High Lymphoid Enhancer-Binding Factor-1 (LEF1) Expression Is Associated with ZAP70 Positivity, Requirement of Treatment, and Fibromodulin (FMOD) Expression In Chronic Lymphocytic Leukemia (CLL)

Abstract 1715

Chronic lymphocytic leukemia has the highest incidence of all leukemias in the western world. It is characterized by accumulation of monoclonal CD5+ B-lymphocytes and has a highly variable clinical outcome. Lymphoid enhancer-binding factor-1 (LEF1) is part of the LEF/TCF transcription factor family which has been shown to play an important role in regulating Wnt-pathway target genes. It is specifically expressed at early stages of B-cell differentiation and is essential for survival and proliferation. LEF1 plays a crucial role in many human cancers and was shown to promote oncogenic transformation and invasiveness of neoplastic diseases. LEF1 has also been shown to be highly overexpressed in CLL. However, the extent of overexpression shows a high variability between patients. The extracellular matrix protein fibromodulin (FMOD) is a tumor-associated antigen and one of the most overexpressed genes in CLL. It was also shown to be associated with p53-mutation and resistance to DNA damage in CLL cells.

The aim of this study was to investigate the prognostic relevance of LEF1 and FMOD expression in CLL. Efficiency-corrected quantitative real-time PCR was used to determine LEF1 and FMOD mRNA expression in 120 and 96 primary CLL samples, respectively. The expression of each sample was calibrated against the mean of 7 healthy B-cell samples. In other words, the calibrated mean relative expression ratio (RER) of the healthy B-cells is 1 by definition, and all other RER values are expressed as fold change compared to healthy B-cells. We assessed correlations with the prognostic markers ZAP70 and CD38 as well as with the percentage of lymphocytes in the peripheral blood. We also investigated correlations of LEF1 and FMOD with each other. Moreover, we compared LEF1 and FMOD expression of patients requiring treatment with the expression of patients in recently diagnosed Binet stage A.

We determined a 40-fold higher mean LEF1 mRNA expression and a 16-fold higher mean FMOD expression in our CLL patients' samples compared to the healthy B-cell samples (p<0.001). Patients requiring treatment showed a mean LEF1 mRNA relative expression ratio (RER) of 85.61 whereas patients in recently diagnosed Binet stage A had a mean of only 22.01 (p<0.001). Moreover, mean LEF1 RERs were 53.72 and 37.10 in ZAP70-positive and ZAP70-negative patients, respectively (p=0.004). Furthermore, we found a highly significant positive correlation of LEF1 with the percentage of lymphocytes in the peripheral blood (Spearman's rho = 0.440, p<0.001). We also performed a LEF1 median split dividing the patients in high and low LEF1 subgroups. We then compared the percentages of lymphocytes in the peripheral blood in both subgroups. We observed mean lymphocyte percentages of 88.45% and 76.28% in the high versus low LEF1 groups, respectively (p<0.001). However, we did not observe a significant difference in LEF1 expression between CD38-positive and CD38-negative patients. We also found LEF1 and FMOD expression to be highly correlated with each other (Spearman's rho = 0.405, p<0.001). In addition, the mean FMOD RER was 34.14 in the high LEF1 group and only 4.19 in the low LEF1 group (p<0.001). Given the fact that FMOD expression, p53 mutation, and DNA-damage resistance have been found previously to be associated, the LEF1–FMOD correlation is an interesting new finding. Eventually, it may lend additional support to the prognostic relevance of LEF1 overexpression in CLL. FMOD was also significantly higher expressed in patients requiring treatment compared to patients in recently diagnosed Binet stage A (p=0.042). However, we could not observe any significant association of FMOD with ZAP70, CD38 or the percentage of lymphocytes in the peripheral blood.

In a nutshell, our results suggest that high LEF1 expression is associated with poor prognosis, high FMOD expression, and the requirement of treatment in CLL. Thus, LEF1 might be involved in the process of disease progression and possibly can serve as a molecular parameter for risk assessment and/or monitoring of CLL.