Enhancing anti-CD20 directed radioimmunotherapy. Preliminary studies using lenalidomide-accelerated internalization

Objectives Recent research found enhanced CD20 antigen internalization by Lenalidomide. We hypothesize an enhanced effect of CD20 directed radioimmunotherapy in combination with Lenalidomide.

Methods EBV-infected human Raji lymphoma cells with transfected luciferase-GFP were used for in vitro experiments. Lenalidomide was purified according to the recent literature and PBS was used as control. Toxicity assays with different concentrations of purified Lenalidomide (0.5uM, 0.25uM, 0.125uM, 0.063uM, 0.031uM) were performed. CD20 expression on cell surface after 2h and 24h incubation with Lenalidomide was analyzed with Flow Cytometry. Visual control of internalization after 24h incubation with Lenalidomide was performed by laser scanning confocal microscopy. The fractions of internalized radiolabeled ¹¹¹Indium-anti-CD20 monoclonal antibodies after 2h & 24h incubation with Lenalidomide was analyzed in the gamma counter after acid disruption of surface bound antibodies.

Results The toxicity assay showed normal growth curves for Raji cells with 0.25uM Lenalidomide and below. Flow cytometry revealed a reduction of the mean fluorescence intensity of 2.35% after 2h and 14.02% after 24h in cells incubated with 0.25uM Lenalidomide. Laser scanning confocal microscopy showed visual accumulation of internalized CD20 fluorescent antibodies within the Raji cells after 24h incubation with 0.25uM Lenalidomide. The fraction of internalized ¹¹¹Indium-anti-CD20 monoclonal antibody was 4.38% after 2h and 12.23% after 24h compared to control.

Conclusions These preliminary results suggest lenalidomide may enhance anti CD20 radioimmunotherapy by accelerated internalization of the radiolabeled antibody.