A novel bispecific aptamer LAG3-HER2 enhances T cellmediated cancer immunotherapy against HER2-positive cancer cells

Zhao Yang^*Rui GuoXiaoyang ChenLu YingWenjing ZhangZufa SabeelLianhui ZhaoJian DongYiyang Dong

• Beijing University of Chemical Technology, Beijing, China

In recent years, immunotherapy has become an emerging treatment method following surgery, radiotherapy, and chemotherapy. The study focused on two critical targets: human epidermal growth factor receptor 2 (HER2), a well-established tumor biomarker overexpressed in malignancies such as non-small cell lung cancer and hepatocellular carcinoma, and lymphocyte activation gene 3 (LAG3), an immune checkpoint molecule predominantly expressed on activated T lymphocytes and natural killer cells. Herein, we developed a novel bispecific aptamer (HLB-apt) to investigate its dual-targeting therapeutic potential in regulating tumor-immune cell interactions.: Firstly, the Moe algorithm was used to simulate the docking results between the aptamer and the corresponding protein. Then, the constructed HLB-apt was validated. In addition, based on A549, HepG2 cells and Jurkat cells, the functions and mechanisms of HLB-apt acting simultaneously with A549, HepG2 cells and Jurkat cells were verified through in vivo and in vitro experiments. Results: HLB-apt demonstrated specific binding capacity to both HER2-expressing tumor cells (A549 and HepG2) and LAG3-positive Jurkat cells. Notably, HLB-apt enhanced the killing effect of Jurkat cells on A549 and HepG2 cancer cells, with a killing rate of up to 29.00% for A549 and 7.46% for HepG2. Mechanistically, HLBapt promotes the expression and secretion of IL-2, TNF-α, and granzyme B in activated Jurkat cells, and increases the expression of BAK1, BIM, and BAX in A549 and HepG2 cells. More importantly, HLB-apt significantly inhibited tumor growth in A549 and HepG2 tumor bearing mice and H&E staining revealed no overt histopathological abnormalities in major organs. Conclusion: Our findings demonstrate that HLB-apt exerts dual antitumor effects likely through simultaneously targeting LAG3 on T cells and HER2 on tumor cells, facilitating T cell recruitment and potentially interfering with immune checkpoint pathways. Therapeutic efficacy was markedly enhanced in tumor-bearing mice receiving combined HLB-apt and Jurkat cell administration, This HLB-apt holds promising clinical potential for malignancies characterized by HER2 overexpression.