Docetaxel induced gene expression patterns in head and neck squamous cell carcinoma using cDNA microarray and PowerBlot.
OBJECTIVE
The purpose is to identify gene expression patterns induced by docetaxelin head and neck squamous carcinoma (HNSCC) cells using high throughput techniques.


METHODS
HNSCC cells were treated with docetaxel or solvent. After mRNA extraction, cDNA fluorescent (Cy3 or Cy5)-labeled probes were synthesized. Then, Cy3 and Cy5-labeled samples were hybridized onto a microarray slide. The fluorescent images were scanned and analyzed for quantification. PowerBlot immunoblotting technique was used to measure protein expression level. Using this dual approach, we focused on genes in established pathways (cell cycle, apoptosis, angiogenesis, and signal transduction) of tumorigenesis and confirmed these results with conventional techniques.


RESULTS
Using cDNA microarray, we found that docetaxel altered the expression of >100 genes in HNSCC cells. A total of 153 of 1191 genes was found to have altered expression in either HN12 (n = 102), HN30 (n = 72), or both (n = 21) by docetaxel. For the PowerBlot analysis, a subset of genes (n = 46) in the cDNA microarray analysis and an additional 98 genes in the cell cycle, apoptosis, angiogenesis, and signal transduction pathways were chosen. We found that PowerBlot data agreed with cDNA microarray in 65% of genes examined. The expression of a cell cycle inhibitor (p19) and promoters (cyclin A, cyclin B1, and cyclin E2F) were increased and decreased, respectively. Apoptosis induced by docetaxel was independent of p53 and, in part, related to increased Fas expression. Both vascular endothelial growth factor secretion and basic fibroblast growth factor expression were inhibited by docetaxel, whereas thrombospondin-1 expression was increased by docetaxel. Epidermal growth factor receptor, activated epidermal growth factor receptor, and activated c-Jun NH(2)-terminal kinase expression was lowered by docetaxel. Activated extracellular signal-regulated kinase was elevated by docetaxel, but not total extracellular signal-regulated kinase levels.


CONCLUSIONS
The identification of altered gene expression induced by docetaxel demonstrates additional biological activity in HNSCC cells, and the altered expression of these genes may serve as potential biomarkers to both predict clinical activity and provide information regarding potential efficacy of adding novel agents.
Yoo
George H
GH
Department of Otolaryngology-Head and Neck Surgery, Wayne State University and Karmanos Cancer Institute, Detroit, Michigan 48201, USA.
Piechocki
Marie P
MP
Ensley
John F
JF
Nguyen
Tam
T
Oliver
Jeffery
J
Meng
Hong
H
Kewson
Danny
D
Shibuya
Terry Y
TY
Lonardo
Fulvio
F
Tainsky
Michael A
MA
D017304:Annexin A5 / Q000378:metabolism; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D002294:Carcinoma, Squamous Cell / Q000235:genetics* / Q000378:metabolism; D017931:DNA Primers / Q000737:chemistry; D000077143:Docetaxel; D016228:Endothelial Growth Factors / Q000378:metabolism; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D006258:Head and Neck Neoplasms / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism; D008222:Lymphokines / Q000378:metabolism; D009363:Neoplasm Proteins / Q000235:genetics* / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D017239:Paclitaxel / Q000031:analogs & derivatives* / Q000494:pharmacology*; D012334:RNA, Neoplasm / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D043823:Taxoids; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D019014:fas Receptor / Q000378:metabolism
D017304:Annexin A5; D000972:Antineoplastic Agents, Phytogenic; D017931:DNA Primers; D016228:Endothelial Growth Factors; D036341:Intercellular Signaling Peptides and Proteins; D008222:Lymphokines; D009363:Neoplasm Proteins; D012334:RNA, Neoplasm; D043823:Taxoids; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D019014:fas Receptor; D000077143:Docetaxel; D017239:Paclitaxel