Expression and function of S100A4 in B-cell lymphomas

Abstract

B-cell lymphomas are clinically, pathologically and molecularly heterogeneous diseases. Multiple studies have been focused on the analysis of the mechanism of B-cell lymphoma progression, search for novel prognostic markers and development of new treatment approaches. In the current project, the research focus was on the analysis of the expression of S100 proteins in B-cell malignancies. S100 proteins are small, calcium-binding proteins, which are expressed in normal and pathological conditions in cell-specific fashion. They comprise a protein family with more than 20 members. S100 proteins are implicated in various biological processes including cell proliferation and migration, cell signalling, and intercellular communications. S100 proteins don’t possess any enzymatic activities, but they participate in transferring calcium signals via interactions with their protein targets. An association between S100 protein expression and cancer was studied in different solid malignancies. Expression of several members of the S100 protein family correlates with the outcome of the disease in different types of epithelial tumours. Therapeutic value of pharmacological targeting of S100 proteins in solid tumours has been discussed. In this study, expression and function of S100 family members was investigated in the peripheral blood and solid tumours of B-cell lymphoma patients. B-cells were purified from the blood samples of healthy volunteers (combined fractions from 5 individuals) and CLL patients. Following RNA isolation, expression of 23 S100 genes was analysed by RT-PCR. Strong activation of S100A4 and S100A6 transcription was detected in malignant B-cells. Western blot analysis of 20 samples prepared from the peripheral blood of the CLL patients confirmed high expression level of S100A4, but not S100A6 in most samples. Therefore, further research was focused on the study of the expression and function of S100A4 protein. Immunohistochemical analysis of 60 blocks of different B-cell lymphomas has demonstrated enhanced expression of S100A4 in the majority of CLL, MCL, DLBCL cases, but not FL. Low or no expression of S100A4 correlated with a longer survival among CLL patients. Although the results were obtained in a relatively small group of patients, data indicate that S100A4 protein may serve as a prognostic marker in CLL. The prognostic significance of S100A4 expression in CLL warrants further investigations. In normal blood and tissue samples, expression of S100A4 was detected exclusively in T-cells and not in B-cells. Immunofluorescent analysis has shown that S100A4 is co-expressed with stem cell markers, CD34, CD133 indicating its potential role in cancer cell stemness. Modulation of S100A4 expression by RNA-interference demonstrated that S100A4 promotes in vitro chemotaxis of B-cells isolated from the peripheral blood of CLL patients. Migratory potential of CLL cells was significantly reduced by two small molecule compounds inhibiting S100A4 function. In conclusion, the data obtained in this study demonstrate for the first time that S100A4 is activated in malignant B-cells, and promotes their migration and invasion. Expression of S100A4 can be considered for the prognostication of CLL patients. Further preclinical studies involving small molecule inhibitors of S100A4 are needed to evaluate their therapeutic potential