Corneal wound healing is crucial for maintaining visual acuity and overall ocular health. To address current therapeutic limitations, we developed a novel cell-penetrating peptide (CPP), specifically aimed at enhancing corneal wound healing. This study presents identification and evaluation of properties of the novel peptide, HP-1, particularly assessing its efficacy in promoting corneal epithelial cell migration and wound closure. The CPP design was guided by computational molecular dynamics and enhanced sampling techniques to assess its interaction with the cellular membrane. Preliminary simulations revealed favorable binding interactions with a cornea-mimicking lipid bilayer, indicating its potential for membrane penetration. In-vitro cell uptake studies were performed using Human Corneal Epithelial (HCE) cells to validate CPP internalization, confirming its internalization ability. Subcellular localization assays were conducted to elucidate the intracellular trafficking of the CPP upon internalization. The data indicated preferential localization in the endosome and nucleus. Functional activity was assessed through in-vitro scratch assays on HCE cell monolayers, demonstrating accelerated cell migration and wound closure. This highlight HP-1’s potential as a therapeutic agent for corneal wound repair. Furthermore, we evaluated its effectiveness in mitigating chemical-induced corneal injury using an in-vitro chemical injury assay, emphasizing its protective role in such injuries. Gene expression analysis using quantitative real-time PCR was performed to gain insights into the underlying molecular mechanisms of the peptide's action. HP-1 treatment upregulated critical genes involved in cell migration, proliferation, and ECM remodeling, supporting its potential as a promising therapeutic strategy for corneal wound healing. Our findings demonstrate favorable membrane interaction, efficient cellular uptake, and enhanced wound healing effects of our novel therapeutic CPP. An in-depth investigation of the  molecular mechanisms of the peptide at in vivo level will provide valuable insights into its potential as a therapeutic agent for corneal wound healing, offering promising prospects for future clinical translation.