Antitumor activity of a newly developed monoclonal antibody against ROR1 in ovarian cancer cells

// Zhengna Yin 1, * , Mengyun Gao 1, * , Sasa Chu 2 , Yiping Su 1 , Chunping Ye 1 , Yiquan Wang 3 , Zhuanqin Pan 4 , Zhuming Wang 5 , Huilin Zhang 1 , Hua Tong 1 and Jin Zhu 6 1 Department of Obstetrics and Gynecology, Nanjing Maternity and Child Health Care Hospital, Obstetrics and Gynecology Hospital Affiliated to Nanjing Medical University, Nanjing 210004, China 2 Department of Infectious Disease, Institute of Liver Disease, Nanjing Jingdu Hospital, Nanjing 210002, China 3 Department of Traditional Chinese Internal Medicine, Longhua Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai 200071, China 4 Department of Nursing, Gaoyou People’s Hospital, Yangzhou 225600, China 5 Department of Pathology, Chinese Ministry of Health-designated Key Laboratory of Antibody Technology, Nanjing Medical University, Nanjing 210029, China 6 Huadong Medical Institute of Biotechniques, Nanjing 210002, China * These authors have contributed equally to this work Correspondence to: Hua Tong, email: thua8@163.com Huilin Zhang, email: zhl068@163.com Keywords: ROR1, monoclonal antibody production, chimeric antibody Fab, binding affinity, antitumor activity Received: July 12, 2017     Accepted: August 29, 2017     Published: October 07, 2017 ABSTRACT Receptor-tyrosine-kinase-like Orphan Receptor 1 (ROR1) is a tyrosine-protein kinase transmembrane receptor and ROR1 overexpression is associated with a poor prognosis in various cancers, including ovarian cancer. Targeting of ROR1 has been evaluated as a novel cancer therapy strategy. This study developed a novel chimeric anti-ROR1 Fab antibody (named ROR1-cFab) and then assessed the antitumor activity of this antibody in ovarian cancer cells, an in vitro model of preclinical cancer therapy. A ROR1-cFab prokaryotic expression vector was constructed from positive fusion cells (splenocytes from mice with high ROR1 immune titers were fused with myeloma cells) after three rounds of sub-clone affinity screening. Then, a variety of assays were employed to assess the binding selectivity and specificity of ROR1-cFab to ROR1 protein. Furthermore, CCK8, flow cytometric apoptosis, wound healing, and Transwell migration assays were used to assess antitumor activity of this newly developed anti-ROR1 antibody in ovarian cancer cells. We demonstrated that ROR1-cFab could specifically bind to ROR1 protein and ROR1-positive ovarian cancer A2780 cells. Functional assays revealed that ROR1-cFab inhibited tumor cell proliferation and migration, as well as inducing apoptosis of ROR1-positive A2780 cells in a dose dependent manner. These effects were not observed in ROR1-negative lose386 cells. In conclusion, ROR1-cFab is a novel anti-ROR1 antibody with a high affinity to ROR1 protein and inhibitory effects on ROR1-positive cells. Future studies will determine whether the ROR1-cFab might be a promising candidate for treatment of ROR1-positive ovarian cancer.