title: Interaction with FcγRIIB is critical for the agonistic activity of anti-CD40 monoclonal antibody.
abstract: A high activatory/inhibitory FcγR binding ratio is critical for the activity of mAb such as rituximab and alemtuzumab that attack cancer cells directly and eliminate them by recruiting immune effectors. Optimal FcγR binding profiles of other anti-cancer mAb, such as immunostimulatory mAb that stimulate or block immune receptors, are less clear. In this study, we analyzed the importance of isotype and FcγR interactions in controlling the agonistic activity of the anti-mouse CD40 mAb 3/23. Mouse IgG1 (m1) and IgG2a (m2a) variants of the parental 3/23 (rat IgG2a) were engineered and used to promote humoral and cellular responses against OVA. The mouse IgG1 3/23 was highly agonistic and outperformed the parental Ab when promoting Ab (10-100-fold) and T cell (OTI and OTII) responses (2- to >10-fold). In contrast, m2a was almost completely inactive. Studies in FcγR knockout mice demonstrated a critical role for the inhibitory FcγRIIB in 3/23 activity, whereas activatory FcγR (FcγRI, -III, and -IV) was dispensable. In vitro experiments established that the stimulatory effect of FcγRIIB was mediated through Ab cross-linking delivered in trans between neighboring cells and did not require intracellular signaling. Intriguingly, activatory FcγR provided effective cross-linking of 3/23 m2a in vitro, suggesting the critical role of FcγRIIB in vivo reflects its cellular distribution and bioavailability as much as its affinity for a particular Ab isotype. In conclusion, we demonstrate an essential cross-linking role for the inhibitory FcγRIIB in anti-CD40 immunostimulatory activity and suggest that isotype will be an important issue when optimizing reagents for clinical use.
authors: Division of Cancer Sciences, Faculty of Medicine, University of Southampton, Southampton SO16 6YD, United Kingdom. alw1@soton.ac.uk
authors: Ann L
authors: AL
authors: White
authors: H T Claude
authors: HT
authors: Chan
authors: Ali
authors: A
authors: Roghanian
authors: Ruth R
authors: RR
authors: French
authors: C Ian
authors: CI
authors: Mockridge
authors: Alison L
authors: AL
authors: Tutt
authors: Sandra V
authors: SV
authors: Dixon
authors: Daniel
authors: D
authors: Ajona
authors: J Sjef
authors: JS
authors: Verbeek
authors: Aymen
authors: A
authors: Al-Shamkhani
authors: Mark S
authors: MS
authors: Cragg
authors: Stephen A
authors: SA
authors: Beers
authors: Martin J
authors: MJ
authors: Glennie
mesh_terms: D000818:Animals; D058846:Antibodies, Monoclonal, Murine-Derived / Q000276:immunology / Q000493:pharmacokinetics / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000276:immunology / Q000493:pharmacokinetics / Q000494:pharmacology; D019013:CD40 Antigens / Q000276:immunology*; D007152:Immunologic Capping / Q000187:drug effects* / Q000276:immunology; D051379:Mice; D018345:Mice, Knockout; D051381:Rats; D017452:Receptors, IgG / Q000276:immunology*; D015398:Signal Transduction / Q000187:drug effects / Q000276:immunology
chemical_list: D058846:Antibodies, Monoclonal, Murine-Derived; D000970:Antineoplastic Agents; D019013:CD40 Antigens; C485700:Fcgr3 protein, mouse; D017452:Receptors, IgG