Asano Teizo, Suzuki Hiroyuki, Kaneko Mika K, Kato Yukinari
Department of Antibody Drug Development and Tohoku University Graduate School of Medicine, Sendai, Japan.
Department of Molecular Pharmacology, Tohoku University Graduate School of Medicine, Sendai, Japan.
Monoclon Antib Immunodiagn Immunother. 2021 Oct;40(5):227-232. doi: 10.1089/mab.2021.0030.
Podocalyxin (PODXL) is a type I transmembrane sialoglycoprotein that is overexpressed in human cancers, including breast, oral, and lung. PODXL promotes tumor progression, and its expression is associated with poor prognosis. Since PODXL is expressed in normal cells, including kidney podocytes and vascular endothelial cells (VECs), cancer-specific monoclonal antibodies (mAbs) are necessary to reduce the adverse effects of antibody therapy on PODXL-expressing cancers. Previously, we established a cancer-specific mAb against PODXL, PcMab-60 (mouse IgM, kappa), by immunizing mice with soluble PODXL produced by LN229 glioblastoma cells. PcMab-60 reacted with PODXL-expressing cancer cells, but did not react with VECs. In this study, we investigated an epitope of PcMab-60 using flow cytometry, surface plasmon resonance (SPR), and enzyme-linked immunosorbent assay (ELISA). The results of SPR revealed that the PcMab-60 epitope consisted of Thr105, Arg109, Gly110, Gly111, Gly112, Ser113, Gly114, Asn115, Pro116, and Thr117. In contrast, the results of ELISA revealed that the PcMab-60 epitope consisted of Arg109, Gly110, Gly111, Gly112, Ser113, Gly114, Asn115, and Pro116. These results demonstrate the cancer-specific epitope, which was recognized by PcMab-60.
足细胞毒素（PODXL）是一种I型跨膜唾液酸糖蛋白，在包括乳腺癌、口腔癌和肺癌在内的人类癌症中过度表达。PODXL促进肿瘤进展，其表达与不良预后相关。由于PODXL在包括肾足细胞和血管内皮细胞（VECs）在内的正常细胞中表达，因此需要癌症特异性单克隆抗体（mAbs）来减少抗体治疗对表达PODXL的癌症的不良反应。此前，我们通过用LN229胶质母细胞瘤细胞产生的可溶性PODXL免疫小鼠，建立了一种针对PODXL的癌症特异性单克隆抗体PcMab-60（小鼠IgM，κ）。PcMab-60与表达PODXL的癌细胞反应，但不与VECs反应。在本研究中，我们使用流式细胞术、表面等离子体共振（SPR）和酶联免疫吸附测定（ELISA）研究了PcMab-60的表位。SPR结果显示，PcMab-60表位由苏氨酸105、精氨酸109、甘氨酸110、甘氨酸111、甘氨酸112、丝氨酸113、甘氨酸114、天冬酰胺115、脯氨酸116和苏氨酸117组成。相比之下，ELISA结果显示，PcMab-60表位由精氨酸109、甘氨酸110、甘氨酸111、甘氨酸112、丝氨酸113、甘氨酸114、天冬酰胺115和脯氨酸116组成。这些结果证明了PcMab-60识别的癌症特异性表位。
