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@proceedings{752116,
author = {Trbušek, Martin and Čejková, Soňa and Ročňová, Ludmila and Potěšil, David and Chumchalová, Jitka and Šmardová, Jana and Malčíková, Jitka and Kuglík, Petr and Doubek, Michael and Brychtová, Yvona and Pospíšilová, Šárka and Mayer, Jiří},
booktitle = {Haematologica, the hematology journal},
keywords = {B-CLL fludarabine rituximab p53 ATM},
language = {eng},
title = {Rituximab senzitizes some B-CLL samples to fludarabine and chlorambucil in vitro, regardless of p53/ATM status},
year = {2007}
}
TY - CONF
ID - 752116
AU - Trbušek, Martin - Čejková, Soňa - Ročňová, Ludmila - Potěšil, David - Chumchalová, Jitka - Šmardová, Jana - Malčíková, Jitka - Kuglík, Petr - Doubek, Michael - Brychtová, Yvona - Pospíšilová, Šárka - Mayer, Jiří
PY - 2007
TI - Rituximab senzitizes some B-CLL samples to fludarabine and chlorambucil in vitro, regardless of p53/ATM status
KW - B-CLL fludarabine rituximab p53 ATM
N2 - Background: Aberrations of two co-operating genes, the p53 and ATM, significantly deteriorate prognosis and treatment options for B-CLL patients. Monoclonal antibody rituximab (anti-CD20) is preferably used in combination regimens in B-CLL, often in those containing fludarabine. Very few in vitro data exist, however, showing an effect of such common treatment on B-CLL cells with aberrant p53 and/or ATM. In this respect, the data are also missing for a potential application of rituximab with chlorambucil. Aims: The aims were to assess the in vitro effect of the above mentioned combinations of drugs on B-CLL cells with various p53/ATM status. Methods: An interphase FISH was used for a determination of p53 and ATM deletions. Functional FASAY analysis coupled to sequencing was employed to supplement the screening also for p53 mutations. A metabolic WST-1 assay monitored the drugs effect on cell viability. An in vitro system lacking active human plasma was used, thus omitting the CDC pathway. After rituximab pre-treatment (10ug/ml, 72h) the chemotherapeutics were applied in four concentrations for additional 48h (F: 25 - 0.4ug/ml; CLB: 50 - 6.25 uM). Two-way analysis of variance (ANOVA) was used for determination of rituximab pre-treatment significance. Results: For the rituximab/fludarabine combination we tested forty samples having a median 85% of B-CLL lymphocytes, with the following characteristics: 13 were wild-type, 10 harbored ATM deletion (median 83% of deleted cells) and 17 exhibited p53 defects of various complexity - both alleles inactivation (del/mut and mut/mut) as well as the separate (one allele) aberrations (del or mut). The sensitivity to fludarabine was determined for the concentration 1.6ug/ml, which provided significant differences among the samples. The sensitivity was assessed as follows: resistant - viability over 60%; medium - viability between 60% and 40%; sensitive - viability under 40%. The p53-affected samples were mostly resistant (71%) and none were sensitive. Among ATM deleted samples, on the contrary, 40% were sensitive, what was more than in wild-type subgroup (23%). Rituximab alone slightly increased a metabolic activity in most of samples, usually to 110-130% compared to untreated controls, while rarely a decrease was also noted (up to 80%). When the viability of fludarabine-tretead and rituximab/fludarabine-treated samples was assessed in relation to fully untreated control, the positive sensitization effect of rituximab pre-treatment (P = 0,05) was noted as follows: within the p53-affected as well as ATM-deleted subgroups in 30% of samples and within the wild-type subgroup in 62% of samples. For the rituximab/chlorambucil testing, which was performed as a pilot study in the same manner in eight samples, the positive effect of antibody pre-treatment was also noted in some samples of all the three subgroups. Summary/Conclusions: Our results indicate that the p53/ATM status is critical for the sensitivity of B-CLL cells to fludarabine. Regardless of the p53 and ATM aberrations, some samples are available for the rituximab-mediated sensitization to this agent. Our pilot data also support a warranty of testing a combined regimen containing rituximab and chlorambucil. Supported by grant IGA MH CR No. 8445-3/2005.
ER -
TRBUŠEK, Martin, Soňa ČEJKOVÁ, Ludmila ROČŇOVÁ, David POTĚŠIL, Jitka CHUMCHALOVÁ, Jana ŠMARDOVÁ, Jitka MALČÍKOVÁ, Petr KUGLÍK, Michael DOUBEK, Yvona BRYCHTOVÁ, Šárka POSPÍŠILOVÁ and Jiří MAYER. Rituximab senzitizes some B-CLL samples to fludarabine and chlorambucil in vitro, regardless of p53/ATM status. In \textit{Haematologica, the hematology journal}. 2007. ISSN~0390-6078.
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