Noninvasive Immunotyping and Immunotherapy Monitoring of Lung Cancers via Nuclear Imaging of LAG-3 and PD-L1

Abstract

Immunotherapy has significantly improved cancer patient survival, while its efficacy remains limited due to the reliance on a single marker like PD-L1 as well as its spatiotemporal heterogeneity. To address this issue, combining lymphocyte activation gene-3 (LAG-3) with PD-L1 is proposed for identifying immunotypes and monitoring immunotherapy through nuclear imaging. In short, ^99mTc-HYNIC-αLAG-3 and ^99mTc-HYNIC-αPD-L1 probes are synthesized using anti-human LAG-3 and PD-L1 antibodies, respectively. With high radiochemical purity and in vitro stability, these probes are confirmed to specifically bind to LAG-3 or PD-L1 in LAG3⁺ A549, LAG3⁻ A549, and H1975 cells. SPECT/CT imaging of both probes showed specific in vivo tumor uptake in multiple lung cancer models, with significant linear correlation with ex vivo tumor uptake and immunohistochemical expression levels of LAG-3/PD-L1. Based on this, dual-index imaging was performed to simultaneously quantify LAG-3 and PD-L1. SPECT/CT imaging of ^99mTc-HYNIC-αLAG-3 and ¹²⁵I-αPD-L1 successfully distinguished four immunotypes. In addition, SPECT/CT imaging revealed LAG-3 upregulation in LLC-bearing LAG-3 humanized mice resistant to immunotherapy. In conclusion, this study demonstrates the feasibility of nuclear imaging of LAG-3 and PD-L1 for both noninvasive immunotyping and immunotherapy monitoring, thus offering novel perspectives on forecasting immunotherapy response, uncovering resistance mechanism, and optimizing combination treatment regimens.