Components of the cell death machine and drug sensitivity of the National Cancer Institute Cell Line Panel.
OBJECTIVE
According to some studies, susceptibility of cells to anticancer drug-induced apoptosis is markedly inhibited by targeted deletion of genes encoding apoptotic protease activating factor 1 (Apaf-1) or certain caspases. Information about levels of these polypeptides in common cancer cell types and any possible correlation with drug sensitivity in the absence of gene deletion is currently fragmentary.


METHODS
Immunoblotting was used to estimate levels of Apaf-1 as well as procaspase-2, -3, -6, -7, -8, and -9 in the 60-cell-line panel used for drug screening by the National Cancer Institute. Sensitivity of the same lines to >80,000 compounds was determined with 48-hour sulforhodamine B binding assays. Additional 6-day assays were performed for selected agents.


RESULTS
Levels of Apaf-1 and procaspases varied widely. Apaf-1 and procaspase-9, which are implicated in caspase activation after treatment of cells with various anticancer drugs, were detectable in all of the cell lines, with levels of Apaf-1 ranging from approximately 1 x 10(5) to 2 x 10(6) molecules per cell and procaspase-9 from approximately 5 x 10(3) to approximately 1.6 x 10(5) molecules per cell. Procaspase-8 levels ranged from 1.7 x 10(5) to 8 x 10(6) molecules per cell. Procaspase-3, a major effector caspase, varied from undetectable to approximately 1.6 x 10(6) molecules per cell. Correlations between levels of these polypeptides and sensitivity to any of a variety of experimental or conventional antineoplastic agents in either 2-day or 6-day cytotoxicity assays were weak at best.


CONCLUSIONS
With the exception of caspase-3, all of the components of the core cell-death machinery are expressed in all of the cell lines examined. Despite variations in expression, levels of any one component are not a major determinant of drug sensitivity in these cells in vitro.
Svingen
Phyllis A
PA
Division of Oncology Research, Mayo Clinic, and Department of Molecular Pharmacology, Mayo Graduate School, Rochester, Minnesota 55901, USA.
Loegering
David
D
Rodriquez
Joe
J
Meng
Xue Wei
XW
Mesner
Peter W
PW
Holbeck
Susan
S
Monks
Anne
A
Krajewski
Stan
S
Scudiero
Dominic A
DA
Sausville
Edward A
EA
Reed
John C
JC
Lazebnik
Yuri A
YA
Kaufmann
Scott H
SH
D017209:Apoptosis / Q000187:drug effects*; D053477:Apoptotic Protease-Activating Factor 1; D020169:Caspases / Q000096:biosynthesis* / Q000235:genetics*; D004354:Drug Screening Assays, Antitumor; D006801:Humans; D015151:Immunoblotting; D009369:Neoplasms / Q000235:genetics* / Q000473:pathology*; D011506:Proteins / Q000032:analysis / Q000235:genetics*; D014407:Tumor Cells, Cultured
C506592:APAF1 protein, human; D053477:Apoptotic Protease-Activating Factor 1; D011506:Proteins; D020169:Caspases