Establishment of an antibody specific for cancer-associated haptoglobin: a possible implication of clinical investigation

// Kimihiro Nishino 1, 2, * , Sayaka Koda 1, * , Naoya Kataoka 1, * , Shinji Takamatsu 1 , Miyako Nakano 3 , Shun Ikeda 1 , Yuka Kamamatsu 1 , Koichi Morishita 1 , Kenta Moriwaki 1 , Hidetoshi Eguchi 4 , Eiko Yamamoto 2, 5 , Fumitaka Kikkawa 2 , Yasuhiko Tomita 6 , Yoshihiro Kamada 1 and Eiji Miyoshi 1 1 Department of Molecular Biochemistry and Clinical Investigation, Osaka University Graduate School of Medicine, Suita, Osaka, Japan 2 Department of Obstetrics and Gynecology, Nagoya University Graduate School of Medicine, Nagoya, Aichi, Japan 3 Graduate School of Advanced Sciences of Matter, Hiroshima University, Higashihiroshima, Hiroshima, Japan 4 Department of Gastroenterological Surgery, Osaka University Graduate School of Medicine, Suita, Osaka, Japan 5 Department of Healthcare Administration, Nagoya University Graduate School of Medicine, Nagoya, Aichi, Japan 6 Department of Pathology, International University of Health and Welfare, Narita, Chiba, Japan * These authors contributed equally to this work Correspondence to: Eiji Miyoshi, email: emiyoshi@sahs.med.osaka-u.ac.jp Keywords: fucosylation; glycosylation; haptoglobin; glycan antibody; cancer biomarker Received: May 24, 2017 Accepted: January 20, 2018 Published: January 29, 2018 ABSTRACT We previously found that the serum level of fucosylated haptoglobin (Fuc-Hpt) was significantly increased in pancreatic cancer patients. To delineate the mechanism underlying this increase and develop a simple detection method, we set out to generate a monoclonal antibody (mAb) specific for Fuc-Hpt. After multiple screenings by enzyme-linked immunosorbent assay (ELISA), a 10-7G mAb was identified as being highly specific for Fuc-Hpt generated in a cell line as well as for Hpt derived from a pancreatic cancer patient. As a result from affinity chromatography with 10-7G mAb, followed by lectin blot and mass spectrometry analyses, it was found that 10-7G mAb predominantly recognized both Fuc-Hpt and prohaptoglobin (proHpt), which was also fucosylated. In immunohistochemical analyses, hepatocytes surrounding metastasized cancer cells were stained by the 10-7G mAb, but neither the original cancer cells themselves nor normal hepatocytes exhibited positive staining, suggesting that metastasized cancer cells promote Fuc-Hpt production in adjacent hepatocytes. Serum level of Fuc-Hpt determined with newly developed ELISA system using the 10-7G mAb, was increased in patients of pancreatic and colorectal cancer. Interestingly, dramatic increases in Fuc-Hpt levels were observed at the stage IV of colorectal cancer. These results indicate that the 10-7G mAb developed is a promising antibody which recognizes Fuc-Hpt and could be a useful diagnostic tool for detecting liver metastasis of cancer.