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Rituximab Kills Freshly Isolated B-NHL and B-CLL More Efficiently by Complement Than by ADCC In Vitro: Role of CD20 Expression Levels and of the CD55 and CD59 Complement Inhibitors

Rituximab Kills Freshly Isolated B-NHL and B-CLL More Efficiently by Complement Than by ADCC In Vitro: Role of CD20 Expression Levels and of the CD55 and CD59 Complement Inhibitors

Current evidence suggests that rituximab (Rituxan) works in vivo mainly through complement-dependent cytotoxicity (CDC) and/or antibody-dependent cellular cytotoxicity (ADCC). Here we have investigated the sensitivity of freshly isolated cells obtained from 33 B-cell chronic lymphocytic leukemia (B-CLL), 5 prolymphocytic leukemia (PLL), and 6 mantle cell leukemia (MCL) patients to be lysed by rituximab and complement in vitro.

The results showed that in B-CLL and PLL, the levels of CD20, measured by standard immunofluorescence or using calibrated beads, correlated linearly with the lytic response (coefficient > 0.9, P < .0001). Furthermore, the correlation remained highly significant when the six MCL patients were included in the analysis (coefficient 0.91, P < .0001), suggesting that CD20 levels primarily determine lysis regardless of diagnostic group.

The role of the complement inhibitors CD55 and CD59 was also investigated. All B-CLL and PLL cells expressed these molecules, but at variable levels (mean fluorescence intensity [MFI] = 20-1,200 vs 20-250, respectively). Although CD55 and CD59 levels did not permit us to predict complement susceptibility, the functional block of these inhibitors demonstrated that they play an important role in regulating CDC. Thus, lysis of poorly responding B-CLL samples was increased five- to sixfold after blocking both CD55 and CD59, whereas that of high responders was essentially complete in the presence of a single blocking antibody.

We have also investigated the lysis through ADCC of five fresh cases of CLL and two of MCL. Of these samples, four expressed relatively high levels of CD20 (MFI > 200) and were lysed significantly with rituximab and complement (40% to 65%). On the contrary, we have found that none of the samples examined was lysed significantly by the natural killer (NK) cell line NKL in the presence of rituximab, whereas the same cells were killed efficiently (by 30% to 70%) in the presence of the anti-CD52 antibody alemtuzumab (Campath). Furthermore, three leukemic B-cell lines could be lysed to a similar extent (48% to 62%) by either rituximab or alemtuzumab and NKL cells.

CONCLUSION: Altogether, these data suggest that, although rituximab is able to activate both CDC and ADCC on B-cell lymphoma lines, CDC is a more efficient mechanism of action of this antibody than ADCC on freshly isolated B-CLL and B-cell non-Hodgkin’s lymphoma cases. Furthermore, they show that CD20, CD55, and CD59 are important factors in determining the response to rituximab and complement in vitro, and indicate strategies to improve the clinical response to this biologic therapy.

Click here to read Dr. Bruce Cheson's commentary on this abstract.

 
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