Overproduction of vascular endothelial growth factor related to von Hippel-Lindau tumor suppressor gene mutations and hypoxia-inducible factor-1 alpha expression in renal cell carcinomas.
PURPOSE The von Hippel-Lindau (VHL) tumor suppressor gene is frequently inactivated in the common type of sporadic clear cell renal cell carcinoma (RCC) as well as RCCs associated with VHL disease. The VHL protein targets hypoxia-inducible factor-1 alpha (HIF-1 alpha), a transcription factor that can induce vascular endothelial growth factor (VEGF) expression, for ubiquitination and degradation. Accumulation of HIF-1 alpha caused by mutant VHL protein in tumor cells may result in VEGF over expression, which has been used to explain the increased vascularity of RCC. However, quantitative analyses of VEGF production and its correlation with VHL mutations and HIF-1 alpha expression in authentic tissues from patients with RCC are lacking.


METHODS
We analyzed VHL gene mutations by direct DNA sequencing and methylation specific polymerase chain reaction in 31 paired RCC tissue samples. HIF-1 alpha protein expression detected by immunoblotting and immunohistochemical staining, and VEGF protein measured by enzyme-linked immunosorbent assay and immunohistochemical staining were performed using tumor and corresponding normal tissues.


RESULTS
VHL gene mutations were detected in 44% of clear cell RCCs but no differences in methylation patterns in the promoter or exon 1 were found. RCCs with VHL gene mutations or of advanced grade produced significantly higher concentrations of VEGF (p <0.0001). HIF-1 alpha protein expression was found in 40% of clear cell RCCs but 80% of them had VHL mutations (p <0.006). HIF-1 alpha expression correlated directly with higher levels of VEGF production (p <0.0001).


CONCLUSIONS
Our findings indicate that VHL gene alterations and HIF-1 alpha protein expression correlate with a significant increase in VEGF production by RCC. In turn it is associated with a more aggressive tumor phenotype.
Na
Xi
X
Department of Urology, University of Rochester Medical Center, New York 14642, USA.
Wu
Guan
G
Ryan
Charlotte K
CK
Schoen
Susan R
SR
di'Santagnese
P Anthony
PA
Messing
Edward M
EM
D000368:Aged; D000369:Aged, 80 and over; D015153:Blotting, Western; D002292:Carcinoma, Renal Cell / Q000235:genetics* / Q000378:metabolism; D004252:DNA Mutational Analysis; D016228:Endothelial Growth Factors / Q000096:biosynthesis* / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D016147:Genes, Tumor Suppressor; D006801:Humans; D007150:Immunohistochemistry; D036341:Intercellular Signaling Peptides and Proteins / Q000096:biosynthesis* / Q000235:genetics; D007680:Kidney Neoplasms / Q000235:genetics* / Q000378:metabolism; D008025:Ligases / Q000235:genetics*; D008222:Lymphokines / Q000096:biosynthesis* / Q000235:genetics; D008297:Male; D008875:Middle Aged; D006899:Mixed Function Oxygenases; D009154:Mutation; D012097:Repressor Proteins / Q000096:biosynthesis* / Q000235:genetics; D014157:Transcription Factors / Q000096:biosynthesis* / Q000235:genetics; D025521:Tumor Suppressor Proteins; D044767:Ubiquitin-Protein Ligases; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D051794:Von Hippel-Lindau Tumor Suppressor Protein
D016228:Endothelial Growth Factors; D036341:Intercellular Signaling Peptides and Proteins; D008222:Lymphokines; D012097:Repressor Proteins; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D006899:Mixed Function Oxygenases; C439790:HIF1AN protein, human; D044767:Ubiquitin-Protein Ligases; D051794:Von Hippel-Lindau Tumor Suppressor Protein; D008025:Ligases; C497446:VHL protein, human