was significantly upregulated in NB cells. Knockdown of ILF3 or DDX3X suppressed NB cell viability, proliferation, invasion, and migration, while promoting M1 macrophage polarization accompanied by increased secretion of inflammatory cytokines. Silencing DDX3X reduced ILF3 mRNA stability and protein expression, whereas ILF3 overexpression negated the suppressive impacts of DDX3X knockdown on malignant behaviors and polarization status. In vivo, DDX3X inhibitor, RK-33, markedly inhibited NB tumor growth and metastasis and enhanced M1 macrophage polarization, which was partially reversed by ILF3 overexpression. DDX3X promotes NB progression by stabilizing ILF3 mRNA, thereby facilitating M2 macrophage polarization.