The dysregulated intracellular cAMP in the kidneys drives cystogenesis and progression in autosomal dominant polycystic kidney disease (ADPKD). Mounting evidence supports that vasopressin V2 receptor (V2R) antagonism effectively reduces cAMP levels, validating this receptor as a therapeutic target. Tolvaptan, an FDA-approved V2R antagonist, shows limitations in its clinical efficacy for ADPKD treatment. Therefore, the pursuit of better-in-class V2R antagonists with an improved efficacy remains pressing. Herein, we synthesized a set of peptide V2R antagonists. Peptide 33 exhibited a high binding affinity for the V2R (Ki = 6.1 ± 1.5 nM) and an extended residence time of 20 ± 1 min, 2-fold that of tolvaptan. This prolonged interaction translated into sustained suppression of cAMP production in washout experiments. Furthermore, peptide 33 exhibited improved efficacies over tolvaptan in both ex vivo and in vivo models of ADPKD, underscoring its potential as a promising lead compound for the treatment of ADPKD.