Preclinical Development of CD38-Targeted [89Zr] Zr-DFO-Daratumumab for Imaging Multiple Myeloma

Multiple myeloma (MM) is a plasma B-cell hematologic cancer that causes significant skeletal morbidity. Despite improvements in survival, heterogeneity in response remains a major challenge in MM. Cluster of differentiation 38 (CD38) is a type II transmembrane glycoprotein overexpressed in myeloma cells and is implicated in MM cell signaling. Daratumumab is a U.S. Food and Drug Administration-approved high-affinity monoclonal antibody targeting CD38 that is clinically benefiting refractory MM patients. Here, we evaluated [Zr-89] Zr-desferrioxamine (DFO)-daratumumab PET/CT imaging in MM tumor models. Methods: Daratumumab was conjugated to DFO-p-benzyl-isothiocyanate (DFO-Bz-NCS) for radiolabeling with Zr-89. Chelator conjugation was confirmed by electrospray ionization-mass spectrometry, and radiolabeling was monitored by instant thin-layer chromatography. Daratumumab was conjugated to Cyanine5 (Cy5) dye for cell microscopy. In vitro and in vivo evaluation of [Zr-89] Zr-DFO-daratumumab was performed using CD38(+) human myeloma MM1.S-luciferase (MM1.S) cells. Cellular studies determined the affinity, immunoreactivity, and specificity of [Zr-89]Zr-DFO-daratumumab. A 5TGM1-luciferase (5TGM1)/KaLwRij MM mouse model served as control for imaging background noise. [Zr-89] Zr-DFO-daratumumab PET/CT small-animal imaging was performed in severe combined immunodeficient mice bearing solid and disseminated MM tumors. Tissue biodistribution (7 d after tracer administration, 1.11 MBq/animal, n = 4-6/group) was performed in wild-type and MM1.S tumor-bearing mice. Results: A specific activity of 55.5 MBq/nmol (0.37 MBq/mu g) was reproducibly obtained with [Zr-89] Zr-daratumumab-DFO. Flow cytometry confirmed CD38 expression (>99%) on the surface of MM1.S cells. Confocal microscopy with daratumumab-Cy5 demonstrated specific cell binding. Dissociation constant, 3.3 nM (+/- 0.58), and receptor density, 10.1 fmol/mg (+/- 60.64), was obtained with a saturation binding assay. [Zr-89] Zr-DFO-daratumumab/PET demonstrated specificity and sensitivity for detecting CD38(+) myeloma tumors of variable sizes (8.5-128 mm(3)) with standardized uptake values ranging from 2.1 to 9.3. Discrete medullar lesions, confirmed by bioluminescence images, were efficiently imaged with [Zr-89] Zr-DFO-daratumumab/PET. Bio-distribution at 7 d after administration of [Zr-89] Zr-DFO-daratumumab showed prominent tumor uptake (27.7 +/- 7.6 percentage injected dose per gram). In vivo blocking was achieved with a 200-fold excess of unlabeled daratumumab. Conclusion: [Zr-89] Zr-DFO-and Cy5-daratumumab demonstrated superb binding to CD38(+) human MM cells and significantly low binding to CD38(low) cells. Daratumumab bioconjugates are being evaluated for image-guided delivery of therapeutic radionuclides.