Abstract 4744: CD22-targeting epratuzumab mediates trogocytosis of multiple cell-surface markers on normal, malignant, and lupus B cells.
Authors: Rossi Edmund A. , Goldenberg David M. , Michel Rosana , Rossi Diane L. , Wallace Daniel J. , Chang Chien-Hsing 
Published: August 2014

Background. Epratuzumab, a humanized anti-CD22 antibody, is currently in clinical trials of B-cell lymphoma and autoimmune diseases, demonstrating therapeutic activities in non-Hodgkin lymphoma (NHL) and systemic lupus erythematosus (SLE). Thus, epratuzumab offers a promising option for CD22-targeted immunotherapy. Although epratuzumab is capable of depleting on average 35% of circulating B cells in patients, its in vivo mechanism of action (MOA) remains incompletely understood. We hypothesized that ligation of epratuzumab to CD22 could modulate other surface molecules involved in regulating B-cell antigen receptor (BCR) signaling, leading to altered B-cell functions that ultimately mitigate symptoms of the underlying diseases. Methods. Peripheral blood mononuclear cells (PBMCs) from either healthy donors or lupus patients with flares, were incubated with epratuzumab, and the relative surface levels of CD22 and selected BCR regulators, including CD19, CD21, and CD79b, were analyzed by flow cytometry. Results. Epratuzumab promptly induced a marked decrease of CD22 (&gt;80%), CD19 (&gt;50%), CD21 (&gt;50%), and CD79b (&gt;30%) on the surface of B cells in PBMCs obtained from normal donors or treatment-naïve lupus patients, and of NHL cells (Daudi and Raji) spiked into normal PBMCs. Although some Fc-independent loss of CD22 is expected from its internalization by epratuzumab, the concurrent and prominent reduction of CD19, CD21, and CD79b is Fc-dependent and results from trogocytosis of epratuzumab-bound B cells to FcγR-expressing effector cells, including monocytes, NK cells, and granulocytes. Following incubation with epratuzumab, but not with an isotype control mAb, reduced staining of surface antigens on B cells coincided with positive CD19 and CD22 staining of the effector cells. In the absence of PBMCs, treatment of NHL cell lines (Daudi and Raji) with epratuzumab resulted in more than 80% reduction of CD22, with little, if any, reduction in CD19, CD21, CD79b or surface IgM observed. Inclusion of a crosslinking second antibody with epratuzumab induced only a minimal (&lt;15%) reduction of CD19 and CD21. When these NHL cells (1 x 105 cells) were mixed with PBMCs (1 x 106 cells), epratuzumab induced a 40 to 70% reduction of CD19 and CD21, with significant down-regulation of surface IgM and CD79b also. Under the conditions examined, rituximab at 10 μg/mL reduced the B-cell count by 50%, whereas epratuzumab did not cause significant B-cell depletion, either at 10 μg/mL or 1 mg/mL. Conclusions. This study revealed a previously unknown, and potentially important, MOA of epratuzumab. Whether the observed trogocytosis could be correlated with the depletion of malignant B cells in lymphoid tissues is currently under investigation.

Citation Format: Edmund A. Rossi, David M. Goldenberg, Rosana Michel, Diane L. Rossi, Daniel J. Wallace, Chien-Hsing Chang. CD22-targeting epratuzumab mediates trogocytosis of multiple cell-surface markers on normal, malignant, and lupus B cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4744. doi:10.1158/1538-7445.AM2013-4744