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Updated: Aug 21, 2025
Development of a 68Gallium-Labeled D-Peptide PET Tracer for Imaging Programmed Death-Ligand 1 Expression
Published on: February 3, 2023
Novel small 99mTc-labeled affibody molecular probe for PD-L1 receptor imaging
Zhigang Liang1, Xianwen Hu1, Hongyu Hu1
1Department of Nuclear Medicine, Affiliated Hospital of Zunyi Medical University, Zunyi, Guizhou, China.
Objective:
The in vivo imaging of programmed death ligand 1 (PD-L1) can monitor changes in PD-L1 expression and guide programmed death 1 (PD-1) or PD-L1-targeted immune checkpoint therapy. A 99mTc-labeled affibody molecular probe targeting the PD-L1 receptor was prepared and evaluated its tracing effect in PD-L1-overexpressing colon cancer.
Methods:
The PD-L1 affibody was prepared by genetic recombineering. The 99mTc labeling of the affibody was achieved by sodium glucoheptonate and an SnCl2 labeling system. The labeling rate, radiochemical purity, and stability in vitro were determined by instant thin-layer chromatography; MC38-B7H1 (PD-L1-positive) and MC38 (PD-L1-negative) colon cancer cells were used to evaluate its affinity to PD-L1 by cell-binding experiments. The biodistribution of the 99mTc-labeled affibody molecular probe was then determined in C57BL/6J mice bearing MC38-B7H1 tumors, and tumor targeting was assessed in C57BL/6J mice with MC38-B7H1, MC38 double xenografts.
Result:
The nondecayed corrected yield of the 99mTc-PD-L1 affibody molecular probe was 95.95% ± 1.26%, and showed good stability both in phosphate-buffered saline (PBS) and fetal bovine serum within 6 h. The affinity of the 99mTc-PD-L1 affibody molecular probe for cell-binding assays was 10.02 nmol/L. Single photon emission-computed tomography imaging showed a rapid uptake of the tracer in PD-L1-positive tumors and very little tracer retention in PD-L1-negative control tumors. The tracer was significantly retained in the kidneys and bladder, suggesting that it is mainly excreted through the urinary system. Heart, liver, lung, and muscle tissue showed no significant radioactive retention. The biodistribution in vitro also showed significant renal retention, a small amount of uptake in the thyroid and gastrointestinal tract, and rapid blood clearance, and the tumor-to-blood radioactivity uptake ratio peaked 120 min after drug injection.
Conclusion:
The 99mTc-PD-L1 affibody molecular probe that we prepared can effectively target to PD-L1-positive tumors imaging in vivo, and clear in blood quickly, with no obvious toxic side effects, which is expected to become a new type of tracer for detecting PD-L1 expression in tumors.
Insights
A novel 99mTc-labeled affibody molecular probe effectively targets programmed death ligand 1 (PD-L1) positive tumors for in vivo imaging. This tracer shows rapid blood clearance and minimal toxicity, offering a promising tool for monitoring PD-L1 expression in cancer immunotherapy.
Area of Science:
- Nuclear medicine
- Molecular imaging
- Cancer diagnostics
Background:
- In vivo imaging of programmed death ligand 1 (PD-L1) is crucial for monitoring expression and guiding immune checkpoint therapies (PD-1/PD-L1 targeted).
- Development of specific molecular probes is essential for accurate non-invasive assessment of PD-L1 expression in tumors.
Purpose of the Study:
- To prepare and evaluate a 99mTc-labeled affibody molecular probe for targeting and imaging PD-L1 receptors.
- To assess the probe's efficacy in tracing PD-L1-overexpressing colon cancer models in vivo.
Main Methods:
- Preparation of PD-L1 affibody via genetic recombineering and subsequent 99mTc labeling using a sodium glucoheptonate and SnCl2 system.
- Assessment of labeling efficiency, radiochemical purity, in vitro stability, and affinity using cell-binding assays with PD-L1-positive (MC38-B7H1) and negative (MC38) colon cancer cells.
- In vivo biodistribution and tumor targeting evaluation in C57BL/6J mice bearing MC38-B7H1 tumors using SPECT imaging.
Main Results:
- The 99mTc-PD-L1 affibody probe achieved a high labeling yield (95.95% ± 1.26%) and demonstrated good in vitro stability.
- The probe exhibited high affinity (10.02 nmol/L) and specific binding to PD-L1-positive cells.
- SPECT imaging revealed rapid uptake in PD-L1-positive tumors with minimal retention in PD-L1-negative tumors; significant renal and bladder excretion was observed, with rapid blood clearance and no major off-target retention in other organs.
Conclusions:
- The developed 99mTc-PD-L1 affibody molecular probe effectively targets PD-L1-positive tumors for in vivo imaging.
- The probe demonstrates rapid blood clearance and no significant toxic side effects.
- This tracer is a promising new tool for detecting PD-L1 expression in tumors, aiding in the management of immunotherapies.
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