ATG-125, a novel B7H3 × PD-L1 bispecific antibody-drug conjugate, demonstrates potent antitumor efficacy by dual targeting of immune evasion and direct tumor killing

Background B7-H3 (CD276) and PD-L1 (CD274) are immune checkpoint molecules overexpressed in a wide range of solid tumors, often associated with immune evasion and poor prognosis. While monoclonal antibodies targeting PD-1/PD-L1 have shown clinical success, resistance remains a major challenge. B7-H3 is an emerging, highly promising target for antibody-drug conjugates (ADCs) due to its broad tumor expression and rapid internalization. Co-expression of B7-H3 and PD-L1 on tumor cells provides a rationale for dual targeting to enhance both direct cytotoxicity and immune activation. ATG-125 is a novel bispecific ADC designed to address this opportunity. Methods ATG-125 is a B7-H3 × PD-L1 bispecific ADC (BsADC) engineered from a human IgG1 anti-B7-H3 antibody with C-terminally fused PD-L1-specific scFv. It is conjugated via a cleavable GGFG linker to the DNA topoisomerase I inhibitor, resulting in a drug-to-antibody ratio (DAR) of ~4. Binding affinity, cytotoxicity, internalization, lysosomal trafficking, bystander killing, and T-cell activation were evaluated in solid tumor cell lines with varying expression levels of B7-H3 and PD-L1. The in vivo antitumor efficacy of ATG-125 against the HCC827 human NSCLC model was evaluated in nude mice. Results ATG-125 bound B7-H3 and PD-L1 with nanomolar affinity and demonstrated strong, antigen-dependent internalization in dual- and single-target positive tumor cells, enabling efficient intracellular release of the Dxd payload. ATG-125 exhibited potent, target-dependent cytotoxicity against multiple cancer cell lines (e.g., lung, breast, ovarian) that co-expressing B7-H3 and PD-L1. The parental naked antibody blocked PD1-PDL1 interaction and induced robust IL-2 and IFNgamma production in a Mixed Lymphocyte Reaction (MLR) experiment. The BsADC induced significantly enhanced T-cell activation, as shown by a higher frequency of CD69 + CD3 + T cells in a co-culture with HCC827 cells and human PBMCs. In vivo , ATG-125 induced marked and sustained tumor regression in HCC827 xenograft models at well-tolerated doses. Its efficacy was significantly superior to that of single-target ADC comparators. Conclusion ATG-125 represents a novel bispecific ADC that co-targets B7-H3 and PD-L1 to achieve synergistic IO+ADC antitumor efficacy. Its unique mechanism combines enhanced, dual-target-mediated internalization for superior payload delivery with potential restoration of anti-tumor immunity. The compelling preclinical profile of ATG-125 underscores its significant therapeutic potential and supports its advancement into clinical trials for patients with solid tumors.