Abstract 2781: "Wheelz": A novel engineered human antibody for possible CAR T-cell therapy
Authors: Whitley Kiara V. , Velazquez Edwin J. , Bennion Kelsey B. , Kingery Brianne M. , Weber Scott K. , O'Neill Kim L. 
Published: August 2018

The purpose of this study is to present a novel engineered human antibody specific to thymidine kinase 1 (TK1). Recent developments in chimeric antigen receptor (CAR) T-cell therapeutic approaches have rocketed into the spotlight in the field of cancer immunotherapy. One of the limitations to CAR T-cell therapy is the need for novel targets on cancer cells. The upregulation of serum TK1 has been shown to be an important biomarker in cancer detection and prognosis. Furthermore, we have previously reported the cell surface expression of TK1 on multiple cancer cell lines and in clinical samples. This data merits an investigation of the use of TK1 as a target for CAR T-cells. We have created an engineered human antibody against TK1. We present “Wheelz,” a single chain fragment variable monoclonal antibody with a constant region (scFv-Fc) to potentially target cancer cells expressing surface TK1.

Our engineered antibody was isolated from a yeast display library (donated by the Wittrup lab at MIT) using magnetic sorting and flow cytometry to find the top 0.1% of TK1-binding single chain fragments. We fused the resulting scFv region to a constant region (Fc) fragment through restriction digestion and ligation. We then transformed the construct with a secretion vector into yeast and induced the transformed yeast to produce the antibody, which we called “Wheelz.” We purified the antibody using His-tag. In order to confirm specificity to TK1, we used dot blotting and saw specific binding to purified TK1. We performed a western blot using chemiluminescence imaging. This indicated that the engineered human scFv-Fc binds to TK1 present in normal serum, cancer serum, and cancer cell extract in TK1's monomeric, dimeric, and tetrameric forms and did not bind to any other human cellular proteins.

In the past, anti-TK1 scFv antibodies of murine origin were used in CAR experiments with some cytotoxic effect. Because “Wheelz” is an engineered human antibody, it could improve previous models by minimizing immune rejection in the patient. Future research will explore the cytotoxic-inducing effects of “Wheelz” before inserting the antibody into a CAR. We will also explore the specificity and cytotoxicity of eight other scFv regions isolated from yeast display that preliminarily appear to be specific for TK1. The specific binding of the engineered scFv-Fc antibody to cell-surface biomarker TK1 makes it a promising approach in cancer immunotherapy. “Wheelz” has the potential to expand the repertoire of tumor targets in CAR T-cell therapy.

Citation Format: Kiara V. Whitley, Edwin J. Velazquez, Kelsey B. Bennion, Brianne M. Kingery, Scott K. Weber, Kim L. O'Neill. "Wheelz": A novel engineered human antibody for possible CAR T-cell therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2781.