Purpose: CD44 is widely expressed on the surface of most tissues and all hematopoietic cells. CD44 plays important roles in cell proliferation, adhesion, and migration. Previously, we developed an anti-CD44 monoclonal antibody, C₄₄Mab-5 (IgG₁, kappa) by immunizing mice with CD44‑overexpressing Chinese hamster ovary (CHO)-K1 cells. In this study, we converted the mouse IgG₁ subclass antibody C₄₄Mab‑5 into an IgG_2a subclass antibody, 5‑mG_2a, and further produced a defucosylated version, 5‑mG_2a‑f and investigate its antitumor activities against oral cancers.
Methods: To generate 5‑mG_2a‑f, appropriate V_H cDNA of mouse C₄₄Mab‑5 and C_H of mouse IgG_2a were subcloned into pCAG‑Neo vector, and light chain of C₄₄Mab‑5 was subcloned into pCAG‑Ble vector. Vectors were transfected into BINDS‑09 (FUT8‑deficient ExpiCHO‑S cells) using the ExpiCHO Expression System. Mouse xenograft models of HSC-2 and SAS (human oral cancer cell lines) were used for examining the antitumor activity.
Results: 5-mG_2a-f demonstrated a sensitive and specific reaction against oral cancer cells in flow cytometry and immunohistochemical analyses. The sensitivity of 5‑mG_2a‑f was similar with that of C₄₄Mab‑5. In vitro analysis demonstrated that 5‑mG_2a‑f showed moderate ADCC and CDC activities against HSC-2 and SAS. 5-mG_2a-f significantly reduced tumor development in HSC-2 and SAS xenografts in comparison to control mouse IgG.
Conclusion: 5‑mG_2a‑f may be a useful antibody-based therapy for patients with CD44-expressing oral cancers.