Cb213: A Half-Life Extended Bispecific Humabody V-H Delivering Dual Checkpoint Blockade To Reverse The Dysfunction Of Lag3+Pd-1+Double-Positive T Cells James

Introduction: Chronic antigen stimulation is known to promote a state of T cell dysfunction whereby cells are unable to mount effector responses such as cytokine release, proliferation and tumour cell killing. Indeed, dysfunctional T cells are found to be located within the tumour microenvironment where they express high levels of inhibitory receptors such as LAG3 and PD-1 on their cell surface. Targeting dysfunctional tumoral T cells and re-invigoration of their effector functions is an attractive strategy for combatting intrinsic and acquired resistance to current checkpoint blockade strategies such as anti-PD-1, PD-L1 or CTLA-4 antagonist antibodies. Methods and Results: Using the Crescendo Mouse™, a transgenic platform which develops fully human VH domains in a background devoid of light chains, we have generated a bispecific Humabody VH which efficiently delivers dual LAG3 and PD-1 checkpoint blockade specifically to LAG3+PD-1+ double positive T cells, thereby reversing T cell inhibition. Selective targeting has been achieved by using an asymmetric 2:1 binding format utilising bivalent LAG3 binding coupled with monovalent PD-1 binding. Using Dynamic Biosensors kinetic binding assessment, high affinity bispecific binding was observed when both ligands were present together whereas the affinity of binding was significantly lower when either ligand was present alone. Humabody bispecific bound to PD1+LAG3− Jurkat cells with a lower EC50 compared to Nivolumab whereas binding to PD-1-LAG3+ Jurkat cells was equivalent to a benchmark anti-LAG3 mAb. The bispecific was also able to robustly enhance IL-2 production from SEB stimulated PBMCs. In order to assess the impact of bispecifics on T cells with a more dysfunctional phenotype we obtained peripheral blood lymphocytes from patients with non-small cell lung cancer (NSCLC). When cultured with a lung carcinoma cell line presenting anti-CD3, patient-derived CD4+ and CD8+ T cells contained a higher proportion of LAG3+PD-1+ than cells obtained from healthy donors and were less proliferative. The Humabody bispecific was able to increase the proliferation of CD4+ and CD8+ cells in cultures from NSCLC donors and suppress the expression of a biomarker of T-cell inhibition. In vivo, in a MC38 syngeneic tumour model grown in a transgenic HuGEMM mouse where the PD-1 and LAG3 receptors have been humanised, CB213 was able to mediate significant anti-tumour effects. Conclusions: Together these data support preclinical and clinical development of CB213, a novel 2:1 asymmetric Humabody dual checkpoint inhibitor targeting LAG3/PD-1 double positive dysfunctional T cells. Citation Format: James William Legg, Brian McGuinness, Hugo Arasanz, Ana Bocanegra, Phillip Bartlett, Giovanni Benedetti, Neil Birkett, Carl Cox, Elena De Juan, Carrie Enever, Emma Hames, Grazyna Kochan, Maria Jesus Garcia-Granda, Angelica Sette, Yumin Teng, Lorraine Thompson, Ruth Vera, Richard Williams, Miren Zuazo, David Escors, Carolyn Edwards. CB213: A half-life extended bispecific Humabody VH delivering dual checkpoint blockade to reverse the dysfunction of LAG3+PD-1+ double-positive T cells [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 930.