Neuroblastoma (NB) is the most common malignant extracranial solid tumor in children, with poor prognosis, inadequate therapeutic responses, and high recurrence rates. Zinc fingers and homeoboxes protein 2 (ZHX2) functions in various cancers, but its role in NB remains unclear. Retinoic acid (RA) has been used as a pro-differentiation agent in NB, but its limited response rate necessitates novel combinations. This study aims to explore key transcription factors (TFs) in improving RA efficacy and NB prognosis.

ZHX2 was identified as the key TF related to RA efficacy via RNA-seq analysis of RA-treated NB cell lines. Based on TARGET and GEO databases, we analyzed the correlation of ZHX2 with NB prognosis, survival, and clinical characteristics. Effects of ZHX2 knockdown/overexpression on NB cell proliferation, migration, colony formation, and apoptosis were assessed, with confirmation in subcutaneous xenografts. Bioinformatic analysis identified ZHX2’s downstream targets, verified by PCR, WB, CHIP-qPCR, and histopathology. ZHX2’s impact on RA efficacy was evaluated via cell proliferation, apoptosis, cell cycle, differentiation markers, spheroid formation, and in vivo tumor growth.

ZHX2 presented as the key TF that potentially influences the sensitivity of NB to RA. Its expression was lower in NB patients with poor prognostic factors. ZHX2 overexpression inhibited NB malignancy (reduced proliferation, migration, tumor growth; increased apoptosis). Notably, ZHX2 enhanced RA efficacy by transcriptionally repressing CRABP1 to modulate retinoid metabolism, augmenting RA-induced neuronal differentiation and reducing tumor growth in mice.

Transcription factor ZHX2 acts as a tumor suppressor in NB, which enhances RA’s therapeutic effects via CRABP1-mediated retinoid metabolism regulation.