CD38 和 CD47 在许多血液恶性肿瘤中表达，包括多发性骨髓瘤 (MM)、B 细胞非霍奇金淋巴瘤 (NHL)、B 细胞急性淋巴细胞白血病 (ALL) 和 B 细胞慢性淋巴细胞白血病 (CLL)。在这里，我们评估了 CD38/CD47 双特异性抗体 (BsAb) 的抗肿瘤活性。使用 2 2“mAb-trap”平台开发了五种合适的抗 CD38 抗体，用于共同靶向 CD47 和 CD38 BsAb。利用体外和体内系统评估CD38/CD47 BsAb的活性特征。利用杂交瘤筛选技术，我们获得了9种合适的抗CD38抗体。所有抗CD38抗体均与CD38肿瘤细胞结合，并通过抗体依赖性细胞毒性（ADCC）和抗体依赖性细胞吞噬作用（ADCP）杀死肿瘤细胞。选择五种抗 CD38 抗体（4A8、12C10、26B4、35G5 和 65A7）用于使用“mAb-trap”平台设计 CD38/CD47 BsAb (IMM5605)。 BsAb 对 CD38 靶点的亲和力和结合活性高于对 CD47 靶点的亲和力和结合活性，从而降低了潜在的靶向潜力和脱瘤效应。 CD38/CD47 BsAb 不与红细胞结合，不诱导红细胞凝集；因此，BsAb 的血液毒性要低得多。 CD38/CD47 BsAb 比 IMM01（SIRPα Fc 融合蛋白）具有更强的阻断 CD38 /CD47 肿瘤细胞中 CD47/SIRPα 信号的能力。通过Fc结构域工程，CD38/CD47 BsAbs被证明可以通过诱导ADCC和ADCP更有效地杀死肿瘤。 IMM5605-26B4对细胞CD38酶活性具有最强的抑制作用。 IMM5605-12C10直接诱导肿瘤细胞凋亡的能力最强。抗CD38抗体26B4与SIRPα-Fc融合蛋白结合显示出很强的抗肿瘤作用，在NCI-H929细胞异种移植模型中优于任何单独使用的单一治疗药物。 CD38/CD47 BsAbs表现出很强的抗肿瘤作用；具体来说，IMM5605-12C10 有效地根除所有小鼠中所有已形成的肿瘤。基于“mAb-tarp”平台开发的一组针对 CD38 和 CD47 的双特异性抗体在体外和体内均显示出强大的肿瘤杀伤能力。由于 BsAb 与 CD47 的结合亲和力较低，与 CD38 的结合亲和力较高，与 RBC 的结合没有亲和力，并且不会诱导 RBC 凝集，因此我们得出结论，CD38/CD47 BsAb 是安全的，并且具有令人满意的耐受性。版权所有 © 2024李、陈、杨、郭、刘、孙、白、王、李、李、杨、张、张、赵、彭、刘、涂、张、田。

CD38 and CD47 are expressed in many hematologic malignancies, including multiple myeloma (MM), B-cell non-Hodgkin lymphoma (NHL), B-cell acute lymphoblastic leukemia (ALL), and B-cell chronic lymphocytic leukemia (CLL). Here, we evaluated the antitumor activities of CD38/CD47 bispecific antibodies (BsAbs).Five suitable anti-CD38 antibodies for co-targeting CD47 and CD38 BsAb were developed using a 2 + 2 "mAb-trap" platform. The activity characteristics of the CD38/CD47 BsAbs were evaluated using in vitro and in vivo systems.Using hybridoma screening technology, we obtained nine suitable anti-CD38 antibodies. All anti-CD38 antibodies bind to CD38+ tumor cells and kill tumor cells via antibody-dependent cellular cytotoxicity (ADCC) and antibody-dependent cellular phagocytosis (ADCP). Five anti-CD38 antibodies (4A8, 12C10, 26B4, 35G5, and 65A7) were selected for designing CD38/CD47 BsAbs (IMM5605) using a "mAb-trap" platform. BsAbs had higher affinity and binding activity to the CD38 target than those to the CD47 target, decreasing the potential on-target potential and off-tumor effects. The CD38/CD47 BsAbs did not bind to RBCs and did not induce RBC agglutination; thus, BsAbs had much lower blood toxicity. The CD38/CD47 BsAbs had a greater ability to block the CD47/SIRPα signal in CD38+/CD47+ tumor cells than IMM01 (SIRPα Fc fusion protein). Through Fc domain engineering, CD38/CD47 BsAbs were shown to kill tumors more effectively by inducing ADCC and ADCP. IMM5605-26B4 had the strongest inhibitory effect on cellular CD38 enzymatic activity. IMM5605-12C10 had the strongest ability to directly induce the apoptosis of tumor cells. The anti-CD38 antibody 26B4 combined with the SIRPα-Fc fusion proteins showed strong antitumor effects, which were better than any of the mono-therapeutic agents used alone in the NCI-H929 cell xenograft model. The CD38/CD47 BsAbs exhibited strong antitumor effects; specifically, IMM5605-12C10 efficiently eradicated all established tumors in all mice.A panel of BsAbs targeting CD38 and CD47 developed based on the "mAb-tarp" platform showed potent tumor-killing ability in vitro and in vivo. As BsAbs had lower affinity for binding to CD47, higher affinity for binding to CD38, no affinity for binding to RBCs, and did not induce RBC agglutination, we concluded that CD38/CD47 BsAbs are safe and have a satisfactory tolerability profile.Copyright © 2024 Li, Chen, Yang, Guo, Liu, Sun, Bai, Wang, Li, Li, Yang, Zhang, Zhang, Zhao, Peng, Liu, Tu, Zhang and Tian.