Persistent antigen exposure and inflammatory signals in tumors induce expression of various co-inhibitory or immune checkpoint receptors on T cells, including programmed death protein 1 (PD-1) and Lymphocyte-Activation Gene 3 (LAG-3). Therapeutic antibodies blocking such co-inhibitory receptors have produced durable antitumor responses as single agents and in combinations. In order to monitor LAG-3 expression and potential changes in expression due to therapeutic intervention, we have developed a radionuclide-conjugated antibody to LAG-3 for immuno-PET. The fully human anti-LAG3 antibody REGN3767 was radiolabeled with the positron-emitting radionuclide Zirconium-89 (89Zr) using the bifunctional chelator p-SCN-Bn-Deferoxamine (DFO). 89Zr-REGN3767 demonstrated high radiochemical purity and immunoreactivity in cell binding assays. The ability of 89Zr-REGN3767 to successfully identify LAG-3 expression in vivo was initially assessed using MC38 mouse tumors expressing human LAG-3 (MC38/hLAG-3) implanted into immune-deficient mice. 89Zr-REGN3767 demonstrated higher uptake in MC38/hLAG-3 tumors compared to an 89Zr-isotype control antibody using immuno-PET, and specificity was confirmed by ex vivo biodistribution at day 6 post radiotracer injection (~35 and ~5 %ID/g for 89Zr-REGN3767 and 89Zr-isotype, respectively). Furthermore, a dose titration study of 89Zr-REGN3767 in immune deficient mice co-implanted subcutaneously with Raji lymphoma cells and human peripheral blood mononuclear cells (hPBMCs) demonstrated the ability of 89Zr-REGN3767 to target LAG-3-expressing intratumoral T-cells. 89Zr-REGN3767 immuno-PET and ex vivo biodistribution demonstrated specific localization to Raji/hPBMC co-implanted tumors; this uptake was significantly higher at antibody doses of 0.03 - 0.3 mg/kg than at 5 mg/kg. Doses of 0.03-0.3 mg/kg 89Zr-REGN3767 were also able to detect LAG-3 positive T cells in the spleen. This study shows the ability of 89Zr-REGN3767 to successfully image LAG-3 expressed on intratumoral and splenic T lymphocytes. This work supports the clinical translation of anti-LAG-3 immuno-PET for the assessment of LAG-3 expression, with the goal to investigate its utility for predicting and monitoring response to checkpoint blockade therapy.

Citation Format: Marcus P. Kelly, Richard Tavare, Jason T. Giurleo, Sosina Makonnen, Carlos Hickey, Makenzie A. Danton, T Cody Arnold, Dangshe Ma, Jie Dai, Jerry Pei, Jessica R. Kirshner, William C. Olson, Gavin. Thurston. Immuno-PET detection of LAG-3 expressing intratumoral lymphocytes using the zirconium-89 radiolabeled fully human anti-LAG-3 antibody REGN3767 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3033.