Development of novel monoclonal antibodies with specific binding affinity for denatured human CD26 in formalin-fixed paraffin-embedded and decalcified specimens

Ryo Hatano; Taketo Yamada; Hiroko Madokoro; Haruna Otsuka; Eriko Komiya; Takumi Itoh; Yuka Narita; Satoshi Iwata; Hiroto Yamazaki; Shuji Matsuoka; Nam H Dang; Kei Ohnuma; Chikao Morimoto

doi:10.1371/journal.pone.0218330

Development of novel monoclonal antibodies with specific binding affinity for denatured human CD26 in formalin-fixed paraffin-embedded and decalcified specimens

PLoS One. 2019 Jun 13;14(6):e0218330. doi: 10.1371/journal.pone.0218330. eCollection 2019.

Authors

Affiliations

¹ Department of Therapy Development and Innovation for Immune Disorders and Cancers, Graduate School of Medicine, Juntendo University, Hongo, Bunkyo-ku, Tokyo, Japan.
² Department of Pathology, Saitama Medical University, Moroyama-machi, Iruma-gun, Saitama, Japan.
³ Department of Pathology, Keio University School of Medicine, Shinjuku-ku, Tokyo, Japan.
⁴ Department of Immunological Diagnosis, Juntendo University Graduate School of Medicine, Hongo, Bunkyo-ku, Tokyo, Japan.
⁵ Division of Hematology/Oncology, University of Florida, Gainesville, FL, United States of America.

Abstract

A 110-kDa type II transmembrane glycoprotein with dipeptidyl peptidase IV (DPPIV) activity in its extracellular region, CD26 has a multitude of biological functions and plays an important role in the regulation of inflammatory responses and tumor biology. Our work has focused on CD26 as a novel therapeutic target for various tumors and immune disorders, and we have recently developed a humanized anti-CD26 monoclonal antibody (mAb), YS110, which has promising safety profile and clinical activity in patients with malignant pleural mesothelioma. The development of an anti-human CD26 mAb that can clearly and reliably detect the denatured CD26 molecule in formalin-fixed paraffin-embedded (FFPE) tissues in the clinical setting is therefore of the utmost importance. To develop novel anti-CD26 mAbs capable of binding to denatured CD26, we immunized mice with urea-treated CD26 protein. Hybridoma supernatants were screened for specific reactivity with human CD26 by immunostaining through the use of a set of FFPE human CD26-positive or negative tumor cell lines. This screening method enables us to develop novel anti-human CD26 mAbs suitable for immunohistochemical staining of CD26 in FFPE non-tumor and tumor tissue sections with reliable clarity and intensity. Specifically, these mAbs display strong binding affinity to denatured human CD26 rather than undenatured human CD26, and are capable of detecting denatured human CD26 in decalcified specimens. These novel anti-CD26 mAbs are potentially useful for the analysis of CD26 expression in cancer patients with bony metastasis, and may help decide the appropriateness of YS110 therapy for future cancer patients.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Antibodies, Monoclonal / immunology
Antibodies, Monoclonal, Humanized / genetics
Antibodies, Monoclonal, Humanized / immunology*
Antineoplastic Agents, Immunological / immunology
Cell Line, Tumor
Dipeptidyl Peptidase 4 / analysis
Dipeptidyl Peptidase 4 / immunology*
Dipeptidyl Peptidase 4 / metabolism
Humans
Hybridomas / metabolism
Mice
Paraffin Embedding
Protein Engineering / methods*

Substances

Antibodies, Monoclonal
Antibodies, Monoclonal, Humanized
Antineoplastic Agents, Immunological
DPP4 protein, human
Dipeptidyl Peptidase 4

Grants and funding

This work was supported by a grant of the Ministry of Health, Labour, and Welfare, Japan (Grant Number 150401-01 (C.M.) and 180101-01 (C.M.)), and JSPS KAKENHI Grant Numbers JP16H05345 (C.M.), JP16H04714 (T.Y.), JP18H02782 (K.O.), and JP17K10008 (R.H.). The funders had no role in study desigh, data collection and analysis, decision to publish, or preparation of the manuscript.