Anti‑EGFR monoclonal antibody 134‑mG2a exerts antitumor effects in mouse xenograft models of oral squamous cell carcinoma

Abstract

The epidermal growth factor receptor (EGFR), a transmembrane receptor and member of the human epidermal growth factor receptor (HER) family of receptor tyrosine kinases, is a critical mediator of cell growth and differentiation. EGFR forms homo- or heterodimers with other HER family members to activate downstream signaling cascades in a number of cancer cells. In a previous study, the authors established an anti-EGFR monoclonal antibody (mAb), EMab-134, by immunizing mice with the ectodomain of human EGFR. EMab-134 binds specifically to endogenous EGFR and can be used to detect receptor on oral cancer cell lines by flow cytometry and western blot analysis; this antibody is also effective for the immunohistochemical evaluation of oral cancer tissues. In the present study, the subclass of EMab-134 was converted from IgG 1 to IgG 2a (134-mG 2a ) to facilitate antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC). The dissociation constants ( K D s) of EMab-134 and 134-mG 2a against EGFR-expressing CHO-K1 (CHO/EGFR) cells were deter-mined by flow cytometry to be 3.2×10 −9 M and 2.1×10 −9 M, respectively; these results indicate that 134-mG 2a has a higher binding affinity than EMab-134. The 134-mG 2a antibody was more sensitive than EMab-134 with respect to antigen detection in oral cancer cells in both western blot analysis and immunohistochemistry applications. Analysis in vitro revealed that 134-mG 2a contributed to high levels of ADCC and CDC in experiments targeting CHO/EGFR, HSC-2, and SAS cells. Moreover, the in vivo administration of 134-mG 2a significantly inhibited the development of CHO/EGFR, HSC-2, and SAS mouse xenografts in comparison to the results observed in response to EMab-134. Taken together, the findings of the present study demonstrate that the newly-formulated 134-mG 2a is useful for detecting EGFR by flow cytometry, western blot analysis and immunohistochemistry. Furthermore, the in vivo results suggested that it may also be useful as part of a therapeutic regimen for patients with EGFR-expressing oral cancer.

Defucosylated mouse‑dog chimeric anti‑HER2 monoclonal antibody exerts antitumor activities in mouse xenograft models of canine tumors

“…Canine mononuclear cells (MNCs) obtained from Yamaguchi University were resuspended in DMEM (Nacalai Tesque, Inc.) with 10% FBS and were used as effector cells (37,38,42). Target cells (CHO-K1, CHO/dHER2, and SNP) were labeled with 10 µg/ml Calcein AM (Thermo Fisher Scientific, Inc.) (31,(39)(40)(41)(43)(44)(45)(46)(47)(48)(49)(50)(51)(52)(53). The target cells (2x10 4 cells) were plated in 96-well plates and mixed with effector canine MNCs (effector/target cells ratio, 50), 100 µg/ml of H77Bf or control dog IgG.…”

“…CDC of H77Bf. Target cells (CHO-K1, CHO/dHER2, and SNP) were labeled with 10 µg/ml Calcein AM (31,(39)(40)(41)(43)(44)(45)(46)(47)(48)(49)(50)(51)(52)(53). The target cells (2x10 4 cells) were plated in 96-well plates and mixed with rabbit complement (final dilution 1:10; Low-Tox-M Rabbit Complement; Cedarlane Laboratories,) and 100 µg/ml of control dog IgG or H77Bf.…”

Defucosylated Mouse–Dog Chimeric Anti-EGFR Antibody Exerts Antitumor Activities in Mouse Xenograft Models of Canine Tumors

“…Additional antibody inoculations were performed on days 14 and 21. This regimen was selected based on prior studies [ 15 , 30 ]. Canine mononuclear cells, which were obtained from Yamaguchi University, were injected surrounding the tumors on days 7, 14, and 21.…”

“…EMab-134 can be used in Western blotting, flow cytometry, and immunohistochemistry. The mouse IgG 2a version of EMab-134 (134-mG 2a ) demonstrates antitumor activities in mouse xenograft models of hEGFR-expressing oral squamous cell carcinoma [ 15 ]. In addition, we produced the defucosylated version of 134-mG 2a (134-mG 2a -f) to augment antibody-dependent cellular cytotoxicity (ADCC) [ 16 ].…”

Current Targeted Therapy for Metastatic Colorectal Cancer

“…To date, we have developed several cancer-specific monoclonal antibodies, including anti-EGFR antibodies using the CasMab method [117][118][119][120][121][122][123][124][125][126][127]. Currently, we are trying to further develop these antibodies for the above-mentioned antibody-based therapy.…”