Aberrant Expression of Trail in B Chronic Lymphocytic Leukemia (B-CLL) Cells.

Chronic lymphocytic leukemia (CLL) is a quintessential example of human malignancies that are caused primarily by defect in apoptosis. Defects in apoptotic pathways contribute also to chemoresistance and can promote resistance to cellular immune responses. TNF-related apoptosis inducing ligand (TRAIL), interacts with four high affinity membrane receptors (TRAIL-R1 – R4): R1 and R2 are thought to transducer apoptotic signals, while R3 and R4 may act as “decoy” receptors, protecting cells from apoptosis. Despite its potential anti–cancer activity, TRAIL alone has a low cytotoxic activity on B-CLL, and no data are available on the expression of TRAIL and its biological potential function in B-CLL.

We examined the expression of TRAIL in peripheral blood CD19+/CD5+ B lymphocytes from 44 patients affected by CLL at diagnosis, the susceptibility of B-CLL cells to recombinant TRAIL and the role of endogenous membrane-bound TRAIL on autologous cell survival.

Each B-CLL patient had a follow up time of 12 to 24 months from diagnosis and was clinically stable. None of the patients received chemotherapy.

Lymphocytes from normal PBMC revealed an absent or dim expression of TRAIL. Surface TRAIL was detected in all 44 B-CLL examined, at variable intensity from patient to patient, but with an overall significantly greater MFI with respect to normal lymphocytes. Higher levels of TRAIL mRNA and protein were documented in purified CLL CD19+ B lymphocytes, confirming that TRAIL was overexpressed at the transcriptional level.

The addition in culture of a TRAIL-R1-Fc chimera, which binds at high affinity to surface TRAIL, significantly decreased the percentage of viable cells with respect to untreated cultures. No effects were noted when TRAIL-R1-Fc chimera was added to normal CD19+ B cells. Preincubation of TRAIL-R1-Fc chimera with recombinant TRAIL significantly counteracted the decrease of viability induced by TRAIL-R1-Fc chimera. These data indicate that surface TRAIL might be involved in a promoting a prosurvival activity, at least in some B-CLL samples.

We then investigated the response in terms of cell viability to exogenously added recombinant TRAIL (1 μg/ml) of all 44 B-CLL samples examined in this study. On the basis of their response to recombinant TRAIL, B-CLL samples could be subdivided in 3 groups. In 11 patients (group 1), B-CLL cells showed a faster decline in the number of viable cells upon treatment with recombinant TRAIL. In 19 samples (group 2),no significant variation in terms of viable cell number in TRAIL-treated cultures was observed. In 14 patients (group 3), TRAIL-treatment results in a significantly higher numbers of viable cells compared to untreated cultures.

At flow cytometry, B-CLL lymphocytes expressed at variable levels TRAIL-R1, TRAIL-R2 and TRAIL-R4, while TRAIL-R3 was never detected. TRAIL-R1 was expressed at low levels in a subset of B-CLL samples, while TRAIL-R2 was expressed in almost the totality of the B-CLL samples. TRAIL-R4 was detected in the majority of B-CLL lymphocytes, but at lower levels than in normal lymphocytes. Of note, the different apoptotic/survival response to recombinant TRAIL among the three groups of B-CLL samples described above, apparently did not depend on differences in death and/or decoy receptor patterns of expression. A progressive increase of TRAIL MFI was noticed from group 1 to group 3.