Methylation of the gamma-catenin gene is associated with poor prognosis of renal cell carcinoma.
OBJECTIVE
Gamma-catenin is a cell adhesion protein, and its functional loss is associated with tumor invasion and metastasis. We hypothesize that (1) promoter CpG methylation regulates the expression and function of the gamma-catenin gene in renal cell carcinoma (RCC) and (2) methylation of the gamma-catenin gene is associated with poor prognosis of RCC. To test these hypotheses, we analyzed the CpG methylation status of the gamma-catenin gene and its correlation with clinical outcome in RCC.


METHODS
Genomic DNA and total RNA were extracted from three renal cancer cell lines (A498, Caki-1, and Caki-2) and 54 RCC tissue samples with their corresponding normal kidney tissue samples. Expression of gamma-catenin gene was analyzed by reverse transcription-PCR and immunostaining. Promoter methylation was analyzed by two different methylation-specific PCR (MSP-A and MSP-B), and the results were verified by DNA sequencing.


RESULTS
The demethylating agent (5-aza-2'-deoxycytidine) increased levels of mRNA transcript of the gamma-catenin gene in three renal cancer cell lines. Gamma-catenin mRNA and protein expression were significantly reduced in RCC samples compared with normal kidney samples, respectively (P < 0.05). MSP-A and MSP-B bands were detected in 45 of 54 (83.3%) and 49 of 54 (90.7%) RCC samples, respectively. In normal kidney, weak products of MSP-A and MSP-B were detected in 5 of 54 (9.3%) and 6 of 54 (11.1%) samples, respectively. Likewise, both MSP-A and MSP-B ratios were significantly higher in RCC samples compared with normal kidney samples, respectively (P < 0.01). Multivariate analysis revealed that the MSP-B ratio was a powerful and independent predictor superior to nuclear grade and Robson stage with respect to survival and disease progression (P = 0.029 and 0.0071, respectively). No mutations in the NH(2)-terminal region of gamma-catenin were found in this study.


CONCLUSIONS
Expression of gamma-catenin is regulated by promoter CpG methylation, and the balance between methylated and unmethylated RCC cell populations could determine its functional role. Because the conventional nuclear grade and/or staging system have some limitations to predict precise clinical outcome, this is the first report demonstrating that promoter CpG methylation of gamma-catenin can be an independent and superior predictor for survival and disease progression.
Breault
Julia E
JE
Department of Urology, Veterans Affairs Medical Center and University of California at San Francisco, 4150 Clement Street, San Francisco, CA 94121, USA.
Shiina
Hiroaki
H
Igawa
Mikio
M
Ribeiro-Filho
Leopoldo A
LA
Deguchi
Masao
M
Enokida
Hideki
H
Urakami
Shinji
S
Terashima
Masaharu
M
Nakagawa
Masayuki
M
Kane
Christopher J
CJ
Carroll
Peter R
PR
Dahiya
Rajvir
R
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001374:Azacitidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D002292:Carcinoma, Renal Cell / Q000235:genetics* / Q000378:metabolism; D018899:CpG Islands / Q000235:genetics*; D003598:Cytoskeletal Proteins / Q000235:genetics*; D019175:DNA Methylation; D004273:DNA, Neoplasm / Q000032:analysis; D000077209:Decitabine; D051180:Desmoplakins; D018450:Disease Progression; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D006801:Humans; D007668:Kidney / Q000378:metabolism / Q000473:pathology; D007680:Kidney Neoplasms / Q000235:genetics* / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D009154:Mutation / Q000235:genetics; D011379:Prognosis; D011401:Promoter Regions, Genetic / Q000235:genetics*; D015996:Survival Rate; D051185:gamma Catenin
D003598:Cytoskeletal Proteins; D004273:DNA, Neoplasm; D051180:Desmoplakins; D004791:Enzyme Inhibitors; C495292:JUP protein, human; D051185:gamma Catenin; D000077209:Decitabine; D001374:Azacitidine