Immunogenic effect of targeted radionuclide therapy-induced tumor cell death

Targeted radionuclide therapy using single domain antibodies

Ertveldt, T.1; Krasniqi, A.2; D’Huyvetter, M.2; Goyvaerts, C.1; Lecocq, Q.1; De Vlaeminck, Y.1; Awad, R.M.1; De Beck, L.1; Devoogdt, N.2; Keyaerts, M.2,3,Δ; Breckpot, K1,Δ.

1Laboratory of Molecular and Cellular Therapy (LMCT), Vrije Universiteit Brussel (VUB), Belgium; 2In Vivo Cellular and Molecular Imaging (ICMI), VUB, Belgium; 3Dienst Nucleaire geneeskunde, UZ Brussel, Belgium; ΔContributed equally to this work.

Introduction: Targeted radionuclide therapy (TRT) is a systemic treatment with radiolabeled cancer-specific probes purposed to selectively hit diseased cells. As radioactive labels, beta-emitters, such as 90Yttrium and 177Lutetium, are studied to cause irreparable DNA damage. Beta-TRT using camelid single domain antibodies (sdAbs) is studied at the VUB, showing high potential in controlling tumor growth. Recently, immune activation after beta-TRT was reported. Therefore, we studied stimulation of CD8+ T cells and upregulation of inhibitory immune checkpoints after sdAb-mediated beta-TRT.

Materials and methods: C57BL/6 mice were inoculated with B16-melanoma cells expressing human CD20 and ovalbumin. After engraftment, mice received fractionated treatment with non-targeting sdAb or anti-CD20 sdAbs, labeled with the radionuclide 177Lutetium. Different readouts were evaluated: (1) tumor volume, measured by caliper, (2) gene expression profiling of the tumor microenvironment (TME), evaluated using RT-qPCR, (3) immune cell composition of the TME, evaluated using flow cytometry and (4) systemic immune responses, evaluated by stimulation of CD8+ splenocytes with tumor antigens.

Results: A reduced tumor progression was observed upon treatment of CD20+ melanoma-bearing mice. Despite this tumor control, we were not able to document immune activation. Gene expression and flow cytometry analysis did not reveal changes in CD8+ T cells or the inhibitory immune checkpoint PD-1/PD-L1 in the TME. Moreover, CD8+ splenocytes did not show specificity for the antigen ovalbumin, which serves as a surrogate tumor antigen.

Conclusion: Although beta-TRT with 90Yttrium coupled to a tumor-targeting alkylphosphocholine in a model non-Hodgking Lymphoma was shown to induce immune responses, we were not able to show similar immune activation with 177Lutetium-coupled anti-CD20 sdAbs in a melanoma model. Further research to confirm and study the reason for these contradicting results is required.