135P - Simultaneous noninvasive imaging of LAG-3 and PD-L1 for noninvasive immunotyping in lung cancers

Background

Immunotherapy has significantly improved the survival of cancer patients, while its efficacy is limited by the reliance on single marker like PD-L1 and its spatiotemporal heterogeneity. To address this issue, the current study introduces a nuclear imaging approach for the simultaneous and noninvasive quantification of PD-L1 and lymphocyte activation gene-3 (LAG-3), aiming to improve immunotherapy response prediction.

Methods

^99mTc-HYNIC-αLAG-3 and ^99mTc-HYNIC-αPD-L1 probes were developed by using anti-human LAG-3 and PD-L1 antibodies, respectively. In vivo and vitro assays were conducted in LAG3⁺A549, H1975, LAG-3^-A549, and LLC cells. Single photon emission computed tomography/computed tomography (SPECT/CT) imaging were performed in various lung cancer models. The correlation between tumor signals in region of interest (ROI), tumor uptake determined by ex vivo γ-counting, and immunohistochemistry (IHC) expression levels was analyzed. Additionally, αPD-L1 was labeled with ¹²⁵I, combined with ^99mTc-HYNIC-αLAG-3, to perform synchronous noninvasive nuclear imaging of immunotypes (LAG-3⁺PD-L1⁺, LAG-3^-PD-L1⁺, LAG-3⁺PD-L1^-, LAG-3^-PD-L1^-).

Results

^99mTc-HYNIC-αLAG-3/PD-L1 probes exhibited a RCP of over 95% and high stability in vitro within 24 hours. The specific binding of probes to LAG-3 or PD-L1 was confirmed. SPECT/CT imaging of ^99mTc-HYNIC-αLAG-3 revealed tumor uptake of 15.5 ± 2.4, 5.3 ± 1.1, and 6.2 ± 2.1 %ID/g in LAG-3⁺A549, LAG-3^-A549 models, and LLC-bearing LAG-3 humanized mice at 24 h, respectively. ^99mTc-HYNIC-αPD-L1 had tumor uptake of 7.4 ± 0.8%ID/g in H1975 models and 2.1 ± 0.8 %ID/g in A549 models at 24 h. The tumor signals within the ROI for both probes demonstrated significant associations with ex vivo tumor uptake and IHC expression levels. Simultaneous SPECT/CT imaging of ^99mTc-HYNIC-αLAG-3 and ¹²⁵I-αPD-L1 at 24 h post-injection successfully differentiated various immunotypes.

Conclusions

This study demonstrates the feasibility of SPECT imaging in visualizing LAG-3 and PD-L1. Moreover, this is the first report of simultaneous nuclear imaging to identify diverse immunotypes in lung cancers, offering new insights into forecasting immunotherapy response and selecting combination therapy.

Legal entity responsible for the study

The authors.

Funding

This study was supported in part by a grant of National Key Research and Development Program of China (2022YFF0705300), National Natural Science Foundation of China (52272281), Shanghai Municipal Science and Technology Major Project (2021SHZDZX0100) and the Fundamental Research Funds for the Central Universities, Shanghai Municipal Health Commission Health Industry Clinical Research Project, Clinical Research Project of Shanghai Pulmonary Hospital (FKLY20010), Young Talents in Shanghai (2019 QNBJ), Shanghai Shuguang Scholar, 2021 Science and Technology Think Tank Youth Talent Plan of China Association for Science and Technology,‘Dream Tutor’ Outstanding Young Talents Program (fkyq1901), National Key Research and Development Program of China (2021YFF1201200 and 2021YFF1200900).

Disclosure

All authors have declared no conflicts of interest.