Different Pattern of CD154 and CD40 Expression On B-CLL and T Lymphocytes In Peripheral Blood, Bone Marrow and Lymph Node Microenvironment In B-Cell Chronic Lymphocytic Leukemia (B-CLL)
Authors: Jaksic Ozren , Gizdic Branimir , Veic Tajana Stoos , Jaksic Vlatka Pandzic , Kusec Rajko , Pejsa Vlatko , Jaksic Branimir 
Published: October 2019

Abstract
Abstract 3607

Background:
In B-cell chronic lymphocytic leukemia (B-CLL) there is a well documented intraclonal and interclonal variability of B-CLL cells in different lymphoid compartments with respect to the expression of a number of surface and intracellular molecules (for example CD38 and ZAP-70). This variability in part may reflect a number of interactions of malignant B-CLL clone with supporting microenviroment including cells (T-cells, nurse-like cells, etc.), cytokines, chemokines and stroma. One of the key interactions of B-CLL clone is with T-cells, through CD154/CD40 system. It is important pathway modulating survival, drug resistance and immunity. It is known that CD154 is transiently expressed on CD4+ T cells, as well as that CD154 can be coexpressed on B-CLL cells with CD40 in a subpopulation of B-CLL patients. Its expression on B-CLL cells can be induced by gene therapy and lenalidomide, being in part responsible for their therapeutic effects. Aim of this study was to determine the level of expression of CD154 and CD40 in vivo on B-CLL cells and T lymphocytes and to evaluate intra and interclonal differences due to different microenvironment, i.e. peripheral blood, bone marrow and lymph nodes.

Methods:
peripheral blood, (PB), bone marrow (BM) and lymph node (LN) samples were taken by conventional techniques (venepuncture and fine needle aspiration) on the same day. The expression level of CD154 and CD40 molecules on CD19+CD5+ B-CLL cells and CD19-CD5+ T cells was analyzed by flow cytometry. Results were expressed as mean fluorescence intensity (MFI) and analyzed by paired tests.

Results:
samples taken from 21 typical B-CLL patients with median age of 72 years were analyzed. There were 9 males and 12 females. Mean beta-2 microglobuin was 4.3mg/l, mean Total Tumor Mass size was 8.9 and mean Tumor Distribution pattern was 0.75. There were 2, 14 and 5 patients in Rai stage 0, I+II and III+IV, respectively. There were 6 previously treated patients (but off therapy 3 months before sampling). The expression level of CD154 was absent/low on T-cells and in 14/21 patients on B-CLL cells. However in 7/21 patients B-CLL cells had higher CD154 expression (“CD154 positive” patients). There was no detectible difference in CD154 expression on T cells between compartments, while on B-CLL cells there was highest expression in lymph nodes and lowest in peripheral blood (p<0.01). CD40 expression on B-CLL cells was significantly higher than CD154, i.e. all cases were positive, and there was no significant difference between lymphoid compartments. There was no significant difference between CD154 positive and negative patients in measured disease parameters.

Conclusions:
our results show that CD154 expression on T-cells is absent/low and not significantly different between lymphoid compartments regardless of different microenvironment milieu. CD40 expression on B-CLL cells is high and comparable through compartments. In subset of patients there is CD154 positivity on B-CLL cells and shows strong association with lymphoid compartments possibly indicating microenviroment influence on CD154/CD40 system in B-CLL in vivo. These results warrant further studies to indentify the role of CD154 expression on B-CLL cells in pathologic process and its regulation and may eventually uncover novel or modulate existing innovative therapeutic approaches (like gene therapy or immunomodulatory agents like lenalidomide).

Disclosures:
No relevant conflicts of interest to declare.