Department of Anatomy, School of Basic Medical Sciences of Dali University, Dali, China (J.W., K.Y.); State Key Laboratory of Toxicology and Medical Countermeasures, Institute of Pharmacology and Toxicology, Beijing, China (J.W., J.P., Y.Zho., Z.W., N.H., D.Z., G.Q., Y.W., J.F., B.S., L.L.); Joint National Laboratory for Antibody Drug Engineering, the First Affiliated Hospital, School of Medicine, Henan University, Kaifeng, China (J.P.); Hunan Normal University School of Medicine, Changsha, China (Y.Zho., R.Z., G.Q.); JOINN Biologics, Co., Ltd, Beijing, China (J.Z.); and Department of Obstetrics and Gynecology, First Medical Center, General Hospital of Chinese PLA, Beijing, China (Y.Zha.).
Department of Anatomy, School of Basic Medical Sciences of Dali University, Dali, China (J.W., K.Y.); State Key Laboratory of Toxicology and Medical Countermeasures, Institute of Pharmacology and Toxicology, Beijing, China (J.W., J.P., Y.Zho., Z.W., N.H., D.Z., G.Q., Y.W., J.F., B.S., L.L.); Joint National Laboratory for Antibody Drug Engineering, the First Affiliated Hospital, School of Medicine, Henan University, Kaifeng, China (J.P.); Hunan Normal University School of Medicine, Changsha, China (Y.Zho., R.Z., G.Q.); JOINN Biologics, Co., Ltd, Beijing, China (J.Z.); and Department of Obstetrics and Gynecology, First Medical Center, General Hospital of Chinese PLA, Beijing, China (Y.Zha.)
Mol Pharmacol. 2022 Sep;102(3):161-171. doi: 10.1124/molpharm.121.000470. Epub 2022 Jun 28.
Sialic acid-binding Ig-like lectin-15 is an important immunosuppressive molecule considered to be a key target in next-generation tumor immunotherapy. In this study, we screened 22 high-affinity antibodies that specifically recognize human Siglec-15 by using a large human phage antibody library, and five representative sequences were selected for further study. The results showed the binding activity of five antibodies to Siglec-15 (EC ranged from 0.02368 μg/mL to 0.07949 μg/mL), and in two Siglec-15-overexpressed cell lines, three antibodies had the strongest binding activity, so the two clones were discarded for further study. Subsequently, the affinity of three antibodies were measured by bio-layer interferometry technology (5-9 × 10E-09M). As the reported ligands of Siglec-15, the binding activity of Siglec-15 and sialyl-Tn, cluster of differentiation 44, myelin-associated glycoprotein, and leucine-rich repeat-containing protein 4C can be blocked by three of the antibodies. Among these, 3F1 had a competitive advantage. Then, the antibody 3F1 showed an obvious antibody-dependent cell-mediated cytotoxicity effect (EC was 0.85 μg/mL). Further, antibody 3F1 can reverse the inhibitory effect of Siglec-15 on lymphocyte proliferation (especially CD4T and CD8T) and cytokine release Interferon-γ. Given the above results, 3F1 was selected as a candidate for the in vivo pharmacodynamics study. In the tumor model of Balb/c Nude mice, 3F1 (10 mg/kg) showed certain antitumor effects [tumor growth inhibition (TGI) was 31.5%], while the combination of 3F1 (5 mg/kg) and Erbitux (5 mg/kg) showed significant antitumor effects (TGI was 48.7%) compared with the PBS group. In conclusion, novel human antibody 3F1 has antitumor activity and is expected to be an innovative candidate drug targeting Siglec-15 for tumor immunotherapy. SIGNIFICANCE STATEMENT: Siglec-15 is considered as an important target in the next generation of tumor immunotherapy. 3F1 is expected to be the most promising potential candidate for targeting Siglec-15 for cancer treatment and could provide a reference for the development of antitumor drugs.
唾液酸结合免疫球蛋白样凝集素 15 是一种重要的免疫抑制分子，被认为是下一代肿瘤免疫治疗的关键靶点。在这项研究中，我们使用大型噬菌体抗体文库筛选了 22 种特异性识别人 Siglec-15 的高亲和力抗体，并选择了五个代表性序列进行进一步研究。结果显示，五种抗体对 Siglec-15 的结合活性（EC 范围为 0.02368 μg/mL 至 0.07949 μg/mL），在两种 Siglec-15 过表达的细胞系中，三种抗体具有最强的结合活性，因此两种克隆被排除在进一步研究之外。随后，通过生物层干涉技术（5-9×10E-09M）测量三种抗体的亲和力。作为 Siglec-15 的报道配体，三种抗体可以阻断 Siglec-15 与唾液酸化-Tn、CD44、髓鞘相关糖蛋白和富含亮氨酸重复蛋白 4C 的结合活性。其中，3F1 具有竞争优势。然后，抗体 3F1 显示出明显的抗体依赖性细胞介导的细胞毒性效应（EC 为 0.85 μg/mL）。此外，抗体 3F1 可以逆转 Siglec-15 对淋巴细胞增殖（尤其是 CD4T 和 CD8T）和细胞因子释放干扰素-γ的抑制作用。鉴于上述结果，选择 3F1 作为体内药效学研究的候选药物。在 Balb/c Nude 小鼠肿瘤模型中，3F1（10mg/kg）表现出一定的抗肿瘤作用[肿瘤生长抑制（TGI）为 31.5%]，而 3F1（5mg/kg）与 Erbitux（5mg/kg）联合使用时，与 PBS 组相比，表现出显著的抗肿瘤作用（TGI 为 48.7%）。综上所述，新型人源抗体 3F1 具有抗肿瘤活性，有望成为针对 Siglec-15 的肿瘤免疫治疗的创新候选药物。 意义：Siglec-15 被认为是下一代肿瘤免疫治疗的重要靶点。3F1 有望成为针对 Siglec-15 的最有前途的潜在候选药物，为抗肿瘤药物的开发提供参考。
