Protease-activated anti-Lewis Y antibody enhances selectivity and safety of glycan-targeting cancer therapy

Int J Biol Macromol. 2025 May;309(Pt 4):143176. doi: 10.1016/j.ijbiomac.2025.143176. Epub 2025 Apr 15.

Abstract

Lewis Y carbohydrate antigen is overexpressed in various epithelial cancers, making it a potential therapeutic target. However, anti-Lewis Y antibodies have faced challenges due to gastrointestinal toxicity from Lewis Y antigen expression in normal tissues. To mitigate the toxicity, we engineered L-HKM4, a protease-activated anti-Lewis Y antibody, by modifying the N-terminal of HKM4 with an IgG1 hinge domain and a matrix metalloproteinase (MMP)-cleavable linker. This design ensures selective activation in the MMP2/9-rich gastric and colon tumor microenvironment, minimizing off-tumor effects. The results demonstrated that L-HKM4 binding to AGS gastric cancer cells decreased 104.6-fold compared to HKM4, accompanied by a 15.82-fold reduction in antibody-dependent cell-mediated cytotoxicity (ADCC) and no activity in complement-dependent cytotoxicity (CDC), all of binding and cytotoxicity were restored upon MMP2 activation. In mice, L-HKM4 showed minimal gastric tissue binding, confirming reduced on-target toxicity in vivo. In an AGS xenograft model, L-HKM4 achieved tumor-specific activation and inhibited tumor growth by 82 % (P < 0.001) compared to saline control. In conclusion, L-HKM4 enables tumor-specific activation while minimizing gastric toxicity, preserving strong tumor inhibition. This innovative approach overcomes key limitations of anti-Lewis Y therapies, making L-HKM4 a promising and safer treatment for gastrointestinal cancers.

Keywords: Anti-carbohydrate antibody; Cancer therapy; Lewis Y; Matrix metalloproteinase activation; Protease-activated antibody; Tumor selectivity.

MeSH terms

  • Animals
  • Antibody-Dependent Cell Cytotoxicity
  • Cell Line, Tumor
  • Humans
  • Lewis Blood Group Antigens* / immunology
  • Matrix Metalloproteinase 2 / metabolism
  • Mice
  • Peptide Hydrolases* / metabolism
  • Polysaccharides* / metabolism
  • Stomach Neoplasms / drug therapy
  • Stomach Neoplasms / immunology
  • Stomach Neoplasms / pathology
  • Xenograft Model Antitumor Assays

Substances

  • Lewis Blood Group Antigens
  • Lewis Y antigen
  • Polysaccharides
  • Matrix Metalloproteinase 2
  • Peptide Hydrolases