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

Zhengna Yin,Zhuanqin Pan,Huilin Zhang,Zhuming Wang,Hua Tong,Mengyun Gao,Jin Zhu,Yiquan Wang,Yiping Su,Chunping Ye,Sasa Chu

doi:10.18632/oncotarget.21618

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.

Highlights

Receptor-tyrosine-kinase-like Orphan Receptor 1 (ROR1) is a tyrosine-protein kinase transmembrane receptor, which consists of FZ, Ig-like C2type, kringle, and protein kinase domains [1], and functions to regulate cell growth in the central nervous system during development [2, 3]
We demonstrated that ROR1cFab could bind to ROR1 protein and ROR1-positive ovarian cancer A2780 cells
To target ROR1, a previous study reported that treatment of chronic lymphocytic leukemia (CLL) cells with monoclonal antibodies against ROR1 were able to induce apoptosis in tumor cells [22], while knockdown of ROR1 expression suppressed proliferation of lung adenocarcinoma cells [23]

Summary

Introduction

Receptor-tyrosine-kinase-like Orphan Receptor 1 (ROR1) is a tyrosine-protein kinase transmembrane receptor, which consists of FZ (frizzled), Ig-like C2type (immunoglobulin-like), kringle, and protein kinase domains [1], and functions to regulate cell growth in the central nervous system during development [2, 3]. To target ROR1, a previous study reported that treatment of chronic lymphocytic leukemia (CLL) cells with monoclonal antibodies against ROR1 were able to induce apoptosis in tumor cells [22], while knockdown of ROR1 expression suppressed proliferation of lung adenocarcinoma cells [23]. In other malignancies, such as ovarian cancer, ROR1 has been shown to be highly expressed and may serve as a useful therapeutic target [14]. Our previous study demonstrated that expression of ROR1 protein in ovarian cancer tissues was significantly higher than in normal ovary tissues and that ROR1 overexpression was associated with poor disease-free survival and overall survival [9]

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