cDNA microarray study to identify expression changes relevant for apoptosis in K562 cells co-treated with amifostine and imatinib.
OBJECTIVE
Chronic myeloid leukemia is a clonal myeloproliferative disorder characterized by the presence of the fusion gene BCR/ABL. We had previously demonstrated an increased proapoptotic effect of imatinib (STI571) in combination with amifostine (AMI) in K562 cell line. In this study, we used genomic scale gene expression profiling to monitor changes at transcriptional level in K562 cells during the treatment with AMI + STI571.


METHODS
cRNA from Control and treated K562 cells were mixed in equal amounts and incubated with a microarray slide for hybridization. RNA from six independent paired experiments was subjected to transcriptional profiling. With the aim to automate the process of biological theme determination, selected genes were further analyzed by EASE. Validation of the expression was carried out by quantitative real-time PCR and western blotting.


RESULTS
As expected, a small percentage of genes accounts for the effects of the combined drug treatment. We identified 61 sequences corresponding to known genes; 17 of the 61 genes were up regulated, such as RHO6, PPP2R5E, PPM1E and BTF that appear to reflect favorable events for apoptosis induction. Between down regulated genes, API5, TUBB2 and TLK1 are also of considerable interest.


CONCLUSIONS
We identified a transcriptional repressor of survival genes, known as BTF, which triggers a proapoptotic signal, potentially helpful to overcome the resistance to STI571. This finding could be particularly useful to design novel therapeutic strategies for leukemia patients. This study demonstrates the importance of in vitro testing of a novel drug combination most likely to predict its potential usefulness for in vivo application.
Bianchini
Michele
M
Departamento de Genética, Instituto de Investigaciones Hematológicas Mariano R. Castex, Academia Nacional de Medicina, Pacheco de Melo 3081, 1425, Capital Federal Buenos Aires, Argentina. mbianchini@hematologia.anm.edu.ar
Martinelli
Giovanni
G
Renzulli
Matteo
M
Gonzalez Cid
Marcela
M
Larripa
Irene
I
D004999:Amifostine / Q000008:administration & dosage; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D001549:Benzamides; D002470:Cell Survival / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000235:genetics; D004338:Drug Combinations; D004354:Drug Screening Assays, Antitumor; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology*; D006801:Humans; D000068877:Imatinib Mesylate; D020014:K562 Cells / Q000187:drug effects / Q000473:pathology; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D010879:Piperazines / Q000008:administration & dosage; D011743:Pyrimidines / Q000008:administration & dosage; D012097:Repressor Proteins / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics; D014158:Transcription, Genetic / Q000187:drug effects*; D025521:Tumor Suppressor Proteins / Q000235:genetics
C119386:BCLAF1 protein, human; D001549:Benzamides; D004268:DNA-Binding Proteins; D004338:Drug Combinations; D010879:Piperazines; D011743:Pyrimidines; D012097:Repressor Proteins; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D000068877:Imatinib Mesylate; D004999:Amifostine