Although we did not observe an independent correlation for short telomere length and overall breast cancer risk in the current study, we have provided population-based evidence that breast cancer risk may be modified by telomere length among certain subgroups of women. First, the shortest telomere length was significantly associated with increased breast cancer risk in younger or pre-menopausal women (ORs ranged from 1.61 to 1.78, P< 0.03). Second, women with the shortest telomere lengths had moderately increased breast cancer risk when they also had poor antioxidant capacity (lower dietary and supplemental β-carotene, vitamin C or E intake). The ORs ranged from 1.39 to 1.57 (P<0.04). However, using urinary 8-oxodG and 15-F2t-IsoP as oxidative damage markers no influence on telomere length and breast cancer risk was observed. Third, among women exposed to lower carcinogen levels (no detectable PAH-DNA adducts), increased breast cancer risks were observed for carrying shorter telomeres (Q2 and Q4), although no significant trend was observed (P trend =0.05).
Previous epidemiological studies to investigate the correlations between telomere lengths in surrogate tissues (including buccal cells, bladder wash samples45
and peripheral blood cells) and overall cancer risks are quite limited, but still provided some positive evidence. Surrogate tissues frequently serve as a proxy for solid target tissues since they are easily obtained from DNA repositories, and can be conveniently used in large sample epidemiological studies.46,47
Short telomeres have been observed to increase risks for bladder, esophageal, head and neck, lung, lymphoma, and renal cell cancers that may be influenced by cigarette smoking.7–13,16,19
Studies on telomere length and breast cancer risk are few and inconclusive. Levy et al
. first observed an association between shorter telomere length in WBC DNA and increased breast cancer risk.14
Another study found breast cancer risk significantly increased with the longest telomere, and short telomeres were associated with a better prognostic outcome for younger women.19
However, not all data reached a significant level 15,16
, including our pilot study conducted in sister-sets from the New York site on the Breast Cancer Family Registry (BCFR) in which carrying short telomeres was associated with a non-significant increased breast cancer risk (OR=2.1, 95%CI=0.8–5.5).17
Also, no statistically significant difference in telomere lengths was detected between breast cancer cases and unaffected controls in another larger cohort study (P
The discrepant results might be due to different methods (terminal restriction fragment (TRF), southern blot, Q-FISH and Q-PCR) used to measure telomere length in dissimilar studies.15–19
Those approaches with different sensitivity, variability and requirements for DNA quality48,49
could be the cause of heterogeneity in the previous observations. Even using the same Q-PCR method, the range of T/S ratio also showed wide variation (from 4.7 to 76.3 folds)7,17,50
. Whether this is due to different reference DNAs used, the influence of the standard curve (covering the potential range of measured DNA concentrations), or variation of studied populations, needs to be further evaluated. These inconsistent results also suggest that telomere length alone might not be sufficiently powerful as an independent biomarker to determine breast cancer risk. Several biologically related pathways (such as hormone and carcinogen metabolism, DNA damage and repair) could interact with telomere length, and would have stronger influences on breast cancer susceptibility. This is consistent with our current observations of significant differences only in certain subgroups (pre-menopausal women and women with a poor anti-oxidative capacity).
The potential biological explanation for the more pronounced association found between the shortest telomere length and breast cancer risk among pre-menopausal or younger women have been discussed previously.17
The first possible reason is the dramatic difference in estrogen level during pre- and post- menopausal periods. Estrogen has anti-inflammatory and antioxidant activities that may contribute to the dynamic of telomere length.51
Another explanation is that the diversity of genomic instability is relative greater between pre-menopausal cases (with an unstable status) and pre-menopausal controls (usually with a stable status), and can be detected easily. While for post-menopausal women, the genome maybe relatively unstable for both cases and controls because of the influence of either carcinogens or aging itself,10,52,53
and the smaller variety in genomic stability is not easy observed.
