Novel single-domain antibodies against the EGFR domain III epitope exhibit the anti-tumor effect
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Journal of Translational Medicine Open Access
RESEARCH
Novel single‑domain antibodies against the EGFR domain III epitope exhibit the anti‑tumor effect Tao Chen†, Xue Liu†, Haifeng Hong and Henry Wei*
Abstract Background: Monoclonal antibodies (mAbs) have been used for cancer therapy. They are large and have some disadvantages limiting their use. Smaller antibody fragments are needed as their alternatives. A fully human singledomain antibody (sdAb) has a small size of only 15 kDa and consists of only the variable domain of the human antibody heavy chain (VH). It has no immunogenicity. It can easily penetrate into tumor tissues, target an epitope inaccessible to mAb and be manufactured in bacteria for a low cost. Epidermal growth factor receptor (EGFR) is overexpressed in many cancer cells and is a good target for cancer therapy. Methods: The EGFR protein fragment located on the EGFR extracellular domain III was chosen to screen a human sdAb library. Five human anti-EGFR sdAbs were identified. Their specific binding to EGFR was confirmed by ELISA, Western blotting and flow cytometry. Their anti-tumor effects were tested. Results: Five novel fully human anti-EGFR sdAbs were isolated. They specifically bound to EGFR, not to the seven unrelated proteins as negative controls. They also bound to the three different human cancer cell lines, but not to the two cell lines as negative controls. They inhibited cell proliferation, migration and invasion and increased apoptosis of these three cancer cell lines. Two of them were tested for their anti-tumor effect in vivo and showed the anti-tumor activity in a mouse xenograft model for human lung cancer. Immunohistochemical staining of xenograft tumors also showed that their anti-tumor effects were associated with the inhibition of cancer cell proliferation and the promotion of cancer cell apoptosis. Conclusions: This study clearly demonstrated that the anti-EGFR sdAbs could inhibit cancer cell growth in vitro and tumor growth in vivo. They could be potential therapeutics for the treatment of different human cancers. Keywords: EGFR, Cancer, Antibody, Single-domain antibody, Antibody phage library Background Epidermal growth factor receptor (EGFR) family comprises the four homologous members: EGFR or HER1, also known as ERBB1, HER2 or ERBB2, HER3 or ERBB3 and HER4 or ERBB4 [1]. Each member consists of an *Correspondence: [email protected] † Tao Chen and Xue Liu contributed equally to this work Department of Cell Biology and Institute of Biomedicine, National Engineering Research Center of Genetic Medicine, Guangdong Provincial Key Laboratory of Bioengineering Medicine, College of Life Science and Technology, Jinan University, Guangzhou 510632, Guangdong, China
extracellular domain (ECD), a transmembrane domain and an intracellular domain (ICD) [2]. EGFR contains an extracellular domain (EGFR-ECD) of 620 amino acids, a transmembrane domain of 23 amino acids and an intracellular domain (EGFR-ICD) of 501 amino acids. EGFRECD comprises four domains, and the domains I and III
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