Abstract 3736: Bridging the Gaps: A Cross-Species Approach to PD-1 and OX40 Combination Immunotherapy

Abstract Monoclonal antibodies (mAbs) targeting immune checkpoints, such as PD-1, PD-L1, and CTLA4, have provided unprecedented clinical benefits to human cancer patients. Novel strategies such as combination therapy with OX40 or 4-1BB agonists are currently under investigation in various pre-clinical studies to enhance the therapeutic benefits of immune checkpoint inhibitors. While syngeneic mouse tumor xenograft models offer valuable insights in pre-clinical testing, they fall short in capturing the nuanced heterogeneity within and between tumors and the dynamic tumor-immune interface. In contrast, canine patients with spontaneous neoplasms possess a closely analogous and intact immune system. These canine patients, who receive sophisticated and advanced medical therapy like their human counterparts, can serve as a vital intermediate pre-clinical model system between rodent studies and human clinical trials, potentially reducing clinical trial failures. In this study, we have developed a novel bispecific fusion protein, denoted BsOXPD, that combines OX40 agonism and PD-1 blockade into a single therapeutic. Our bispecific fusion protein employs an anti-PD-1 nanobody (Nb) to bind and block the PD1 pathway and the extracellular domain (ECD) of the OX40 ligand to stimulate the OX40 receptor. We have developed murinized, caninized, and humanized versions of this protein. All versions of the bispecific fusion protein retain their biochemical and functional properties following the murinization, caninization, and humanization processes. Importantly, all versions effectively disrupt the PD-1/PD-L1 axis in murine, canine, and human systems while concurrently acting as potent OX40 agonists. To further improve the humanized version of the BsOXPD, we partially humanized the anti-PD1 Nb by introducing amino acid substitutions in various framework regions. The humanized anti-PD1 Nb still completely inhibited the binding of human PD-L1 to the PD-1 receptor. The caninized version of BsOXPD exhibits no toxicities or immune-related adverse events (irAEs) following intravenous injection at doses of 1 mg/kg or 3 mg/kg body weight in healthy beagle dogs. In summary, we have designed a cross-species Nb-based fusion protein for PD1 blockade and OX40 agonism. Our next goal is to evaluate the therapeutic efficacy of caninized BsOXPD in pet dogs with oral melanoma in neoadjuvant settings. Therefore, our cross-species approach will offer comprehensive insights into the safety, pharmacokinetic (PK) profile, and clinical benefits associated with the concurrent application of PD1 blockade and OX40 agonism. Citation Format: Maninder Sandey, Damien Ruiz, Jonathan Marable, Deepa Bedi, Payal Agarwal. Bridging the gaps: A cross-species approach to PD-1 and OX40 combination immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3736.