Radiolabelling rituximab with ^99mTc in three steps procedure

Abstract

Lymphomas are the most frequent haematological malignancy. In non-Hodgkin's lymphomas (NHL), more than 90% of tumor cells express the cluster of differentiation (CD) 20 antigen. At the end of frontline therapy, the evaluation of remission is based on computed tomography (CT) and positron emission tomography coupled with computer tomography (PET/CT) with [(18)F]-fluorodeoxyglucose ([(18)F]FDG). Unfortunately, these techniques are not specific and cannot distinguish residual active tumor from inflammation. The aim of this study was to develop a specific radiotracer of NHL CD 20+ cells for clinical applications. The radiolabelling technique presented, based on the use of tricarbonyl compound, does not include an antibody reduction because this step could damage the protein. Actually, rituximab, an anti-CD 20 chimeric antibody used for the treatment of these NHL, was radiolabelled with Isolink® (99m)Tc-tricarbonyl compound in a three-step procedure without using a specific antibody reducer. Radiolabelling yield was greater than 97%. In vitro experiments showed a conservation of antibody integrity. In vivo experiments using Single-photon emission computed tomography/CT showed significant tumor targeting 24 h after injection of the radiotracer. It was consequently possible to develop an immunoradiolabelling method to specifically detect the residual disease. As this procedure is fast, reproducible and gentle, it will be possible to comply with Good Manufacturing Practices.

Technetium-99m- or Cy7-Labeled Rituximab as an Imaging Agent for Non-Hodgkin Lymphoma

“…To radiolabel rituximab with 99m Tc, we used NHS-HYNIC-Tfa as a bifunctional chelating agent due to our expertise in radiolabeling antibodies with it [27,28,29,30], highlighting the time it takes to complete the labeling (30 min) compared to other methods [17,18,20,21]. Abrams et al [37] were the first to describe the synthesis and in vivo evaluation of HYNIC bioconjugates.…”

“…This last step creates thiol residues that facilitate technetium binding, but could modify the protein structure and decrease antigen recognition. Labeling of rituximab using 99m Tc-carbonyl core ([ 99m Tc(CO) 3 ] + ) has also been performed with interesting results, although heating is required in order to achieve quantitative labeling at micromolar concentrations [17,20,21]. …”

“…In this way, technetium-99m ( 99m Tc)-labeled compounds have better dosimetric properties (half-life = 6 h, gamma energy = 140 keV) than 18 FDG, being widely available at lower cost. In the last years, several methods for radiolabeling rituximab with 99m Tc have been described [17,18,19,20,21]. All of them include 2 reduction steps - the first step dedicated to the reduction of 99m Tc, and the second step specifically targeting the antibody, either with photoactivation [18] or 2-mercaptoethanol [19,20].…”

Lymphoma: current status of clinical and preclinical imaging with radiolabeled antibodies

“…Due to the short half-life of 64 Cu (12.7 h), the biodistribution of the tracer ( 64 Cu-DOTA-rituximab) was monitored for only 48 h. The spleen could be easily delineated at 24 h after injection in mice without blocking (16.5 ± 0.45 %ID/g), and the mice with blocking showed minimal uptake (16.5 ± 0.45 %ID/g). In addition, rituximab has also been labeled with 99m Tc for SPECT imaging [98] and the near-infrared dye Cy5.5 for optical imaging [99]. …”