Abstract B058: Antitumor activity and bystander killing effect of HER2 antibody-drug conjugates (ADCs) against tumors with different HER2 expression levels

Abstract Introduction Antibody-drug conjugates (ADCs) are a groundbreaking therapeutic strategy, and human epidermal growth factor receptor 2 (HER2) is a prominent target antigen for ADCs. Trastuzumab emtansine (T-DM1) was the 1st FDA approved HER2 ADC for patients with metastatic HER2-positive breast cancer. Outperforming T-DM1, Trastuzumab deruxtecan (T-DXd) has achieved remarkable success in multiple cancer types and demonstrated surprising efficacy in metastatic breast cancer with variable HER2 expression. To support next-generation HER2 ADC development, we established a preclinical panel of cell lines and cell line-derived xenograft (CDX) with varying HER2 expression levels. Methods CDX models of breast, colorectal, gastric, pancreatic cancer, NSCLC and glioma and a model established with the mixture of NCI-N87 (HER2 positive) and U87MG-Luc (HER2 negative) cells for bystander effect studies were established subcutaneously. HER2 protein levels were determined by Flow cytometry and Immunohistochemistry (IHC) staining. Trastuzumab, IgG-ADC control, T-DXD and T-DM1 were evaluated (3 mg/kg) in these models. in vitro IC50 of T-DXD was assessed in cell lines using CellTiterGlo (CTG) assay. A correlation analysis of T-DXD response status was conducted using efficacy data from CDXs and cell lines. Results Flow cytometry and IHC analysis revealed that HER2 expression levels in CDX tumors were consistent with their corresponding cell lines. In HER2 high expressing (IHC, 3+) models, T-DXD showed potent anti-tumor activities with TGI of 95% in KPL-4 model, 99% in HCC1954 model, 100% in NCI-N87 model and 100% in Calu-3 model. It induced complete tumor regression in 3 out of 8 mice in KPL-4 model, 7 out of 8 mice in HCC1954 model, 6 out of 8 mice in NCI-N87 model and 8 out of 8 mice in Calu-3 model. Compared to T-DXD, T-DM1 showed less potent tumor inhibition with TGI of 77% in KPL-4 model, 90% in HCC1954 model, 78% in NCI-N87 model and 59% in Calu-3 model. However, T-DXD demonstrated less pronounced anti-tumor effects in HER2 low expressing breast cancer (ZR-75-1, IHC, 1+, TGI of 49%), colorectal cancer (HCT-116, IHC, 0, TGI of 50%; HT-29, IHC, 1+, TGI of 55%), and glioma (U87MG, IHC, 0, TGI of 53%) models, comparing to the HER2 high expressing CDX models. Unsurprisingly, T-DM1 was not effective against these models. Consistently, in vitro data demonstrated similar results. Moreover, in vivo assessment of the bystander killing showed that T-DXD, not T-DM1 effectively eliminated U87MG-Luc cells, as evidenced by a marked reduction in luciferase signal in mice injected with a mixture of NCI-N87 and U87MG-Luc cells, despite the Her2-negative status of the U87MG-Luc population. Besides, T-DXD could also lead to tumor regression with TGI of 121% while T-DM1 showed less potency with TGI of 77%. Conclusion This panel of CDX-cell line models provides a robust preclinical platform for investigating mechanisms of action (MOA) such as bystander effect and developing next-generation HER2-targeting therapies. Citation Format: Aaron Li. Hua, Jian Feng, Chenpan Nie, Jun Zhou, Ludovic Bourre, Jessie Jingjing. Wang. Antitumor activity and bystander killing effect of HER2 antibody-drug conjugates (ADCs) against tumors with different HER2 expression levels [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2025 Oct 22-26; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2025;24(10 Suppl):Abstract nr B058.