Abstract 3291: The expression of CD20 on malignant B cells is regulated by chemokine signaling through the CXCR4/SDF-1 axis: implications for targeting the microenvironmental interactions
Authors: Pavlasova Gabriela , Borsky Marek , Seda Vaclav , Cerna Katerina , Osickova Jitka , Doubek Michael , Brychtova Yvona , Mayer Jiri , Pospisilova Sarka , Davids Matthew S. , Brown Jennifer R. , Mraz Marek 
Published: July 2016

Abstract
The introduction of anti-CD20 antibodies has significantly improved the outcome of patients with chronic lymphocytic leukemia (CLL) and B-cell non-Hodgkin lymphomas. The aim of this study was to analyze molecular pathways that influence expression of CD20 since this is largely unknown. This is of a great clinical interest, as combinatorial therapy of novel BCR-signaling inhibitors ibrutinib and idelalisib currently focuses mainly on the use with anti-CD20 antibodies.
Firstly, we analyzed samples obtained from CLL patients treated with ibrutinib and showed that administration of ibrutinib in vivo leads to CD20 down-modulation (P&lt;0.0001). That implies that CD20 expression might be regulated by a yet unknown mechanism in the context of immune niches. Therefore, we focused on determining the effect of microenvironmental interactions on CD20 expression on malignant B cells. We co-cultured primary CLL cells with the bone marrow stromal cell line HS-5. This revealed that CLL cells co-cultured with HS-5 had higher surface CD20 expression compared to control cells cultured on plastic (P&lt;0.01). Moreover, CLL cells that have been recently released from the lymph node microenvironment to the peripheral blood (CXCR4dimCD5bright) had ∼2-fold higher surface CD20 expression when compared to CLL cells circulating in blood of the same patient for a longer time (CXCR4brightCD5dim cells; P&lt;0.0001). Further, sorted CXCR4dimCD5bright CLL cells had also ∼2 times higher CD20 mRNA expression (P = 0.002) suggesting that changes in CD20 expression were rather due to changes in its mRNA levels than surface modulation. As CD20 expression was decreasing with the transition from CXCR4dim to CXCR4bright CLL cells, we hypothesized that the CXCR4/SDF-1 axis is directly implicated in CD20 regulation. Indeed, CLL cells treated with SDF-1α (CXCL12), a ligand for CXCR4 produced by stromal cells, significantly up-regulated surface CD20 (P&lt;0.05). On the contrary, the treatment with plerixafor, CXCR4 antagonist, inhibited this SDF-1α mediated up-regulation of CD20 which proves that SDF-1α binding to CXCR4 is directly involved in CD20 regulation.
Altogether, the CD20 levels are up-regulated within the context of microenvironmental interactions through the CXCR4/SDF-1 axis, and the impairment of microenvironmental interactions mediated by ibrutinib down-regulates CD20 expression. This study reveals a novel regulation of CD20 expression in the context of immune niches, which has important implications for CD20-targeting antibodies and the use of BCR-inhibitors in combination.
Supported by: SoMoPro II-no.4SGA8684; NGS-PTL(306242); EHA Fellowship award; Ministry of Health of CR (16-29622A); Academy of Sciences of CR (16-13334Y); Ministry of Education, Youth and Sports, grant LD15144 (COST CZ); MUNI/A/1028/2015; CZ.1.05/1.1.00/02.0068; G.P. is supported by Ostrava city.
Citation Format: Gabriela Pavlasova, Marek Borsky, Vaclav Seda, Katerina Cerna, Jitka Osickova, Michael Doubek, Yvona Brychtova, Jiri Mayer, Sarka Pospisilova, Matthew S. Davids, Jennifer R. Brown, Marek Mraz. The expression of CD20 on malignant B cells is regulated by chemokine signaling through the CXCR4/SDF-1 axis: implications for targeting the microenvironmental interactions. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3291.