Abstract 2282: Discovery and characterization of novel TNFR2 antibodies to modulate T cell activities in immunosuppressive environment
Authors: Wei Shuo , Yang Guang , Li Juying , Zhang Qian , Delince Matthieu , Rose Greg , Ortega Charina , Mary Pascaline , Zhou He , Lee Dean , Kumar Roshan , Beq Stephanie , Beltraminelli Nicola , Adrian Francisco , Schweizer Liang 
Published: September 2020

Abstract
TNFR2 has recently emerged as a promising therapeutic target for Immuno-Oncology. TNFR2 expression on regulatory and effector T cells in the tumor microenvironment has been associated with T cell exhaustion and resistance to immune-checkpoint blockade. In light of this, we set out to discover antibodies against human TNFR2 that could be developed as anti-cancer agents. Mouse immunization with the recombinant extracellular domain of human TNFR2 and subsequent single B cell screening using our proprietary microfluidic-based single cell platform, CelliGO™, identified a series of diverse antibodies that were characterized for binding, cross-reactivity, selectivity and functional activity. Two antibodies, HFB3-1 and HFB3-14, with sub- or single-digit-nanomolar binding affinities for human TNFR2 were selected for further characterization and humanization. Epitope mapping experiments show that the two antibodies recognize different domains of TNFR2: HFB3-1 binds to a region within the CRD2 domain and HFB3-14 binds within the CRD3 region. Despite their different binding sites, both antibodies are selective for TNFR2, cross-react with cynomolgus and rhesus monkey orthologs, and enhance the binding of human recombinant TNFα to TNFR2, as well as stimulate CD8 and conventional CD4 T cells. Humanized variants of these antibodies, HFB3-1hz6 and HFB3-14hz1c, retaining the binding and cross-reactivity profiles of the parent antibodies, were assessed for their functional activities, developability and pharmacokinetic profiles. The humanized antibodies preferentially bind to TCR-activated primary CD8 and CD4 T cells as compared to unstimulated T cells and enhance CD3/CD28-induced activation and proliferation. This co-stimulatory mechanism of action is cross-linking independent and is consistent with the antibodies' ability to enhance NFκB signaling and induce upregulation of NFκB downstream target genes. Both antibodies demonstrated good developability profile and stability under high temperature, low pH conditions and following several freeze/thaw cycles. Good plasma exposures for lead antibodies were observed in mice models. The in vivo efficacy evaluation of both antibodies in mouse tumor models as well as initial toxicology analysis are in progress. The functional profile of these antibodies along with their favorable developability and pharmacokinetic profiles support their development as a potential novel immune-therapeutic option for cancer patients.
Citation Format: Shuo Wei, Guang Yang, Juying Li, Qian Zhang, Matthieu Delince, Greg Rose, Charina Ortega, Pascaline Mary, He Zhou, Dean Lee, Roshan Kumar, Stephanie Beq, Nicola Beltraminelli, Francisco Adrian, Liang Schweizer. Discovery and characterization of novel TNFR2 antibodies to modulate T cell activities in immunosuppressive environment [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2282.