Triggering of CD40 antigen inhibits fludarabine-induced apoptosis in B chronic lymphocytic leukemia cells.
We analyzed the effect of CD40 triggering on the fludarabine-induced apoptosis of B chronic lymphocytic leukemia (B-CLL) cells. Peripheral blood samples obtained from 15 patients were incubated with fludarabine in the absence or the presence of the anti-CD40 monoclonal antibody (MoAb) G28-5. In 12 patients a significant proportion of apoptotic cells, ranging from 22% to 38% (mean +/- SE: 28.5 +/- 1.6), were detected after 3 days of culture. In 9 of these samples, the addition of G28-5 reduced apoptosis by at least 30.1% and by 57.1% +/- 7.8% on average (P = .0077). Because the CD40 antigen activates NF-kappaB/Rel transcription factors in B cells, and NF-kappaB/Rel complexes can inhibit cell apoptosis, we investigated whether the antiapoptotic effect of G28-5, in our system, could be related to modulation of NF-kappaB/Rel activity. As expected, B-CLL cells displayed significant levels of nuclear NF-kappaB/Rel activity; p50, RelA, and c-Rel components of the NF-kappaB/Rel protein family were identified in these complexes. After exposure to fludarabine, NF-kappaB/Rel complexes were decreased in the nuclei. The addition of G28-5 upregulated the NF-kappaB/Rel levels. To determine the involvement of NF-kappaB/Rel activity in the G28-5-mediated inhibition of apoptosis, we blocked the transcription factor with a decoy oligonucleotide, corresponding to the NF-kappaB/Rel consensus sequence. Cells incubated with the anti-CD40 MoAb in the presence of the decoy oligonucleotide but not a control oligonucleotide displayed a complete impairment of the G28-5 antiapoptotic effect, indicating that NF-kappaB/Rel activity was required for the inhibition of apoptosis. These results suggest that CD40 triggering in vivo could counteract the apoptotic effect of fludarabine on B-CLL cells and that its neutralization, or the use of NF-kappaB/Rel inhibitors, could improve the therapeutic effect of fludarabine.
Dipartimento di Biochimica e Biotecnologie Mediche, Università Federico II, Napoli, Italia.
M F
MF
Romano
A
A
Lamberti
P
P
Tassone
F
F
Alfinito
S
S
Costantini
F
F
Chiurazzi
T
T
Defrance
P
P
Bonelli
F
F
Tuccillo
M C
MC
Turco
S
S
Venuta
D000911:Antibodies, Monoclonal / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D019013:CD40 Antigens / Q000276:immunology / Q000502:physiology*; D015973:Gene Expression Regulation, Leukemic / Q000187:drug effects*; D006801:Humans; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000473:pathology*; D016328:NF-kappa B / Q000378:metabolism; D009363:Neoplasm Proteins / Q000378:metabolism; D014411:Neoplastic Stem Cells / Q000187:drug effects* / Q000473:pathology; D009841:Oligonucleotides / Q000494:pharmacology; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D020569:Proto-Oncogene Proteins c-rel; D014158:Transcription, Genetic / Q000187:drug effects; D014407:Tumor Cells, Cultured; D014740:Vidarabine / Q000031:analogs & derivatives* / Q000037:antagonists & inhibitors / Q000494:pharmacology
D000911:Antibodies, Monoclonal; D019013:CD40 Antigens; D016328:NF-kappa B; D009363:Neoplasm Proteins; D009841:Oligonucleotides; D011518:Proto-Oncogene Proteins; D020569:Proto-Oncogene Proteins c-rel; D014740:Vidarabine; C024352:fludarabine