Abstract A26: Tetravalency endows anti-CD27 antibodies with enhanced co-stimulatory activity and anti-tumour immune responses

Abstract Immunostimulatory antibodies targeting co-stimulatory TNFRSF members have shown promise in pre-clinical cancer models, but this has not translated into clinical success, primarily due to lack of efficacy. Oligomerisation of the co-stimulatory molecule CD27 by its trimeric membrane-bound ligand activates NF-κB and AP1 signalling leading to enhanced CD8 T cell responses in both humans and mice. Targeting CD27 with bivalent mAb may lead to sub-optimal receptor oligomerisation and inefficient T cell activation. Here we present a general approach to improve the potency of anti-CD27 mAb. We engineered tetravalent forms of anti-CD27 antibodies targeting both mouse and human CD27 by attaching two additional Fab fragments to the N-terminus of the of the IgG VH domain. These tetravalent mAb had greater avidity to CD27 and were more potent than their bivalent counterparts in stimulating NF-κB activation and T cell proliferation in vitro, with optimal responses requiring antibody crosslinking by FcγRIIb. In addition, confocal microscopy of hCD27-GFP fusion protein expressing Jurkat T cells showed that tetravalent anti-hCD27 mAb triggered receptor clustering more efficiently than the equivalent bivalent mAb, as evidenced by the increased number of receptor clusters on the cell surface. Importantly, tetravalent anti-CD27 mAb induced greater CD8 T cell responses than the bivalent mAb in a vaccination model and was more efficacious in suppressing tumour growth in vivo. This work demonstrates that the agonistic activity of anti-CD27 mAb can be significantly improved by tetravalency and suggest that this approach could be utilised to enhance the therapeutic activity of mAb targeting other members of the TNFRSF. Citation Format: Marcus A Widdess, H. T. Claude Chan, Christine A Penfold, C Ian Mockridge, Tatyana Inzhelevskaya, Hannah J Metcalfe, Mark S Cragg, Aymen Al-Shamkhani. Tetravalency endows anti-CD27 antibodies with enhanced co-stimulatory activity and anti-tumour immune responses [abstract]. In: Proceedings of the AACR Special Conference: Tumor Immunology and Immunotherapy; 2022 Oct 21-24; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2022;10(12 Suppl):Abstract nr A26.