Evaluation of Radiolabeled Girentuximab In Vitro and In Vivo

Abstract

Girentuximab (cG250) targets carbonic anhydrase IX (CAIX), a protein which is expressed on the surface of most renal cancer cells (RCCs). cG250 labeled with 177Lu has been used in clinical trials for radioimmunotherapy (RIT) of RCCs. In this work, an extensive characterization of the immunoconjugates allowed optimization of the labeling conditions with 177Lu while maintaining immunoreactivity of cG250, which was then investigated in in vitro and in vivo experiments. cG250 was conjugated with S-2-(4-isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecane tetraacetic acid (DOTA(SCN)) by using incubation times between 30 and 90 min and characterized by mass spectrometry. Immunoconjugates with five to ten DOTA(SCN) molecules per cG250 molecule were obtained. Conjugates with ratios less than six DOTA(SCN)/cG250 had higher in vitro antigen affinity, both pre- and postlabeling with 177Lu. Radiochemical stability increased, in the presence of sodium ascorbate, which prevents radiolysis. The immunoreactivity of the radiolabeled cG250 tested by specific binding to SK-RC-52 cells decreased when the DOTA content per conjugate increased. The in vivo tumor uptake was < 10% ID/g and independent of the total amount of protein in the range between 5 and 100 µg cG250 per animal. Low tumor uptake was found to be due to significant necrotic areas and heterogeneous CAIX expression. In addition, low vascularity indicated relatively poor accessibility of the CAIX target.

Isolation and characterization of monoclonal antibodies against human carbonic anhydrase-IX

“…Although native cG250 failed to show anti-tumor efficacy in clinical trials, 29 it has been used successfully as a carrier for radioisotopes. 30 Furthermore, Xu et al . 31 isolated a series of single-chain antibody fragments of which one (BAY 79–4620) was further developed as a monomethyl auristatin E (MMAE) antibody-drug-conjugate (ADC).…”

In Search of Advanced Tumor Diagnostics and Treatment: Achievements and Perspectives of Carbonic Anhydrase IX Targeted Delivery

“…Another group recently tried to optimize the production of radioconjugate lutetium-177 attached to cG250 via DOTA chelator. 95 Antibodies cG250 with less than six DOTA groups were shown to have the highest immunoreactivity in vitro on SK-RC-52 cells. However, in vivo biodistribution studies showed that only less than 10% of injected 177 Lu-DOTA(SCN)-cG250 was found in tumor 48 h after administration.…”

“…94 Many issues should be considered in developing radioconjugates, including optimal half-life time of the radionuclide, conjugate stability, and postmodification changes in affinity toward the target. 95 Radioconjugates used to visualize tumors by positron emission tomography (PET) or single-photon emission computed tomography (SPECT) scans also suffer from the above mention obstacles. Radiopharmaceutic 18 F fludeoxyglucose ( 18 F-FDG) that was registered some decades ago for tumor visualization is still not surpassed by new ADC or SMDC products, even though these are broadly investigated in clinical trials.…”

“…Small molecule inhibitor–radionuclide complexes face the reverse problem, as rapid removal from blood hinders their ability to reach targets . Many issues should be considered in developing radioconjugates, including optimal half-life time of the radionuclide, conjugate stability, and postmodification changes in affinity toward the target . Radioconjugates used to visualize tumors by positron emission tomography (PET) or single-photon emission computed tomography (SPECT) scans also suffer from the above mention obstacles.…”

Alpha emitting nuclides in nuclear medicine theranostics

“…Here, for tumor tissue microscopic conditions become complex since both microscopic heterogeneity of target accumulation and heterogeneity of radiation response play a role. The negative impact of heterogeneity in tumor tissue is further augmented when large carrier molecules like monoclonal antibodies are used [20]. Therefore, with better diffusion of smaller molecules into tumor tissue small molecules as carrier nuclides for alpha emitters have shown great potential for systemic treatment of solid tumors whereas monoclonal antibodies have been shown to be suitable for treatment of hematological diseases with readily accessible tumor cells within the blood or for solid tumors within body cavities [21,22,23,24].…”