Abstract
Anti-CD20 monoclonal antibodies (mAbs) represent an effective treatment for a number of B cell malignancies and autoimmune disorders. Glycoengineering of anti-CD20mAb may contribute to increased anti-tumor efficacy through enhanced antibody-dependent cellular cytotoxicity (ADCC) and phagocytosis (ADP) as reported by in vitro studies. However, where and how glycoengineered Ab may potentiate therapeutic responses in vivo is yet to be elucidated. Here, we have performed mouse liver transplants to demonstrate that the liver is sufficient to mediate systemic B cells depletion after anti-CD20 treatment. Relying on intravital two-photon imaging of human CD20-expressing mice, we provide evidence that ADP by Kupffer cells (KC) is a major mechanism for rituximab-mediated B cell depletion. Notably, a glycoengineered anti-mouse CD20 Ab but not its wild-type counterpart triggered potent KC-mediated B cell depletion at low doses. Finally, distinct thresholds for KC phagocytosis were also observed for GA101 (obinutuzumab), a humanized glycoengineered type II anti-CD20 Ab and rituximab. Thus, we propose that enhanced phagocytosis of circulating B cells by KC represents an important in vivo mechanism underlying the improved activity of glycoengineered anti-CD20 mAbs.
Highlights
Anti-CD20 monoclonal antibodies represent an effective treatment for a number of B cell malignancies and autoimmune disorders
We have previously shown using partial hepatectomy that the liver is necessary for optimal B cell depletion by a mouse anti-CD20 Ab (5D2)[5], a mechanism shown to be dependent on activating FcRs5,16
We and others have shown that splenectomy has little effect on anti-CD20-mediated B cell depletion[5,7]
Summary
To prepare mouse donor livers, ligaments, esophageal, pyloric, right adrenal veins and the hepatic artery were cut. Liver were perfused and harvested after cutting the inferior vena cava, the portal vein, and the superior vena cava. For mouse recipient liver removal, sutures were placed on the portal vein and the superior vena cava, the portal vein, and the inferior vena cava were clumped before being cut. Anastomoses were performed by suture or by sliding recipient vessels over cuffed donor vessels. Wild type C57BL/6 mice were purchased from Charles River France. HCD20Tg mice[7] (Genentech, USA), C57BL/6 GFP+ and FcRγ−/− mice were bred in our facility. Anti-CD20 mAbs included 5D2 (mIgG2a, Genentech), WT and GE 18B12 (mIgG2a, Roche), rituximab and GA101 (Hôpital Necker), HY1.2 (mIgG2a) isotype control and mg[053] (hIgG1) isotype control. A coversplit was placed on the top of the liver maintained by silicone paste
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