Clonal variability in CD95 expression is the major determinant in Fas-medicated, but not chemotherapy-medicated apoptosis in the RPMI 8226 multiple myeloma cell line.
CD95 (Fas/APO-1) is a member of the TNFR superfamily that induces apoptosis following cross-linking with its cognate ligand, CD95L (FasL/APO-1L) or agonist antibody. The human myeloma cell line, RPMI 8226, has limited sensitivity to CD95-mediated apoptosis, with a maximum of 65% of the population responding. To determine the source of the limited sensitivity to CD95-mediated apoptosis, we isolated multiple clones from the RPMI-8226 cell line by limiting dilution. Analysis of these clones demonstrated that sensitivity to CD95-mediated cell death directly correlated with CD95 expression. Clones with high levels of CD95 expression had greater than 90% cell death, whereas cells with low levels of expression had less than 10% cell death. In contrast, no correlative differences were identified for other members of the DISC complex, or for members of the anti-apoptotic Bcl-2 family. We further examined the sensitivity of the 8226 clones to various cytotoxic agents. Although modest clonal variability was demonstrated in response to the chemotherapeutic drugs, doxorubicin, etoposide (VP-16), and vincristine, there was no correlation between CD95 function and sensitivity to chemotherapeutic drugs. These results indicate that in this cell line, receptor expression is rate limiting in CD95-mediated apoptosis, whereas CD95 expression was not a determinant in drug-induced programmed cell death.
Shain
K H
KH
Clinical Investigations Program, H Lee Moffitt Cancer Center and Research Institute, University of South Florida, Tampa 33612, USA.
Landowski
T H
TH
Buyuksal
I
I
Cantor
A B
AB
Dalton
W S
WS
D000970:Antineoplastic Agents / Q000633:toxicity*; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000235:genetics / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002999:Clone Cells; D004317:Doxorubicin / Q000633:toxicity; D005047:Etoposide / Q000633:toxicity; D005434:Flow Cytometry; D006801:Humans; D009101:Multiple Myeloma / Q000473:pathology*; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D014750:Vincristine / Q000633:toxicity; D019014:fas Receptor / Q000235:genetics* / Q000502:physiology
D000970:Antineoplastic Agents; D019014:fas Receptor; D014750:Vincristine; D005047:Etoposide; D004317:Doxorubicin; C505407:CASP3 protein, human; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases