Anti-CD40L immune complexes potently activate platelets in vitro and cause thrombosis in FCGR2A transgenic mice.
Anti-CD40L immunotherapy in systemic lupus erythematosus patients was associated with thromboembolism of unknown cause. We previously showed that monoclonal anti-CD40L immune complexes (ICs) activated platelets in vitro via the IgG receptor (FcgammaRIIa). In this study, we examined the prothrombotic effects of anti-CD40L ICs in vivo. Because mouse platelets lack FcgammaRIIa, we used FCGR2A transgenic mice. FCGR2A mice were injected i.v. with preformed ICs consisting of either anti-human CD40L mAb (M90) plus human CD40L, or a chimerized anti-mouse CD40L mAb (hMR1) plus mouse CD40L. ICs containing an aglycosylated form of hMR1, which does not bind FcgammaRIIa, were also injected. M90 IC caused shock and thrombocytopenia in FCGR2A but not in wild-type mice. Animals injected with hMR1 IC also experienced these effects, whereas those injected with aglycosylated-hMR1 IC did not, demonstrating that anti-CD40L IC-induced platelet activation in vivo is FcgammaRIIa-dependent. Sequential injections of individual IC components caused similar effects, suggesting that ICs were able to assemble in circulation. Analysis of IC-injected mice revealed pulmonary thrombi consisting of platelet aggregates and fibrin. Mice pretreated with a thrombin inhibitor became moderately thrombocytopenic in response to anti-CD40L ICs and had pulmonary platelet-thrombi devoid of fibrin. In conclusion, we have shown for the first time that anti-CD40L IC-induced thrombosis can be replicated in mice transgenic for FcgammaRIIa. This molecular mechanism may be important for understanding thrombosis associated with CD40L immunotherapy. The FCGR2A mouse model may also be useful for assessing the hemostatic safety of other therapeutic Abs.
Robles-Carrillo
Liza
L
Center for Thrombosis Research, Florida Hospital, Orlando, FL 32804, USA.
Meyer
Todd
T
Hatfield
Meghan
M
Desai
Hina
H
Dávila
Mónica
M
Langer
Florian
F
Amaya
Mildred
M
Garber
Ellen
E
Francis
John L
JL
Hsu
Yen-Ming
YM
Amirkhosravi
Ali
A
D000818:Animals; D000911:Antibodies, Monoclonal / Q000008:administration & dosage / Q000096:biosynthesis / Q000633:toxicity; D000936:Antigen-Antibody Complex / Q000008:administration & dosage / Q000502:physiology* / Q000633:toxicity; D001323:Autoantibodies / Q000008:administration & dosage / Q000627:therapeutic use / Q000633:toxicity*; D023201:CD40 Ligand / Q000276:immunology*; D006801:Humans; D006825:Hybridomas; D051379:Mice; D018345:Mice, Knockout; D008822:Mice, Transgenic; D015539:Platelet Activation / Q000235:genetics / Q000276:immunology*; D017452:Receptors, IgG / Q000172:deficiency / Q000235:genetics* / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000008:administration & dosage / Q000502:physiology / Q000633:toxicity; D013927:Thrombosis / Q000097:blood / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D001323:Autoantibodies; C445323:FCGR2A protein, human; C080568:Fc gamma receptor IIA; D017452:Receptors, IgG; D011993:Recombinant Fusion Proteins; D023201:CD40 Ligand