title: Cytokine release assays for the prediction of therapeutic mAb safety in first-in man trials--Whole blood cytokine release assays are poorly predictive for TGN1412 cytokine storm.
abstract: The therapeutic monoclonal antibody (mAb) TGN1412 (anti-CD28 superagonist) caused near-fatal cytokine release syndrome (CRS) in all six volunteers during a phase-I clinical trial. Several cytokine release assays (CRAs) with reported predictivity for TGN1412-induced CRS have since been developed for the preclinical safety testing of new therapeutic mAbs. The whole blood (WB) CRA is the most widely used, but its sensitivity for TGN1412-like cytokine release was recently criticized. In a comparative study, using group size required for 90% power with 5% significance as a measure of sensitivity, we found that WB and 10% (v/v) WB CRAs were the least sensitive for TGN1412 as these required the largest group sizes (n = 52 and 79, respectively). In contrast, the peripheral blood mononuclear cell (PBMC) solid phase (SP) CRA was the most sensitive for TGN1412 as it required the smallest group size (n = 4). Similarly, the PBMC SP CRA was more sensitive than the WB CRA for muromonab-CD3 (anti-CD3) which stimulates TGN1412-like cytokine release (n = 4 and 4519, respectively). Conversely, the WB CRA was far more sensitive than the PBMC SP CRA for alemtuzumab (anti-CD52) which stimulates FcγRI-mediated cytokine release (n = 8 and 180, respectively). Investigation of potential factors contributing to the different sensitivities revealed that removal of red blood cells (RBCs) from WB permitted PBMC-like TGN1412 responses in a SP CRA, which in turn could be inhibited by the addition of the RBC membrane protein glycophorin A (GYPA); this observation likely underlies, at least in part, the poor sensitivity of WB CRA for TGN1412. The use of PBMC SP CRA for the detection of TGN1412-like cytokine release is recommended in conjunction with adequately powered group sizes for dependable preclinical safety testing of new therapeutic mAbs.
authors.affiliation: National Institute for Biological Standards and Control, Biotherapeutics Group, Blanche Lane, South Mimms, Potters Bar, Hertfordshire EN6 3QG, United Kingdom. Electronic address: Sandrine.vessillier@nibsc.org.
authors.forename: S
authors.initials: S
authors.lastname: Vessillier
authors.affiliation: National Institute for Biological Standards and Control, Biotherapeutics Group, Blanche Lane, South Mimms, Potters Bar, Hertfordshire EN6 3QG, United Kingdom.
authors.forename: D
authors.initials: D
authors.lastname: Eastwood
authors.affiliation: National Institute for Biological Standards and Control, Biotherapeutics Group, Blanche Lane, South Mimms, Potters Bar, Hertfordshire EN6 3QG, United Kingdom.
authors.forename: B
authors.initials: B
authors.lastname: Fox
authors.affiliation: MRC Centre for Drug Safety Science, Department of Molecular & Clinical Pharmacology, University of Liverpool, Sherrington Buildings, Ashton Street, Liverpool L69 3GE, United Kingdom.
authors.forename: J
authors.initials: J
authors.lastname: Sathish
authors.affiliation: MRC Centre for Drug Safety Science, Department of Molecular & Clinical Pharmacology, University of Liverpool, Sherrington Buildings, Ashton Street, Liverpool L69 3GE, United Kingdom.
authors.forename: S
authors.initials: S
authors.lastname: Sethu
authors.affiliation: National Institute for Biological Standards and Control, Biotherapeutics Group, Blanche Lane, South Mimms, Potters Bar, Hertfordshire EN6 3QG, United Kingdom.
authors.forename: T
authors.initials: T
authors.lastname: Dougall
authors.affiliation: National Institute for Biological Standards and Control, Biotherapeutics Group, Blanche Lane, South Mimms, Potters Bar, Hertfordshire EN6 3QG, United Kingdom.
authors.forename: S J
authors.initials: SJ
authors.lastname: Thorpe
authors.affiliation: National Institute for Biological Standards and Control, Biotherapeutics Group, Blanche Lane, South Mimms, Potters Bar, Hertfordshire EN6 3QG, United Kingdom.
authors.forename: R
authors.initials: R
authors.lastname: Thorpe
authors.affiliation: National Institute for Biological Standards and Control, Biotherapeutics Group, Blanche Lane, South Mimms, Potters Bar, Hertfordshire EN6 3QG, United Kingdom; MRC Centre for Drug Safety Science, Department of Molecular & Clinical Pharmacology, University of Liverpool, Sherrington Buildings, Ashton Street, Liverpool L69 3GE, United Kingdom; Medimmune, Aaron Klug Building, Granta Park, Cambridge CB21 6GH, United Kingdom. Electronic address: stebbingsr@medimmune.com.
authors.forename: R
authors.initials: R
authors.lastname: Stebbings
mesh_terms: D000074323:Alemtuzumab; D000911:Antibodies, Monoclonal / Q000008:administration & dosage; D061067:Antibodies, Monoclonal, Humanized / Q000009:adverse effects* / Q000494:pharmacology / Q000627:therapeutic use; D000970:Antineoplastic Agents / Q000009:adverse effects / Q000627:therapeutic use; D016207:Cytokines / Q000097:blood* / Q000378:metabolism; D000077561:Daclizumab; D004912:Erythrocytes / Q000378:metabolism; D015200:Fluoroimmunoassay / Q000379:methods; D006021:Glycophorins / Q000378:metabolism; D006801:Humans; D007074:Immunoglobulin G / Q000494:pharmacology / Q000627:therapeutic use; D007963:Leukocytes, Mononuclear / Q000187:drug effects / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents; D016207:Cytokines; C409972:GYPA protein, human; D006021:Glycophorins; D007074:Immunoglobulin G; D000074323:Alemtuzumab; D000077561:Daclizumab; C509303:TGN-1412
keywords: Cytokine release assay; Group size; PBMC; TGN1412; Therapeutic mAb; Whole blood