Establishment of a novel anti-TROP2 monoclonal antibody TrMab-29 for immunohistochemical analysis

Yusuke Sayama; Mika K Kaneko; Junko Takei; Hideki Hosono; Masato Sano; Teizo Asano; Yukinari Kato

doi:10.1016/j.bbrep.2020.100902

Establishment of a novel anti-TROP2 monoclonal antibody TrMab-29 for immunohistochemical analysis

Biochem Biophys Rep. 2021 Jan 12:25:100902. doi: 10.1016/j.bbrep.2020.100902. eCollection 2021 Mar.

Authors

Yusuke Sayama¹, Mika K Kaneko¹, Junko Takei¹, Hideki Hosono¹, Masato Sano¹, Teizo Asano¹, Yukinari Kato^{1

2}

Affiliations

¹ Department of Antibody Drug Development, Tohoku University Graduate School of Medicine, 2-1 Seiryo-machi, Aoba-ku, Sendai, Miyagi, 980-8575, Japan.
² New Industry Creation Hatchery Center, Tohoku University, 2-1 Seiryo-machi, Aoba-ku, Sendai, Miyagi, 980-8575, Japan.

Abstract

TROP2 is a type I transmembrane glycoprotein originally identified in human trophoblast cells that is overexpressed in several types of cancer. To better understand the role of TROP2 in cancer, we herein aimed to develop a sensitive and specific anti-TROP2 monoclonal antibody (mAb) for use in flow cytometry, Western blot, and immunohistochemistry using a Cell-Based Immunization and Screening (CBIS) method. Two mice were immunized with N-terminal PA-tagged and C-terminal RAP/MAP-tagged TROP2-overexpressed Chinese hamster ovary (CHO)-K1 cells (CHO/PA-TROP2-RAP-MAP), and hybridomas showing strong signals from PA-tagged TROP2-overexpressed CHO-K1 cells (CHO/TROP2-PA) and weak-to-no signals from CHO-K1 cells were selected using flow cytometry. We demonstrated using flow cytometry that the established anti-TROP2 mAb, TrMab-29 (mouse IgG₁ kappa), detected TROP2 in MCF7 breast cancer cell line as well as CHO/TROP2-PA cells. Western blot analysis showed a 40 kDa band in lysates prepared from both CHO/TROP2-PA and MCF7 cells. Furthermore, TROP2 was strongly detected by immunohistochemical analysis using TrMab-29, indicating that TrMab-29 may be a valuable tool for the detection of TROP2 in cancer.

Keywords: ADC, antibody-drug conjugates; ADCC, antibody-dependent cellular cytotoxicity; BSA, bovine serum albumin; Breast cancer; CAR-T, chimeric antigen receptor T-cell; CBIS method; CBIS, Cell-Based Immunization and Screening; CDC, complement-dependent cytotoxicity; CHO, Chinese hamster ovary; DAB, 3,3′-diaminobenzidine tetrahydrochloride; Monoclonal antibody; P3U1, P3X63Ag8U.1; PBS, phosphate-buffered saline; PIT, photoimmunotherapy; PVDF, polyvinylidene difluoride; RIT, radioimmunotherapy; TROP2; TROP2, trophoblast cell-surface antigen; mAb, monoclonal antibody.