Membrane microdomain sphingolipids are required for anti-CD20-induced death of chronic lymphocytic leukemia B cells

Abstract

The lipid organization of membranes of B cells from patients with chronic lymphocytic leukemia differs from one patient to another. In practice, given the relevance of the membrane lipid distribution to the efficacy of biotherapies, this observation is of potential importance.

Resistance to complement activation, cell membrane hypersialylation and relapses in chronic lymphocytic leukemia patients treated with rituximab and chemotherapy

“…Next we compared CD20 (target antigen), GM1 and sphingomyelin (lipid raft markers) in vitro expression between the two patient groups as the ability of CD20 to translocate into lipid rafts is suspected to be important for complement activation by anti-CD20 mAb [ 31 ], and that lipid raft integrity may be compromised in CLL cells [ 25 , 26 , 32 ] (Figure 3A and Table 2 ). No differences were observed when comparing in both groups (i) the numbers of CD20 molecules between groups; (ii) the mean fluorescence intensity (MFI) of the FITC-conjugated cholera toxin subunit B recognizing GM1; and (iii) the MFI of lysenin-bound-sphingomyelin.…”

“…No differences were observed when comparing in both groups (i) the numbers of CD20 molecules between groups; (ii) the mean fluorescence intensity (MFI) of the FITC-conjugated cholera toxin subunit B recognizing GM1; and (iii) the MFI of lysenin-bound-sphingomyelin. As we have previously observed that sphingomyelin overexpression in CLL cells induced by rifampicin affects the type II anti-CD20 mAb (B1) capacity to kill CLL cells [ 32 ], the experiment was repeated with RTX instead of B1 (tositumomab), revealing that sphingomyelin overexpression had no effect on the RTX capacity to induce CDC (data not shown).…”

“…It's important to note that four patients from the CDC-resistant group have received Ofatumumab at relapse and that such administration was associated with an important C4 complement exhaustion [ 38 ]. A part of the patients from our cohort (47%) was also tested in vitro with the type II anti-CD20 mAb B1 (tositumomab) that triggers cell death by altering lipid organization in a mechanism independent from CDC and CLL hypersialylation ([ 32 ] and Figure 4C ). Accordingly, future research is necessary to appreciate whether determining hypersialylation might be useful to propose a new generation anti-CD20 mAb as an alternative to RTX.…”

“…The percentage of CDC decreased survival was calculated by using the formula: 100x [(% of AV-PI- cells without RTX in HSAB)-(% of AV-PI- cells with RTX in HSAB)]/[% of AV-PI- cells without RTX in HSAB]. In selected experiments, direct apoptosis and ADCC [ 25 , 32 ] were monitored as previously described; and cells were additionally treated with 0.05U of neuraminidase (Sigma-Aldrich) for 30 minutes at 37°C, and two washes were then performed to avoid any side effects of neuraminidase on RTX or B1 (10μg/ml, generous gift of Cragg MS, Southampton, UK) activity.…”

Gene expression profiling identifies ARSD as a new marker of disease progression and the sphingolipid metabolism as a potential novel metabolism in chronic lymphocytic leukemia

“…Specific changes in the composition and metabolism of GSLs occur during cell proliferation, cell cycle phases, brain development, differentiation, and neoplasia in various cell types, indicating that the ganglioside composition responds to changes in the morphology and function of cells [43]. Hammadi et al showed that B cells are diverse from one CLL patient to another with respect to their membrane lipid organization [11]. In particular, they observed that CLL B cells were characterized by high GM1 gangliosides and SM (Sphingomyelin) levels in their plasma membrane, they possessed active P-gp (Pglycoprotein) pumps, recruited Cbp (Csk-binding protein), and ultimately they were sensitive to B1 mAb-induced apoptosis.…”