The Generation of Dual-Targeting Fusion Protein PD-L1/CD47 for the Inhibition of Triple-Negative Breast Cancer

Triple-negative breast cancer (TNBC) is a highly aggressive subset of breast cancer with limited therapeutic options. However, its immune evasion mechanisms, characterized by the over-expression of the immune checkpoint molecules PD-L1 and CD47, can be targeted in order to facilitate cancer elimination by cells of innate and adaptive immunity. In this paper, we describe the design, preparation, and evaluation of three novel dual-targeting fusion proteins that were based on the structure frame of prototype IAB (innate and adaptive dependent bispecific fusion protein) and the Orcutt-type IgG-scFv molecular model. Three molecules with different spatial conformations were designed to improve antigen-antibody affinity by the addition of Ag-Ab binding sites from the variable region sequences of the anti-PD-L1 monoclonal antibody (mAb) atezolizumab and CV1, a high-affinity receptor of CD47. The results showed that the best-performing among the three proteins designed in this study was protein Pro3; its CV1 N-terminus and Fc domain C-terminus were not sterically hindered. Pro3 was better at boosting T cell proliferation and the engulfment of macrophages than the IAB prototype and, at the same time, retained a level of ADCC activity similar to that of IAB. Through improved design, the novel constructed dual-targeting immunomodulatory protein Pro3 was superior at activating the anti-tumor immune response and has thus shown potential for use in clinical applications.

Automated summary

In this study, three novel dual-targeting fusion proteins were designed and constructed to enhance antigen-antibody affinity. Among these proteins, Pro3 showed the best performance in activating the anti-tumor immune response. Pro3 demonstrated superior T cell proliferation and macrophage engulfment compared to the prototype IAB, while retaining comparable ADCC activity. The improved design of the dual-targeting immunomodulatory protein Pro3 has potential for clinical applications.