Extensive crosslinking of CD22 by epratuzumab triggers BCR signaling and caspase-dependent apoptosis in human lymphoma cells

Abstract

(2015) Extensive crosslinking of CD22 by epratuzumab triggers BCR signaling and caspase-dependent apoptosis in human lymphoma cells, mAbs, 7:1, 199-211, DOI: 10.4161/19420862.2014.979081 To link to this article: https://doi.org/10. 4161/19420862.2014 Abbreviations: Anti-IgM, F(ab') 2 fragment of affinity-purified goat anti-human IgM, Fc 5m fragment; BCR, B-cell antigen receptor; BSA, bovine serum albumin; CM-H 2 DCF-DA, 2 0 ,7 0 -dichlorodihydrofluorescein diacetate; Dc m , mitochondria membrane potential; DNP, 2,4-dinitrophenyl; EC, endothelial cells; ERKs, extracellular signal-regulated kinases; FBS, fetal bovine serum; FITC-DNase I, fluorescein isothiocyanate-conjugated DNase I; 488-annexin V, Alexa Fluor 488-conjugated annexin V; GAH, F(ab 0 ) 2 fragment of affinity-purified goat anti-human IgG Fcg fragment-specific; HUV-EC, human umbilical vein endothelial cells; ITIM, immunoreceptor tyrosine-based inhibition motif; JNKs, c-Jun N-terminal kinases; JP, jasplakinolide; LatB, latrunculin B; Lyn, Lck/Yes novel tyrosine kinase; MAP kinases, mitogen-activated protein kinases; mIgM, membrane IgM; MTS, (3-(4, 5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium; PARP, poly(ADP-ribose) polymerase; PBS, phosphate-buffered saline; PLCg2, phospholipase C, isotype gamma 2; Rhodamine-anti-IgG, rhodamine-conjugated F(ab 0 ) 2 fragment of affinity-purified goat anti-human IgG, F(ab 0 ) 2 fragment-specific; ROS, reactive oxygen species; 7-AAD, 7-aminoactinomycin D, Syk, spleen tyrosine kinase; TMRE/tetramethylrhodamine/ethyl ester Epratuzumab has demonstrated therapeutic activity in patients with non-Hodgkin lymphoma, acute lymphoblastic leukemia, systemic lupus erythematosus, and Sj€ ogren's syndrome, but its mechanism of affecting normal and malignant B cells remains incompletely understood. We reported previously that epratuzumab displayed in vitro cytotoxicity to CD22-expressing Burkitt lymphoma cell lines (Daudi and Ramos) only when immobilized on plates or combined with a crosslinking antibody plus a suboptimal amount of anti-IgM (1 mg/mL). Herein, we show that, in the absence of additional anti-IgM ligation, extensive crosslinking of CD22 by plate-immobilized epratuzumab induced intracellular changes in Daudi cells similar to ligating B-cell antigen receptor with a sufficiently high amount of anti-IgM (10 mg/mL). Specifically, either treatment led to phosphorylation of CD22, CD79a and CD79b, along with their translocation to lipid rafts, both of which were essential for effecting caspase-dependent apoptosis. Moreover, such immobilization induced stabilization of F-actin, phosphorylation of Lyn, ERKs and JNKs, generation of reactive oxygen species (ROS), decrease in mitochondria membrane potential (Dc m ), upregulation of pro-apoptotic Bax, and downregulation of anti-apoptotic Bcl-xl and Mcl-1. The physiological relevance of immobilized epratuzumab was implicated by noting that several of its in vitro effects, including apoptosis, drop in Dc m , and generation of ROS, could be obse...

Advances in targeted therapy for malignant lymphoma

“…The crosslinking of CD22 by epratuzumab triggers BCR signaling and caspase-dependent apoptosis in human lymphoma cells. 50 Preclinical studies demonstrated that CD22 mAbs had independent lymphomacidal properties. 51 Single-agent epratuzumab has been investigated in both indolent and aggressive NHLs.…”

Modulating carbohydrate–protein interactions through glycoengineering of monoclonal antibodies to impact cancer physiology

“…Second, the Fab can target siglec-sialic acid interactions of inhibitory siglecs (e.g., CD22) that normally prevent B-cell over-stimulation by inhibiting B-cell receptor signaling [15]. For example, the anti-CD22 antibody (epratuzumab) targets siglec-sialic acid interactions between B cells with endothelial cells, leading to downstream B-cell receptor signaling suppression [16]. Third, the Fab can interfere with angiogenesis by blocking interactions between glycans and VEGF, a key inducer for several downstream signaling pathways (e.g., MAPK and PI3K/AKT [17]) that lead to vascular and stromal cell ablation [18].…”

Development and characterization of a camelid single‐domain antibody directed to human CD22 biomarker

“…Traditional treatment of the leukemic patients is based on combination chemotherapy, which has considerable nonspecific cytotoxicity. Molecular immunology improvements have brought encouraging diagnostic and therapeutic advances to hematologic malignancies including leukemia and the first antibody target has been CD20 .…”

“…In addition, different studies have shown the effects of Epratuzumab, a humanized mAb which targets CD22, on the B cells and this mAb is under clinical investigation for treatment of leukemia .…”