SignificanceCD8+ T lymphocytes, which are typically devoted to eliminate malignant and infected cells, have been described in the central nervous system (CNS) of patients and mice with amyotrophic lateral sclerosis (ALS). However, their role in ALS pathogenesis has yet to be unraveled. Here, we show that ablation of CD8+ T cells in ALS mice increased the number of surviving motoneurons. CD8+ T cells expressing the ALS-causing superoxide dismutase-1 mutant protein recognize and selectively kill motoneurons in vitro. To exert their cytotoxic function, mutant CD8+ T cells required presentation of the antigen-MHC-I complex at the surface of the motoneurons. Analysis of T cell receptor diversity supports the evidence that self-reactive CD8+ T lymphocytes infiltrate the CNS of ALS mice to exert cytotoxic function.

Purpose TEM-1 (tumor endothelial marker-1) is a single-pass transmembrane cell surface glycoprotein expressed at high levels by tumor vasculature and malignant cells. We aimed to perform a preclinical investigation of a novel anti-TEM-1 scFv-Fc fusion antibody, 1C1m-Fc, which was radiolabeled with 177 Lu for use in soft tissue sarcomas models. Methods 1C1m-Fc was first conjugated to p-SCN-Bn-DOTA using different excess molar ratios and labeled with 177 Lu. To determine radiolabeled antibody immunoreactivity, Lindmo assays were performed. The in vivo behavior of [177Lu]Lu-1C1m-Fc was characterized in mice bearing TEM-1 positive (SK-N-AS) and negative (HT-1080) tumors by biodistribution and single-photon emission SPECT/CT imaging studies. Estimated organ absorbed doses were obtained based on biodistribution results. Results The DOTA conjugation and the labeling with 177 Lu were successful with a radiochemical purity of up to 95%. Immunoreactivity after radiolabeling was 86% ± 4%. Biodistribution showed a specific uptake in TEM-1 positive tumor versus liver as critical non-specific healthy organ, and this specificity is correlated to the number of chelates per antibody. A 1.9-fold higher signal at 72 h was observed in SPECT/CT imaging in TEM-1 positive tumors versus control tumors. Conclusion TEM-1 is a promising target that could allow a theranostic approach to soft-tissue sarcoma, and 1C1m-Fc appears to be a suitable targeting candidate. In this study, we observed the influence of the ratio DOTA/antibody on the biodistribution. The next step will be to investigate the best conjugation to achieve an optimal tumor-to-organ radioactivity ratio and to perform therapy in murine xenograft models as a prelude to future translation in patients.

1C1m-Fc, an anti-tumor endothelial marker 1 (TEM-1) scFv-Fc fusion protein antibody, was previously successfully radiolabeled with 177Lu. TEM-1 specific tumor uptake was observed together with a non-saturation dependent liver uptake that could be related to the number of dodecane tetraacetic acid (DOTA) chelator per 1C1m-Fc. The objective of this study was to verify this hypothesis and to find the best DOTA per 1C1m-Fc ratio for theranostic applications. 1C1m-Fc was conjugated with six concentrations of DOTA. High-pressure liquid chromatography, mass spectrometry, immunoreactivity assessment, and biodistribution studies in mice bearing TEM-1 positive tumors were performed. A multi-compartment pharmacokinetic model was used to fit the data and a global pharmacokinetic model was developed to illustrate the effect of liver capture and immunoreactivity loss. Organ absorbed doses in mice were calculated from biodistribution results. A loss of immunoreactivity was observed with the highest DOTA per 1C1m-Fc ratio. Except for the spleen and bone, an increase of DOTA per 1C1m-Fc ratio resulted in an increase of liver uptake and absorbed dose and a decrease of uptake in tumor and other tissues. Pharmacokinetic models correlated these results. The number of DOTA per antibody played a determining role in tumor targeting. One DOTA per 1C1m-Fc gave the best pharmacokinetic behavior for a future translation of [177Lu]Lu-1C1m-Fc in patients.