Genetic variation in IL-6 and TNF-α genes with risk of breast cancer in a Saudi population-based case-control study.

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LETTER TO THE EDITOR
Genetic Variation in IL-6 and TNF-a Genes with Risk
of Breast Cancer in a Saudi Population-based
Case–Control Study
To the Editor:
Breast cancer is the second most common cancer in
the world, and the most common cancer among
women which accounts for 22% of all cancers of
women. In Saudi-Arabian population 21% of total
female cancer diagnosis refers to breast cancer (1).
The 2002 annual report of Saudi National Cancer
Registry enlightens the fact that breast cancers devel-
oped before the age of 40 comprise 26.4% of all
female breast cancers compared to 6.5% in the USA
since, young age in women had been reported as an
independent risk factor for breast cancer in the Saudi-
Arabian population (2). Hence, it seems with increase
in longevity, breast cancer is also going to be a very
serious medical challenge in Saudi Arabia.
Circulating IL6 titer has been found increased in
breast cancer patients compared to that in healthy
women. IL-6 up-regulation is associated with invasive-
ness and increased metastatic potential of ER negative
tumors (3). TNF-a was identified for its capacity to
induce necrosis of solid tumors but later it had also
been recognized for its involvement in priming, prolif-
eration, recruitment, function T cell (4). The titer of
TNF-a in body fluid is higher in patients with solid
tumors including breast cancer (5).
In the present study, we analyzed the TNF-a
(rs1800629) -308 A⁄G and IL-6 (rs1554606) 1888G⁄T
polymorphisminassociation
patients of a Saudi Population. To the best of our
knowledge, the association between polymorphisms of
TNF-a (rs1800629) -308 A⁄G and IL-6 (rs1554606)
with breastcancer
1888G⁄T and the development of breast cancer in
Saudi population has not yet been investigated.
A total of 100 breast cancer patients who were
clinically and histopathologically diagnosed at King
Khalid University Hospital, King Saud University from
January 2009 to October 2009 were included. The
study was approved by the Hospital Ethical-Committee
Board, College of Applied Medical Sciences, King
Saud University. Written
obtained from all patients
included in this study. About 5 mL of venous blood
was drawn using sterile heparinized syringe from
healthy individuals as control samples after clinical
checkup and from malignant cases after diagnosis
from the pathology laboratory for genotyping using
standard protocol. Genomic DNA was extracted from
the sample using QIAamp mini blood DNA extraction
kit as per the instructions of the manufacturer. Geno-
typing was carried out using TaqMan assays (Applied
Biosystems, Foster City, CA) according to the manu-
facturer’s instructions. Primers and probes were sup-
plied directly by Applied Biosystems as Assays-by-
Design. All assays were carried out in 96-well plates.
Statistical significance of the genotype frequencies
compared to the expected values was evaluated
according to Hardy–Weinberg rule. The allelic varia-
tions among the cases and controls were analyzed
using chi-squared test, odd’s ratio evaluation at 95%
confidence interval. A p-value less than 0.05 was con-
sidered to be statistically significant.
Genotypic and allelic frequencies of the TNF-a
(rs1800629) A⁄G and IL6 (rs1554606) G⁄T polymor-
phism in patients and controls are presented in
Tables 1 and 2, respectively. Genotype frequencies
were distributed in accordance with the Hardy-
Weinberg equilibrium. For TNF-a (rs1800629) A⁄G
there is no significant difference between patients and
control in allelic distribution (p = 0.404) but geno-
typicdistributiondiffers
informed
and
consent
control
was
subjects
significantly (p = 0.026),
Address correspondence and reprint requests to: Ali A Alshatwi, PhD,
Molecular Cancer Biology Research Laboratory (MCBRL), Department of
Food Science and Nutrition, King Saud University, Riyadh-11451, Saudi
Arabia, or e-mail: alialshatwi@gmail.com.
DOI: 10.1111/j.1524-4741.2012.01260.x
ª 2012 Wiley Periodicals, Inc., 1075-122X/11
The Breast Journal, 2012 1–3

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whereas, IL6 (rs1554606) G⁄T genotypic as well as
allelic distribution differ significantly (p = 0.006 and
p = 0.016, respectively).
