<p>Tamibarotene, a synthetic retinoid used in the treatment of acute promyelocytic leukemia, has been reported to induce differentiation in the SH-SY5Y cell line into neurons. However, the underlying mechanisms remain unclear. This study aimed to determine the optimal concentration of Tamibarotene (Am80) for promoting neuronal differentiation and to elucidate the underlying molecular mechanisms. SH-SY5Y cells were treated with Am80 at various concentrations, and the effects on cell morphology, gene expression, cell proliferation and apoptosis assessed using immunofluorescence, Western blotting, qPCR, and RNA sequencing. Results indicated that that 1µM Am80 effectively promoted neuronal differentiation, upregulating neuronal markers and the <i>KCNT1</i> gene, while downregulating tumor-related genes <i>MYC</i> and <i>CXCR4</i>. The differentially expressed genes are predominantly enriched in the PI3K-Akt signaling pathway, with upregulation of genes related to neuronal development such as <i>NTRK2</i>, <i>RET</i>, and <i>CNR1</i>, and downregulation of tumor-related genes including <i>MYC</i> and <i>CXCR4</i>. Inhibition of the PI3K/Akt signaling pathway using LY294002 resulted in a decreased efficacy of AM80-induced differentiation in SH-SY5Y cells, along with downregulation of neuronal marker expression. These findings suggest that Am80 can effectively promote the differentiation of SH-SY5Y cells into neurons and reduce the proliferation of neuroblastoma cells, which is related to the PI3K/AKT pathway, providing a good model for the study of nervous system diseases.</p> Graphical abstract <p></p>