Single-domain antibodies for targeting, detection and in vivo imaging of human CD4+ cells

• Published in: bioRxiv
• Main Authors: Traenkle, Bjoern, Kaiser, Philipp D, Pezzana, Stefania, Richardson, Jennifer, Gramlich, Marius, Wagner, Teresa R, Seyfried, Dominik, Weldle, Melissa, Holz, Stefanie, Parfyonova, Yana, Schaller, Martin, Nueske, Stefan, Scholz, Armin, Zeck, Anne, Jakobi, Meike, Schneiderhan-Marra, Nicole, Maurer, Andreas, Gouttefangeas, Cecile, Kneilling, Manfred, Pichler, Bernd J, Sonanini, Dominik, Rothbauer, Ulrich
• Format: Paper
• Language: English
• Published: Cold Spring Harbor Cold Spring Harbor Laboratory Press 02.09.2021
• Subjects: Animal models; Antibodies; Autoimmune diseases; CD4 antigen; Cell proliferation; Cytokines; Epitope mapping; Immunotherapy; Inflammatory diseases; Lymphocytes T; Nanobodies; Patent applications; Probes; Xenografts
• DOI: 10.1101/2021.07.02.450848

The advancement of new immunotherapies necessitates appropriate probes to monitor the presence and distribution of distinct immune cell populations. Considering the key role of CD4+ T cells in regulating immunological processes, we generated novel single-domain antibodies (nanobodies, Nbs) that specifically recognize human CD4. After in depth analysis of their binding properties, recognized epitopes, and effects on T cell proliferation, activation and cytokine release, we selected CD4 Nbs that did not interfere with crucial T cell processes in vitro and converted them into immune tracers for non-invasive molecular imaging. By optical imaging, we demonstrate the ability of a high-affinity CD4-Nb to specifically visualize CD4+ cells in vivo using a xenograft model. Furthermore, time-resolved immune positron emission tomography (immunoPET) of a human CD4 knock-in mouse model showed rapid accumulation of 64Cu-radiolabeled CD4-Nb in CD4+ T cell-rich tissues. We propose that the CD4 Nbs presented here could serve as versatile probes for stratifying patients and monitoring individual immune responses during personalized immunotherapy in both cancer and inflammatory diseases. Competing Interest Statement D.S., M.K., B.P., B.T., P.D. K., U.R. are named as inventors on a patent application claiming the use of the described nanobodies for diagnosis and therapeutics filed by the Natural and Medical Sciences Institute and the Werner Siemens Imaging Center. The other authors declare no competing interest. Footnotes * The revised version now includes additional PET/MR data for the radiolabeled CD4-Nb1 in a CD4 mouse knock-in model. Detailed analyses and results are shown in new Figure 6 and Supplementary Figure 12