ZAP-70 Expression in T Cells of B-Cell Chronic Lymphocytic Leukaemia: Correlation with Negative Prognostic Factors of the Disease.

Several studies demonstrated that ZAP-70, a tyrosine kinase which plays a critical role in T cell antigen receptor signaling, is expressed in cells of B cell chronic lymphocytic leukemia (B-CLL) in correlation with unmutated status of immunoglobulin genes, progression of the disease and overall survival. Recently, it has been reported a correlation among ZAP-70 expression in T cells of CLL and CLL Zap-70 levels, clinical stage and disease progression. To analyse the functional capacity of the T residual component in the B-CLL in this study we evaluated ZAP-70 expression in peripheral T cells from patients with B-CLL and from healthy volunteers using a direct immunofluorescence technique by flow cytometry (FACSCAN, Becton Dickinson). Analysis was performed using CellQuest software and results were expressed using mean fluorescence index (MFI) calculated as ratio between mean channel of sample and mean channel of negative control.

Sixty two B-CLL patients and ten normal subjects entered the study, 35/62 patients were males and 27 females. Our results showed a higher ZAP-70 expression in T cells from B-CLL compared to normal T cells, with a median MFI value of 14 (range 4,2–65) and 11 (range 9,2–14,6) respectively (p=0,045). Moreover, using a cut-off of 11 (that represents the median MFI value in normal T cells) we defined two groups of B-CLL cases: patients with high expression (MFI > 11) and patients with low expression (MFI ≤ 11). No differences among the various stages were recorded in the two groups, in fact 25/50 (50%) ZAP-70^high T-cell case were stage A, 8/50 (16%) were stage B and 17/50 (34%) were stage C, and 9/12 (75%) ZAP-70^low T-cell cases were stage A, 1/12 (8%) were stage B and 2/12 (16%) were stage C. High expression of ZAP-70 in T cells seems to correlate with negative prognostic factors, in fact our results showed a higher incidence of CD38 and of unmutated immunoglobulin genes in ZAP-70^high T-cell group with respect to ZAP-70^low T-cell cases: 24/45 (53%) vs 3/11 (27%) and 22/29 (76%) vs 3/7 (42%) however these associations were not of statistical significance, probably due to the small numbers of patients analysed. On the contrary a significant correlation was recorded with level of expression in T cells and presence of ZAP-70 in B cells, indeed ZAP-70 was expressed in B lymphocytes of 37/50 (79%) patients from ZAP-70^high T-cell group, while only in 5/12 (41%) cases among ZAP-70^low T-cell group (p=0,04).

The meaning of ZAP-70 over expression in T cells of B-CLL is unknown. Some studies suggest that it may be indicative of a state of cell activation, however very preliminary data on 4 patients show a notably lower constitutive phosphorilation state compared to the one observed in T cells of healthy volunteers. Besides, our data don’t show a significant increase of the global phosphorilation pattern after T cell stimulation with PHA, even if the CD3 e resulted strongly phosphorilated, suggesting a reduced responsiveness of T cells to mitogenic stimuli.

In conclusion, ZAP-70 over-expression in T cells from B-CLL seems to identify a subgroup of patients with a more aggressive disease, and moreover we hypothesize that it may be the expression of alteration of T cells functionality associated to the disease.