Carcinogenesis vol.23 no.2 pp.257–264, 2002
Cyclin D1 gene polymorphism is associated with an increased risk
of urinary bladder cancer
Lizhong Wang1,*, Tomonori Habuchi1,*,
Takeshi Takahashi3, Kenji Mitsumori1,
Toshiyuki Kamoto3, Yoshiyuki Kakehi3,
Hideaki Kakinuma1, Kazunari Sato1,
Akira Nakamura2, Osamu Ogawa3and
1Department of Urology and2Medical Information Science, Akita
University School of Medicine, Akita 010-8543 and3Department of
Urology, Kyoto University Graduate School of Medicine, Kyoto 606-8507,
4To whom correspondence should be addressed at: Department of Urology,
Akita University School of Medicine, 1-1-1 Hondo, Akita 010-8543, Japan
Cyclin D1 is believed to play an important role in the
genesis and/or progression of transitional cell cancer (TCC)
of the urinary bladder. Cyclin D1 gene (CCND1) mRNA is
alternatively spliced to produce two transcripts, and the
splicing pattern may be modulated by a G to A single
nucleotide polymorphism within the splice donor site of
exon 4. This study was conducted to explore the associa-
tion between the polymorphism and the susceptibility to
and disease status of TCC of the bladder in 222 cases
and 317 native Japanese controls. The relationship
between the CCND1 polymorphism and the mRNA splicing
pattern in TCC cells was evaluated by semi-quantitative
reverse-transcription PCR. The CCND1 A allele was more
frequently observed in the TCC group than the control
group (P ? 0.032) with a significant difference in the
genotype frequency between the two groups (P ? 0.029).
The AA genotype was associated with a significantly higher
risk of TCC compared with the AG?GG genotypes
(adjusted odds ratio (aOR) ? 1.76, 95% confidence interval
more significantly in nonsmoking cases (aOR ? 2.53; 95%
CI ? 1.28–4.51, P ? 0.008). Looking at tumor grade, the
presence of the A allele was associated with higher grade
(? grade 3) tumors with a gene dosage effect (aOR ? 1.77,
CI ? 1.16–2.69, P ? 0.008). In tumor stage, although not
significant, the AA ? AG genotypes tended to be more
frequently observed in cases with T1-4 tumors than those
with Ta tumors (aOR ? 1.94, 95% CI ? 0.98–3.82,
P ? 0.057). The genotype seemed to influence the two
alternatively spliced forms of the CCND1 mRNA because
the ratio of the CCND1 transcript-b/transcript-a was signi-
ficantly higher in cases with the AA genotype compared
Abbreviations: CCND1, the cyclin D1 gene; TCC, transitional cell carcinoma;
RT-PCR, reverse transcription-PCR; GAPDH, glyceraldehyde-3-phosphate
dehydrogenase; OR, odds ratio; aOR, adjusted odds ratio; CI, confidence
*The authors contributed equally to this work.
© Oxford University Press
with those with the AG ? GG genotypes. These data
suggest that the CCND1 variant A allele may be associated
with an increased risk of TCC of the bladder, especially in
men without a history of smoking, and it may also have
an effect on its disease status.
Transitional cell carcinoma (TCC) of the urinary bladder is
the most common cancer of the urinary tract. Recent molecular
genetic studies have shown that accumulation of activated
and progression of TCC (1,2). A frequent target in transitional
cell carcinogenesis is the deregulation of G1-S phase progres-
sion in the cell cycle, whose transition through the G1 into S
phase is regulated by cyclins, cyclin-dependent kinases and
their inhibitors (1,2). Allelic deletions of the chromosomal
region at 9p21 and 13q are frequent in TCCs, and the targets
of such alterations are presumably p16 and Rb (3–8), which
play an important role in the regulation of the G1/S phase
transition (9). Cyclin D1 is an important positive regulator of
the G1/S phase, which has been shown to be involved in
various types of human cancer (10). The cyclin D1 gene
(CCND1) is located at chromosome 11q13, and amplification
of the chromosomal region is frequently detected in TCCs
of the bladder (11,12). Furthermore, CCND1 overexpression
occurs frequently in TCCs of the bladder (13–16) and may be
associated with growth of low-grade papillary tumors (13,14).
These findings suggest that CCND1 is a significant proto-
oncogene in genesis and progression of TCC.
Recently, it has been reported that CCND1 mRNA is
alternatively spliced to produce two transcripts (transcript-a
and transcript-b), which are present simultaneously in a variety
of normal tissues and cancer cells (17–19). The alternative
splicing pattern seems to be modulated by a G to A single
nucleotide polymorphism at codon 241, which corresponds to
a conserved splice donor site of exon 4 (17,18). Studies using
several kinds of cancer cells have demonstrated that the
variant allele encoding A may be a major source of variant
transcript-b (17,18). The CCND1 genotype was significantly
correlated with clinical outcome in patients with non-small
cell cancer of the lung, squamous cell carcinoma of the head
and neck, squamous cell carcinoma of the oral/pharyngeal
cavity, hereditary nonpolyposis colorectal cancer, and epithelial
ovarian cancer, although the proposed risk allele was not
consistent among these cancer types (17,20–23). These data
suggest that the difference in the levels of the alternate CCND1
transcripts caused by the A/G polymorphism may influence
the biological behavior in a variety of cancers.
This study was conducted to explore the association between
the CCND1 polymorphism and the susceptibility to TCC
or its disease status. We further evaluated the association
stratified by patients’ smoking history since epidemiological
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Received May 11, 2001; revised October 18, 2001; accepted October 24, 2001