Esophageal squamous cell carcinoma (ESCC) is a common malignancy worldwide with a low survival rate due to a lack of therapeutic targets. Here we found that the mitochondria-related gene GrpE-like 2 (GRPEL2) transcript levels are significantly upregulated in ESCC patient samples, and its high expression predicts poor prognosis. Knockdown of GRPEL2 aggravated suppressed cell proliferation and colony formation. Conversely, overexpression of GRPEL2 promotes ESCC cell proliferation both In Vitro and In Vivo. We delved deeper into the effects of GRPEL2 on mitochondrial function and found that the depletion of GRPEL2 induced mitochondrial dysfunction and cellular apoptosis. Mechanistically, our RNA-Seq analysis revealed that suppression of GRPEL2 expression triggers activation of the MAPK/JNK signaling pathway. Additionally, the apoptosis induced by GRPEL2 loss can be largely reversed by treatment with SP600125, a JNK inhibitor. To further enhance the feasibility of targeting GRPEL2 for inhibiting ESCC proliferation in practical applications, we conducted computer-based drug screening to identify potential GRPEL2 inhibitors. We identified Vandetanib, a known antitumor agent, as a promising molecule that not only exhibits robust binding activity but also effectively reduces GRPEL2 protein levels. In conclusion, the data presented herein implicate GRPEL2 as a pivotal regulator in ESCC, modulating the MAPK/JNK signaling cascade to potentiate apoptosis, thereby offering a specific therapeutic vulnerability for targeting ESCC.© 2025 Wiley Periodicals LLC.