Matrix metallopeptidase 7 (MMP7) plays a crucial role in cancer metastasis and progression, making it an attractive target for therapeutic development. However, the development of selective MMP7 inhibitors is challenging due to the conservation of active sites across various matrix metalloproteinases (MMPs). Here, we have developed mirror-image random nonstandard peptides integrated discovery (MI-RaPID) technology to discover innate protease-resistant macrocyclic peptides that specifically bind to and inhibit human MMP7. One identified macrocyclic peptide againstD-MMP7, termedD20, was synthesized in its mirror-image form,D'20, consisting of 12D-amino acids, one cyclicb-amino acid, and a thioether bond. Notably, it potently inhibited MMP7 with an IC50value of 90 nM, and showed excellent selectivity over other MMPswith similar substrate specificity. Moreover,D'20inhibited the migration of pancreatic cell line CFPAC-1, but had no effect on the cell proliferation and viability.D'20exhibited excellent stability in human serum, as well as in simulated gastric and intestinal fluids.This study highlights that MI-RaPID technology can serve as a powerful tool to developin vivostable macrocyclic peptides for therapeutic applications.