The tumor suppressor gene WWOX at FRA16D is involved in pancreatic carcinogenesis.
OBJECTIVE
WWOX (WW domain containing oxidoreductase) is a tumor suppressor gene that maps to the common fragile site FRA16D. We showed previously that WWOX is frequently altered in human lung and esophageal cancers. The purpose of this study was to delineate more precisely the role of WWOX in pancreatic carcinogenesis.


METHODS
We analyzed 15 paired pancreatic adenocarcinoma samples and 9 pancreatic cancer cell lines for WWOX alterations. Colony assay and cell cycle analysis were also performed to evaluate the role of the WWOX as a tumor suppressor gene.


RESULTS
Loss of heterozygosity at the WWOX locus was observed in 4 primary tumors (27%). Methylation analysis showed that site-specific promoter hypermethylation was detected in 2 cell lines (22%) and treatment with the demethylating agent 5-aza-2'-deoxycytidine demonstrated an increase in the expression of WWOX. In addition, 2 primary tumor samples (13%) showed promoter hypermethylation including the position of site-specific methylation. Transcripts missing WWOX exons were detected in 4 cell lines (44%) and in 2 tumor samples (13%). Real-time reverse transcription PCR revealed a significant reduction of WWOX expression in all of the cell lines and in 6 primary tumors (40%). Western blot analysis showed a significant reduction of the WWOX protein in all of the cell lines. Furthermore, transfection with WWOX inhibited colony formation of pancreatic cancer cell lines by triggering apoptosis.


CONCLUSIONS
These results indicate that the WWOX gene may play an important role in pancreatic tumor development.
Kuroki
Tamotsu
T
Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
Yendamuri
Sai
S
Trapasso
Francesco
F
Matsuyama
Ayumi
A
Aqeilan
Rami I
RI
Alder
Hansjuerg
H
Rattan
Shashi
S
Cesari
Rossano
R
Nolli
Maria L
ML
Williams
Noel N
NN
Mori
Masaki
M
Kanematsu
Takashi
T
Croce
Carlo M
CM
D000230:Adenocarcinoma / Q000378:metabolism; D000483:Alleles; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D017209:Apoptosis; D001374:Azacitidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D015153:Blotting, Western; D002453:Cell Cycle; D045744:Cell Line, Tumor; D002469:Cell Separation; D002471:Cell Transformation, Neoplastic; D019175:DNA Methylation; D004252:DNA Mutational Analysis; D018076:DNA, Complementary / Q000378:metabolism; D000077209:Decitabine; D005091:Exons; D005434:Flow Cytometry; D016147:Genes, Tumor Suppressor; D006801:Humans; D019656:Loss of Heterozygosity; D010088:Oxidoreductases / Q000235:genetics / Q000502:physiology*; D010190:Pancreatic Neoplasms / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D025521:Tumor Suppressor Proteins; D000074584:WW Domain-Containing Oxidoreductase
D000964:Antimetabolites, Antineoplastic; D018076:DNA, Complementary; D012333:RNA, Messenger; D025521:Tumor Suppressor Proteins; D000077209:Decitabine; D010088:Oxidoreductases; D000074584:WW Domain-Containing Oxidoreductase; C486071:WWOX protein, human; D001374:Azacitidine