Angiogenesis and lymphangiogenesis are some of the routes that cause metastasis. Vascular Endothelial Growth Factors (VEGFs) stimulate angiogenesis and lymphangiogenesis through VEGF receptors. Especially, VEGF-D and its receptor, VEGFR-3, play a pivotal role in regulating cellular processes such as survival, proliferation, and migration, thereby influencing lymphangiogenesis. The aim of this research is to clarify the molecular characteristics of VEGF-D and VEGFR-3 proteins and identify the key residues that are essential for the interaction between VEGF-D and VEGFR-3. Experiments, including size exclusion chromatography and GST pull-down assay analysis, reveal that specific residues, particularly D103 and Q110, are essential for VEGF-D/VEGFR-3 binding. Mutations in these residues induce structural alterations, resulting in reduced binding affinity and impaired activation of VEGFR-3. Moreover, this study suggests that a synthesized peptide, designed based on key residues of VEGF-D involved in binding to VEGFR-3, may act as a metastasis suppressor by competitively inhibiting the interaction between VEGF-D and VEGFR-3. Understanding these molecular interactions is expected to have significant potential to develop therapeutic peptides that can inhibit cancer cell-induced lymphangiogenesis and resolve metastasis via lymphangiogenesis across various cancer types.