Bioluminescence imaging for monitoring therapeutic response of CD38-specific nanobody-Fc fusion proteins in a lymphoma model

Zielsetzung:

CD38 is overexpressed by lymphomas and other hematological tumors and represents a promising target for immunotherapy. We aimed to develop CD38-specific nanobody-Fc fusion proteins for imaging and treatment of hematological tumors and to test them in a bioluminescent lymphoma model.

Material und Methodik:

We generated CD38-specific nanobodies (15kDa) by phage-display technology from immunized llamas. Specific binding, affinity and epitope specificity of selected nanobodies were analyzed by flow cytometry. Nanobodies were fused to the Fc-domains of human IgG1. The capacity of Nb-Fc fusion proteins (60kDa) to induce complement-dependent cytotoxicity (CDC) of human lymphoma cell lines and bone marrow cells of multiple myeloma patients were analyzed by flow cytometry. In vivo effects of Nb-Fc fusion proteins on tumor growth were assessed in SCID-mice injected with luciferase-transduced human Burkitt lymphoma cells. Therapeutic response was determined via bioluminescence imaging of disseminated tumors (IVIS200,PerkinElmer) and survival curves.

Ergebnisse:

We selected 22 families of CD38-specific nanobodies. Crossblockade analyses indicate that most of these bind to 3 non-overlapping epitopes. While single Nb-Fc fusion proteins showed little if any capacity to induce CDC, the combination of two Nb-Fc-fusion proteins recognizing distinct epitopes showed very potent CDC toward CD38-expressing cell lines as well as toward primary CD38+ tumor cells from bone marrow samples of myeloma patients. In vivo bioluminescence imaging of luciferase-transduced CD38+ Burkitt lymphoma cells confirmed the therapeutic efficacy of combinations of two Fc-fusion proteins recognizing distinct epitopes. Such combinations significantly reduced tumor growth and significantly prolonged survival of treated mice compared to controls.

Schlussfolgerungen:

Bioluminescence imaging is a reliable tool for monitoring diffuse tumor growth and therapy response. CD38-specific nanobody-Fc fusion proteins hold promise as therapeutics for CD38-expressing lymphomas.