In TNF-a (rs1800629) A⁄G, the homozygous GG
genotype was seen same percentage of patients with
breast cancer and (10%) healthy women, GA geno-
type was found to have higher percentage of patients
(87%) in comparison of normal healthy women, while
AA genotype accounted for lower percentage of breast
cancer patients (3%) compared to healthy control
(8%). In IL6 (rs1554606) G⁄T, percentage of GG
genotype in patients is higher (12%) than control
(5%); that of GT genotype is almost same for patients
and control (76% and 77%, respectively), whereas for
TT genotype, percentage is lower in patients (12%)
compared to control (18%). Our data showed that in
TNF-a (rs1800629) A⁄G, only genotype AA⁄GG +
GA (p = 0.007) differs significantly from patients to
that of control with an OR = 2.80 (1.22–6.61), In IL6
(rs1554606)G⁄T,Only
(p = 0.0004) differs significantly from patients to that
of control with an OR = 0.34 (0.18–0.66). TNF-a
(rs1800629) A⁄G recessive model genotype GG versus
GA + AA and genotype AA versus GA of patients dif-
fer significantly (p = 0.006 and p = 0.0005, respec-
tively) from that of control along with OR = 3.57
(1.53–8.13) and 0.35 (0.15–0.80), respectively. IL6
(rs1554606) G⁄T genotypes of patients GG versus
genotypeGG⁄GT + TT
GT, GG versus TT, dominant model GG versus GT +
TT and allelic genotype G versus T differ significantly
fromthatofcontrol
p = 0.0004 and p = 0.016, respectively) along with the
OR = 2.74 (1.42–5.34), OR = 3.87 (1.79–8.46), OR =
2.90 (1.51–5.63), and OR = 1.31 (1.04–1.64; Table 3),
respectively.
Previous studies on TNF-a (rs1800629) A⁄G had
shown no significant association of variants AA, AG,
and GG with the risk of breast and prostate cancer
(6). Boraska et al., (7) had reported from a family
based study on a Croatian population that TNF-a
(rs1800629) A⁄G polymorphism is associated with
risk of diabetes Type-I. Although, our study accounts
for a small population, we found that there is no sig-
nificant difference between the allelic frequencies of
A and G, but the genotypic frequencies of AA is
markedly low (3%) in patients than those in control
(8%) whereas there is borderline difference in geno-
typic frequencies of GA of patients (87%) and control
(82%) for TNF-a (rs1800629) A⁄G. And again the
genotype of recessive model AA versus GA + GG dif-
ferssignificantlybetween
(p = 0.006). Hence,it seems
(p = 0.0009,p = 0.0001,
patientsand
the
control
lossthatof
Table 1. Distribution of TNF-a Genotypes And
Allelic Frequencies of the Study Population
Study
group (%)
TNF-a genotypes
Allele
frequency
GG GAAA TotalGA
Patients
Control
10
10
87
82
3
8
100
100
0.53
0.51
0.46
0.48
v2(2 d.f.) = 7.23, p = 0.026 for genotypes; v2(1 d.f.) = 0.70, p = 0.404 for allelic
frequency. Odds ratio for GA genotype = 0.95 (0.55–1.63), AA genotype = 2.67 (0.99–7.31)
and A allele = 1.10 (0.88–1.37) as an independent risk factor for breast cancer.
Table 2. Distribution of
Allelic Frequencies of the Study Population
IL-6 Genotypes and
Study group
IL-10 genotypesAllele frequency
GGGTTT TotalGT
Patients
Control
1276
77
12
18
100
100
(0.5)
(0.43)
(0.5)
(0.56)5
v2(2 d.f.) = 14.77, p = 0.0006 for genotypes; v2(1 d.f.) = 5.73, p = 0.016 for allelic
frequency. Odds ratio for GT genotype = 2.74 (1.42–5.34), TT genotype = 3.89 (1.87–8.46)
and T allele = 1.31 (1.04–1.64) as an independent risk factor for breast cancer.
