The regulation and function of CD20: an "enigma" of B-cell biology and targeted therapy

Abstract

The introduction of anti-CD20 monoclonal antibodies such as rituximab, ofatumumab, or obinutuzumab improved the therapy of B-cell malignancies even though the precise physiological role and regulation of CD20 remains unclear. Furthermore, CD20 expression is highly variable between different B-cell malignancies, patients with the same malignancy, and even between intraclonal subpopulations in an individual patient. Several epigenetic (EZH2, HDAC1/2, HDAC1/4, HDAC6, complex Sin3A-HDAC1) and transcription factors (USF, OCT1/2, PU.1, PiP, ELK1, ETS1, SP1, NFκB, FOXO1, CREM, SMAD2/3) regulating CD20 expression (encoded by MS4A1) have been characterized. CD20 is induced in the context of microenvironmental interactions by CXCR4/SDF1 (CXCL12) chemokine signaling and the molecular function of CD20 has been linked to the signaling propensity of B-cell receptor (BCR). CD20 has also been shown to interact with multiple other surface proteins on B cells (such as CD40, MHCII, CD53, CD81, CD82, and CBP). Current efforts to combine anti-CD20 monoclonal antibodies with BCR signaling inhibitors targeting BTK or PI3K (ibrutinib, acalabrutinib, idelalisib, duvelisib) or BH3-mimetics (venetoclax) lead to the necessity to better understand both the mechanisms of regulation and the biological functions of CD20. This is underscored by the observation that CD20 is decreased in response to the "BCR inhibitor" ibrutinib which largely prevents its successful combination with rituximab. Several small molecules (such as histone deacetylase inhibitors, DNA methyl-transferase inhibitors, aurora kinase A/B inhibitors, farnesyltransferase inhibitors, FOXO1 inhibitors, and bryostatin-1) are being tested to upregulate cell-surface CD20 levels and increase the efficacy of anti-CD20 monoclonal antibodies. Herein, we review the current understanding of CD20 function, and the mechanisms of its regulation in normal and malignant B cells, highlighting the therapeutic implications.

Figure 1.

Summary of the known mechanisms of action of anti-CD20 monoclonal antibodies and an overview of potential factors affecting resistance to anti-CD20 therapy in malignant B cells. Anti-CD20 monoclonal antibodies act through several mechanisms, including complement-dependent cytotoxicity (CDC), complement-dependent cellular cytotoxicity (CDCC), antibody-dependent cellular phagocytosis (ADCP), antibody-dependent cellular cytotoxicity (ADCC), and induction of direct apoptosis.

Figure 2.

A schematic view of interacting partners of CD20 on cell membrane and mechanisms of CD20 gene (MS4A1) regulation in malignant B cells. mAbs: monoclonal antibodies; BCR: B-cell receptor; HDACi: histone deacetylase inhibitors; DNMTi: DNA methyl-transferase inhibitors; NICD: NOTCH1 intracellular domain; TGFβ: transforming growth factor β.

Figure 3.

A schematic of the proximal region of MS4A1 promoter with transcription factor binding sites. Several regulatory elements differ in nucleotide sequence when comparing data from literature and the TRANSFAC database. Shimizu et al. described the GC-box binding SP1 as being located in a region between bases -548 and -539, but the TRANSFAC database identified a GC-box in a position -234 to -224 bp [based on the SP1 chromatin immunoprecipitation sequencing data (ENCODE ID: ENCSR000BHK)]. A BAT-box was identified by Thévenin et al. in the proximal promoter region located between bases -214 and -202 (in bold), while others describe it between bases -214 and -200.^, The PU.1/PiP binding site was originally defined as a sequence between -161 and -148 bp, but TRANSFAC predicts it in the region from -120 to -107 bp (in italics). The E-box sequence is usually described as the CACCTG sequence between -44 and -39 bp (in bold)^,, but the TRANSFAC database suggests (based on ENCODE ID: ENCSR000BGI) a longer sequence from -48 to -35 bp (in italics). FOXO1 was suggested as being recruited to the MS4A1 promoter indirectly by the DNA-binding element between bases -182 and -88. NFκB binds into the region of the MS4A1 promoter between -425 and -417 bp.