An approach for chemical evaluation of immunoconjugates of “cold” 177lutetium-rituximab

Gjorgieva Ackova, Darinka and Smilkov, Katarina and Stafilov, Trajče and Kiprijanovska, Sanja and Sukarova Stefanovska, Emilija and Janevik-Ivanovska, Emilija (2014) An approach for chemical evaluation of immunoconjugates of “cold” 177lutetium-rituximab. In: XXIII Congress of Chemists and Technologists of Macedonia, 8-11 Oct 2014, Ohrid, Macedonia.

[thumbnail of poster-70x100_DGA.pdf]

Download (539kB) | Preview


Various radiolabeled monoclonal antibodies have been developed for the treatment and diagnosis of malignancies. Rituximab is a chimeric mouse-human monoclonal antibody. Rituximab selectively binds with high affinity to the CD20 antigen (human B-lymphocyte restricted differentiation antigen, Bp35), a hydrophobic transmembrane protein, which is expressed on B-lymphocytes and on >90% of B cell non-Hodgkin’s lymphomas. These properties make the CD20 receptor a suitable target for radioactive therapy. 177Lu has been considered as a promising radionuclide for developing therapeutic radiopharmaceuticals, esp. radiolabeled monoclonal antibodies, owing to its suitable decay characteristics, as well as the feasibility of large-scale production in adequate specific activity and radionuclidic purity.
In order to obtain 177Lu anti-CD20 radioimmunoconjugates for using in therapeutic studies, different bifunctional chelating agents-antiCD20 (rituximab) (BFCA-rituximab) were labeled by “cold” 177Lu chloride for preliminary chemical characterization. Rituximab, conjugated with three different BFCA, p-SCN-Bn-DOTA, p-SCN-Bn-DTPA and 1B4M-DTPA in a form of freeze-dried preparation was dissolved with sterile saline, and subsequent labelled with 1.0709 µg Lu chloride which is equievalent to MTD for 177Lu (118.3 mCi) in a total volume of 1mL at pH 7.0, and incubation for 30 min at room temperature (p-SCN-Bn-DTPA-rituximab and 1B4M-DTPA-rituximab) and 60 min at 40°C (p-SCN-Bn-DOTA-rituximab). Characterization of the conjugated BFCA-MoAb and determination of the average number of BFCA attached to each antibody molecule was performed by Matrix-Assisted Laser Desorption Ionization time-of-flight (MALDI-ToF) mass spectrometry. MALDI-TOF experiments revealed the presence of two major peaks corresponding to a MW of conjugated and unconjugated rituximab equivalent to an average of 7.7 (p-SCN-Bn-DOTA), 11 (p-SCN-Bn-DTPA) and 9.8 (1B4M-DTPA) groups per molecule of antibody. In order to examine relevant quality parameters, including detection, and purity assessment, SDS-PAGE experiments, under reducing conditions, were performed. All three BFCA-rituximab conjugates (labeled and non-labeled) were resolved in two distinct Mr species which migrated in two bands (upper band at ~50 kDa and lower band ~30 kDa) confirming the migration behavior typical for IgG antibodies which are composed of two identical subunits each composed by two polypeptide chains: two heavy and two light chains, linked via disulfide bonds.
The next step is radio labeling of BFCA-rituximab conjugates and their evaluation in order to find the most appropriate form for potential radiopharmaceutical for treatment of patients of relapsed and refractory non Hodgkin's lymphoma.

Item Type: Conference or Workshop Item (Poster)
Subjects: Natural sciences > Chemical sciences
Medical and Health Sciences > Other medical sciences
Divisions: Faculty of Medical Science
Depositing User: Darinka Gorgieva Ackova
Date Deposited: 25 Nov 2014 14:15
Last Modified: 26 Nov 2014 08:19

Actions (login required)

View Item View Item