483 Dual blockage of PD-1 and TIGIT with rilvegostomig inhibits tumor growth in various in vitro/in vivo/ex vivo tumor models through effector CD8 activation and immune modulation

<h3>Background</h3> Rilvegostomig is a monovalent, bispecific humanized IgG1 monoclonal antibody that binds to human TIGIT and PD-1 with high affinity. Rilvegostomig has shown promising antitumor activity in adult patients in a phase I/II clinical trial (NCT04995523) and is currently being investigated in two Phase III clinical trials (NCT06109779, NCT06357533). Here, we report in vitro/in vivo/ex vivo antitumor and pharmacodynamic effects of rilvegostomig and its murine surrogate in preclinical cancer models. <h3>Methods</h3> In vitro, CD8-mediated killing of human cancer cell lines was monitored using xCELLigence real time cell analysis when treated with rilvegostomig, or aPD1 and aTIGIT monotherapies/combination. The resulting secretion of IFNg and various analytes was quantified by electrochemiluminescence assays. In vivo antitumor efficacy was determined in the syngeneic flank CT26 colon cancer model and the orthotopic EMT6 breast cancer model treated with rilvegostomig surrogate (mAZD2936) or the valency-matched amount of aPD1 and aTIGIT. Pharmacodynamic effects were evaluated using flow cytometry and single cell RNA (scRNA)seq analysis. The antitumor activity of rilvegostomig and its control arms were evaluated ex vivo using commercially attainable non-small cell lung cancer and head and neck squamous cell carcinoma patient-derived fresh tumor samples. <h3>Results</h3> Compared to isotype, CD8-mediated killing of A375 cancer cells in vitro was enhanced using Rilvegostomig, which was two-fold higher than aPD1 or aTIGIT monotherapy. The corresponding amount of secreted IFNg was directly correlated to the killing activity. In testing three distinct cell lines, we found that the ratio of PDL1 and CD155 expression dictated whether the PD-1 or TIGIT arm of rilvegostomig drove the killing activity. In vivo, we observed enhanced overall response and cure rate following treatment with mAZD2936 or combination treatment compared to aPD-1 or aTIGIT monotherapy. ScRNAseq revealed that mAZD2936 invigorated effector CD8 and reduced the number of exhausted T-cells, which indirectly resulted in the transcriptomic modulation of the myeloid compartment into an antitumoral phenotype. Ex vivo, rilvegostomig induced higher cytokine production and killing activity of human tumors compared to aPD-1. <h3>Conclusions</h3> In preclinical models, rilvegostomig reinvigorated CD8-mediated killing of various cancer cells more effectively than aPD-1 or aTIGIT monotherapy; Rilvegostomig demonstrated antitumor efficacy in both ex vivo NSCLC and HNSCC patient derived tumors. These pre-clinical studies demonstrate the potential of rilvegostomig for the treatment of various solid tumors in support of the ongoing clinical trials. <h3>Acknowledgements</h3> We thank Amy Plotkin, Jennifer Coleman, and the University of Maryland School of Medicine Center for Innovative Biomedical Resources Pathology Biorepository Shared Service (Baltimore, MD), for providing patient NSCLC and HSNCC samples. <h3>Consent</h3> This study was conducted in accordance with international, national, and institutional guidelines as well as all relevant laws and regulations regarding research involving animals. Studies performed in mice received Institutional Animal Care and Use Committee approval under AUC protocol number 22-25. All patient samples were obtained with fully informed consent and their use was approved by AstraZeneca HBS.’