ImmunoPET imaging of human CD8 T cells with novel Ga-labeled nanobody companion diagnostic agents.

BACKGROUND

Although immunotherapy has revolutionized treatment strategies for some types of cancers, most patients failed to respond or obtain long-term benefit. Tumor-infiltrating CD8 T lymphocytes are closely related to the treatment outcome and prognosis of patients. Therefore, noninvasive elucidation of both systemic and tumor-infiltrating CD8 T lymphocytes is of extraordinary significance for patients during cancer immunotherapy. Herein, a panel of Ga-labeled Nanobodies were designed and investigated to track human CD8 T cells in vivo through immuno-positron emission tomography (immunoPET).

RESULTS

Among the screened Nanobodies, SNA006a showed the highest binding affinity and specificity to both human CD8 protein and CD8 cells in vitro, with the equilibrium dissociation constant (K) of 6.4 × 10 M and 4.6 × 10 M, respectively. Ga-NOTA-SNA006 was obtained with high radiochemical yield and purity, and stayed stable for at least 1 h both in vitro and in vivo. Biodistribution and Micro-PET/CT imaging studies revealed that all tracers specifically concentrated in the CD8 tumors with low accumulation in CD8 tumors and normal organs except the kidneys, where the tracer was excreted and reabsorbed. Notably, the high uptake of Ga-NOTA-SNA006a in CD8 tumors was rapid and persistent, which reached 24.41 ± 1.00% ID/g at 1.5 h after intravenous injection, resulting in excellent target-to-background ratios (TBRs). More specifically, the tumor-to-muscle, tumor-to-liver, and CD8 to CD8 tumor was 28.10 ± 3.68, 5.26 ± 0.86, and 19.58 ± 2.70 at 1.5 h, respectively. Furthermore, in the humanized PBMC-NSG and HSC-NPG mouse models, Ga-NOTA-SNA006a accumulated in both CD8 tumors and specific tissues such as liver, spleen and lung where human CD8 antigen was overexpressed or CD8 T cells located during immunoPET imaging.

CONCLUSIONS

Ga-NOTA-SNA006a, a novel Nanobody tracer targeting human CD8 antigen, was developed with high radiochemical purity and high affinity. Compared with other candidates, the long retention time, low background, excellent TBRs of Ga-NOTA-SNA006a make it precisely track the human CD8 T cells in mice models, showing great potential for immunotherapy monitoring and efficacy evaluation.