Table 3. Odds Ratio with 95% CI of TNF-a and
IL-6 Gene in Breast Cancer Patients
Genotypes Odds ratio (CI 95%)
v2(1 d.f.) p-value
TNF-a
AA ⁄GG + GA
GG ⁄ GA + AA
GA ⁄ GG + AA
IL-6
AA ⁄GG + GA
GG ⁄ GA + AA
GA ⁄ GG + AA
TNF-a
AA versus GG
GG versus AA
AA versus GA
Dominant model
AA versus GA + GG
Recessive model
GG versus GA + AA
Allele
A versus G
IL-6
GG versus GT
GG versus TT
AA versus GA
Dominant model
GG versus GT + TT
Recessive model
TT versus GT + GG
Allele
G versus T
2.80 (1.22–6.61)
1.04 (0.60–1.78)
0.70 (0.44–1.10)
7.18
0.02
2.68
0.007
0.89
0.101
1.55 (0.98–2.45)
0.34 (0.18–0.66)
1.31 (0.88–1.93)
3.90
12.42
1.96
0.048
0.0004
0.161
0.95 (0.55–1.63)
2.67 (0.99–7.31)
0.35 (0.15–0.80)
0.4
4.65
7.50
0.833
0.031
0.006
1 (0,58–1.71)1.711
3.57 (1.53–8.13)11.800.0005
1.10 (0.88–1.37) 0.700.404
2.74 (1.42–5.34)
3.87 (1.79–.46)
0.71 (0.45–1.13)
10.89
14.38
2.36
0.0009
0.0001
0.124
2.90 (1.51–5.63)12.420.0004
0.65 (0.41–1.02) 3.900.048
1.31 (1.04–1.64)5.73 0.016
2 • letter to the editor

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heterozygocity is closely associated with breast cancer
pathogenesis in targeted population.
IL6 has been found that IL6 (rs6949149) was mar-
ginally associated with breast cancer risk (8). We stud-
ied IL6 (rs1554606) G⁄T SNP and found that,
genotypic frequencies of GG is more (12%) among
patients compared to those in control (5%) and that
for genotype TT is less (12%) as compared to control
(18%) as well as allelic frequency of G is more (0.5)
in patients in comparison to control (0.43) and that of
T is less (0.5) as compared to control (0.56). Further-
more, genotype dominant model GG versus GT + TT
and allelic genotype G versus T both differ signifi-
cantly from patients to control (p = 0.0004 and
p = 0.016; Table 3). Thus, it can be concluded as the
loss of heterozygocity (GT) since both TT and GG are
associated to the breast cancer pathogenesis. If the
allele G of heterozygote GT transform as allele T, the
risk of disease is less whereas if allele T of heterozy-
gote reversed as G, the risk of breast cancer is high.
In summary, our study suggests that Genotype AA
for TNF-a (rs1800629) A⁄G and both GG as well as
TT genotype for IL6 (rs1554606) G⁄T may be risk
factors due to loss of heterozygosity (LOH) in case of
breast cancer in a Saudi population. Since our sample
size is small, so before certifying these findings for the
use of prognostic and diagnostic marker, additional
investigations are warranted with metapopulation.
Ali A. Alshatwi, PhD*
Tarique N. Hasan, MS*
Gowhar Shafi, MS*
Adulaziz. A. Alsaif, MD?
Abdulrahman A. Aldiab, MD?
Mohammed A. Alsaif, PhD?
Amal A. Al-Hazzani, PhD§
and Kai. Y. Lei, PhD–
*Molecular Cancer Biology Laboratory,
Department of Food Science and Nutrition,
College of Food and Agricultural Sciences,
King Saud University, Riyadh, Saudi Arabia;
?Department of Surgery, College of Medicine, King
Saud University, Riyadh, Saudi Arabia;
?Department of Community Health Sciences,
College of Applied Medical Sciences,
King Saud University, Riyadh, Saudi Arabia;
§Department of Botany and Microbiology,
King Saud University, Riyadh, Saudi Arabia;
and–Department of Nutrition and Food Science,
University of Maryland, College park, Maryland
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