Preparation and characterization of anti‐tissue factor single‐chain variable fragment antibody for cancer diagnosis

• Published in: Cancer science 2014-12, Vol.105 (12), p.1631-1637
• Main Authors: Sato, Ryuta, Obonai, Toshifumi, Tsumura, Ryo, Tsumoto, Kouhei, Koga, Yoshikatsu, Yasunaga, Masahiro, Matsumura, Yasuhiro
• Format: Article
• Language: English
• Subjects: Affinity; Animals; Antigens; Biochemical characteristics; Blood coagulation; Brain cancer; Brain tumors; Cancer diagnosis; Cell Line, Tumor; Clinical medicine; Cloning; Diagnosis; Female; Flow cytometry; Gastric cancer; Humans; Immunoglobulin G; Labeling; Laboratories; Medical diagnosis; Medical research; Mice; molecular imaging; Monoclonal antibodies; monoclonal antibody; Neoplasms, Experimental; Neuroimaging; Original; Pancreatic cancer; Pancreatic Neoplasms - diagnosis; Peptides; Proteins; R&D; Recombinant DNA; Research & development; Single-Chain Antibodies; single‐chain variable fragment; Skin Neoplasms - diagnosis; Solid tumors; Surface Plasmon Resonance; Thermal cycling; Thromboplastin - metabolism; Tissue factor; Tumors
• ISSN: 1347-9032; 1349-7006
• DOI: 10.1111/cas.12557

Tissue factor (TF), which serves as the initiator of the extrinsic blood coagulation cascade, has been found to be overexpressed in various solid tumors, especially brain tumors, pancreatic cancer, and gastric cancer. Overexpression of TF is considered to contribute to the high incidence of thrombotic complications and poor prognosis in patients with such cancers. Therefore, detection or targeting of TF may be a promising approach for the diagnosis and treatment of solid tumors that are known to overexpress the protein. Here, we used the recombinant DNA technology to develop an anti‐TF single‐chain Fv (scFv) of small size and high affinity for its target. The biochemical characteristics of the anti‐TF scFv were evaluated using surface plasmon resonance (SPR) sensing and flow cytometry. The data obtained showed that the affinity of the anti‐TF scFv was 2.04 × 10−8 (KD), and that the protein showed significant binding to the cancer cells. Then, Alexa 647‐labeled anti‐TF scFv and anti‐TF IgG were administered to mice bearing chemically induced spontaneous tumors. The maximum tumor to background ratios of anti‐TF scFv and anti‐TF IgG were obtained 3 and 24 h after the injections, respectively. This study indicates anti‐TF scFv may be suitable as an imaging probe for the diagnosis of solid tumors. Our anti‐mTF scFv exhibited rapid renal clearance and faster selective intratumor accumulation after the injection. Thus, anti‐TF scFv may be a suitable imaging tool for the diagnosis of solid tumors.