Structure-apoptotic potency evaluations of novel sterols using human leukemic cells.
Three oxidized analogs of cholesterol have been characterized for their ability to cause apoptotic cell death in CEM-C7-14 human leukemic cells. In addition to testing 15-ketocholestenol (K15), 15-ketocholestenol hydroxyethyl ether (CK15), and 7-ketocholesterol hydroxyethyl ether (CK7), an oxysterol of known apoptotic response, 25-hydroxycholesterol (25OHC), served as a standard for comparison. Growth studies based on dye exclusion by viable cells while using a sublethal concentration of oxysterols ranked their potency for cell kill as 25OHC > K15 > CK15 > CK7. Both the TUNEL assay (terminal deoxynucleotidyl transferase-mediated dUTP-X nick end labeling), which quantifies the amount of DNA nicks caused by a toxic agent, and the MTT assay, which measures cell metabolism and thus reflects cell viability, substantiated the same rank order. An ELISA assay for evaluating release of DNA fragments into the cytosol after treatment gave a similar potency order. The oncogene c-myc mRNA was suppressed by all three oxysterols, with 25OHC and K15 being the most potent suppressors. Hoechst and Annexin V staining documented that these oxysterols kill cells by an apoptotic pathway as evidenced by condensation of nuclear chromatin and plasma membrane inversion, respectively. From these in vitro studies, we believe that 25OHC, K15, and possibly CK15 have the potential to be chemotherapeutic agents.
Johnson
B H
BH
Department of Human Biological Chemistry and Genetics, The University of Texas Medical Branch, Galveston 77555-0645, USA.
Russell
M J
MJ
Krylov
A S
AS
Medh
R D
RD
Ayala-Torres
S
S
Regner
J L
JL
Thompson
E B
EB
D017209:Apoptosis / Q000187:drug effects*; D002455:Cell Division / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002784:Cholesterol / Q000031:analogs & derivatives* / Q000494:pharmacology*; D053938:DNA Fragmentation; D004797:Enzyme-Linked Immunosorbent Assay; D016259:Genes, myc / Q000187:drug effects; D006801:Humans; D020287:In Situ Nick-End Labeling; D007700:Kinetics; D007938:Leukemia; D014407:Tumor Cells, Cultured
D002784:Cholesterol