Single-Domain Antibody Nuclear Imaging Allows Noninvasive Quantification of LAG-3 Expression by Tumor-Infiltrating Leukocytes and Predicts Response of Immune Checkpoint Blockade

Quentin Lecocq,Geert Raes,Nick Devoogdt,Wout De Mey,Robin Maximilian Awad,Yannick De Vlaeminck,Thomas Ertveldt,Karine Breckpot,Cleo Goyvaerts,Kris Thielemans,Marleen Keyaerts

doi:10.2967/jnumed.120.258871

Abstract

Recent advances in the field of immune-oncology led to the discovery of next-generation immune checkpoints (ICPs). Lymphocyte activation gene-3 (LAG-3), being the most widely studied among them, is being explored as a target for the treatment of cancer patients. Several antagonistic anti-LAG-3 antibodies are being developed and are prime candidates for clinical application. Furthermore, validated therapies targeting cytotoxic T-lymphocyte–associated protein-4, programmed cell-death protein-1, or programmed cell-death ligand-1 showed that only subsets of patients respond. This finding highlights the need for better tools for patient selection and monitoring. The potential of molecular imaging to detect ICPs noninvasively in cancer is supported by several preclinical and clinical studies. Here, we report on a single-domain antibody to evaluate whole-body LAG-3 expression in various syngeneic mouse cancer models using nuclear imaging. Methods: SPECT/CT scans of tumor-bearing mice were performed 1 h after injection with radiolabeled single-domain antibody. Organs and tumors of mice were isolated and evaluated for the presence of the radiolabeled tracer and LAG-3–expressing immune cells using a γ-counter and flow cytometry respectively. PD-1/LAG-3–blocking antibodies were injected in MC38-bearing mice. Results: The radiolabeled single-domain antibody detected LAG-3 expression on tumor-infiltrating lymphocytes (TILs) as soon as 1 h after injection in MC38, MO4, and TC-1 cancer models. The single-domain antibody tracer visualized a compensatory upregulation of LAG-3 on TILs in MC38 tumors of mice treated with PD-1–blocking antibodies. When PD-1 blockade was combined with LAG-3 blockade, a synergistic effect on tumor growth delay was observed. Conclusion: These findings consolidate LAG-3 as a next-generation ICP and support the use of single-domain antibodies as tools to noninvasively monitor the dynamic evolution of LAG-3 expression by TILs, which could be exploited to predict therapy outcome.

Highlights

INTRODUCTIONA frequently exploited immunotherapy strategy in cancer is blockade of inhibitory immune checkpoints (ICPs) [1]
Analysis of tumor uptake showed that only radiolabeled Lymphocyte activation gene-3 (LAG-3) nanobody accumulated in MC38-tumors with little increase in larger tumors (p=0.0364 and p=0.0091 for day 11 and 17 tumors, resp.) (Fig. 2b)
LAG-3 nanobody tumor uptake was evaluated in mice bearing TC-1 or MO4 tumors

Summary

Introduction

INTRODUCTIONA frequently exploited immunotherapy strategy in cancer is blockade of inhibitory immune checkpoints (ICPs) [1]. LAG-3 is a CD4-like molecule belonging to the immunoglobulin superfamily and is expressed on activated CD4+ and CD8+ T-cells [6], regulatory T-cells [7], B-cells [8], natural killer (NK)-cells [9], plasmacytoid dendritic cells [10] and myeloid cells like macrophages [11]. Molecules such as MHC-II [12], Galectin-3 [13], LSECtin [14], α-synuclein [15] and fibrinogen-like protein-1 [16] can interact with LAG-3, with MHC-II being the canonical ligand. LAG-3 signaling is co-opted in the tumor microenvironment (TME) to enable tumor cell escape

Methods

Results

Conclusion