In Vitro Cytotoxicity of Alemtuzumab on B-CLL Cells: Differential Effect on B and T Lymphocytes.

Alemtuzumab, a monoclonal anti-CD52 antibody, has shown high efficacy against lymphoproliferative disorders such as B-cell chronic lymphocytic leukemia (B-CLL). However, its use results in a profound immunosuppression with decrease of T lymphocytes leading to increased susceptibility to infections. The aim of our study was to assess the in vitro cytotoxic effect of alemtuzumab on normal T and neoplastic B lymphocytes obtained from B-CLL patients.

Peripheral blood mononuclear cells (PBMC) from 12 B-CLL patients (5 at diagnosis and 7 previously treated) were collected and treated in vitro with alemtuzumab (10 μg/ml) and autologous serum in the culture as complement source. Spontaneous and alemtuzumab-induced apoptosis were quantified in T (CD3+) and B (CD20+) lymphocytes after 24 hours using an annexin-V flow cytometry based asssay.

Alemtuzumab was able to induce apoptosis on both B and T lymphocytes of CLL patients. However, the cytotoxic activity on neoplastic B cells was comparable in all cases analyzed, irrespective of stage or previous treatments. Spontaneous apoptosis of B CLL cells was also comparable in all studied samples.

On the contrary, the activity of alemtuzumab on normal T lymphocytes varied according to stage of disease and previous treatment. In early stages (0–I) alemtuzumab induced apoptosis in 19 % of T cells vs 60% of advanced stages (II–IV) (p=0,009). Differences were also relevant when patients were split by treatment: in fact, alemtuzumab induced apoptosis in 15% of T cells of untreated patients vs 52% of T cells of previously treated patients (p=0,005). Moreover, spontaneous apoptosis of T cells was more pronounced in treated vs. untreated patients (15% vs 2%, p=0.03) while the difference was not statistically significant in early vs advanced stage (p=0.06).Our in vitro data confirm that alemtuzumab is active against B-CLL cells in all stage of disease. However, in advanced stages and in previously treated patients, T cell compartment seems to be more fragile and susceptible to both spontaneous and alemtuzumab-induced apoptosis. This observation supports the hypothesis that using Alemtuzumab earlier in the treatment of B-CLL might result in less immunosuppression. The study is still ongoing with accrual of more patients samples.