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1192 A novel MSLN×4–1BB bispecific antibody for solid tumor
  1. Dayan Zhang1,
  2. Wenting Liu1,
  3. Guodong Shen2,
  4. Weiming Zhou1,
  5. Xiaoli Zeng3 and
  6. Liansheng Cheng1
  1. 1Hefei Hankemab Biotechnology CO., LTD, Anhui, China
  2. 2The First Affiliated Hospital of USTC, Hefei, Anhui, China
  3. 3Hefei Hankemab Biotechnology Co.,LTD, Hefei City, Shushan District, Hong Kong
  • Journal for ImmunoTherapy of Cancer (JITC) preprint. The copyright holder for this preprint are the authors/funders, who have granted JITC permission to display the preprint. All rights reserved. No reuse allowed without permission.


Background 4–1BB (CD137) is not only expressed on the surface of activated T cells and NK cells, but also a marker for Treg cells. Mesothelin (MSLN) is a ~71 kDa cell surface glycoprotein that is rarely expressed in normal tissues but overexpressed in many types of tumors. Here, we developed a bispecific antibody (bsAb) targeting both MSLN and 4–1BB with an intact Fc fragment from human IgG1. It can simultaneously exert the cytotoxic effect of CD8+T cells and and NK cells on tumor cells expressing MSLN to achieve better antitumor efficacy.

Methods HK013-G1 with IgG-scFv format was constructed with IgG1 and its affinity was optimized to making it highly effective in tumor localization. Next, we evaluated the binding activities of HK013-G1 to tumor cells with different MSLN expression levels and its effects on 4–1BB+ cell activation mediated by MSLN-crosslinking. Subsequently, the cell-killing abilities of NK induced by HK013-G1 were quantified. The risk of cytokine release storm (CRS) of HK013-G1 was detected in vitro. And the anti-tumor activity was evaluated in CT26/MSLN tumor model. Finally, PK and safety analysis of HK013-G1 were undertaken in cynomolgus monkeys following intravenous infusion (IV) at 3 or 30 mg/kg.

Results Affinity-optimized HK013-G1 has an order of magnitude greater affinity for MSLN than 4–1BB. HK013-G1 is able to bind different MSLN-expressing cancer cells and bridge MSLN+ cells and 4–1BB+ cells. In luciferase reporter assay, the bsAb-induced 4–1BB activation is dependent on expression level of MSLN. While incubated with CD8+T cells, HK013-G1 increased IFN-γ production only in the presence of MSLN+ cells. Furthermore, HK013-G1 could exert its affects via NK cell. Also, HK013-G1 was shown no stronger ability to inducing CRS in vitro. In CT26-hMSLN tumor models, HK013-G1 showed a dose-dependent anti-tumor activity and more significant growth inhibition effect than HK013-G4. In addition, HK013-G1 could can protect mice against tumor re-challenge. For the IV administration of HK013-G1, it displays prolonged half-life and no CRS and hepatotoxicity were observed in NHP.

Conclusions HK013-G1, a MSLN×4–1BB bsAb with human IgG1 Fc fragment prevents tumor development by killing tumor cells directly via effector functions mediated by NK and cytotoxic T cells. Moreover, HK013-G1 is well tolerated in cynomolgus monkeys. These results show that this bsAb has the potential to develop into a new clinical therapy for cancer types with high-level MSLN expression.

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