8-OxodG and 15-F2t
-IsoP as oxidative damage biomarkers have been associated with several cancers. Our previous study based on a subset of 801 breast cancer cases and controls observed a significantly increased breast cancer risk at higher levels of urinary 15-F2t
-IsoP, and this relationship remained after excluding the effect of radiation therapy.38
It was agreement with the observations that smokers54
and alcohol drinkers55
had higher levels of 15-F2t
-IsoP, a situation of favorable to carcinogenesis. However, the current analysis of the expanded case-control study (1,061 cases and 1,108 controls) found no statistically significant associations between the two biomarkers (15-F2t
-IsoP and 8-OxodG) and breast cancer risk, although the direction of relationship was consistent. Previous studies have indicated that DNA oxidative damage and repair capacity was another important pathway significantly influencing telomere shortening56
and potentially modulating breast cancer risk. Von Zglinicki reviewed 22 independent experiments, and found an increased telomere shortening rate under conditions of increased oxidative stress in 19 reports from seven different laboratories.20
The DNA excision repair proteins (XPC, XPE and ERCC2 etc
.) were found to be up-regulated in cell lines with long telomeres, while integrin, its ligands, and other interacting proteins were down-regulated possibly related to telomere dysfunction.57
Measuring both telomeric-shortening rate and antioxidant capacity (using DCF fluorescence as an indicator of intracellular peroxide levels), Von Zglinicki observed a significant inverse correlation between telomere-shortening rate and anti-oxidative capacity in more than 20 human fibroblast cell lines.58
The role of antioxidant defense in affecting telomere length in cell lines was also confirmed for different tissues of the same donor in two independent studies.56,59
In an in vitro
study, telomere attrition was accelerated by chronic oxidative stress.60,61
The rate of telomere shortening in human vascular endothelial cells could be slowed down by Asc2P, an oxidation-resistant derivative of vitamin C.62
Chondrocytes treated with Asc2P displayed a tendency to protect telomeres from erosion compared with untreated controls.63
An in vivo
study also found a relationship between oxidative stress and telomere shortening in male and female rats of the same age. Female rats exhibited longer telomeres and higher expression of the antioxidant enzyme (MnSOD) than male rats.64
Only one population study confirmed the relationship between higher oxidative stress and telomere shortening.65
The oxidative stress index used was the ratio of urine 15-F2t
-IsoP and vitamin E level. Healthy pre-menopausal women who were the mothers of children with a chronic illness were considered a high oxidative stress group and were found to have a significantly shorter telomeres length (3,110bp) compared with controls (3,660bp) who were age-matched mothers with healthy children.65
The major mechanisms underlying the link between oxidative damage-telomere shortening and breast cancer risk include 1) the direct damage of ROS on telomeric DNA; 2) the telomeric high content of guanines (G triplet) is particularly susceptible to accumulate oxidative stress-induced 8-OxodG; 3) the repair capacity for oxidative lesions is less efficient in telomeres than in the rest of the genome; and 4) a high level estrogen enhanced gene expression of MnSOD, an antioxidant enzyme.20,23,24,26,27,56,66–69
All these data support the theory that telomere length is determined by the balance between oxidative damage and antioxidant defense capacity.56
Our current results indicate that women with a poor anti-oxidative capacity and the shortest telomere length are at significantly increased breast cancer risk, and are the first population evidence to endorse this theory.
The advantages of current study include the population-based study design with a relatively large sample size; the quantitative PCR assay with high through-put (96-well plates used) and high sensitivity (ng quantities of DNA can be analyzed) to measure telomere length; the available oxidative status obtained from both urinary biomarkers and the FFQ; and the extensive information on co-variables allowing us to comprehensively control for potential confounders and evaluate effect modifications in the data analysis. A major limitation is samples were collected after breast cancer diagnosis, which restricts our ability to confirm the etiological temporal sequence between telomere length, oxidative biomarkers and breast cancer risk. This limitation provides another possible explanation for the discrepant results obtained with the two urinary oxidative biomarkers (representing current oxidative damage) and the FFQ data (indicating anti-oxidative capacity prior to cancer diagnosis) on telomere-breast cancer relationship. The former data may be influenced by intake of more fruits, vegetables and antioxidants after cancer diagnoses. Our previous data that did not show an association between fruit and vegetable consumption and 15-F2t-IsoP or 8-oxodG levels supports this possibility.38
Second, we have evaluated available confounders in the analytic models, but we cannot exclude residual confounding caused by unknown factors related to both the dynamics of telomere length and breast cancer risk, including chronic inflammation, telomerase activity, and epigenetic and genetic changes in telomere-related genes.
In summary, our data are the first to suggest that carrying shorter telomeres is associated with a significantly increased breast cancer risk for pre-menopausal women and women with lower dietary and supplemental β-carotene vitamin C or E intake. Additional population data will greatly accelerate our understanding of the role of shortened telomere length, oxidative damage and antioxidants in the development of breast cancer.