Calcein release assay as a method for monitoring serum complement activity during monoclonal antibody therapy in patients with B-cell malignancies

Grzegorz Stasiłojć , Anna Felberg , Aleksandra Urban , Daria Kowalska , Shuo Ma , Anna Maria Blom , Jeanette Lundin , Anders Österborg , Marcin Okrój


Monoclonal antibodies ofatumumab (anti-CD20) and alemtuzumab (anti-CD52) which are approved for usage in patients with chronic lymphocytic leukemia (CLL), efficiently activate the classical complement pathway. However complement is an exhaustible component and high doses of its activators may deplete complement-dependent cytotoxicity (CDC) potential, thus reducing the effect of repeated mAb dosing. Widely used method to measure CDC activity of patients' serum is hemolytic assay (CH50) on sheep erythrocytes. Despite its simplicity, such CH50 assay may not reflect pivotal interactions between patient serum and human complement inhibitors on the surface of target cells. We propose calcein release assay performed on tumor cells similar to those targeted by therapeutic antibodies as an alternative method. We analyzed serum samples collected from 12 patients participating in the clinical study, receiving s.c. 30 mg alemtuzumab three times per week combined with i.v. ofatumumab at an initial dose of 300 mg in week 3 further escalated to 2000 mg every other week. All serum samples were measured by hemolytic assay on sheep erythrocytes as well as using calcein release assay on CD20-positive Raji cells. Our data show that results obtained from both assays are related to each other at the level of the whole group (n = 96 samples, Spearman r = 0.504, p < .001) but may substantially differ when analyzing individual patients. Furthermore, by using CDC assay on Raji cells, we found that in the presented clinical study CDC serum potential was not significantly affected when measured before consecutive administrations in most of the patients.
Author Grzegorz Stasiłojć (IFB/DMedB)
Grzegorz Stasiłojć,,
- Department of Medical Biotechnology
, Anna Felberg (IFB/DMedB)
Anna Felberg,,
- Department of Medical Biotechnology
, Aleksandra Urban (IFB)
Aleksandra Urban,,
- Intercollegiate Faculty of Biotechnology UG
, Daria Kowalska (IFB)
Daria Kowalska,,
- Intercollegiate Faculty of Biotechnology UG
, Shuo Ma
Shuo Ma,,
, Anna Maria Blom
Anna Maria Blom,,
, Jeanette Lundin
Jeanette Lundin,,
, Anders Österborg
Anders Österborg,,
, Marcin Okrój (IFB/DMedB)
Marcin Okrój,,
- Department of Medical Biotechnology
Journal seriesJournal of Immunological Methods, ISSN 0022-1759, e-ISSN 1872-7905, (N/A 70 pkt)
Issue year2020
Publication size in sheets0.50
Article number112675
Keywords in EnglishOfatumumab, alemtuzumab, CLL, complement system
ASJC Classification2723 Immunology and Allergy; 2403 Immunology
Languageen angielski
LicenseOther; published final; Uznanie Autorstwa (CC-BY); with publication
Score (nominal)70
Score sourcejournalList
ScoreMinisterial score = 70.0, 14-02-2020, ArticleFromJournal
Publication indicators Scopus SNIP (Source Normalised Impact per Paper): 2018 = 0.648; WoS Impact Factor: 2018 = 1.913 (2) - 2018=2.105 (5)
Citation count*
Share Share

Get link to the record

* presented citation count is obtained through Internet information analysis and it is close to the number calculated by the Publish or Perish system.
Are you sure?