Purine-Rich Box-1–Mediated Reduced Expression of CD20 Alters Rituximab-Induced Lysis of Chronic Lymphocytic Leukemia B Cells

Abstract

The anti-CD20 monoclonal antibody rituximab has been less successful in treating chronic lymphocytic leukemia (CLL) than lymphoma, possibly due to the lower density of CD20 on B lymphocytes from CLL patients than on those from lymphoma patients. This lowering may result from insufficiency of one of the transcription factors of cd20. Of these, purine-rich box-1 (PU.1) is poorly expressed in CLL. To estimate its weight in CD20 expression, pu.1 cDNA was transfected into CLL B cells and shown to raise the membrane expression of CD20 and to improve the rituximabinduced lysis of transfected cells. Granulocyte macrophage colony-stimulating factor and all-trans-retinoic acids were not involved in the defective expression of PU.1 or the excessive methylation of the pu.1 gene, because 6 of 14 CLL samples tested were normally methylated. This was confirmed by the failure of DNA methyltransferase inhibitors to restore pu.1 transcription in hypermethylated CLL, and, in fact, the expression of PU.1 was down-regulated by excessive expression of the FMS proto-oncogene-like tyrosine kinase 3 (Flt3) receptor. This abnormality is consistent with our finding of elevated levels of Flt3 ligand (FL) in 20 of 23 CLL sera tested. We propose that FL-dependent increased Flt3 signaling prevents the expression of PU.1, which down-regulates that of CD20, and accounts for resistance of leukemic B cells to rituximab-induced lysis.

Prenyltransferases Regulate CD20 Protein Levels and Influence Anti-CD20 Monoclonal Antibody-mediated Activation of Complement-dependent Cytotoxicity

“…In CLL, rituximab-mediated down-modulation of CD20 is associated with reduced levels of CD20 mRNA both in vitro (16) and in vivo (17), indicating regulation at the level of transcription. For example, activated Flt3 signaling cascade has been reported to inhibit expression of PU.1, a transcription factor involved in the expression of CD20 gene (18). Down-regulation of CD20 mRNA has been also observed in CD20-negative cells obtained from patients after relapse of rituximab-treated B-cell malignancies (15).…”

A new approach to comparing anti-CD20 antibodies: importance of the lipid rafts in their lytic efficiency

“…However, resistance to RTX was identified in certain diseases, and especially in CLL. Mankai et al and others have established that this is related to a lower density of CD20 on B-cells in CLL compared to B-cells from other lymphomas, 65 associated with a down-regulation of the purine-rich box-1 (PU.1) transcription factor, so that transfection of B-cells with its cDNA restores RTX-induced lysis. Similarly, stimulation of B-cells with CpG increases the expression of CD20 and ameliorates the CDC by RTX.…”

“…[72][73][74] When plasma membrane lipids and protein are clustered in the LRs, they can mediate transduction pathways which favor cellular adhesion, transmembrane signaling, virus budding, control of ionic pumps and channels, and mediation of vesicle fusion. 75,65 Proteins connected to LRs have been described as insoluble in the nonionic detergent Triton X-100 at low temperature, in contrast to the other membrane lipids. 70 This means we should be careful when investigating proteins linked to rafts, as analyses of fluorescence microscopy or fluorescence resonance energy transfer confirm these data.…”

B-cell Chronic Lymphocytic Leukemia/Small Lymphocytic Lymphoma, without the development of Richter's Syndrome, with Neoplastic Cells Lacking CD20 Antigen Expression after Rituximab Treatment

“…The prognosis of patients who develop RS is very poor, with survival rate of 7 to 8-month, even in the patient treated with R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisolone), hyper-CVAD (cyclophosphamide, vincristine, doxorubicin, and dexamethasone) or a purine analogue regimen [1]. Aggressive cases of B-CLL/SLL, which develop into RS with cells that lack CD20 antigen expression have also been reported [8][9][10]. However, the cases of B-CLL/SLL with cells that lack CD20 antigen expression without RS development, are not fully understood.…”

“…Furthermore, the bone marrow neoplastic cells were positive for CD5, CD19 ( Figure 3C), CD23, CD25, and λ-chain of immunoglobulin light chain and negative for not only CD3 and CD10, but also for CD20 ( Figure 3D), which was found to be positive at the initial diagnosis ( Figure 3B) by the cell surface marker analysis using flow cytometry. Complicated chromosomal abnormalities including abnormalities in chromosomes 3,6,7,8,9,13,15,16,17, and 19, were detected in 15 of 20 analyzed BMCs by the G-banding method. The abnormalities in chromosome 11, which were detected at the initial diagnosis, were not included.…”