ImmunoPET of Malignant and Normal B Cells with ⁸⁹Zr- and ¹²⁴I-Labeled Obinutuzumab Antibody Fragments Reveals Differential CD20 Internalization <i>In Vivo</i>.

Abstract

<b>Purpose:</b> The B-cell antigen CD20 provides a target for antibody-based positron emission tomography (immunoPET). We engineered antibody fragments targeting human CD20 and studied their potential as immunoPET tracers in transgenic mice (huCD20TM) and in a murine lymphoma model expressing human CD20.<b>Experimental Design:</b> Anti-CD20 cys-diabody (cDb) and cys-minibody (cMb) based on rituximab and obinutuzumab (GA101) were radioiodinated and used for immunoPET imaging of a murine lymphoma model. Pairwise comparison of obinutuzumab-based antibody fragments labeled with residualizing (⁸⁹Zr) versus non-residualizing (¹²⁴I) radionuclides by region of interest analysis of serial PET images was conducted both in the murine lymphoma model and in huCD20TM to assess antigen modulation <i>in vivo</i><b>Results:</b>¹²⁴I-GAcDb and ¹²⁴I-GAcMb produced high-contrast immunoPET images of B-cell lymphoma and outperformed the respective rituximab-based tracers. ImmunoPET imaging of huCD20TM showed specific uptake in lymphoid tissues. The use of the radiometal ⁸⁹Zr as alternative label for GAcDb and GAcMb yielded greater target-specific uptake and retention compared with ¹²⁴I-labeled tracers. Pairwise comparison of ⁸⁹Zr- and ¹²⁴I-labeled GAcDb and GAcMb allowed assessment of <i>in vivo</i> internalization of CD20/antibody complexes and revealed that CD20 internalization differs between malignant and endogenous B cells.<b>Conclusions:</b> These obinutuzumab-based PET tracers have the ability to noninvasively and quantitatively monitor CD20-expression and have revealed insights into CD20 internalization upon antibody binding <i>in vivo</i> Because they are based on a humanized mAb they have the potential for direct clinical translation and could improve patient selection for targeted therapy, dosimetry prior to radioimmunotherapy, and prediction of response to therapy. <i>Clin Cancer Res; 23(23); 7242-52. ©2017 AACR</i>.