Little information is available on long-term immune reconstitution after therapy with alemtuzumab in B-CLL patients. We present long-term follow-up data for blood lymphocyte subsets analysed by flow cytometry in previously untreated B-CLL patients who received alemtuzumab subcutaneously as firstline therapy. All lymphoid subsets were significantly (Po0.001) and profoundly reduced; the median end-of-treatment counts þ and CD8 þ levels in blood had reached 4100 cells/ll in 450% of the patients at 4 months after the end of treatment. One patient had a cytomegalovirus reactivation and one patient developed Pneumocystis carinii pneumonia during therapy. No opportunistic or other major infections were recorded during unmaintained, long-term follow-up. There was no correlation between the cumulative dose of alemtuzumab and the severity or length of immunosuppression. CD52À T-cell subsets occurred during the treatment and comprised 480% of all CD4 þ and CD8 þ cells in the blood at the end of therapy. These subpopulations declined gradually during unmaintained followup. The relationship between these observations and the safety/antitumour effects of alemtuzumab is discussed.

Fibromodulin is an extracellular matrix protein normally produced by collagen-rich tissues; the fibromodulin gene has been found to be the most overexpressed gene in B-cell chronic lymphocytic leukemia. In this study, fibromodulin was expressed at the gene level (

Dendritic cells (DC) are professional (specialised) antigenpresenting cells that can capture antigen from apoptotic tumour cells and induce MHC class I-and II-restricted responses. Also, DC fused with tumour cells may be effective for immune response induction. Both cell preparations may be considered as vaccine candidates in a therapeutic approach. We examined autologous T-cell activation by DC that had endocytosed leukaemic B-cell apoptotic bodies (Apo-DC) and compared it to the T-cell stimulatory capacity of DC that were fused with tumour cells. Following incubation, 22.676.2 (mean7s.e.m.) of DC had endocytosed leukaemic cells, while the frequency of DC-leukaemic cell hybrids was 10.572.6%. Apo-DC and hybrid cells both demonstrated the ability to stimulate a tumour-specific T-cell immune response in vitro. A T-cell proliferation response was also observed in four out of five CLL patients when using Apo-DC. However, fusion hybrids lacked the ability to elicit a proliferative response. Apo-DC also induced an IFN-c response, as did hybrid cells. The cytokine response induced by Apo-DC was significantly higher than that induced by fusion (Po0.05). This study shows that endocytosed apoptotic tumour cells induced a significantly stronger T-cell response than DC hybrids; and as such should be a better candidate for vaccine production.

TCRBV (T-cell receptor B variable) gene usage and CDR3 size distribution were analyzed using reverse transcription polymerase chain reaction (RT-PCR) to assess the T-cell repertoire of 10 patients with B-cell chronic lymphocytic leukemia (B-CLL) and in nine age-matched healthy control donors. When the usage of each TCRBV gene within the CD8+ T cells of the patients was compared with that of the controls, no statistically significant difference was noted except for BV 6S1-3. In contrast, within the CD4+ T cells of the CLL patients, a statistically significant overexpression for four BV families (2, 3, 5S1, 6S1-3) was seen while an underrepresentation was noted for five BV families (10, 11, 15, 16, 19). Based on the criterion that a value of any BV higher than the mean + 3 standard deviation (SD) of healthy controls indicated an overexpression, individual patients were shown to overexpress several TCRBV genes compared with the controls. Analyses of the CDR3 length polymorphism showed a significantly higher degree of restriction within CD4+ and CD8+ T cells of the patients, as compared with the corresponding control T-cell population. There was a significant difference in the CDR3 size distribution pattern with a more polymorphic CDR3 length pattern in the age-matched controls as compared with CLL patients, suggesting different mechanisms driving the T cells towards a clonal/oligoclonal TCRBV usage in patients and controls, respectively. The results show major perturbations of T cells in CLL patients, more frequently seen in the CD4+ T-cell subset, indicating that nonmalignant CD4+ T cells may be involved in the pathogenesis of CLL, but also CD8+ T cells.